7.14.22 DS Eligible Parent Grants Publish List

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Cheng,Guang-Shing

Longitudinal Impact of Respiratory Viruses on Bronchiolitis Obliterans Syndrome in Allogeneic Hematopoietic Cell Transplant Recipients

Bronchiolitis obliterans syndrome after allogeneic hematopoietic cell transplant is a highly morbid manifestation
of chronic graft-versus-host disease which is poorly understood, hampering our ability to intervene effectively.
We hypothesize that cumulative burden of symptomatic and asymptomatic respiratory viral infections contributes
to the development of BOS and progressive lung dysfunction, leading to poor outcomes. In this proposal we will
conduct a multicenter longitudinal prospective study on the association of respiratory viral infection and lung
function trajectory in patients at risk for and with a clinical diagnosis of BOS, by utilizing home spirometry, self-
collection of viral PCR, and a novel serosurvey (VirScan), to fully characterize the natural history of BOS.

Bronchiolitis obliterans syndrome (BOS) is the most severe manifestation of chronic graft-versus-host disease
(cGVHD) in survivors of allogeneic hematopoietic cell transplant (alloHCT), leading to irreversible pulmonary
impairment, poor quality of life, and 5-year survival of 40%. Fundamental gaps in knowledge of the pathogenic
events that contribute to progressive lung dysfunction in BOS have not been well characterized, hampering our
ability to intervene effectively. Our preliminary data suggest that respiratory viruses, including respiratory
syncytial virus (RSV), parainfluenza (PIV), human metapneumovirus (HMPV), and influenza (FLU), are
independent risk factors for the development of BOS. Additionally, we show that asymptomatic respiratory viral
infections (RVI) are common posttransplant. We have shown that mobile wireless home spirometry is feasible
in patients with cGVHD and can enable early diagnosis and a granular understanding of the trajectory of lung
function decline. Our overarching hypothesis is that cumulative respiratory viral exposure leads to the
development of BOS and poor outcomes in the context of alloimmunity. The overall aim of this proposal is
to establish the temporal relationship between RVI along the continuum of disease presentations, from
asymptomatic to symptomatic upper respiratory tract to lower tract disease, and the lung function trajectory of
BOS. We propose to conduct a multicenter prospective longitudinal study of the natural history of RVI and lung
function with an innovative home monitoring approach that overcomes the barriers to understanding clinical
events that lead to BOS and severe BOS phenotypes. Aim 1 investigates the role of RVI as triggers BOS. We
will enroll alloHCT recipients at risk for BOS (Cohort 1, n=200), including those with a diagnosis of cGVHD or a
history of high-risk RVI (RSV/PIV/HMPV/Flu/SARS-CoV2). Patient will perform weekly home spirometry and
protocolized surveillance and symptom-prompted self-collected nasal swab viral PCR. In addition, serum will be
collected quarterly via a needle-less home blood collection kit and assayed with VirScan, a novel comprehensive
serosurvey that detects epitopes of >1000 virus strains, in order to assess the impact of cumulative respiratory
viral burden on BOS outcomes. Aim 2 examines the role of RVI on pulmonary exacerbations in BOS, as well as
the association of cumulative RVI exposure (as determined by VirScan) on accelerated FEV1 decline in patients
with a severe BOS phenotype. Patients with a clinical diagnosis of BOS (Cohort 2, n=80), will perform the same
procedures as Cohort 1. For both aims, viral PCR and VirsScan results will be compared and analyzed as
predictors for BOS development or accelerated FEV1 decline. The critical data generated by this study will
improve recognition of early BOS in the context of RVI, risk stratify patients at highest risk for intensive
monitoring, and identify tangible endpoints and biologic rationale for testing early interventions and novel
therapies. Importantly, this proposal will also establish a unique adult and pediatric multicenter Consortium with
the specific goal of addressing lung disease in HCT recipients, an area of significant and urgent unmet need.

NHLBI

11/30/26

R01 HL161037

FHCRC

Peters,Ulrike

Interdisciplinary Epidemiologic Consortium to Investigate T-cell Response in Colorectal Cancer

In this large, collaborative effort, we will examine how germline genetics, lifestyle factors, and tumor
characteristics relate to the profile of T cells in colorectal cancers, and how those T cell profiles relate to cancer
survival. The results of this work may ultimately inform strategies to harness the immune system for the
prevention and treatment of colorectal cancer.

The immune system has pivotal influence in the evolution and progression of many tumor types, including
colorectal cancer (CRC). In particular, the presence of a strong T cell response in CRC, indicating activation of
the adaptive immune system, has been associated with better patient outcomes. As such, recently developed
immunotherapeutic approaches often attempt to harness the adaptive immune response. Immune cells are an
integral component of the tumor microenvironment, and dynamically interact with neoplastic cells. However,
our understanding as to the complexity of the T cell response and the factors that drive this response remains
limited. The objective of this proposal is to identify genetic, lifestyle, and tumor factors associated with the T
cell response in CRC, and to characterize the survival implications of that response. Specifically, in Aim 1 we
will examine the relationship of personal characteristics with T cell response in CRC, including the role of (1a)
germline genetic variation within human leukocyte antigen (HLA) and killer-cell immunoglobulin-like receptor
(KIR) genes, and (1b) lifestyle factors (e.g., aspirin use, smoking, alcohol consumption). In Aim 2 we will focus
on several colorectal tumor characteristics as they relate to T cell response, including (2a) the presence of
Fusobacterium nucleatum and bacterial toxin genes in CRC, and (2b) somatic mutations in key signaling
pathways (e.g., WNT signaling and RAS/RAF). In Aim 3, we will evaluate the associations of different aspects
of T cell response with CRC survival, accounting for known prognostic factors and the relationships identified in
Aims 1-2. To achieve these Aims, we propose to assess the density and spatial distribution of specific T cell
subsets using multiplexed immunofluorescence (mIF) to quantify expression levels and co-expression patterns
of CD3, CD4, CD8, CD45RO, and FOXP3 at the single cell level. This multiplexed assessment will allow us to
examine the epidemiologic and prognostic relevance of numerous metrics of T cell response in CRC. This
research will leverage the resources of the Genetics and Epidemiology of Colorectal Cancer Consortium
(GECCO) and the Colon Cancer Family Registry (CCFR). GECCO-CCFR is a large collaborative effort
between observational studies of CRC. We have completed genome-wide germline genotyping and have
harmonized epidemiologic data regarding a variety of lifestyle factors and personal characteristics for all
participating studies. We are conducting DNA sequencing with a panel of 205 human genes and a small
number of bacterial genes in CRC tumor tissue. Prospective follow-up for survival is ongoing, and we have
harmonized existing survival data. Through this project, we will add information on T cell response in CRC for
>2,500 CRC cases to the GECCO-CCFR resource. This project provides an unprecedented opportunity to
investigate the epidemiology of the T cell response in CRC and the relationship of that response with personal
and tumor characteristics. Insights gained through this novel study could ultimately inform the development
and targeted implementation of emerging immunotherapeutic and immunopreventative strategies.

NCI

04/30/26

R01 CA248857

FHCRC

Smith,Gerald R

Molecular analysis of genetic recombination and DNA break repair

Program Narrative (Relevance)
The DNA of our cells is frequently broken and must be faithfully repaired for life to continue. Failure of faithful
DNA break-repair by genetic recombination leads to birth defects, infertility, and cancer. Inhibitors of DNA
break-repair in bacteria can be developed into novel, sorely needed antibiotics to combat the ever-threatening
emergence of drug-resistant bacteria.

The long-term goal of the research proposed here is to determine the molecular mechanism of homologous
genetic recombination and DNA break repair. This objective is approached by a combination of genetic
analysis of mutants and biochemical analysis of proteins and DNA from cells. This research uses the fission
yeast Schizosaccharomyces pombe as well as the bacterium Escherichia coli and its phage lambda. All are
widely studied, highly tractable model organisms with features common to all organisms, including humans.
The studies are focused on meiotic recombination in S. pombe, whose high rates of recombination facilitate
both genetic and biochemical analyses, and on the major pathway of recombination and DNA break repair in
bacteria, promoted by RecBCD enzyme, a complex DNA repair machine, whose 3D structure allows us to
determine at atomic level how recombination initiation is regulated.
Building on past achievements, the research is currently focused on the following areas. 1) Studying
how meiotic DNA double-strand break (DSB) hotspots form clusters, and how these clusters impart DSB
interference and, consequently, crossover interference important for proper chromosome segregation. This
research promises to solve the 100-year-old problem of crossover interference, a major genetic puzzle for
which we have proposed a molecular mechanism and supported with many data. 2) Studying how RecBCD
enzyme controls its potentially rampant nuclease activity and appropriately activates it by interaction with Chi
hotspots of recombination (5’ GCTGGTGG 3’). This research promises to solve at near-atomic level the
molecular mechanism of RecBCD enzyme, the principal controller of the major pathway of E. coli
recombination, first observed 75 years ago, and a paradigm for chromosomal site control of other complex
DNA enzymes. 3) Seeking more potent RecBCD inhibitors, which are promising antibiotics against a novel
(unused) target. New antibiotics are needed to counter ever-more-frequent drug-resistant bacteria. These
goals will be attacked by a combination of genetic analysis of mutants, fluorescence microscopy of intracellular
proteins and chromosomal sites, physical analysis of DNA intermediates from meiotic cells, and enzymatic and
biophysical analyses of isolated proteins. The results of these studies will elucidate the molecular mechanism
of recombination and DNA break repair as well as the controls on recombination that ensure that it occurs at
the proper time and place along chromosomes.
Recombination is important for faithful meiotic chromosome segregation, error-free repair of frequently
arising DNA double-strand breaks, and generation of cellular and organismal diversity. Aberrancies of
recombination can generate chromosomal rearrangements, such as translocations, duplications, and deletions,
which are often associated with or the cause of infertility, birth defects, and cancers.

NIGMS

04/30/26

R35 GM118120

FHCRC

Berger,Alice H

Understanding and exploiting novel therapeutic vulnerabilities of RIT1-driven lung cancer

Despite recent advances, lung cancer five-year survival rates remain dismal. One barrier to progress is the
identification of new treatment strategies for specific genotype-defined subsets of lung cancer. In this work, we
characterize the mechanism of a little-studied new cancer oncogene, RIT1, and test two new therapeutic
strategies for the treatment of RIT1-mutant lung cancer.

Recent advances in targeted therapies have revolutionized lung cancer clinical practice. Lung adenocarcinomas
harbor frequent mutations/amplifications/fusions in receptor tyrosine kinase (RTK) and RAS pathway
oncogenes, many of which can be targeted by FDA-approved therapies. However, the majority of patients do
not have targeted treatment options. Our previous work identified somatic RIT1 mutations in lung
adenocarcinomas and discovered that RIT1 variants act as gain-of-function mutations to promote cellular
transformation and drug resistance. RIT1 amplification and overexpression may play a similar pathogenic role.
RIT1 mutations also are found in myeloid leukemias and in the germline of individuals with Noonan Syndrome.
In all diseases, mutations in RIT1 are mutually exclusive with other RAS-pathway mutations, implicating RIT1 as
a RAS-pathway driver gene. However, our recent preliminary data show that RIT1 and KRAS substantially differ
in the downstream effectors needed to promote tumorigenesis. Further understanding the cellular consequences
of RIT1 mutations will open up new strategies for treatment of RIT1-mutant cancers.
In this proposal, we define the mechanism of action of RIT1 mutations in lung cancer and test the efficacy of two
new treatment strategies. Building on our preliminary studies that constitute the first global profiling of RIT1
function, we now will: (1) Identify the mechanism of RIT1-YAP1 synergy in lung cancer, (2) Determine how a
USP9X-RIT1 axis regulates the spindle assembly checkpoint and sensitivity to anti-mitotic therapies, and (3)
Define the therapeutic potential of anti-YAP1/TEAD and anti-mitotic therapies in RIT1-mutant lung cancer.
Ultimately, this work will advance our understanding of the role and mechanism of RIT1 mutations in cancer and
contribute the rationale and pre-clinical data needed to translate these findings into new clinical trials. Our access
to novel patient-derived and genetically-engineered mouse models, coupled with our expertise in both functional
genomics and pre-clinical studies, make our laboratory uniquely well-suited to discover new therapeutic options
and improve outcomes for patients with RIT1-mutant cancers.

NCI

03/31/26

R37 CA252050

FHCRC

Ghajar,Cyrus M

Chemotherapy-driven evolution of the vascular secretome and its role in therapeutic resistance

Our continued inability to develop selective therapies that prevent the emergence of disseminated breast tumor
cells into lethal metastases necessitates new approaches. We have shown that disseminated tumor cells
(DTCs) occupy the perivascular niche (PVN), that this niche promotes chemotherapeutic resistance, and that
targeting interactions between DTCs and the PVN sensitizes DTCs to chemotherapy. Translating this
approach to pre-clinical models of breast cancer prevented metastases in over 60% of at-risk mice.
Questioning the mechanism(s) of resistance in the remainder of mice led us to consider the dynamic response
of the vascular niche to chemotherapy. Our preliminary data indicate that genotoxic agents trigger the
secretion of pro-survival factors into the PVN, and that the major trigger of this chemotherapy-associated
vascular secretome is the DNA damage response (DDR). Therefore, here our goal is to target the cause (i.e.,
the DDR) and the consequence (i.e., the chemotherapy-associated vascular secretome) of vascular evolution,
which we suspect promotes therapeutic resistance of DTCs. We will pursue this goal through 2 specific aims:
Specific Aim 1. To determine whether individual extracellular factors elicited from vascular endothelium by
chemotherapy protect DTCs. We have defined a global vascular response to DNA damaging agents. Netrin-1
is a prominent component of this secretome with documented pro-survival functions. Our data show that endo-
thelial-derived Netrin-1 protects DTCs from chemotherapy. Using organotypic, transgenic and preclinical mod-
els, we will determine: i) whether endothelial cells are the relevant source of chemotherapy-elicited Netrin-1 in
vivo; ii) whether targeting Netrin-1 as an adjuvant sensitizes DTCs to chemotherapy; iii) whether this syner-
gizes with targeting pre-existing protective factors in the PVN; and iv) the safety of these approaches.
Specific Aim 2. To identify DNA damage driven signaling pathways in vascular endothelium that induce the
chemotherapy-associated vascular secretome. Our preliminary data demonstrate that genotoxic therapy elicits
a stereotypic DDR from quiescent endothelium, and that targeting a nucleator of this response mutes the ma-
jority of the chemotherapy-associated vascular secretome. This approach may be a more robust alternative to
targeting a single pro-survival factor such as Netrin-1. Here, we will identify and target the signaling pathway
that connects DDR to the vascular secretome, and measure the efficacy and toxicity of this approach.
The significance and innovation of this work lie in the discovery of origins and outputs of chemoprotective
factors in the DTC niche. Targeting both will result in the first approach to specifically target DTCs, impacting
breast cancer survival in a positive and lasting fashion.

NCI

12/31/25

R01 CA249528

FHCRC

Tsukiyama,Toshio

Molecular mechanisms and functions of global chromatin control

Proper chromatin regulation is essential for all DNA-dependent processes. How
chromatin can be regulated in a genome-wide scale is not well understood. We will use
yeast quiescence as a model to elucidate molecular mechanisms and functions of
global chromatin reprogramming.

The long term goal of the proposed study is to determine, at the molecular level, mechanisms and
functions of chromatin regulation at a global level. Chromatin regulation profoundly affects a wide
variety of DNA-dependent processes, including transcription, DNA replication, recombination, DNA
repair, and DNA damage response. Therefore, elucidating the mechanisms of chromatin regulation is a
necessary prerequisite for understanding how these essential processes are controlled. One of the
major challenges the chromatin field is to elucidate how chromatin is globally reprogrammed during
processes like cell fate determination, development and cell-cycle control. This is a particularly
important challenge, because it was recently determined that mutations in chromatin regulators
represent one major class of so called cancer driver mutations, yet how these mutations drive cancer
remains unknown. Therefore, elucidating the mechanisms of chromatin regulation impacts not only the
researchers who study fundamental principles of DNA-dependent processes, but also cancer biologists.
We have previously elucidated how chromatin regulation affects transcription, DNA replication,
S phase checkpoint and recombination using budding yeast as a model organism. Like most studies in
the field, we did our work during the mitotic cell-cycle. However, yeast cells in the wild, like other
eukaryotic cells, spend most of their time in quiescence. Quiescence is associated with massive
chromatin reprogramming for global condensation. Because the vast majority of work on chromatin
regulation has been done during mitotic cell-cycle, we have little idea of how chromatin is regulated
during the time cells spend most of their time. In order to understand the whole picture of chromatin
regulation in vivo, it is essential to understand mechanisms and functions of chromatin regulation during
quiescence. In the next funding period, we will ask the following questions in quiescent state: 1) How is
chromatin globally reprogrammed by ATP-dependent chromatin remodeling factors? 2) How are
chromatin domains and nucleosome array folding regulated? 3) How is gene expression regulated
post-transcriptionally at a global scale? We will use the combination of genomics, molecular genetics,
EM, modeling and biochemistry to identify novel mechanisms by which highly conserved chromatin
regulators function to massively reprogram chromatin in a genome-wide scale. In the long run, these
studies will allow us to compare and integrate the principles of chromatin regulation throughout the
mitotic cell-cycle and quiescence, such that we can obtain the full picture of chromatin regulation.

NIGMS

12/31/25

R35 GM139429

FHCRC

Paddison,Patrick J

Defining cellular states of quiescence in human brain tumors

Solid tumor recurrence is likely driven by slow dividing or quiescent tumor subpopulations in the primary tumor
which survive treatment regimens and give rise to secondary tumors. However, we currently lack a basic
understanding of molecular features of these populations and, as a result, do not have effective therapeutic
strategies to neutralize them. The experiments described in this proposal are directly relevant to public health
because they will identify and dissect molecular vulnerabilities that can eliminate tumor subpopulations resistant
to conventional therapies.

The most aggressive type of brain tumor, grade IV glioma known as glioblastoma (GBM), is one of the few tumor
types with both a poor outcome and minimal improvement in survival in the past decades. For GBM, like other
solid cancers, intratumoral heterogeneity is likely an important factor in mediating therapeutic response. In
particular, quiescent, G0-like subpopulations may engender tumors with more robust responses to treatment
regimens and allow for tumor regrowth after standard of care (SOC). However, G0-like tumor populations are
currently ill-defined, even after application of single cell genomics to GBM. Our failure to fully comprehend and
experimentally model quiescent/G0-like states represents a critical knowledge gap, but also a key opportunity,
for glioma and other cancers, as neutralizing G0 cells could effectively prevent chemoradiotherapy resistance
and tumor recurrence.
The purpose of this grant is to provide a functional and molecular definition of G0-like states in GBM tumors and
their responses to SOC. In Aim 1, we will define molecular networks governing long- and short-term quiescent
states in GBM patient tumors using a novel G0 reporter system in combination with single cell genomic analysis.
In Aim 2, we will test the hypothesis that dormant G0 GBM cells have unique RNA and chromatin signatures
required for SOC survival and tumor regrowth. In Aim 3, we will study and nominate the NuA4/KAT5 lysine
acetyltransferase complex as a key regulator of G0-like states in GBM and candidate therapeutic target.
The Aims are built on strong preliminary data, including: the creation of a machine learning-based method for
identifying G0-like cells in gliomas, integrated analysis of single cell RNA and chromatin analysis of primary and
PDX GBM tumors with standard of care, a functional genomic screen to identify regulators of GBM G0, and key
experimental models to functionally dissect G0 states in GBM tumor models.
If successful, this grant will produce a new working model for GBM G0-like states, provide key genes and gene
networks associated with G0, and analysis tools for identifying G0-like states in clinical samples. It will also
define how these populations respond to SOC and shift tumor dynamics during recurrence. Finally, it will provide
data for a new therapeutic strategy, "downgrading", where grade IV tumors are made less aggressive by
triggering extended or permanent G0-like states in tumor cells.

NINDS

11/30/25

R01 NS119650

FHCRC

Eisenman,Robert N

The MYC Transcription Factor Network and the Path to Cancer

The Myc protein is an essential regulator of growth in all normal cells, but in many cancers Myc is escapes
normal controls and supports high rates of growth needed for tumor progression. In this proposal we will
determine what events contribute to activation of Myc in tumors and devise approaches to inhibit Myc function.
This research is relevant to public health because it will ultimately lead a better understanding of the causes for
cancer and provide new approaches to cancer therapy.

Myc proteins are essential for normal cellular growth and proliferation. However, when its normal regulation
is compromised (i.e. deregulated) Myc promotes initiation and progression of a broad spectrum of human
cancers. Myc has been long known to be a transcription factor that heterodimerizes with the Max protein in
order to specifically recognize DNA. When deregulated, Myc-Max alters gene expression programs resulting
in metabolic and growth related changes that in turn support tumor progression. Recent studies show Myc-
Max does not function alone, but is part of a larger transcriptional “network” of related, yet functionally dis-
tinct, factors that heterodimerize with either Max or the Max-like protein MLX, or both. In order to understand
and control Myc's role in the etiology of cancer it will be essential to define how Myc both depends on and
influences the extended network. This application builds on 3 broad aspects of our ongoing studies:
Transcriptional reprogramming of metabolism: We had earlier uncovered a critical role for Mlx, and its hetero-
dimeric partner MondoA, in the metabolism and survival of several Myc-driven tumors. Focusing on pancre-
atic adenocarcinoma we will examine cross-talk and functional dependencies involving Myc in the context of
its extended network that may be exploited to identify new therapeutic strategies. Moreover, Myc and the
other network proteins are transcription factors and we will determine their shared target genes and their co-
operative effects on chromatin modifications and higher order structure as well as gene expression.
Tumor suppression mediated by Mga, a member of the Myc Network: Mga is a large and unusual transcrip-
tion factor with two distinct DNA binding domains, one of which dimerizes with Max, binds DNA, and is fre-
quently subject to deletion or mutation in a wide range of neoplasms. However, little is known about Mga's
oncogenic functions. Our very recent findings that Mga loss of function results in altered cell motility in vitro,
and rapid lung adenocarcinoma formation in mice provide a biological system to elucidate Mga's capacity to
suppress cancer. We will define regions in Mga essential for DNA binding, identify transcriptional complexes
associated with Mga, and assess how loss of Mga leads to tumor initiation, progression and metastasis.
Molecular alterations driving Myc oncogenicity: we introduced a point mutation (T58A), associated with B cell
lymphomas and AML, within the phosphodegron of the endogenous murine myc gene. In these mice, Myc-
T58A is regulated normally with no overt changes in tissue growth or proliferation. Yet we find that myc-T58A
mice display increased hematopoietic progenitor cell self-renewal and resistance to apoptosis, and develop
long-latency AML or lymphoma. Our data show that the Myc-T58A mutation alters the association of Myc
with a specific co-regulatory complex. We hypothesize that this altered binding modifies expression of a sub-
population of Myc target genes during hematopoiesis, resulting in production of tumor initiating cells. We plan
to elucidate the underlying molecular basis for the T58A phenotype in these tumor-prone mice.

NCI

08/31/25

R35 CA231989

FHCRC

Shen,Megan

Planning Advance Care Together (PACT) to Improve Engagement in Advance Care Planning among Advanced Cancer Patients

The proposed research is consistent with broader public health goals focused on improving patient end-of-life
care and the College of Surgeons Commission on Cancer and Centers for Medicare and Medicaid Services
(CMS) call for increased patient engagement in advance care planning (ACP) as part of standard care. The
proposed study will address this public health issue by developing and pilot testing a mobile application
(Planning Advance Care Together, PACT) designed to improve advanced cancer patients’ levels of
engagement in ACP, documented ACP conversations with doctors and family members, completion of
advance directives, and receipt of goal-concordant care by integrating family members/loved ones (i.e.,
“caregivers”) and providers into the ACP decision-making process. Therefore, this study is aligned with the
NCI’s long-term goals of improving quality of care at the end of life and the CMS’s goal of improving
engagement in ACP.

Engagement in advance care planning (ACP), which includes having end-of-life (EoL) conversations and
completing advance directives (ADs) (e.g., living will, health care proxy), has been shown to improve the
quality of care and reduce suffering at the end of life. However, less than half of advanced cancer patients
engage in ACP or complete ADs. One commonly overlooked barrier to ACP engagement and AD completion is
a lack of acknowledgment that the majority of cancer patients report ACP as a social process in which they
want to engage multiple loved ones and family members in their decision-making process. Dr. Shen’s work, in
collaboration with her Co-Investigator Dr. Prigerson, suggests that patients frequently report involvement of
family members as a critical concern and need for engaging in ACP and that 70.7% report their EoL care
treatment preferences being shaped around concerns about their family members. Based on this pilot work
and prior work indicating a patient preference to engage in ACP as a social process, this study aims to develop
and pilot test a mobile application [Planning Advance Care Together (PACT)] designed to improve advanced
cancer patients’ and caregivers’ engagement in patients’ ACP, presence of ACP discussions, completion of
ADs, and receipt of goal-concordant care. The goals of this study are to: (1) refine and field-test a mobile
application intervention (PACT) using an iterative design approach, “Think Aloud” exercises, and usability
protocols; (2) evaluate the feasibility, acceptability, usability, satisfaction, and user engagement of the
intervention among advanced cancer patients and their caregivers; (3) test the preliminary efficacy of the
intervention on patients’ and caregivers’ level of engagement in ACP, documented ACP conversations, and
patients’ completion of ADs (primary outcomes); and patients’ and caregivers’ perceived social support and
family functioning as well as patients’ receipt of goal-concordant care (secondary outcomes); and (4) evaluate
process measures in a post-intervention interview. To meet these goals, we will collect feedback from
advanced cancer patients (n=10), caregivers (n=10), and providers (n=10) to improve and refine the
intervention prototype. Next, we will conduct iterative field-testing with two sets of patient-caregiver dyads
(n=15 per group) to refine the mobile application. Then, we will randomize n=100 patient-caregiver dyads to
the intervention group and n=100 dyads to the control group (usual care) and assess outcomes at baseline
and 3 and 6 months post-intervention to determine the feasibility, acceptability, usability, satisfaction, user
engagement, and preliminary efficacy of the intervention. Finally, we will conduct a brief post-intervention
interview with a subset of patients (n=15) and caregivers (n=15) to understand processes of using the PACT
application. Grounded in established theories of decision-making science, the proposed project takes the novel
approach of utilizing mobile health technology to integrate loved ones into patients’ ACP decision-making.

NCI

08/31/25

R37 CA246703

FHCRC

Houghton,A McGarry

Liquid biopsy of the lung to profile lung cancer

Correlative measures of clinical tumor behavior for targeted, immune and cell based therapies in lung cancer
are required for optimal treatment plans. A liquid biopsy of the lung using bronchoalveolar lavage (BAL) can
augment tumor and blood immunogenomic profiling but pre-analytic variables will alter results. We propose
experiments herein to standardize collection and processing of BAL for the immunogenomic profiling of lung
cancer.

A tumor biopsy is traditionally performed for lung cancer diagnosis using either bronchoscopy through the airway
or a needle aspiration through the chest wall. Advances in targeted and immune therapies now often require
more tissue for molecular and immune profiling to optimally manage lung cancer. The yield for cancer diagnosis
using modern bronchoscopic tools approaches only 50% across the spectrum of lung tumors biopsied, and the
additional requirement for molecular and immune profiling erodes this yield further. This leads to delayed and
suboptimal care, increased healthcare costs, and increased patient morbidity since patients can often require
multiple procedures to obtain the correct information for treatment. Our group has recently demonstrated that
targeted Bronchoalveolar Lavage (BAL) – or a washing of the lung cancer performed during a procedure – is a
reservoir of genomic and cellular biomarkers in the lung tumor macroenvironment (TMaE). Further, we have
demonstrated that molecular analyses of BAL from the lung TMaE recapitulate cancer biology in the lung tumor
microenvironment (TMiE). While BAL is very safe and routinely performed during bronchoscopy, to date, it is a
pauci-cellular fluid that is of limited clinical utility for cancer diagnosis. Beyond cytology that is low yield, there
are no molecular or cellular assays that are used in the clinic to fully inform providers who treat lung cancer.
Because of this, rigorous attention to how methods of collection, patient host factors and processing of BAL will
alter genomic and high dimensional cell based assays is lacking. Our central hypothesis is: BAL globally
samples the tumor microenvironment (TMiE) to overcome limitations of tumor heterogeneity and is more
sensitive than blood for immunogenomic profiling due to increased quantities of tumor specific
biomarkers. To realize our goal and prove our hypothesis, in depth analysis of the conditions affecting BAL for
high dimensional genome and cell assays is required. Here, we propose studying how basic conditions in the
lung, variations in acquisition of BAL, and storage and processing of BAL affect its utility for comprehensive
genome profiling and analysis of the T cell repertoire. Following identification of key pre-analytic variables, we
propose a standard operating procedure for implementation in observational biomarker and first-in-man clinical
trials to demonstrate the clinical utility of our approach. Proposal success will facilitate the introduction of novel
molecular assays into the clinic that augment extant and developing blood and tumor assays. This approach will
be particularly relevant as we move into the era of precision guided therapies for lung cancer treatment, which
have begun to reduce mortality in even the most advanced stages, over the coming years and decades.

NCI

08/31/25

U01 CA253166

FHCRC

Tapscott,Stephen Justice

SMCHD1 Pathways as Candidate Targets for FSHD

The proposed research will identify the fundamental molecular mechanisms that
epigenetically repress the D4Z4 region through the production, modification, stability and
interacting partners of SMCHD1. The health relevance of this research is that the
failure of these mechanisms results in facioscapulohumeral muscular dystrophy and the
proposed studies will provide the basis for future therapeutic development.

Facioscapulohumeral dystrophy (FSHD) affects ~1/10,000 people and is caused by decreased
epigenetic repression of the DUX4 retrogene with subsequent mis-expression of DUX4 in skeletal
muscle. As increasing the SMCHD1-mediated epigenetic repression at the D4Z4 locus silences DUX4
in FSHD1 and FSHD2 muscle cells, this application will take a direct molecular biology approach to
identify the diversity of SMCHD1 complexes and the role of each component in establishing and
maintaining repressive chromatin structure at the D4Z4 and preventing DUX4 expression in skeletal
muscle. The broad and long-term goal is to determine the functional components of the SMCHD1
complexes at the D4Z4 locus as a basis for future therapies directed at increasing epigenetic
repression. The major hypothesis is that SMCHD1 forms different interactions depending on post-
translational modification, chromatin association, and developmental state of the cell, and that
understanding the functional roles of the different complexes will provide simple reductionist models for
testing candidate interventions. Aim 1 will determine the proteins complexed with SMCHD1 at the
D4Z4 locus and their functional significance, chromatin association, and SUMO dependence. Aim 2 will
determine the composition of SMCHD1 complexes at autosomal single copy loci in the genome
compared to repetitive regions and to subdomains of the D4Z4. Aim 3 will identify the relative roles of
SMCHD1 complex components in the establishment and maintenance of epigenetic modifications
during stem cell reprogramming and differentiation. Together, these aims will add clarity to the
components of SMCHD1 complexes and their functional roles in D4Z4 epigenetic repression, and
provide new opportunities to design interventions to suppress DUX4 expression as a treatment for
FSHD.

NIAMS

06/30/25

R01 AR066248

FHCRC

Peters,Ulrike

Advancing equity in colorectal cancer genetic risk prediction through expansion of racial/ethnic minority representation

This study will develop a comprehensive risk prediction model based on genetic and environmental/lifestyle risk
factors in a racially/ethnically diverse population which will correct the Euro-centric bias. To translate this
transethnic risk prediction model into clinical practice, we will use microsimulation modeling to personalize
screening recommendations based on genetic and environmental risk factor information and we will develop a
web-based risk communication tool which we will evaluate in racially/ethnically diverse participants. Results from
our study will ensure that targeted personalized interventions will confer equitable benefits for all Americans
regardless of their ethnicity and race.

As colorectal cancer (CRC) remains one of the leading causes of cancer death new approaches are of vital
importance to make critical inroads in reducing the burden of this lethal disease. Precision medicine holds
enormous promise as advances in genomic research that were previously unimaginable now offer vast
commercial potential and are rapidly being moved into clinical practice, even though critical questions remain to
be addressed. Polygenic risk scores (PRS) that aggregate common genetic risk variants into a single score to
predict disease are a key example. Broad accessibility, plummeting genotyping costs, and the need to account
for the patient's individual risk profile to improve screening have provided transformative opportunities in
personalized prevention. However, wide-scale clinical adoption of PRS raises key ethical and scientific
challenges. Arguably one of the most critical challenge is the fact that current PRS are substantially more
effective in predicting risk in individuals of European ancestry compared with other populations due to the Euro-
centric bias in genetic research. Accordingly, we need to develop an unbiased PRS that predicts CRC risk in all
major racial/ethnic groups. However, this is only the first step towards implementation, which also requires the
evaluation of the optimal risk-stratified screening approach and development of risk communication tool among
others. To address these needs, we will develop and validate an unbiased PRS for CRC across ethnic/racial
minority groups to inform risk-stratified CRC screening (Aim 1a). We will augment the PRS with an environmental/
lifestyle risk score (ERS) to account for other risk factors (Aim 1b) and examine differential effects by age of
onset (Aim 1c), given the alarming increase of early onset CRC. In Aim 2a we will determine the optimal CRC
screening strategy given an individual's risk defined in Aim 1 using our microsimulation modeling by incorporating
differences in CRC incidence and mortality rates by age, sex and race/ethnicity, and in risk factor distributions
across racial/ethnic groups. In Aim 2b we will evaluate the cost-effectiveness of risk stratified screening
compared with current screening guidelines. Finally, we will develop a risk communication tool (Aim 3a) and
investigate potential dissemination issues of risk-stratified CRC screening across ethnic/racial minority groups
(Aim 3b). Our trans-disciplinary research team, which includes a community advisory board, is uniquely set up
to address these critical questions as we 1) have brought together all known racially/ethnically diverse CRC
studies totaling to over 120,000 CRC cases and controls, 2) have strong expertise in developing comprehensive
genetic and environmental risk scores in racially/ethnically diverse populations, 3) lead one of the most
comprehensive decision models for cost-effectiveness analysis which has consistently been used to inform US
screening guidelines and 4) have expertise in implementation science for genetic research in minorities.
Addressing all aims is critical to avoid a sequential science in this rapidly moving field of precision medicine.

NCI

06/30/25

R01 CA244588

FHCRC

Bricker,Jonathan B

Telephone Delivered Acceptance & Commitment Therapy for Weight Loss

The overall aim of this project is to determine the effectiveness of Acceptance and Commitment Therapy
telephone coaching for weight loss. If successful, this intervention can be cost-effectively scaled on a
population level, thereby making a high public health impact.

More than 2 in 3 adults in the US are overweight or obese [1, 2]. Both conditions, and especially
obesity, contribute to many health conditions including diabetes, heart disease, and cancers [3-5]. National
costs due to obesity are high: $342 billion dollars (2013) in medical costs, which is 28% of all adult healthcare
spending [6]. One form of behavioral intervention for weight loss is telephone-delivered coaching [7-9].
Telephone coaching has clear advantages: high population level reach and individually tailored training from a
coach [10-12]. Telephone coaching for weight loss reaches over 1.2 million US adults per year [7, 12-14].
However, a critical barrier to progress in the field is that standard behavioral therapy (SBT) telephone coaching
interventions have small effect sizes that are rarely evaluated against active treatment controls. Needed now
are telephone coaching programs with the potential to boost weight loss success over and above SBT. We
propose a new behavioral intervention for telephone coaching: Acceptance and Commitment Therapy (ACT)
[15]. Unlike SBT, ACT interventions address the fundamental challenge of weight loss: overeating in response
to internal (e.g., stress) and external (e.g., high calorie foods) cues [16-19]. ACT for weight loss addresses
disinhibition by focusing on (1) increasing willingness to experience physical cravings, emotions, and thoughts
that cue eating and impede physical activity while (2) making healthy diet and physical activity choices guided
by deeply held values [16, 20]. While ACT has been applied to many behaviors and in a variety of delivery
modalities [21-24], for weight loss it has only been tested in RCTs for in-person interventions [23, 25-30]. Dr.
Bricker’s team recently conducted a multi-step design process that yielded an ACT telephone coaching
protocol. We tested the protocol in a pilot RCT (N = 105), comparing it with telephone coaching SBT.
Compared to SBT, ACT participants had greater success on the 10% or more weight loss main outcome, at
both the 3- and 6-month follow-up. Building on these encouraging results, we propose to conduct a fully
powered randomized controlled trial of ACT telephone coaching (n = 199) versus SBT telephone coaching (n =
199), in order to determine if telephone coaching ACT: (1) has significantly higher weight loss at 12 months
post randomization, and (2) has 12-month weight loss on the main outcome (and secondary outcomes)
mediated by these ACT-consistent psychological processes: (a) acceptance of food cravings, (b) acceptance
of discomfort from physical activity, (c) mindful eating, and (d) values guided motivation to change. We will
explore whether the 12-month weight loss main outcome for ACT, versus SBT, differs by these baseline
factors: (a) age, (b) sex, (c) race/ethnicity, (d) BMI, (e) depression, (f) anxiety. If successful, telephone
coaching ACT will offer a more effective, broadly scalable weight loss treatment—thereby making a high public
health impact.

NIDDK

06/30/25

R01 DK124114

FHCRC

Parkhurst,Susan M

Cellular mechanisms of nucleocytoplasmic export through Nuclear Envelope Budding

The transport of molecular complexes from the nucleus to the cytoplasm is essential for most
developmental processes and has been thought to occur only through specific openings in the nuclear
envelope/membrane. A new alternate export pathway was recently described wherein large molecular
complexes exit the nucleus by ‘budding’ through the nuclear envelope/membrane, a process that we
find requires the highly-conserved WASH subfamily of Wiskott-Aldrich Syndrome (a disease with clinical
symptoms including immune deficiency and cancer) proteins. These studies are expected to provide
new insight into the rules governing this new nuclear export pathway in normal cellular processes, the
consequences of inappropriate regulation leading to disease conditions such as laminopathies,
neurological disorders, aging, herpesvirus infections, and/or cancer onset/progression, as well as
provide a fresh basis for new potential therapeutic developments.

Transport of nucleic acids and proteins from the nucleus to the cytoplasm is essential for nearly all cellular
processes, and when mis-regulated, is associated with diseases, tumor formation/growth, and cancer
progression. Canonically, this indispensable process has been thought to occur exclusively via Nuclear
Pore Complexes, which span the nuclear envelope’s double membranes and provide a critical regulatory
step in what exits (and enters) the nucleus. Recently, Nuclear Envelope (NE-) budding was shown to
provide an alternative pathway for nuclear exit, particularly for large ribonucleoprotein (RNP) complexes
that would otherwise need to unfold/remodel to fit through the pores. In this pathway, large
macromolecule complexes are encapsulated by the inner nuclear membrane, cross the perinuclear
space, fuse with the outer nuclear membrane, and are released into the cytoplasm, a mechanism
strikingly similar to herpesvirus nuclear egress. Thus, NE-budding elegantly allows for large RNP
complexes to exit the nucleus together and be delivered as a package for specific cellular functions.
Despite its clear biological importance and clinical relevance, very little is yet known about the regulatory
or structural machineries that allow NE-budding to occur in any system. Recently, we found that the
Wiskott Aldrich Syndrome family actin nucleation protein, WASH, its four subunit regulatory complex
(SHRC), and Arp2/3 are necessary for NE-budding. Using WASH/SHRC as a new entry point, in tandem
with strategies to discover novel genes/proteins involved in this process, our long-term goal is to
understand the molecular and cellular mechanics that govern NE-budding. The specific aims of this
proposal are to determine the mechanism(s) of WASH/SHRC function in NE-budding, and to
identify/analyze the infrastructural components/machineries governing the dynamic NE-budding process
using a combination of genetic, biochemical, cell biological, time-lapse live imaging, and super-
resolution/EM microscopy approaches. Drosophila provides an excellent, genetically amenable,
organism for studying this conserved process due to its amenability for imaging and the wealth of cutting
edge cell/molecular techniques and reagents. The information gathered in these studies will help to
elucidate the mechanisms governing this exciting new nuclear export pathway in normal development or
when mis-regulated in disease conditions, and may inform the study of herpesvirus nuclear egress as
well.

NIGMS

06/30/25

R01 GM143186

FHCRC

Bricker,Jonathan B

Quit2Heal: Rigorous Randomized Trial of a Smartphone Application to Help Cancer Patients Stop Smoking

The overall aim of this project is to determine the effectiveness of a smartphone app designed to help cancer
patients quit smoking. If successful, this intervention would provide an effective and highly accessible public
health intervention for cancer patients.

In the US, 15% to 54% of cancer patients are cigarette smokers at the time of their diagnosis. Unfortunately,
up to 80% of smokers with cancer continue to smoke after their diagnosis. Persistent smoking after a cancer
diagnosis leads to worse clinical outcomes including decreased treatment effectiveness, increased risk of
disease recurrence, increased risk of developing second cancers, and higher mortality rates. The National
Comprehensive Cancer Network (NCCN) recommends that every cancer patient who smokes be offered an
evidence-based cessation intervention. However, limited access to effective tobacco cessation interventions is
a key barrier for promoting cessation in the context of cancer care. Needed now is a method for all smokers
with cancer to access effective and low-cost interventions designed to help this special population stop
smoking. A smartphone application (“app”) could address the access need by providing a widely available
tobacco intervention targeted for cancer patients. Apps have potentially high population level reach to cancer
patients—especially given that over three quarters (76%) of all smokers own smartphones. Despite their high
potential value, there is no evidence about: (1) the effectiveness of smartphone apps targeted to cancer
patients who smoke and (2) why these apps would be effective. Also unexplored is who is most likely to benefit
from smartphone apps targeted to cancer patients who smoke. We recently developed the first known
smartphone app (called “Quit2Heal”) specifically designed to help cancer patients quit smoking. We compared
Quit2Heal with the NCI’s QuitGuide, a widely used smoking cessation app for the general population, in a pilot
randomized controlled trial with 59 adult cancer patients who smoke. With 92% retention at the 2-month follow-
up, 30-day point prevalence quit rate was 20% for Quit2Heal vs. 7% for QuitGuide (OR=5.16; 95% CI: .71,
37.29; p=.104). Building on these knowledge gaps and encouraging results, this application proposes to
conduct a fully powered randomized controlled trial of Quit2Heal (n = 211) versus QuitGuide (n = 211), in order
to determine whether: (1) Quit2Heal has significantly higher biochemically verified 30-day point prevalence
smoking cessation at 12 months post-randomization relative to QuitGuide, and (2) Quit2Heal’s (but not
QuitGuide’s) 12-month smoking cessation outcome is significantly mediated by improvements in cancer-
related shame, stigma, depression, anxiety, and knowledge about consequences of smoking vs. quitting after
cancer diagnosis. The project will also explore whether these baseline factors moderate the cessation
outcome: (a) cancer type, (b) stage of cancer, (c) time since diagnosis, (d) demographics, (e) having a
partner/spouse who smokes, (f) whether participant was advised to quit by oncologist, and (g) recruitment
source. If the aims are achieved, this project will advance scientific knowledge about the effectiveness of apps
for helping cancer patients quit smoking. Positive results would provide an effective and highly accessible
public health intervention for cancer patients.

NCI

05/31/25

R01 CA253975

FHCRC

Orozco,Johnnie J

Combining Targeted RIT and Synergistic Novel Agents to Eradicate AML

Although favorable responses have been attributed to recently approved targeted agents for the treatment of
hematologic malignancies, such as acute myeloid leukemia (AML), the diversity of genetic mutations within blood
cancers limits the curative potential of targeted monotherapies. However, hematologic malignancies are
especially vulnerable to radiation, and thus genetically diverse sub-clones may all be eliminated by targeting
radiation directly to the sites of disease through the powerful technology of radioimmunotherapy. This grant
proposes to improve outcomes for AML by: 1) identifying synergistic combinations of radioimmunotherapy
approaches with highly effective, but not yet curative, targeted agents; 2) correlating efficacy with the extent of
DNA damage achieved by synergistic combinations; and 3) by comparing these radioimmunotherapy
combinations as preparative chemotherapy prior to allogeneic bone marrow transplantation.

Potentially curative treatments for acute myeloid leukemia (AML) are limited to intensive systemic
chemotherapy with or without allogeneic bone marrow transplantation (BMT). However, not every patient is
healthy enough to tolerate intensive treatments, and not every patient may have a suitably HLA-matched stem
cell donor, especially patients from ethnic minority groups. Targeted agents have recently been approved to
treat AML, but these usually require intensive systemic chemotherapy to optimize efficacy. Furthermore, AML
is genetically heterogeneous with distinct genetic mutations and chromosomal alterations that makes targeted-
agent monotherapy unlikely to be curative. AML, like most hematologic malignancies, is very sensitive to
radiation therapy but even involved field radiation may be too toxic and ineffective for disseminated systemic
disease. However, radioimmunotherapy (RIT) mitigates the off-target toxicity by using monoclonal antibodies
conjugated to radioactive isotopes to deliver radiation payloads directly to sites of disease by virtue of the
antibody specificity. We have shown that RIT using 90Y- and 131I-radiolabeled anti-CD45 antibody targets
radiation to sites of leukemia while minimizing radiation to uninvolved organs. We have improved upon our
approach without increasing toxicity by targeting higher energy alpha-emitting radionuclides (astatine-211;
211At) to sites of disease and by developing a pre-targeted RIT (PRIT) approach using bispecific antibodies
targeting CD45 and 90Y-DOTA.
Using preclinical murine models, we now propose to identify synergistic combinations of 211At- and 90Y-anti-
CD45 RIT with novel targeted therapies that interfere with DNA repair or promote apoptosis. We will do this by
first assessing for synergy between alpha- or beta-emitting radionuclides (211At- and 90Y-) employed in anti-
CD45 directly labeled RIT with recently approved targeted agents (PARP and BCL2 inhibitors) in both
disseminated syngeneic and xenograft leukemia murine models. Second, we will improve therapeutic efficacy
of anti-CD45 PRIT via bispecific antibody constructs targeting CD45 and 90Y-DOTA by assessing for synergy
with targeted therapies (PARP and BCL2 inhibitors) in leukemia murine models. We will characterize the extent
of DNA damage achieved with these two approaches as a means to elucidate the mechanism of efficacy.
Finally, we will compare these two approaches as part of conditioning prior to allogeneic BMT using
haploidentical, or partially matched donors, as all patients should have haploidentical donors.
These preclinical studies should readily translate into clinical trials given our infrastructure for NIH funded and
pharmaceutical-sponsored clinical trials, using anti-CD45 RIT prior to bone marrow transplantation for
aggressive hematologic malignancies. These studies will add effective, well-tolerated treatment options for
patients with AML by identifying synergistic combinations of targeted agents with anti-CD45 RIT approaches
and by identifying the optimal RIT approach prior to haploidentical BMT.

NCI

05/31/25

R37 CA252070

FHCRC

Eisenman,Robert N

Investigating Max as a tumor suppressor gene in small cell lung cancer and other neuroendocrine tumors

This proposal is focused on new genetic and molecular pathways that cause and sustain small cell lung cancer
and potentially other related tumors. This research is relevant to public health because it will ultimately lead to
a better understanding of the causes for a deadly cancer and provide new approaches to cancer therapy.

Dysregulated expression of the MYC family of transcriptional regulators is a common denominator in a wide
spectrum of human cancers, including small cell lung carcinoma (SCLC), a highly aggressive neuroendocrine-
type tumor that is among the leading causes of US cancer mortality. MYC family proteins specifically
heterodimerize with MAX in order to bind genomic DNA and stimulate widespread transcription. Surprisingly,
recent reports show that MAX is inactivated through deletions and truncating mutations in a significant subset of
SCLC and other neuroendocrine cancers. The paradox that MAX may act as a tumor suppressor, but yet be
crucial for MYC oncogenicity in SCLC, has important implications for our understanding of the etiology of these
tumors but has not been systematically investigated. This proposal is based on the findings from our two
laboratories that (i) a whole-genome CRISPR inactivation screen in pre-neoplastic SCLC (preSCs) revealed
MAX-targeting sgRNAs to be highly growth promoting; and (ii) deletion of MAX dramatically accelerates SCLC
in an autochthonous mouse model. These results provide, for the first time, highly relevant biological systems to
elucidate MAX's tumor suppressor function.
In Aim 1 we will characterize the biological properties of MAX-deleted SCLC including proliferation, apoptosis
and genomic stability. Moreover, we will use ChIP-Seq and RNA-Seq to determine the genomic landscape of
MYC/MYCL and MAX binding and target gene expression in SCLC and MAX-deleted SCLC. We will functionally
interrogate the importance of key MAX target genes identified through integrative genomic analyses. Targets to
be studied will include MAX-dependent regulators of one carbon metabolism already identified. Aim 2 is based
on the hypothesis that MAX deletion not only alters MYC activity but disrupts the broader MYC- MAX
transcriptional network of activators and repressors. We will determine if MYC/MYCL have MAX independent
oncogenic functions in our SCLC models and in human SCLC cell lines, and examine whether network members
that antagonize or cooperate with MYC (such as MXD/MNT, MLX, and MondoA), act to influence SCLC
progression. In Aim 3 we propose to determine core MAX-regulated genes and pathways common to
neuroendocrine tumor suppression by MAX. This will entail molecular and genetic characterization of our new
models of thyroid medullary carcinomas and pheochromocytomas resulting from MAX loss (in an Rb/p53
deficient background) and identification of pathways shared with MAX-null SCLC. This research will extend the
breadth of our studies to uncover how MAX suppresses neuroendocrine cancers. We anticipate that these
studies will deepen our understanding of the complex role of the MYC network in both driving and suppressing
neoplasia and identify novel tumorigenic pathways that may have the potential to serve as therapeutic targets.

NCI

04/30/25

R01 CA248762

FHCRC

Ghajar,Cyrus M

Defining and characterizing microenvironmental drivers of disseminated tumor cell dormancy in brain

Brain metastases are a growing problem across cancers. They can take many years to manifest, but are lethal
once they do. Here, we propose to investigate breast tumor cells upon their arrival to the brain in order to
define the cues that make them dormant, and to unravel the consequent signaling. We believe that solving
these mysteries will inspire approaches to prevent brain metastasis by leveraging our discoveries for
prophylactic therapies.

Brain metastases arise later than metastases at other sites. Once they do, they are rapidly debilitating and
lethal. The time it takes for brain metastases to emerge suggests that a dormancy phase is involved. This
notion is supported by clinical and experimental data. Indeed, our own preliminary data show that breast
cancer cells become dormant upon entering the brain, and that emerging from this state is the rate-limiting step
of metastasis. These data indicate that targeting dormant disseminated tumor cells (DTCs) is a logical
approach to brain metastasis prevention. However, despite a growing understanding of dormancy mechanisms
in common metastatic sites like lung and bone marrow, a parallel understanding of how DTCs are driven into a
dormant state in brain has not developed. The overarching goal of this proposal is to address this issue. We
will formulate a basic framework for how the brain microenvironment drives DTCs into a dormant state, with
support from clinical specimens, so that we can leverage this understanding for therapies that keep DTCs
dormant indefinitely.
Our recent investigations have revealed that dormant DTCs occupy the brain’s vascular niche, where
perivascular astrocytes suppress their outgrowth. We suspect that astrocytic contributions to the parenchymal
basement membrane are responsible for DTC suppression, and that these contributions converge on a
common receptor: dystroglycan. Therefore, our hypotheses are that: (i) astrocytic basement membrane is a
key driver of DTC dormancy, and (ii) dystroglycan function must remain intact for DTCs to interpret these
signals. We will test these hypotheses through two specific aims:
Aim 1. Determine whether astrocytic basement membrane promotes and sustains DTC dormancy.
Aim 2. Elucidate the dystroglycan-mediated signaling axis that effects DTC quiescence in brain.
We have brought every relevant resource to bear in order to address these aims. These resources span: (i)
long-term intravital imaging to determine the fate of DTCs following ablation of DTC-associated astrocytes; (ii)
transgenic mice to measure the outcome of ablating astrocyte derived basement membrane molecules on DTC
fate; (iii) rare clinical specimens to establish whether astrocytes and astrocytic basement membrane are asso-
ciated with dormant DTCs in humans; and (iv) a host of mutant, over- and under- expression constructs to
solve how dystroglycan functions from the outside-in to drive DTC quiescence.
The significance and innovation of this work lie in the identification of the first dormancy drivers in brain,
ultimately to unravel dystroglycan-driven signaling that effects disseminated breast tumor cell quiescence. This
work will set the stage for agonists of dystroglycan function that serve as prophylactics for brain metastasis
prevention.

NCI

04/30/25

R01 CA252874

FHCRC

Bricker,Jonathan B

Full Scale Randomized Trial of an Innovative Conversational Agent for Smoking Cessation

The overall aim of this project is to determine the effectiveness of a conversational agent for smoking
cessation. If successful, this intervention can be cost-effectively scaled on a population level, thereby making a
high public health impact.

Cigarette smoking accounts for 480,000 premature deaths and one third of all cancer deaths annually in the
US. There is enormous need for high-impact, cost-effective population-level interventions for smoking
cessation. For the past 15 years, mobile phone-delivered text messaging interventions such as the NCI’s
SmokefreeTXT have been a prominent technology addressing this need. However, very much like all widely
available technologies for smoking cessation (e.g., websites), text messaging interventions have modest quit
rates, driven largely by low engagement. Fortunately, a new technology provides a therapeutic conversation to
address the problem of engagement that impacts text messaging and other current digital technologies for
smoking cessation. Advances in machine learning, natural language processing, and cloud computing are now
making it possible to create and widely disseminate conversational agents (CAs), which are computer-powered
digital coaches designed to form long-term social-emotional connections with users through conversations.
CAs are supportive, empathic, reflectively listen, provide personalized responses, and offer goal setting and
advice appropriately timed to the needs of the user. Regarding CAs for smoking cessation, the major
knowledge gaps are: (1) their efficacy, (2) theoretical mechanisms, and (3) the cost-effectiveness. Also
unexplored are the potential baseline moderators of CAs for smoking cessation. We recently developed a CA
for smoking cessation, called “QuitBot,” evaluated it in a diary study, and then tested it in a pilot randomized
controlled trial (N = 306), comparing it with the NCI’s SmokefreeTXT. The pilot RCT design was very feasible
with 93% three-month follow-up. QuitBot had: (a) high participant engagement and (b) high quit rates at the
three-month follow-up—very promising in comparison with SmokefreeTXT. Addressing these knowledge gaps
and building on the promising results of our QuitBot research, the project will conduct a randomized controlled
trial of QuitBot (n = 760) versus SmokefreeTXT (n = 760) with 12-month follow-up in order to determine
whether QuitBot: (1) provides higher quit rates than SmokefreeTXT, (2) has smoking cessation outcomes
significantly mediated by therapeutic alliance processes and engagement, and (3) is cost-effective vs.
SmokefreeTXT. In addition, this study will explore whether these baseline factors moderate the effectiveness
of QuitBot: trust, social support, and demographics (e.g., sex). This innovative project will advance the fields of
research on CAs both for smoking cessation in particular and for health behavior change in general—
regardless of whether the results are positive or null. Positive results could have high population-level impact
and stimulate new lines of research into CA dissemination and implementation, and the adaptation of CAs for
multiple subpopulations of smokers, languages, and community and medical settings.

NCI

03/31/25

R01 CA247156

FHCRC

Hsu,Li

Integrative Genomics into Genetic Association Studies of Blood Pressure and Stroke in African Americans

While stroke is the third leading cause of death among African Americans (AA) and blood-pressure is a major
risk factor of stroke, genetic susceptibility to stroke and hypertension of AAs is less well studied than that of other
ethnic groups. The objective of this study is to identify variants predicting various types of genomic features in
AAs, and to integrate the functional regulation information into AA-specific genetic association analysis of stroke
and blood pressure. Discovery of genetic variants that predispose to stroke and blood pressure is a crucial step
toward understanding genetic mechanisms that may lead to novel prevention and treatment strategies in this
under-studied ethnic group.

Stroke is the third leading cause of death among African Americans (AAs): they are twice as likely to die from
stroke as European Americans (EAs), and their incidence rate is almost double that of EAs. Recent genome-
wide studies (GWAS) suggest there is a substantial genetic contribution to stroke risk in African ancestry
populations, with heritability estimates of about 35%. However, to date, genetic studies in AAs are greatly lagging
behind those in EAs despite their increased stroke burden. Among the risk factors for stroke, blood pressure is
a major contributor: 4 in 10 AAs suffer from hypertension, 50% more than EAs. These disparities have been
considered to be mediated by environmental and social determinants, yet they remain after adjusting for
demographics, socioeconomic status, clinical characteristics, and modifiable health behaviors. Heritability
analysis suggests African ancestry is associated with hypertension, with heritability estimates from 30–40% for
systolic and diastolic blood pressure. However, genetic susceptibility to hypertension among AAs is less well
studied compared to other ethnic groups. Therefore, there is considerable motivation for identifying the genetic
components of stroke and high blood pressure in AAs.
Discovery of genetic variants that predispose to blood pressure and stroke is a crucial step toward
understanding genetic mechanisms that may lead to novel prevention and treatment strategies. Yet, GWAS have
thus far identified genetic loci that together account for only a small proportion of the heritable risk. Substantial
efforts have been devoted to studying the association of genetic variation with gene expression and other
molecular characteristics through large collaborative initiatives such as Genotype-Tissue Expression (GTEx) and
Encyclopedia of DNA Elements. These initiatives have provided a deeper understanding of functional elements
across the genome, which have been used to inform genetic association and identified many novel loci. However,
most of the data in these studies have focused on European ancestry and little has been done in AAs. Our recent
work in Nature shows that genetic discoveries in one population do not readily transfer to other populations.
The objective of this study is to identify variants predicting various genomic features (gene expression,
methylation and protein) in AA samples that have been recently collected through Trans-Omics for Precision
Medicine, the CommonMind Consortium, and GTEx, and to integrate this functional information into genetic
association analysis of blood pressure and stroke in AAs. Insight into both molecular activity and genetic variation
can inform association analysis and enable novel genome-wide discoveries. In particular, we propose to develop
methods that leverage the data for EAs to improve power for identifying genetic variants that regulate various
types of genomic features in AAs, and for integrating the genomic regulation models into GWAS with the ultimate
goal to identify novel loci for stroke risk and blood pressure in AAs. To facilitate these aims we have assembled
the largest number of AAs for genomic studies and AA stroke cases and blood pressure data for GWAS.

NHLBI

03/31/25

R01 HL152439

FHCRC

Kooperberg,Charles L

Integrative Genomics into Genetic Association Studies of Blood Pressure and Stroke in African Americans

NHLBI

03/31/25

R01 HL152439

FHCRC

Cheung,Kevin J

Role of tumor cell cluster-induced signaling in breast cancer metastasis

The ultimate cause of most breast cancer-related deaths is metastasis, the spread of cancer to distant organs.
Recent research indicates that in breast cancer, as well as many other tumor types, circulating clusters of
tumor cells are associated with therapy resistance, widespread metastases, and significantly worse prognosis.
By learning more about these micrometastases, this proposal seeks to identify molecular vulnerabilities that
can be used to eradicate them, thereby extending survival in patients with metastatic breast cancer.

The root cause of most breast cancer deaths is metastasis. By dissecting the molecular events driving it, the
research community can develop new therapeutic approaches to eradicate and prevent metastatic disease.
One promising avenue of research involves the cooperative behavior of tumor cells. Conventionally,
metastasis is conceptualized as the dissemination of individual tumor cells to distant organs. However, recent
studies by the Cheung research group and others have established that clusters of tumor cells metastasize to
distant organs more efficiently than single cells in mouse models, and that circulating tumor cell clusters are
associated with poor patient outcomes and therapy resistance in humans. The molecular mechanisms
responsible for aggression in tumor cell clusters and the optimal therapeutic strategies to eliminate clusters
have remained obscure. Recently, the Cheung laboratory has found that clustered tumor cells display
heightened levels of apoptosis resistance, cell proliferation, and changes in molecular expression that indicate
that the cells are cooperating with one another. These studies reveal that the tyrosine kinase EGFR is
activated at cell-cell contacts in clustered tumor cells, and they establish that EGFR and the low-affinity EGFR
ligand Epigen are necessary for cluster-dependent proliferation and metastatic colonization. The proposed
project will test the hypothesis that tumor cell clusters are highly metastatic because they contain a private
signaling environment involving EGFR, Epigen, and the transcription factor Fra-1, and that disrupting this
signaling environment will neutralize clusters’ metastatic potential. The Cheung lab has already developed
technically innovative organoid and murine models to study cluster-based signaling and its impact on
metastasis in vivo. Using these models, the lab will first determine whether cluster-induced metastatic
efficiency depends specifically on local activation by Epigen. Second, the lab will determine the impact of Fra-1
transcriptional programs and signaling feedback loops on metastatic processes specific to tumor cell clusters,
as well as whether this program depends on the presence of Epigen. Third, the lab will supplement its
experimental findings by studying the association between EGFR, Epigen, and long-term recurrence and
mortality data from human breast cancer datasets. Through this integrated approach, the Cheung lab will
develop an understanding of the cooperative molecular mechanisms that underlie the propensity of tumor cell
clusters to metastasize. As described in the proposal, this understanding is likely to reveal molecular
vulnerabilities that can be exploited to develop new anti-metastatic therapies. Although the work proposed here
focuses on uncovering therapeutic strategies to target tumor cell clusters in breast cancer, the findings will
potentially be relevant to a wide range of tumor types.

NCI

11/30/24

R37 CA234488

FHCRC

Kooperberg,Charles L

Physical Activity to Improve CV Health in Older Women: A Pragmatic Trial

WHISH is a landmark, pragmatic trial testing the hypothesis that a centralized, public health, multi-component
physical activity (PA) intervention designed to increase and/or maintain PA and reduce sedentary behavior, will
reduce major cardiovascular events in older women. We propose to extend the WHISH intervention and follow-
up for 4 additional years to ensure a definitive test of the benefits and risks of the program, and to determine
PA effects on physical function, sleep, depressive symptoms and biomarkers of healthy cardiovascular aging.
The outcomes of WHISH are likely to have immense translational impact on the future of physical activity
programs for healthy cardiovascular aging.

This is a competitive renewal application for the Women’s Health Initiative (WHI) Strong & Healthy (WHISH)
trial. America’s 65-and-older population is projected to double in size from 49 million today to 95 million by
2060, with women far outnumbering men, particularly among adults aged 85-and-older. Compelling evidence
supports the hypothesis that physical activity (PA) reduces cardiovascular (CV) disease (CVD), preserves
physical function (PF) and promotes other aspects of CV health in older adults. WHISH is a landmark,
pragmatic randomized controlled trial testing whether a centralized, public health intervention designed to
increase and/or maintain PA levels and reduce sedentary behavior will reduce major CVD (MI, stroke, CV
death) in older women. Using a randomized consent design to simulate real-world programmatic
implementation, WHISH randomized 49,333 eligible participants in the WHI Extension Study to a behavioral
intervention versus usual follow-up in May 2015. A passive consent process in the Intervention group
(n=24,663) resulted in <4% of women “opting out” of receiving intervention materials. WHISH delivers a
targeted, adaptive, instructional intervention, based on 2008 and 2018 DHHS PA guidelines and designed to
complement the National Institute on Aging’s (NIA) Go4Life® campaign, using seasonal newsletters, manuals,
pedometers, resistance bands, telephone and e-mail motivational messages, and a website designed for older
women. The intervention adapts to participant feedback from annual surveys and other input regarding activity
preferences and is customized and targeted to their current PF and PA levels. Primary effectiveness and safety
outcomes (CVD, fracture) are evaluated using intention-to-treat in the entire randomized cohort. By the end of
the current grant period (Feb. 2020), 4 years of follow-up will be available. Based on WHISH observed CVD
event rates, intervention effects on PA and sedentary behavior, and new WHI data relating PA behaviors to
CVD endpoints, revised power calculations suggest we will have only 65% power to evaluate the impact of the
WHISH intervention on CVD events. We estimate that 4 additional years of follow-up (8 years overall) will yield
85-89% power to provide a definitive test of the primary WHISH hypothesis. This application proposes to
extend the WHISH intervention and follow-up of outcomes for 4 additional years so that the trial can reach a
definitive conclusion on the benefits and risks of the PA intervention. We also propose to leverage data and
biospecimen collections in a planned WHI Extension Study home visit to enable evaluation of key markers of
healthy CVD aging including physical performance, sleep duration and insomnia symptoms, and depressive
symptoms. Pilot studies will be executed to explore long-term effects of the WHISH intervention on changes in
established and novel biomarkers known or postulated to be influenced by PA levels and indicative of various
underlying mechanisms related to CVD aging. The outcomes of WHISH, whether positive, null, or adverse,
could have immense translational impact on the future of PA dissemination programs for healthy CV aging.

NHLBI

08/31/24

R33 HL151885

FHCRC

Anderson,Garnet L

Life and Longevity After Cancer (LILAC): The Women's Health Initiative Cancer Survivor Cohort

To address compelling issues of cancer and aging, we propose to enhance the Life and Longevity After
Cancer (LILAC) cohort by continuing to enroll older (70+ year old) cancer survivors from the Women’s Health
Initiative (WHI), by establishing measures of frailty, creating age-matched cancer-free controls, and in a subset
of participants, conducting a geriatric assessment and collecting a new post-diagnosis blood collection in a
subset of participants to assess biomarkers of aging. Further, by making these resources broadly available,
LILAC will provide the research infrastructure needed to examine many emerging questions around
accelerated aging and resilience in cancer patients.

There are 16 million cancer survivors in the United States, and approximately 60% are age 65 years and over.
By 2040, it is estimated that the number of cancer survivors will grow to 26 million with 73% age 65 and older
and almost 50% age 75 and older. Despite this so-called silver tsunami of older cancer survivors, there are
significant knowledge gaps regarding how cancer and cancer treatment impacts the aging process. There is
also a growing consensus regarding the need for studies of cancer in older adults because of the prevalence of
comorbidities, functional losses, cognitive impairment, and frailty, and particularly for those age ≥ 75 for whom
there are virtually no data. Studies are needed to better understand how cancer and its treatment interact with
underlying vulnerabilities, which in turn impacts the feasibility, safety, and efficacy of interventions in this
population. The Life and Longevity After Cancer (LILAC) project is a cancer survivor cohort embedded within
the Women’s Health Initiative to support studies of cancer survivorship in an aging population. During the initial
funding period, we developed the LILAC cohort of 13,453 WHI cancer survivors who were diagnosed with one
of eight LILAC designated cancers: invasive breast, colorectal, endometrial, melanoma, leukemia, lung,
lymphoma, melanoma, or ovarian. We collected information on first course of treatment, clinical and patient
reported outcomes, as well as archival tissue from 4351 solid tumors. With a minimum of 20 years of data on
these women and a current age-range of 70 to101 years, the LILAC population is poised to provide key
information on cancer and aging to advance our cancer and aging research agenda. To accomplish this we
propose to enhance the LILAC resource through several mechanisms designed to support analyses of
emerging questions. Specifically, we propose: 1) To fill critical gaps in knowledge regarding the self-reported
physical, mental and social health of older female cancer survivors by continuing to enroll newly diagnosed
survivors (N=2685) and follow the LILAC cohort; 2) To develop the analytic framework to assess trajectories of
aging, including an accelerated aging phenotype in the WHI/LILAC database; 3) To establish cohorts of age-
matched WHI participants with similar data who have remained cancer free, to help us understand the
diagnosis of cancer and its treatment on the trajectories of aging, the accelerated aging phenotype and age-
related comorbidities; 4) To obtain performance-based measures of physical function and new post-treatment
blood samples to assess potential biomarkers of accelerated aging; and 5) To maximize the use and impact of
this resource. Given the aging of the US population, the increase in the number of cancer survivors and the
association between cancer and aging, these strategic and timely investments in the LILAC infrastructure will
help to fill many of these critical research gaps.

NCI

08/31/24

U01 CA173642

FHCRC

Sorror,Mohamed L

Novel Intervention Approaches to Alleviate Allogeneic Transplant-Related Morbidity and Mortality

The proposed study will determine in a seamless phase II/III design whether older and/or medically infirm
patients with hematological malignancies will see an improvement in their quality of life following allogeneic
hematopoietic cell transplantation when treated with one of three novel peri-transplant interventions: supportive
and palliative care, clinical management targeting specific comorbidities, and both interventions combined.
Overall survival, non-relapse mortality, other patient-reported outcomes, and use of resources will also be
compared between the intervention arms.

BACKGROUND: Hematological malignancies (HM) are rare cancers that affect the blood and lymph system
and can only be cured with transplantation of donor stem cells; namely allogeneic hematopoietic cell
transplantation (HCT). While advancements have improved outcomes for younger and otherwise healthy
patients, HCT can have devastating effects on the health-related quality of life (HRQOL) for older and
medically infirm (vulnerable) patients with HM. We were the first to develop an HCT-specific comorbidity index
(HCT-CI) that specifically showed that those with high scores can suffer from significant impairments in
HRQOL and higher rates of morbidity and mortality after HCT compared to patients with lower comorbidity
scores. Guided by our and others’ preliminary studies, we propose here to randomize those vulnerable patients
between supportive and palliative care, clinical management targeting specific comorbidities, both approaches
combined, vs. standard of care (SOC) to see which intervention can improve HRQOL of those patients after
HCT. OBJECTIVES: (1) Evaluate in a randomized phase II study the effectiveness of the four approaches
mentioned above in improving day-90 HRQOL for vulnerable recipients of allogeneic HCT; (2) determine in a
phase III study whether the winner arm from phase II definitively improves HRQOL vs. SOC; and (3) compare
the interventions with respect to survival, additional patient-reported outcomes, and the use of resources.
METHODS: We will conduct a multi-center seamless phase II/III randomized clinical trial in five large transplant
centers. The seamless feature means patients used in the phase II analysis will be included in the phase III
analysis but that will only happen if phase II produces an intervention with a clear advantage over SOC. In Aim
1, we will enroll 300 patients who either have age of ≥65 years, HCT-CI scores of ≥3, and/or slow walk speed
as indication of frailty, two weeks before they start their allogeneic HCT. Each intervention arm will be
implemented over 10 weeks period, 2 weeks before and 8 weeks after HCT, to achieve the maximum benefit in
preparing patients for HCT and guiding them through the early phases of the procedure. Patients will be
randomly assigned to one of the four arms described above. If there is an intervention arm from phase II that
improves HRQOL, then we will continue to test that arm only against SOC in a phase III study. We will only
need an additional 300 vulnerable patients for phase III (total of 600 for phase II/III). In Aim 3, we will see if any
of the interventions can improve survival, other patient-reported outcomes, and/or the use of resources.
PATIENT OUTCOMES: This proposal is the first to compare these peri-transplant interventions in vulnerable
HM patients given HCT. Results can minimize the suffering and, if possible, prolong the lives of similar patients
in the future and world-wide. Results could also encourage physicians to offer transplants to more vulnerable
patients, who are currently being denied the procedure for fear of its morbidity and mortality risks. Finally, the
national overall use of healthcare resources could be improved for the benefit of other patients.

NCI

07/31/24

R01 CA227092

FHCRC

Bradley,Robert K

Genetic and molecular basis for SRSF2 mutations in myelodysplasia

Myelodysplastic syndromes (MDS) are a heterogeneous group of blood diseases, all of which are
characterized by ineffective production of blood by the bone marrow. Here, we will investigate why mutations in
the SRSF2 gene, which are commonly found in MDS, cause molecular changes within blood cells that result in
ineffective blood production. We also seek to find new ways to treat MDS with SRSF2 mutations.

Mutations in genes encoding RNA splicing factors are the most common class of genetic alterations in
myelodysplastic syndromes (MDS), a group of blood disorders that are characterized by clonal, dysplastic, and
ineffective hematopoiesis. One of the most commonly mutated genes is SRSF2, which encodes a regulator of
alternative splicing and is subject to recurrent missense mutations primarily affecting a single “hotspot” residue.
During the initial funding period of this grant, work by our labs and others led to a consensus model for how
SRSF2 mutations promote MDS: MDS-associated hotspot SRSF2 mutations alter SRSF2’s RNA-binding
affinity, driving mis-splicing of key hematopoietic regulators to cause dysplastic hematopoiesis. Importantly,
SRSF2 mutations may confer therapeutically actionable vulnerabilities. We identified specific compounds that
modulate RNA splicing to preferentially kill SRSF2-mutant cells over their wild-type counterparts, helping to
motivate the earliest clinical trials of new drugs targeting MDS with splicing factor mutations.
Here, we propose to refine and extend our current understanding of SRSF2 mutations. While useful, our
current model is not sufficient to fully explain the genetic spectrum of SRSF2 mutations, interactions between
SRSF2 mutations and other co-occurring genetic lesions, and the functional roles and therapeutic implications
of SRSF2 mutations in MDS. Our interdisciplinary team consists of a physician-scientist with expertise in MDS
and patient care (Abdel-Wahab) and a basic scientist with expertise in RNA splicing and functional genomics
(Bradley). In preliminary studies, we identified diverse phenomena that are not explained by our current model
of SRSF2 mutations: rare, non-hotspot SRSF2 mutations may be pathogenic; although multiple co-occurring
splicing factor mutations are generally thought to be incompatible with cell survival, a subset of MDS patients
carry two such mutations; SRSF2 mutations cause profound changes in RNA processing beyond mis-splicing
of cassette exons; and SRSF2 mutations induce sensitivity to multiple classes of compounds that modulate
RNA splicing via distinct mechanisms of action. We propose to build on these preliminary studies as follows:
Aim 1, Determine the molecular basis and functional consequences of widespread intron retention in SRSF2-
mutant MDS; Aim 2, Determine the biological and molecular basis for allele-specific interactions between
SRSF2 mutations and additional genetic alterations in MDS; Aim 3, Identify and test therapeutic strategies for
targeting cells with spliceosomal gene mutations. The significance of these studies is that they will give insight
into the molecular and functional basis for SRSF2 mutations in MDS. The health relatedness of this effort is
that the proposed work may identify new treatment modalities that specifically target SRSF2-mutant MDS,
which is associated with particularly poor prognosis.

NHLBI

07/31/24

R01 HL128239

FHCRC

Houghton,A McGarry

Fred Hutchinson Cancer Research Center Lung SPORE

The Fred Hutch Lung SPORE consists of four innovative translational research projects, three supportive
cores, and the required Developmental Research Program and Career Enhancement Program. We have
leveraged the strengths of the investigators and our Center to tackle three critical barriers precluding
meaningful improvements in lung cancer survival rates: facilitation of pulmonary nodule evaluation for lung
cancer early detection and screening, lack of effective therapies for small cell lung cancer (SCLC), and the
sub-optimal response rates of non-small cell lung cancer (NSCLC) patients to novel immune-based therapies.

The Fred Hutch Lung SPORE consists of four innovative projects, three supportive cores, and the required
Developmental Research Program and Career Enhancement Program. We have leveraged the strengths of
the investigators and our Center to tackle three critical barriers precluding meaningful improvements in lung
cancer survival rates: facilitation of pulmonary nodule evaluation for lung cancer early detection and
screening, lack of effective therapies for small cell lung cancer (SCLC), and the sub-optimal response rates of
non-small cell lung cancer (NSCLC) patients to novel immune-based therapies. To overcome these barriers,
we propose the following four projects: 1) Targeting the Neutrophil Lineage to Enhance Immune Checkpoint
Inhibitor Efficacy in NSCLC, 2) Targeting Neoantigens for Lung Cancer Immunotherapy, 3) Identifying
Determinants of Sensitivity to LSD1 Inhibition in SCLC, and 4) Risk Stratification for Pulmonary Nodules
Detected by CT Imaging Using Plasma and Imaging Biomarkers. These projects will be supported by an
Administrative Core, a Biostatistics and Bioinformatics Core (BBC), and a Histopathology and Biospecimen
Core (HBC).

NCI

05/31/24

P50 CA228944

FHCRC

OhAinle,Molly O

Discovery and Characterization of Capsid-Targeting Lentiviral Restriction Factors

Eradication of HIV infection and disease remains a significant obstacle in biomedical research.
Here we will further our basic understanding of the adaptation of primate lentiviruses to humans
and the mechanisms by which human cells block infection. We foresee that defining host
genetic barriers to cross-species transmission and intra-host evolution of lentiviruses will have
implications for efforts at achieving a functional HIV cure and for improved HIV therapies.

As an obligate intracellular parasite HIV, like all viruses, relies on host-encoded factors to
complete its life cycle inside the host cell. However, the virus must also evade recognition by
specialized host factors that have evolved specifically to defend cells against viral invasion.
Some of the antiviral genes are part of an Interferon-induced transcriptional program that is
deployed by the host on sensing of a viral invader. A comprehensive description of host genes
that block primate lentiviruses from successfully infecting human cells has evaded
characterization with current methodologies. In Aim 1 we will describe the mechanism of
inhibition of HIV-1 and related primate Simian Immunodeficiency Virus (SIV) variants by
TRIM34, a TRIM5 paralog we have discovered to specifically target HIV-1 capsid variants and
SIVs. In Aim 2 we will explore the role of TRIM34 as a broadly acting antiviral gene in primates
as well as the potential role of TRIM34 in contributing to limiting the evolution of HIV-1 infections
within the host. In Aim 3 we will perform a whole-genome HIV-CRISPR screening approach to
find the Interferon-Stimulated Genes (ISGs) that inhibit HIV-1 capsid mutants. In addition to
defining a novel HIV restriction by TRIM34 in human cells, this work has the potential to
discover novel host antiviral genes that limit SIV and HIV strains from replicating in human cells.
Follow-up studies will be focused on furthering our understanding the mechanism of action of
genes of interest as well as mechanisms of viral antagonism or escape. Ultimately, manipulation
of these factors could be important for approaches aimed at achieving a functional HIV cure.

NIAID

05/31/24

R01 AI147877

FHCRC

Beronja,Slobodan

Selective mRNA translation in developmental disorders

RASopathies, a family of developmental disorders marked by abnormal tissue growth, represent the leading
cause of congenital abnormalities associated with significant morbidity. Using a novel mouse model and state-
of-the-art genetic and biochemical approaches we show that aberrant growth is driven by a specific subset of
translationally regulated mRNAs, which fundamentally impact stem cell biology. Our proposal will
mechanistically dissect the interplay between RAS, the translation apparatus, and progenitor cell regulators,
which will provide invaluable insights into the causes of RASopathies and inform new therapeutic strategies.

RASopathies cause the majority of congenital disorders affecting nearly 1 in 1000 individuals. In particular,
mutations in RAS-MAPK pathway genes lead to distinct pathologies including craniofacial dysmorphology,
mental impairment, musculoskeletal defects, and a predisposition to cancer. Although presentations may vary
between different mutations, nearly all RASopathies share common skin growth abnormalities. At a genetic
level, germline mutations to RAS pathway members including Hras and Kras are known to cause these defects
which are best exemplified by Costello, Noonan, and Cardiofaciocutaneous syndromes. For years,
comprehensive interrogation of RAS in development has been limited to genome-wide studies of DNA and
RNA. While important, these investigations have left translation-based mechanisms largely untouched. This is
remarkable in light of emerging evidence that developmental disorders, such as Diamond-Blackfan Anemia
and Schwachman-Diamond and Treacher Collins syndromes (reviewed in Tahmasebi et al., 2018), are
causally linked to impairments in the translation apparatus. Thus, our current knowledge of the mechanistic
basis of RASopathies is incomplete, which is a barrier to therapeutic innovation. Our long-term goal is to
uncover the mechanism of Ras-mediated tissue growth, which will ultimately yield innovative therapies to
restore normal tissue homeostasis without compromising housekeeping functions during development. Using
skin as a defined model of tissue development we have discovered that hyperactive Hras simultaneously
drives specialized proliferation and differentiation programs by rewiring the translation initiation machinery
through eIF2B5. Utilizing state-of-the-art in vivo genetic screens pioneered in our laboratory, we have
determined the regulon of genes that eIF2B5 governs to impact self-renewal and cell fate choice. Remarkably,
these mRNA networks are clearly demarcated by their function with ubiquitination emerging as a key regulator
of cellular differentiation. As such, we hypothesize that activation of Ras promotes translation of a subset of
mRNAs that support non-physiological tissue growth during development, where increased stem cell
proliferation is balanced by their loss through differentiation into post-mitotic progeny. In this proposal we will
use a confluence of in vivo models, intra-vital microscopy, and newly developed cellular and molecular assays
to delineate how the interplay between RAS and eIF2B5 influences tissue dynamics. We will accomplish the
following Aims: 1) Uncover how eIF2B5-dependent ubiquitin ligases directs progenitor renewal and fate choice;
and 2) Elucidate how activated Hras and eIF2B5 direct mRNA specific translation to regulate progenitor
renewal. Collectively, the successful completion of our Aims will provide a new understanding of cellular and
molecular principles that support Ras-driven non-physiological growth during development. Ultimately, these
new insights will inform the development of novel therapeutics, which can differentially inhibit the pathologic
impact of Ras mediated tissue imbalance while maintain homeostasis which is essential for life.

NIGMS

05/31/24

R01 GM135362

FHCRC

Bradley,Robert K

Functional and molecular basis of ineffective erythropoiesis in SF3B1-mutant myelodysplastic syndromes

Myelodysplastic syndromes (MDS) are diseases that are characterized by ineffective production of blood.
Many patients with MDS carry a mutation affecting the SF3B1 gene, which encodes a protein that is important
during a molecular process called RNA splicing. We propose to study SF3B1 mutations in order to determine
why they cause MDS and discover novel opportunities for treating this disease.

The majority of patients with myelodysplastic syndromes (MDS), a heterogeneous group of blood disorders
characterized by ineffective and clonal hematopoiesis, carry a somatic mutation affecting an RNA splicing
factor. The most commonly mutated splicing factor is SF3B1, a core component of the spliceosome that is
preferentially mutated in MDS with ring sideroblasts (MDS-RS). Although SF3B1 mutations are among the
most common genetic lesions in MDS, they are nonetheless relatively poorly understood. Our incomplete
understanding of SF3B1 mutations is due in part to the absence of a model system that recapitulates hallmark
disease phenotypes, including ring sideroblast formation and ineffective erythropoiesis. As a consequence, it is
unclear how SF3B1 mutations alter RNA splicing mechanisms, which specific mis-spliced genes drive hallmark
disease phenotypes, and whether SF3B1-mutant cells can be killed by targeted therapies.
Here, we propose to elucidate the functional basis as well as mechanistic and phenotypic consequences of
SF3B1 mutations in MDS-RS. Our team consists of a stem cell biologist with expertise in hematologic disease
modeling (Doulatov), a basic scientist with expertise in RNA splicing and functional genomics (Bradley), and a
physician-scientist with expertise in erythropoiesis and heme biology (Abkowitz). In preliminary studies, we
generated MDS-RS patient-derived induced pluripotent stem cells (iPSCs) that recapitulate hallmark disease
phenotypes during erythroid differentiation, identified specific mis-spliced genes that contribute to ineffective
erythropoiesis, and performed functional genomic screens to identify molecular vulnerabilities of SF3B1-mutant
cells. We propose to build on those preliminary studies as follows: Aim 1, Define the molecular consequences
of SF3B1 mutations for mRNA splicing, stability, and translation; Aim 2, Determine the functional basis of ring
sideroblast formation and ineffective erythropoiesis in SF3B1-mutant MDS-RS; Aim 3, Identify therapeutic
opportunities for treating MDS-RS with SF3B1 mutations. The significance of these studies is that they will
elucidate the mechanistic and functional consequences of SF3B1 mutations in MDS-RS. The health
relatedness is that the proposed work may identify new opportunities for treating MDS by specifically targeting
SF3B1-mutant cells. As the incidence of MDS is rising and patients with SF3B1-mutant MDS-RS face life-long
transfusion burdens and associated morbidity and mortality, there is a public health need to develop new
therapies for this disorder.

NHLBI

05/31/24

R01 HL151651

FHCRC

Warren,Edus Houston

Dissecting the Kaposi Sarcoma Tumor Microenvironment at the Single Cell Level

Kaposi sarcoma (KS) is a cancer that occurs frequently in individuals with HIV infection, and often leads to death
despite treatment. It is known that the various cell types that comprise a tumor – including the immune cells that
play a role in fighting the cancer – play an important role in determining tumor response for several cancers, but
little is known about how these cells in KS tumors and how HIV infection might influence the make-up and
function of these cells. This study will define the composition of the cells that comprise KS tumors, their function,
and the targets of effective immune responses against KS tumors, which could ultimately lead to new therapies
for the treatment of KS.

Kaposi sarcoma (KS) is among the most common of HIV-associated malignancies worldwide, and a leading
cause of cancer morbidity and mortality in sub-Saharan Africa (SSA). Although KS can occur in HIV-uninfected
individuals, particularly in SSA – termed “endemic” KS – HIV infection increases the risk of KS several thousand-
fold. KS is a unique tumor whose pathogenesis involves chronic infection with HHV-8, disordered angiogenesis,
and inflammation. Increasing evidence points to the importance of the interaction of these various components
of the tumor microenviroment (TME) in driving tumorigenesis and response to treatment in a range of
malignancies. However, our understanding of the composition and function of the KS TME is limited. Detailed
study of the KS TME in vivo will elucidate mechanisms of KS pathogenesis, including the unique role of HIV
infection, could elucidate fundamental aspects of tumor biology and help identify new therapeutic approaches.
Since 2011 our team has been conducting a comprehensive prospective cohort study (“HIPPOS”) of KS in
HIV+ and HIV- adults initiating treatment at the Uganda Cancer Institute in Kampala, Uganda. Subjects enrolled
on this study provide blood samples and serial tumor biopsies and are followed for up to 1 year to rigorously
define treatment response and clinical outcomes. Our results to date, based on study of over 160 KS subjects,
suggest possible differences in the clinical outcome of HIV+ and HIV- KS subjects in Uganda, and have provided
provocative preliminary data on the virology and immunology of KS in Uganda. Our team has recently developed
and implemented a protocol for the isolation of viable single cells from KS tumors – including both tumor cells,
tumor-infiltrating lymphocytes (TIL), and macrophages – that offers the unique opportunity to extend our studies
of the role of HIV in the development and progression of KS to the single-cell level. In this application we propose
to define the role of HIV in the KS tumor niche by performing comprehensive molecular profiling of isolated single
tumor cells, TIL, and macrophages from serially acquired KS tumors from 20 HIV+ and 20 HIV- adults. The
specific aims are:
1) To compare the transcriptional profile of KS tumor cells from HIV+ and HIV- subjects.
2) To perform serial transcriptional profiling of immune cells in KS tumors from HIV+ and HIV- adults, and to
determine if changes in their transcriptional profiles are correlated with suppression of HIV and with treatment
response.
3) To define the antigenic specificity of candidate CD8+ “public” HHV-8-specific infiltrating T cells in KS tumors
that are associated with superior treatment response.
The results of these studies will provide unprecedented insights into the role of HIV in the pathogenesis of KS
and its response to treatment and could lay a foundation for antigen-specific immunotherapy of KS.

NCI

04/30/24

R01 CA239287

FHCRC

Walter,Roland B

Development of 211Astatine-Conjugated Anti-CD45 Antibody-Based Conditioning for Hematopoietic Stem Cell Gene Therapy and Editing

Gene replacement and editing therapies have emerged as promising strategy to treat hemoglobinopathies but
existing approaches depend on myeloablative doses of total body irradiation or alkylating agents, both carrying
the risk of significant short-and long-term toxicities, for conditioning to facilitate engraftment of hematopoietic
stem and progenitor cells (HSPCs). To address this critical limitation, we here propose to develop a less toxic
conditioning regimen that is based on radioimmunotherapy (RIT) with anti-CD45 antibodies conjugated to the a-
emitting radionuclide astatine-211, exploiting Fc engineering to further minimize non-specific RIT toxicities. As
we are interested in rapid clinical translation of our findings, we will test this novel transplantation approach in
our established nonhuman primate model of autologous transplantation of gene-edited HSPCs.

The inherited disorders of hemoglobin (Hb) are the most common monogenic diseases worldwide and, even in
developed countries, associated with substantial morbidity and shortened life expectancy. Allogeneic
hematopoietic cell transplantation (HCT) is clinically pursued as a means to treat the underlying cause of these
disorders – the genetic defect in the patients’ hematopoietic stem and progenitor cells (HSPCs). However, this
approach is limited by the availability of HLA-matched donors in the majority of patients and associated
immunological complications. Use of autologous HSPCs either transduced with a functional b-hemoglobin gene
or modified with recently-developed genome-editing technologies would overcome the current limitations of
allogeneic HCT. In particular, the recapitulation of naturally-occurring hereditary persistence of fetal hemoglobin
(HPFH) mutations in HSPCs using gene editing can, in principle, reverse the clinical phenotype of these
disorders. However, just like with allogeneic HCT, there is still need for conditioning to facilitate engraftment of
these cells. To date, this is accomplished with g-beam total body irradiation (TBI) or alkylating agents such as
busulfan which carry the risk of significant toxicities including infertility, growth retardation, and – as has already
been reported – secondary malignancies. Thus, a critical remaining factor for next-generation transplant
approaches and gene therapy/editing will be the development of nongenotoxic conditioning regimens that have
minimal toxicity and allow robust engraftment of allogeneic or modified autologous HSPCs. One promising
strategy is the use of radioimmunotherapy (RIT) with a-emitting radionuclides conjugated to antibodies targeting
CD45, an antigen expressed on almost all hematopoietic cells except platelets and erythrocytes and some of
their progenitors. Compared to b-emitters, a-emitters deliver a higher amount of energy over just a few cell
diameters for potent, precise, and efficiently targeted cell kill and minimized toxicity to non-targeted surrounding
cells. With a half-life of 7.2 hours, astatine-211 (211At) is ideal for patient application. Based on our previous
studies in dogs demonstrating that 211At-anti-CD45 RIT can replace g-beam TBI as conditioning before allogeneic
HCT, we are currently using 211At-anti-CD45 RIT in patients with active hematologic malignancies. We now plan
to develop 211At-anti-CD45 RIT as conditioning before autologous transplantation of gene-modified HSPCs for
people with hemoglobinopathies, exploiting Fc engineering of antibodies to further minimize non-specific
toxicities associated with RIT. We hypothesize that optimized 211At-anti-CD45 RIT will enable engraftment of
autologous HSPCs edited with CRISPR/Cas9 at the g-globin gene locus to reproduce HPFH mutations and have
significantly less off-target toxicities and better tolerability than the standard conditioning with high-dose g-beam
TBI. As we are interested in rapid clinical translation of our findings and have already collected substantial data
demonstrating feasibility, we will test this hypothesis in our established nonhuman primate model of autologous
HCT for hemoglobinopathies.

NHLBI

04/30/24

R01 HL151765

FHCRC

Till,Brian

Mechanisms of treatment failure in chimeric antigen receptor T cell therapy

Patients with non-Hodgkin lymphoma (NHL) who relapse or have refractory disease have a poor prognosis
with standard treatments, but immunotherapy using genetically modified T cells that recognize and kill
lymphoma cells holds great promise. However, fewer than half of patients with relapsed NHL achieve durable
complete remissions following treatment with chimeric antigen receptor (CAR) T cells, and the reasons for this
are not understood. We propose a plan to discover why this treatment fails in many patients, using specimens
collected in a recently initiated CAR T cell clinical trial, insights that will help to make this treatment more
effective in the future.

Adoptive immunotherapy with genetically modified T cells expressing a chimeric antigen receptor (CAR) is
highly promising for B-cell malignancies. However, fewer than half of patients with relapsed non-Hodgkin
lymphoma (NHL) achieve durable remissions following treatment with CD19-targeted CAR T cells. In some
cases this results from target antigen loss or rejection of the infused cells due to immunogenic murine CAR
components, but in most cases the causes of treatment resistance or relapse after an initial response remain
poorly understood. We propose a plan to elucidate the reasons for treatment failure in a recently initiated CAR
T cell clinical trial by carefully evaluating biological features of the tumor and tumor microenvironment before
and after CAR T cell therapy as well as phenotypes of patient T cells and infused CAR T cells.
As part of our phase I/II clinical trial of a fully human 3rd generation CD20-specific CAR in patients with
relapsed or refractory B-cell NHL (funded by a separate source), all patients undergo mandatory tumor
biopsies before and after treatment. This will allow us to discover biological characteristics predictive of
responsiveness to treatment, and to evaluate adaptive changes in the tumor over time to reveal the
mechanisms of immune escape leading to relapse. We will employ a step-wise approach using state-of-
the-art methodologies, including multicolor flow cytometry, single-cell RNA sequencing and gene expression
profiling, and multiplex immunohistochemistry. We have assembled a world class team of investigators that will
evaluate the potential obstacles to successful therapy, including tumor entry barriers, tumor infiltration by
suppressive cells, CAR T-cell exposure to inhibitory ligands or secreted proteins, and CAR T-cell exhaustion.
There is a robust body of preclinical data demonstrating that less-differentiated T cell subsets impart
superior in vivo expansion, persistence, and anti-tumor efficacy, compared with more differentiated T cell
subsets. Recent data from a small trial suggests that the frequency of these less-differentiated CD8+ cell
subsets before and after CAR T cell manufacturing correlates with clinical responses in patients with chronic
lymphocytic lymphoma receiving CD19-targeted CAR T cells. These results have important implications, but
must be validated in other settings. We will quantify less-differentiated T cell subtypes prior to leukapheresis,
as well as in the infused CAR T cell products, and correlate these characteristics with anti-tumor responses
and in vivo expansion and persistence.
We anticipate that these correlative studies will yield critical insights into the reasons why CAR T cell
therapy is successful for some NHL patients but not others. We are hopeful that our findings will help to guide
patient selection and counseling, and inform future strategies to overcome these obstacles through improved
cell manufacturing technologies, CAR vector design, and/or combinatorial adjuvant therapies, not only for
CD20-specific CAR T cells, but also for CAR T cell therapy for other targets.

NCI

01/31/24

R01 CA230520

FHCRC

Moens,Cecilia B

Discovery of sensorimotor connectivity mechanisms in a continuous topographic map

Reflexes are simple, unconscious and highly specific motor responses to sensory stimuli. Sensory and motor
neurons that participate in different reflexes are spatially intermingled in our complex brain, and it is unclear
how specific sensory and motor neurons are selectively connected into the same circuit inside the brain. We will
address how reflex-specific sensory-motor circuit is built by using the vagus system of the larval zebrafish, a
powerful system for neuroscience research due to its genetic tractability that permits efficient gene manipulation,
and optic clarity that enables live-neuron imaging.

The long-term goal of this project is to understand the developmental mechanisms underlying neural connectiv-
ity within sensorimotor reflex circuits in our brain. Reflex circuits enable individual sensory inputs to elicit func-
tionally appropriate stereotyped motor outputs, suggesting fine-scale connection specificity between the sensory
and motor systems. However, in the brain, neurons responsible for different functions are often continuously
aligned on topographic maps, with functionally different neurons being intermingled at the boundary regions
between functional groups. It is poorly understood how functionally different neighbors on a topographic map
are distinguished during reflex circuit development so that they can invariably generate appropriate responses
to sensory information. We have established the vagus nerve in larval zebrafish as an efficient system in which
to address this long-standing mystery in brain circuit development. The vagus nerve exits the hindbrain and
branches widely to innervate the pharynx, larynx, stomach, heart and other visceral organs. This nerve carries
both sensory and motor axons, each of which participates in one of several polysynaptic reflex circuits including
the pharyngeal reflex and baroreflex. Our group has discovered that vagus motor neurons and sensory axons are
co-organized in a continuous topographic map that is detectable within the larval zebrafish hindbrain. Our pre-
liminary data support that local sensory inputs to the vagus sensory system selectively activate functionally ap-
propriate groups of vagal motor neurons with a strikingly fine-scale connection specificity that distinguishes
adjacent functionally different neurons. In order to investigate the mechanism underlying this functional sepa-
ration, we will investigate the contribution of neural activity in vagal motor neurons for fine-scale connection
specificity (Aim 1), and we will determine the structural basis of vagal reflex circuit refinement via transsynaptic
labeling (Aim 2). The successful outcome of these aims will provide neurophysiological and neuroanatomical
insights into fine-scale connection specificity at the level of entire sensorimotor reflex circuits in the vertebrate
brain. The larval zebrafish has emerged as a premiere system in which to study developmental neurobiology, and
the tools we develop in the vagus system will be generally applicable to questions about the role of neural activity
in other aspects of nervous system development.

NINDS

01/31/24

R21 NS124191

FHCRC

Stamatatos,Leonidas

Evaluating anti-idiotypic antibodies as novel vaccine candidates against HIV-1

n/a

No abstract available

NIAID

11/30/23

P01 AI138212

FHCRC

Lund,Jennifer

Tissue Regulatory T Cells in Mucosal Infection

While tissue Tregs have been studied in a number of tissues and disease settings, we propose here to for the
first time systematically define the characteristics and function of tissue Tregs in the context of mucosal infection
in both mice and humans. We predict that such cells have high clinical significance, as vaccine designs aimed
at inducing local T cell responses to combat infections and tumors may be subject to local Treg-mediated
regulation, and so a clear understanding of how such tissue-resident Tregs function is required in order to
rationally design vaccines and therapeutics best able to provide immune-mediated protection.

Regulatory T cells (Tregs) are a subset of CD4 T cells that are essential for the maintenance of peripheral
tolerance, yet their precise roles during infections remain an active area of investigation. It is well-documented
that following certain infections, Tregs are required to attenuate an overly robust immune response to prevent
collateral damage to self-tissues. However, we have demonstrated that removal of Tregs prior to infection with
Herpes Simplex Virus, type 2 (HSV-2), among other infections, results in delayed clearance of the pathogen,
suggesting that the presence of Tregs can be critical to facilitating an appropriately robust and protective immune
response. These differing results emphasize that the role played by Tregs during infections is context-dependent,
and thus we propose here to focus on the location of the cells and the time post-infection as key factors that
influence the role that Tregs play during mucosal virus infection.
Recent evidence suggests that there exists a distinct subset of Tregs known as tissue Tregs. These cells have
been best-characterized in skin and visceral adipose tissue, where they function to limit inflammation, though it
has been suggested that tissue Tregs in other locations function to prevent autoimmunity, to promote fetal and
graft tolerance, and to impair anti-tumor immune responses in various non-lymphoid tissues. However, despite
the hypothesized role of tissue Tregs in controlling local inflammation to prevent autoimmunity and
immunopathology, local immune responses are routinely and beneficially generated against mucosal infections,
often without excessive tissue destruction at the infection site, and we thus hypothesize that tissue Tregs are
involved in mediating this balance. Additionally, as effector T cell immune memories remain following infection
clearance, we hypothesize that regulatory memory also persists such that these tissue memory T cell
responses can be controlled under both homeostatic conditions as well as upon pathogen re-encounter to
promote local tissue integrity. Therefore, we propose to extend our investigations of the role of Tregs during
mucosal virus infection, now with a focus on the presence and consequences of tissue Tregs on anti-viral
immune responses in mice and humans. Tissue-resident memory T cells have been intensely studied in recent
years, and are now the basis for a promising new vaccine platform, so it is imperative that we understand how
such tissue T cell responses might be regulated in order to support tissue protection in the face of a robust
immune response.

NIAID

10/31/23

R01 AI141435

FHCRC

Li,Christopher I

Project 1: Discovery of novel tumor-tissue based predictors of lethal colorectal cancer by race/ethnicity

This first in class study will comprehensively evaluate differences in the molecular profiles of colorectal cancer
across four racial/ethnic groups, and a particularly unique feature of this study is its inclusion of Alaska Natives.
The data generated may inform novel primary and secondary prevention strategies, including the development
of novel therapeutics, that have the potential to reduce persistent racial/ethnic disparities in colorectal cancer
outcomes.

While improvements in colorectal cancer (CRC) outcomes have been made over the past several decades,
pronounced disparities in CRC incidence and mortality rates by race/ethnicity persist in the United States. In
particular, CRC incidence and mortality rates among Alaska Native people (a particularly understudied
population) are 124% and 148% higher, respectively, compared to the overall national rates, and among
African Americans they are 21% and 39% higher, respectively. Our primary overarching goal is to reduce
disparities in CRC mortality, particularly those experienced by Alaska Natives and African Americans, through
improving approaches to identify patients with aggressive CRC at diagnosis and discovering novel potential
therapeutic targets relevant to these populations. To address this goal, we will conduct a nested case-control
study that utilizes highly clinically annotated tumor tissue specimens that are readily available from several
resources. Cases are CRC patients who died of CRC within 5 years of their diagnosis, and controls are
matched CRC patients who survived at least as long as the duration between diagnosis and death of the case
that they are matched to. We will include four groups of patients defined by race/ethnicity: Alaska Native
people, African Americans, Hispanics, and non-Hispanic whites with 70 cases of lethal CRC and 140 CRC
controls from each group (total n=840). To discover novel prognostic markers, we will perform RNAseq
analyses on RNA extracted from formalin-fixed paraffin embedded tumor material. The specific aims of this
proposal are as follows: 1. Assess differences in gene expression profiles across four racial/ethnic groups and
their relationships to selected CRC risk factors; 2. Discover predictors of CRC mortality across and within four
different racial/ethnic groups; and 3. Characterize and evaluate differences in the immune and stromal cell
populations in the tumor microenvironment associated with CRC mortality, overall and by race/ethnicity. This
study will generate novel multi-ethnic high-dimensional data that we envision feeding into larger studies with
translational goals focused on advancing novel intervention targets for primary and secondary prevention, and
developing clinically useful predictors of poor CRC outcomes that can be used to guide clinical decision
making. This study is designed to generate findings that will be of particular benefit to underserved populations
as this work is aimed directly at reducing long-standing racial/ethnic disparities in CRC outcomes.

NCI

08/31/23

P20 CA252733

FHCRC

Peters,Ulrike

Project 1: Discovery of novel tumor-tissue based predictors of lethal colorectal cancer by race/ethnicity

NCI

08/31/23

P20 CA252733

FHCRC

Baker,Kevin Scott

An INteractive Survivorship Program to Improve Healthcare REsources [INSPIRE] for Adolescent and Young Adult (AYA) Cancer Survivors

Adolescent and young adult (AYA) cancer survivors, live with emotional distress and health care demands that
exceed those of their peers, but can have difficulty accessing the care they need. In a 5-site national study,
this research will adapt a tested, personalized, interactive digital and telehealth survivorship program to the
needs of AYA survivors. We will test the AYA-focused survivorship intervention in a randomized controlled trial
to see if it improves emotional distress and adherence to health care guidelines after 3 and 12 months when
compared to AYA survivors who receive access to a website of annotated links to existing survivorship
resources followed by delayed access to the digital survivorship program after 12 months.

The proposed adolescent and young adult (AYA) mixed methods project will meet the goals and scientific scope
of the U01 by refining and testing a scalable intervention to mitigate and manage adverse long-term outcomes
in AYA cancer survivors. By improving access to survivorship resources, health literacy, self-management skills
and support, the intervention is designed to improve adherence to AYA healthcare guidelines and reduce cancer-
related distress. The project leverages an existing survivorship informatics infrastructure, an established
multicenter survivorship research network, and our experiences with a randomized controlled trial (RCT) with
AYAs and a digital and telehealth program initially developed for hematopoietic cell transplant survivors. This
research fills a critical gap in care for AYA survivors by providing a novel, personalized, technology-based self-
management program with stepped care telehealth. To be scalable, it uses a risk-based delivery model targeted
to AYA survivors with demonstrated need. The two-arm RCT will test the AYA-adapted INSPIRE digital program,
including an interactive mobile app/website and social media, adding stepped care telehealth for those with
continued distress, lack of survivorship healthcare literacy, or who are not using the digital program at 1 month.
In the active control arm, survivors receive access to a study website with existing AYA survivor resource links
followed by delayed access to the INSPIRE program. Specific aims are: 1) Among AYA survivors with elevated
cancer-related distress, determine the efficacy of a self-management program, INSPIRE, delivered by interactive
digital cross-device options and stepped care telehealth coaching, compared to the active control arm. 2) Among
AYA survivors with inadequate adherence to healthcare guidelines, determine the efficacy of the INSPIRE digital
and telehealth self-management program in improving overall healthcare adherence and cardiometabolic or
subsequent malignancy surveillance compared to the active control arm. 3) Examine mechanisms associated
with intervention efficacy including engagement with the online program, use of telehealth contacts, and
improved health self-efficacy, health literacy, AYA-specific impact of cancer, and barriers to healthcare; and
examine moderators of intervention engagement or response including cancer clinical factors, health status, and
sociodemographic and environmental factors. 4) Define costs of maintaining and delivering components of the
INSPIRE program if delivered in a scaled-up model of care. An exploratory aim will assess dissemination
potential and define risk models associated with distress, poorer health status or healthcare nonadherence. Aims
will be accomplished with a two-arm, multicenter, risk-stratified RCT of N=980 AYA survivors of leukemia,
lymphoma, colorectal cancer, melanoma, or sarcoma, age 18-39, and 1-5 years post-diagnosis. The primary
outcome is reduced distress at 3 months; secondary outcomes are improved adherence and reduced distress
at 12 months. If effective, the program is positioned for accelerated dissemination/implementation to improve care
for AYA survivors by using a scalable informatics-based administration and largely digital intervention program.

NCI

08/31/23

U01 CA246659

FHCRC

Simon,Julian A

Project 3: Engaging Navajo Elementary Schools in a Randomized Controlled Trial: Yéego! Healthy Eating & Gardening

n/a

Engaging Navajo Elementary Schools in a Randomized Controlled Trial: Yéego! Healthy Eating &
Gardening
Project Leaders: Kevin Lombard, NMSU; Shirley Beresford, India Ornelas; Fred Hutch
[The content of this Pilot Proposal is identical in the NMSU and Fred Hutch proposals]
/
Navajo families are at increased risk for obesity, diabetes and cancer in part due to low fruit and vegetable
consumption. The proposed study aims to increase fruit and vegetable consumption among Navajo families
through a school-based intervention focused on gardening and healthy eating. The study builds on our
longstanding collaboration with the Navajo Nation through which, we have shown that gardening is a culturally
appropriate strategy for improving healthy eating in this population. Our previous research has highlighted the
importance of family in shaping health behaviors, and a desire to protect the health of the next generation by
encouraging healthy eating habits. We have developed and are currently pilot testing a healthy eating and
gardening curriculum to promote both gardening and healthy eating among Navajo elementary school children.
The intervention was developed based on social cognitive theory, literature on previous school-based
gardening interventions, and our own formative work in Navajo communities. The integrated intervention
comprises 1) a 24-unit curriculum on both healthy eating and gardening, and 2) a school garden, and aims to
build students’ self-efficacy to grow and eat fruits and vegetables, as well as making other healthy food
choices. In the new proposed study, we will estimate the efficacy of the intervention using a small, randomized
delayed intervention comparison trial in two Navajo Nation Chapters (towns), involving about six schools in all.
During the first year, we will identify and engage schools in the Navajo Division of Education system interested
in participating in the trial. We will match schools within each town, and randomize schools within towns to
intervention and delayed intervention groups. In the second year, we will implement the integrated intervention
in the intervention schools, and will assess changes in outcomes over the school year among a sample of the
elementary school children in grades 3 and 4 at each school (in both the intervention and comparison schools)
as well as their parent. The primary outcomes are child measures: a) tendency to choose vegetable and fruit
items to eat; b) healthy eating score from a Navajo foods picture sort; c) gardening at home and d) self-efficacy
to garden. Analyses estimating the intervention effect will use the differential change over this school year
between the two groups of schools, adjusting for school level clustering. We will also assess process
outcomes, including recruitment, retention and intervention fidelity. After the primary evaluation is complete,
the delayed intervention group will receive the curriculum intervention in the second school year. An additional
assessment for secondary analysis purposes will occur at the end of that year. Continued engagement and
skill strengthening will occur through quarterly parent workshops for all participating schools. This proposal is
timely as the Navajo Nation consider their policy on dietary and activity benchmarks for schools as part of a
rounded curriculum.

NCI

08/31/23

U54 CA132381

FHCRC

Diab,Ahmed M

Targeting the DNA Damage Response in HPV+ Head and Neck Cancer

HPV+ HNSCC is a growing health burden in the US with numerous treatment-related morbidities, highlighting
the need for novel targeted therapies. Inhibition of the cell cycle kinase WEE1 is a promising therapeutic strategy
for HNSCC, and for HPV+ HNSCC in particular. This proposal seeks to investigate how HPV compromises DNA
damage responses to WEE1 inhibition in order to identify novel synthetic lethal interactions with WEE1 in HPV+
HNSCC.

Human papilloma virus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) is a growing public
health burden and has already surpassed cervical cancer as the most common HPV-related malignancy in
the United States. While HPV+ HNSCC patients have generally good survival, they suffer from life-long
chemoradiotherapy-related morbidities. Current data is insufficient to inform de-intensification of standard
chemoradiotherapy or the development of targeted therapies. My ultimate goal is to understand the
mechanisms by which HPV disrupts DNA damage response (DDR) signaling during HNSCC development,
and to thereby inform the rational design of new targeted therapies. In considering new strategies to effectively
control HPV+ HNSCC, I noted that HPV’s oncogenic E6 and E7 proteins abrogate tumor suppressor pathways
and impair DDR signaling to cause genomic instability. The Mendez lab and others have established DDR
kinase WEE1 inhibition via the specific inhibitor AZD1775 (WEE1i) as a new therapeutic strategy in HNSCC,
and that HPV+ HNSCC tumors are hypersensitive. WEE1 inhibition causes S-phase replication stress (RS)
and irreparable DNA damage. Combined with genotoxic chemotherapy (e.g., cisplatin), WEE1i abrogates the
G2/M checkpoint and causes premature mitosis. I recently showed that HPV16 E6/E7 oncoproteins sensitize
HNSCC cells to WEE1i monotherapy through activation of a FOXM1-CDK1 circuit that drives mitotic gene
expression and DNA damage. I also showed that elevated basal FOXM1 activity predisposes HPV+ HNSCC
to WEE1i-induced toxicity. Next, I used an RNAi genetic screen to identify RS and DDR targets that synergize
with WEE1i; based on my findings to date, I hypothesize that disruption of RS and DDR pathways by E6/E7
provide exploitable vulnerabilities for a combination targeted therapy that also includes WEE1i. I plan to clarify
the mechanisms by which HPV sensitizes cancer cells to WEE1i-induced replication failure (Aim 1) and
compromises DNA repair pathways upon WEE1 inhibition (Aim 2). I will use murine cancer models to test
novel therapeutic combinations for targeting RS/DDR defects in HPV+ HNSCC and identify the situations in
which they are most effective. In parallel, I will use a targeted quantitative proteomics approach to determine
the E6/E7-specific RS and DDR responses to WEE1i, and multi-panel flow cytometry to determine the WEE1i-
associated changes in the immune landscape of E6/E7-driven tumors in immunocompetent mice. This award
will help me develop my scientific ideas and increase my competency in working with the mouse models that
faithfully recapitulate human cancer. The scientific advances that I make during this training period will be
critical to my ultimate goal of establishing an independent research program that focuses on how HPV drives
HNSCC development and how HPV+ HNSCC might be more safely and effectively treated.

NIDCR

06/30/23

K99 DE030194

FHCRC

Moens,Cecilia B

Topographic mapping by cranial motor neurons

We propose to study a fundamental problem in brain development: how neurons project their axons to a target
field such that the spatial relationships amongst the neuron cell bodies are reflected in the spatial arrangement
of their targets; a process called topographic mapping. The neuronal projection whose topographic map we
study is that of the vagus efferent neurons in the brainstem, which control speech and swallowing as well as
autonomic functions like heartbeat, breathing and digestion. There is growing interest in the vagus nerve as a
target for treating a variety of cardiac and gastrointestinal disorders, so it is important to understand the
spatial relationships between vagus neurons and their targets.

Topographic neural maps are ordered connections between the brain and the periphery in which
spatial coordinates in the projecting field are represented in the target field. Topographic maps
are a common motif in vertebrate nervous system organization and are critical for our ability to
perceive the world and accurately respond to it, so their development is of fundamental interest
to neurobiology. Examples of topographic maps are in the ordered projections of retinal neurons
to visual centers in the brain and in the projections motor neurons in the spinal cord to specific
target muscles in the limb. Cranial motor neurons in the vertebrate hindbrain exhibit a topo-
graphic relationship with the pharyngeal arch-derived muscles in the head periphery that they
innervate, whereby more anterior neurons innervate more anterior pharyngeal arches. Using the
transparent zebrafish model for live imaging and transplantation of single motor neurons, we
have found that a topographic map is detectable within the vagus (cranial nerve X) motor pro-
jections to the posterior pharyngeal arches in the 3-day embryo, and have discovered two paral-
lel strategies that govern its formation: a Hox-regulated molecular mechanism and a novel tem-
poral mechanism in which timing of vagus axon initiation is regulated to match the sequential
development of the pharyngeal arch targets. We call this a “temporal matching” model as distin-
guished from classical spatial matching (chemoaffinity) models of topographic mapping. The
overall aim of this proposal is to discover how the Hox-regulated and temporal matching mech-
anisms together regulate topographic mapping. We will identify the molecular mechanism by
which timing of vagus axon initiation is spatially regulated in vivo and how it is matched to the
timing of pharyngeal arch development in Aim 1. We will identify the guidance pathway that is
regulated by hox5 genes in Aim 2. Finally, we will determine how the two mechanisms are coor-
dinately regulated by spatial cues in Aim 3. Ultimately our goal is to elucidate novel mechanisms
of topographic mapping, their regulation and integration during development.

NINDS

06/30/23

R01 NS109425

FHCRC

Li,Christopher I

Precision disease prevention via somatic mutagenesis enumeration (PREDICTION)

Monitoring the frequency of DNA mutations in humans has the potential to serve as early markers of exposure
and increased risk of developing disease, but at present we lack validated, readily measured predictors that
are clinically useful. The overarching goals of this proposal are to characterize the spectrum and burden of
spontaneous and induced somatic DNA mutations in humans at unprecedented depth and fidelity, and to
evaluate the utility of mutagenesis as a marker of exposure and predictor of risk for developing a disease.

In order to realize the promise of precision medicine, better biomarkers are needed to guide clinical decisions.
New markers are needed to predict individuals at risk for developing a disease and thus may benefit from a
particular intervention. Focusing on markers of mutagenesis is a promising strategy given that mutation is the
ultimate source of all genetic variation, and mutated genes drive a number of important pathogenic processes,
such as cancer. However, key elements are missing for evaluating mutagenesis in relation to disease risk.
Absent are large-scale, well-characterized cohorts with high quality exposure data; populations with serial
samples that have been collected and stored using uniform protocols; and the ability to robustly monitor
somatic mutation in humans. We overcome these issues through a synergistic collaboration among clinicians,
basic, computational, biostatistical, and population scientists that leverages an exceptionally sensitive next
generation sequencing (NGS)-based mutational assay, which we recently developed; and its application to
biological samples of the highest quality from both a clinical trial and the landmark Women’s Health Initiative
(WHI) study. Our overarching goal is to contribute to the realization of the promise of precision prevention
through completion of the following specific aims: 1) Monitor the kinetics of mutagenesis and selection across
the human genome to identify robust mutational targets; 2) Examine the utility of somatic mutation induction as
a biomarker of mutagenic exposure and its potential to stratify smokers that develop lung cancer versus those
that do not; and 3) Test the utility of monitoring somatic mutation rate as a susceptibility/risk biomarker to
identify individuals who will develop cancer. Overall, our proposal is innovative with respect to the technology
used and its application to highly curated human samples. This project will highlight the potential utility of
monitoring in vivo mutation induction to stratify cancer risk, providing a basis for directing medical intervention,
lifestyle changes (i.e. limiting mutagen exposure), early diagnosis, and/or the application of chemopreventive
measures with the potential to ultimately save lives.

NIEHS

06/30/23

U01 ES029516

FHCRC

Corey,Lawrence

Harnessing Tissue Resident Memory T Cells for Immunotherapy of Herpes Virus Infections

This proposal is directed at extending the observations that tissue resident memory (TRM) CD8+ T cells to
HSV-2 are the first line of defense for containing reactivated HSV-2 in genital skin. The antigenic targets of
these as yet uncultivable cells are currently undefined. Identifying these and including them in an
immunotherapeutic vaccine would likely enhance potential efficacy of an HSV-2 vaccine. We will identify and
construct HSV TRM cells from genital skin and cervix, and compare their antigenic specificity and functional
characteristics to PBMC derived T cells. These studies will help define if the TRM system can be harnessed to
treat/cure chronic viral infections.

Recent work has shown that tissue resident memory (TRM) CD8+ T cells are the first line of defense for
containing reactivated HSV-2 in genital skin. Deep sequencing of these unique cells has shown their TCRs
differ from the TCRs detected in PBMC of the same person sampled over time, suggesting they may be
directed to as yet unidentified HSV-2 antigens. As all current immunotherapeutic vaccines for HSV-2 are
designed based upon antigens derived from PBMC, defining the antigenic targets of TRM cells may enhance
the efficacy of novel immunotherapeutic approaches to HSV. We have recently identified TCRs of the α and β
chain of laser capture purified TRM from genital skin biopsies and utilized synthetic biology to create autologous
TCRs, and shown that some of these resident CD8+ T cells found in genital skin are HSV-2 specific and are
directed at targets not currently emphasized in immunotherapeutic approaches to HSV-2. This proposal utilizes
single cell emulsion based bar coding technology to identify in a high-throughput unbiased approach full-length
TCR α and β chains of CD8+ and CD4+ T cells with RNA/protein signatures of TRM cells in cervical and vulvar
tissue, as well as PBMCs of HSV-2 infected persons. We will use synthetic biology to create “reporter” T cells
that can then be evaluated for their HSV specificity using a novel HSV-1/HSV-2 vireome. Identification of these
cognate antigens of the TRM population is likely to yield new approaches for vaccine design.

NIAID

05/31/23

R01 AI134878

FHCRC

Salama,Nina R

Mechanisms and consequence of helical shape generation in Helicobacter pylori

Evident in its name, Helicobacter pylori is a helical rod shaped bacterium that colonizes the human stomach
causing clinical outcomes that range from mild gastritis to peptic ulcer and gastric cancer. We discovered
genes that promote helical cell shape through modification of the cell wall. Study of how these cell wall
modifications promote helical cell shape and the ability of the bacterium to live in its niche will help us better
understand how this bacteria causes disease and uncover new ways to inhibit infection.

Bacteria come in many shapes, which may enhance motility, biofilm formation, nutrient uptake, and
pathogenesis. However, these functional consequences of shape have not been well studied, owing in part to a
paucity of tools to manipulate bacterial cell shape. To probe how form (cell shape) drives function (radiation to
diverse niches), we must first understand how shape is generated. Bacterial shapes varying from spheres to
rods to helices all arise from the same cell wall polymer: peptidoglycan (PG). The PG wall surrounds the cell to
contain turgor pressure. The major hypothesis in the field holds that diverse shapes arise from different patterns
of PG synthesis. Indeed Escherichia coli, a straight rod, and Caulobacter crescentus and Vibrio cholerae, curved
rods, require cytoskeletal proteins to modulate their PG synthesis patterns. Mechanisms that create helical cells,
seen in multiple lineages of bacteria, have not been elucidated.
Helicobacter pylori has emerged as the leading model for the study of helical shape. This bacterium persistently
colonizes the human stomach causing chronic inflammation and clinical pathologies ranging from peptic ulcers
to gastric cancer, the world’s third leading cause of cancer mortality in 2012 [2]. We isolated mutants with stable
non-helical shapes, and our work demonstrating their defects in stomach colonization presented the first
experimental evidence for a link between cell shape and bacterial infectivity that has now been extended to other
bacteria (Vibrio, Campylobacter) [3-5]. However, we only have a cursory understanding of the importance of
shape in initial infection and do not understand how altered shape impacts long-term colonization, niche
acquisition, or host immune responses.
Furthermore, H. pylori’s strategy for maintaining helical shape differs significantly from bacteria studied thus far.
Five of our shape mutants map to confirmed PG hydrolases suggesting a model whereby helical shape arises
from structural modification of PG rather than modulation of PG synthesis [5-7]. Homologues of these hydrolases
can be found in several Proteobacteria classes, most of which are curved/helical, indicating that other bacteria
may also employ direct modification of the PG to achieve curvature and twist [5, 8, 9].
The main hypothesis that guides this proposal is that spatially localized PG hydrolases promote H. pylori
helical shape, which allows colonization of distinct niches from non-helical bacteria and underlies
persistent infection. Our collection of non-helical mutants provides unique opportunities to explore the
mechanisms of helical cell shape generation and maintenance in bacteria as well as the functional role(s) of cell
shape in niche acquisition and persistent colonization.
A more complete understanding of the causes and consequences of helical cell shape could elucidate
new therapeutic targets in H. pylori and other curved and helical pathogens, and will thus further the
mission of NIAID to understand and treat infectious diseases.

NIAID

05/31/23

R01 AI136946

FHCRC

Kemp,Christopher

Exploiting WEE1/p53 synthetic lethality as a novel therapy in head and neck cancer

Despite overwhelming evidence implicating p53 functional derangement in the biology and clinical outcome of
head and neck squamous cell carcinoma (HNSCC), there are no targeted therapies that capitalize on this
knowledge. Our preclinical studies using an in vitro/in vivo pipeline have identified a p53 synthetic lethal
interaction with several regulators of the G2/M cell cycle checkpoint. Our proposal will advance our
understanding of these mechanisms as we seek to credential AZD1775, a first-in-class WEE1 inhibitor, as a
novel targeted therapy in HNSCC.

This proposal addresses one of the most urgent needs in the clinical management of head and neck squamous
cell carcinoma (HNSCC), the development of targeted and less toxic therapies. Recent studies have shown that
50-60% of HNSCC tumors harbor mutations in the TP53 tumor suppressor gene with other studies
demonstrating that disruptive mutations in TP53 are associated with worse prognosis and survival. Additional
studies have linked HPV infection as a second pathway to p53 inactivation in HNSCC. Despite overwhelming
evidence implicating p53 functional derangement in the biology and clinical outcome of HNSCC, there are no
targeted therapies that capitalize on this knowledge. Towards this goal, we hypothesized that the driver
oncogenic/tumor suppressor mutations (i.e., TP53 loss) that confer dominant malignant phenotypes in cancer
cells also engender unique, exploitable vulnerabilities. Since p53 mutant HNSCCs are aggressive tumors
incapable of G1 arrest and with higher levels of genomic instability, these tumors rely on a functional G2/M cell
cycle checkpoint to repair the DNA damage that might occur as a result of this instability or through genotoxic
therapy. In support of this hypothesis, we identified p53 synthetic lethal interactions with several G2/M checkpoint
regulators using high throughput arrayed siRNA gene silencing against human kinases in p53-mutated HNSCC.
Moreover, treatment with AZD1775, a specific WEE1 inhibitor, blocked tumor growth as a single agent and
caused tumor regression when used in combination with cisplatin in p53 mutant HNSCC xenografts. To translate
these findings to the clinic, we opened a phase I clinical trial with AZD1775 in combination with neoadjuvant
weekly cisplatin and docetaxel in previously untreated, metastatic HNSCC patients. Building on these successes,
this proposal will provide needed mechanistic understanding on the biology of p53 alterations and G2/M reliance
in HNSCC, while providing translational data to advance WEE1 inhibition with AZD1775 as a novel therapy for
HNSCC. Therefore, in Aim 1 we will determine the mechanism(s) of growth arrest upon WEE1 inhibition in
HNSCC and determine how p53 inactivation affects this response. In Aim 2 we will identify novel sensitizers to
the WEE1 inhibitor AZD1775 to unveil novel synergistic and less toxic partners for combinatorial therapy. Lastly,
in Aim 3, we will leverage our ongoing phase I clinical trial to functionally evaluate the effects of AZD1775 on
tumor biopsies and relevant preclinical models established from patient biopsies before and after treatment to
correlate markers of WEE1 inhibition to p53 status and clinical efficacy for the first time in HNSCC. This new
knowledge will help inform which HNSCC might benefit most from this line of therapy and help advance AZD1775
into phase II clinical trials.

NCI

05/31/23

R01 CA215647

FHCRC

Paddison,Patrick J

Epitranscriptomic control of erythropoiesis

This grant will define the roles for mRNA methylation in controlling normal red blood cell development
and determine whether altered mRNA methylation contributes to the anemia of myeloid dysplasia syndrome.
The grant is directly relevant to public health because it will identify novel ways in which adult humans regulate
red blood cell development and whether these mechanisms can cause red blood cell loss in anemia-
associated human diseases.

We recently found that N6-methyladenosine (m6A) mRNA regulation plays an essential role in human red blood
cell development. We observed that genes coding for the m6A mRNA methyltransferase (MTase) complex,
including, METTL14, METTL3, and WTAP, are required for promoting erythroid gene expression programs in
human erythroid leukemia (HEL) cells and for the erythroid lineage specification in human primary, bipotent
hematopoietic progenitor cells (HPCs). Critically, we find that m6A mRNA marks promote the translation of >200
genes, many of which play a role in erythroid lineage gene expression, and differentiation. Both Diamond-
Blackfan anemia, and Myelodysplastic Syndromes are associated with mutations in these genes. The purpose
of this grant is to elucidate how this novel form of gene regulation contributes to normal and abnormal
erythropoiesis. To this end, in Aim 1, we test the hypothesis that m6A-mRNA marking promotes selective
translation of genes required for human erythropoiesis. In Aim 2, we test the hypothesis that m6A marking of
mRNAs coding for a network of SET domain histone/protein methyltransferase genes is critical for erythroid
lineage specification. We will create a resource of all m6A mRNA marking events during human erythropoiesis,
while examining the effect of m6A methylation on key erythroid transcriptional regulators (e.g., GATA1, IKZF1,
KLF2, and ZFPM1) and whether specificity factors play roles in lineage-specific marking events. In Aim 3, we
define the m6A-mRNA "methylome" for human adult erythropoiesis and hematopoiesis and ask whether m6A
mRNA methylation is altered in myelodysplastic syndrome (MDS).
Our interest in MDS stems from several observations. First, we found that m6A mRNA methylation occurs in 70
out of 104 genes associated with MDS, including 8 of the 10 most frequently mutated genes (e.g., TET2, SF3B1,
ASXL1, RUNX1, DNMT3A, ZRSR2, and STAG2). Second, the blockage of erythroid lineage formation in human
adult progenitors by inhibition of m6A MTase copies the phenotype of MDS-related anemias. Third, m6A-mRNA-
sequencing pilot studies of multiple MDS patient samples provided evidence of significantly lower mRNA marking
for networks of splicing genes and other key regulators of erythropoiesis (e.g., U2AF2 and KLF1). Approximately,
one-third of patients with MDS progress to acute myeloid leukemia (AML), with poor outcomes. Although multiple
somatic events have been implicated in MDS, including chromosomal abnormalities, mitochondrial (metabolic)
dysfunction, abnormal expression of pro-inflammatory cytokines, and dysregulated erythropoiesis, none have
been shown unequivocally to initiate or drive disease progression.
The proposed studies will define the role that m6A-mRNA methylation plays in regulating normal erythropoiesis
and determine whether altered m6A-mRNA contributes to the anemia of MDS.

NIDDK

05/31/23

R01 DK119270

FHCRC

Parkhurst,Susan M

Mechanoregulation of Cell Functions during Embryogenesis

Constructing complex embryo body plans requires the precise orchestration of a wide range of cell
and tissue behaviors. Mechanical signals and stresses have emerged as important regulators in this
process, although the means by which they do this are not well known. The proposed studies are
expected to provide new insight into the ways in which cells perceive and then respond to mechanical
cues during normal development, and the consequences of inappropriate regulation leading to
developmental disorders.

Sculpting complex morphological body plans requires the precise orchestration of biophysical and
biochemical cues to control the wide range of behaviors demanded of individual cells, as well as groups
of cells. Biomechanical forces at the cell level affect the functions of the cell cortex: the plasma
membrane and its underlying cortical cytoskeleton. These mechanical cues must be sensed by the cell,
then properly acted upon, with the failure to do so leading to abnormal development. Thus, cell
mechanoregulation is of fundamental developmental and cell biology interest and significant clinical
relevance. The general aim of this proposal is to delineate the contribution of mechanoregulatory
signals to cell functions required for normal development, and the consequences of their mis-regulation
leading to aberrant cell functions and/or developmental disorders. Drosophila provides an excellent,
genetically amenable, model in which to investigate these fundamental processes due to its
accessibility to dynamic in vivo imaging and the wealth of state-of-the-art developmental/cell/molecular
techniques and reagents available. Our long-term goal is to understand how mechanical cues are
sensed, then acted upon, by cells to guide their functions during embryogenesis. To this end, we
propose to use the forces generated by cellular wounding as an inducible system in which to study the
mechanical properties of the cell cortex, including membrane tension, cortical cytoskeleton dynamics
and the integration of these properties. The specific aims of this proposal are: 1) to determine the
nature of the scaffold at the embryo cortex involved in tension regulation, and 2) to elucidate the
mechanisms regulating actomyosin organization necessary for generating contractile forces within
cells. The information gathered in these studies will provide new insight into the mechanical
characteristics of the cell, as well as provide a better understanding of how the cell interprets the
intrinsic and extrinsic forces acting upon it to orchestrate complex functions and interactions.

NICHD

05/31/23

R01 HD095798

FHCRC

Lund,Jennifer

Immunogenetic determinants of HSV-2 infection and disease

Infection with genital Herpes Simplex virus-2 (HSV-2) is life-long, and there is currently no cure or vaccine despite
decades of effort. Through our proposed studies using the Collaborative Cross in conjunction with a mouse
model of vaginal HSV-2 infection, we expect to uncover novel genetic regions associated with HSV-2 shedding
and disease phenotypes, as well as with tissue-specific immune responses to infection. A deeper understanding
of how host genetics regulate HSV-2 disease outcomes and immunity at the mucosal pathogen portal of entry,
the genital tract, will not only pave the way for the generation of novel immunotherapeutic and prevention
strategies for HSV-2 infection, but also for other sexually-transmitted infections.

Infection with genital Herpes Simplex virus-2 (HSV-2) is life-long, and there is currently no cure or preventative
vaccine despite substantial efforts. Furthermore, current anti-viral drugs such as acyclovir do not fully eliminate
viral shedding or symptomatic genital ulcers for all patients, underscoring the need for new prevention and
therapeutic strategies. There is considerable variation in rates of symptomatic and asymptomatic shedding as
well as symptomatic disease between HSV-infected individuals, yet little understanding of the reasons underlying
this variability. It is hypothesized that environmental and life-style factors such as stress, as well as genetic
factors could play roles. Thus, we propose to use the Collaborative Cross in conjunction with a mouse model of
vaginal HSV-2 infection to uncover novel genetic regions associated with HSV-2 shedding and disease, as well
as with tissue-specific immune responses to infection. By defining host genetic regions that regulate these
infection and disease-related phenotypes, we hope to pave the way for the identification and subsequent
development of novel host-targeted and/or immune-based HSV therapies and prevention strategies that could
lessen the burden of this global infectious disease as well as other infections transmitted via a mucosal surface.
The Collaborative Cross (CC) is a population of recombinant inbred mouse strains with high levels of standing
genetic variation, and was designed to allow for studies of the association between allelic variation in one or
more genes and a phenotype of interest. We have successfully used the CC to screen for genetic loci involved
in West Nile virus infection susceptibility and disease as well as immune phenotypes. We now propose to
leverage our expertise with the CC as well as with the mouse model of HSV-2 to perform a screen of CC mouse
strains for HSV-2 shedding, disease, and tissue inflammation phenotypes. Further, we will assess post-infection
immune response phenotypes within both lymphoid tissues and tissue sites that are viral targets of disease, such
as the genital tract and the central nervous system (CNS). We will use this data to perform quantitative trait loci
(QTL) mapping to identify chromosomal regions associated with vaginal viral shedding rates and levels, virus-
associated disease such as the formation of genital lesions and mortality, and immune cell responses at different
times and in different tissues post-infection. Through this proposed work, we expect to identify novel HSV
susceptibility alleles that could inform the rational design of host-targeted HSV treatments and prevention
strategies. Additionally, it is increasingly recognized that immune cell phenotype and function can vary widely
based on tissue location. This study will identify genetic regions associated with immune responses to infection
in distinct tissue locations, including both lymphoid and mucosal tissues, to thus increase knowledge of the host
genetic regulation of tissue-specific immune cell function following infection.

NIAID

05/31/23

R21 AI152559

FHCRC