Projects

Overview

Li-Fraumeni Syndrome (LFS) is a cancer predisposition syndrome caysed by inherited mutations in TP53 which encodes the tumor suppressor protein p53. Classic TP53 mutations confer an 80-90% lifetime risk of multiple types of cancer. Currently, surveillance for malignancy involves frequent laboratory and radiologic studies that are imperfect measures of disease onset; therefore, more specific, less invasive biomarker-driven screening methods are needed, particularly in patients with lower risk or mutations. LFS research in our laboratory is multi-institutional; we have an ongoing prospectively enrolling biobank of LFS pediatric and adult patients from whom we collect tumor and normal tissues, peripheral blood mononuclear cells, and longitudinal plasma samples. 

Projects

1. Characterizing hypomorphic TP53 variants that lead to attenuated LFS phenotype: 

There are several TP53 mutations that are considered lower risk variants that lead to a more attenuated clinical phenotype, with fewer typical LFS tumors that have a later age of onset compared to classic loss of function variants. Our laboratory and others have described the lower risk Ashkenazi jewish founder mutation p.G334R. We have also characterized the clinical and functional phenotype of hypomorphic R181H and R181C variants of TP53 by identifying p53 tumor suppressive functions that are lost versus retained by the R181 mutants. There are several other variants that have conflicting reports of pathogenicity with unclear mechanisms of tumorigenesis in LFS that our laboratory is interested in studying. 

2. Genomic analysis of cell-free DNA from LFS patients for early tumor detection:

The high rate of cancer development in LFS patients coupled with frequent standard of care imaging make LFS patients an ideal cohort that will greatly benefit from multi-cancer early detection tests. Tumors at multiple stages shed small amounts of DNA fragments into the bloodstream that are present in cell-free DNA (cfDNA) as circulating tumor DNA (ctDNA). Our lab is implementing enzymatic methylation-sequencing (EM-seq) on cfDNA from LFS patient plasma, which provides data on DNA methylation, copy number profiles, nucleosome occupation, and other mutations from a single sequencing experiment. We are leveraging our longitudinally collected blood specimens from LFS patients to validate the genomic profile of cfDNA as biomarkers for early tumor development.

3. Understanding the immune response in LFS patients:

The immune system can produce T-cell responses to novel antigens (neoantigens) produced or stabilized by tumor alterations. Tumors in Lynch syndrome patients have shared neoantigens based on similar mechanisms of tumorigenesis and are proposed to produce shared clonal T-cell responses, which can be identified by T-cell receptor (TCR) sequencing of lymphocytes from the bloodstream. Whether LFS tumors produce shared neoantigens to induce a common T-cell response is presently unknown. Our laboratory aims to determine if common neoantigens and TCRs exist in LFS tumors and tumor infiltrating lymphocytes (TILs) respectively, and determine whether these T-cell alterations can be detected in the circulating lymphocytes in the bloodstream.

4. Delineating mutant p53 mediated tumorigenesis:

Approximately half of all human tumors in the general population develop a TP53 mutation as the cancer progresses. In LFS patients, the TP53 mutation is the initial hit that induces tumorigenesis. The exact mechanism by which p53-mutant initiated tumors occur is not understood. Using fresh biopsies and other surgical specimens, we aim to use single-cell RNA sequencing and organoid development to determine the mechanism of LFS cancer development in breast, prostate, and colon tissues.

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Funding

AnchorOverview

Recent studies have shown that 4-20% of prostate cancers are associated with germline mutations in DNA repair genes. Currently, the majority of data on DNA repair gene mutational rates has been studied in metastatic prostate cancer patients.  As a result, the National Cancer Care Network supports germline genetic testing in metastatic prostate cancer patients with a family history suggestive of the associated syndrome, but there is currently no indication for testing in patients with localized prostate cancer.  Prior studies suggest a lower rate of DNA repair gene mutation positivity in localized prostate cancer patients, but few explore the differences in gene mutation rate for subsets of localized prostate cancer, for example by Gleason score.

Our access to clinical genetic testing data and patient samples obtained from multiple patient populations in Penn Medicine and across multiple VA Health care systems, allows us to study mechanisms known genetic mutations (BRCA1/2, ATM, CHEK2, HOXB13, etc.) that increase the risk of developing prostate cancer in various patient populations. As we have expanded our efforts to improve clinical care of PCa patients with these mutations and across self-identified race. Identification of these alterations may have clinical implications for personalized treatment of patients with prostate cancer. In addition, identification of an inherited mutation in a cancer affected patient can have far-reaching beneficial effects on reducing morbidity and mortality in his family members via implementation of cancer screening and prevention strategies.

Projects

1. Mechanisms of Prostate Cancer Development in BRCA carriers:

Our current work aims to characterize prostate cancer development in BRCA2 carriers with PCa, through the use of our tissue biobanks (PTBB and VA-MAPP) we collect fresh and archival tissue from BRCA2 carriers as well as from clinically matched control patients. BRCA2 plays a critical role in the establishment of RAD51 filaments onto DNA resected ends to initiate strand repair and patients with BRCA2 germline mutations can lose protein function and therefore have genomic instability. Prostate cancer patients with BRCA2 mutations have more aggressive disease and increased risk of developing metastasis. Prostate cancer is mediated by androgen receptor (AR) signaling as perturbations in the pathway leads to prostate cancer development, however, how this pathway is regulated in BRCA2 carriers with prostate cancer is unknown. Through genomic approaches and patient derived disease modelling we hope to uncover mechanisms of cancer development and screen for targeted therapies to improve patient care in BRCA2 carriers.

  • Utilizing paired tumor-normal genomic profiling by amplicon based DNA sequencing to study the relationship of DNA repair gene mutations to Gleason score in localized prostate cancer patients in the Penn Medicine Biobank. Additionally, we are determining the association of a number of candidate genes with prostate cancer risk.
  • Utilizing fresh prostate tissues from localized PCa patients undergoing prostate needle biopsy or robotic-assisted laparoscopic prostatectomy to perform single cell RNA sequencing comparing cell type specific transcriptomic profiles of patients with and without BRCA2 germline mutations. This data will be used to compare normal and tumor prostate tissues from BRCA2 carriers to clinically matched non-carriers across self-identified race.
  • Utilizing archival prostate tissues from PCa patients to perform bulk RNA sequencing and targeted DNA sequencing of BRCA2 tumor and surrounding normal prostate tissue vs clinically matched control tissues.

2. Patient Derived Disease Modelling:

  • Utilizing fresh metastatic biopsies from PCa patients to generate patient derived prostate organoid models. Organoids developed from prostate cancer patients are characterized by western blot, RT-qPCR, and immunofluorescence for prostate specific markers (AR, KLK3, NKX3-1) and will be sequenced to validate prostate origin. Transfection of prostate organoids will be performed to model BRCA2 related prostate cancer and other prostate cancer risk genes.

3. Prostate Cancer Genotype-Phenotype Studies:

Utilizing large genetic biobanks (including MVP, AllofUs and Penn Medicine Biobank), we are interested in understanding genotype phenotype correlations of known prostate cancer genetic risk factors and discovery of novel prostate cancer risk genes, especially in individuals of African genetic ancestry.

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Overview

We leverage the resources of Penn Medicine Biobank, the Veterans' Affairs and Prostate Cancer Foundation Partnership, the VA National Precision Oncology Program and the VA Million Veterans' Program to understand how to better utilize germline genetic data and tumor genomic data in clinical practice.

Projects

1. Clinical utility of mutational signatures:

Despite advances in sequencing, current tumor genomic profiling still fails to identify a precision oncology target in the majority of cancer patients. In addition to tumor-driving mutations, somatic signatures that are consequence of aging, mutagen exposure, and germline deficient DNA-repair become part of the tumor genome. These signatures do not drive the tumor but are instead a genomic scar of oncogenic drivers. We have described unique signatures from whole genome sequencing of 400 tumor specimens from a diverse population cohort at the Veterans Administration (VA). Our laboratory is interested in

  • Assessing viability of attaining mutational signatures in whole genome sequenced tumor-only formalin-fixed, paraffin-embedded (FFPE) samples
  • Study known and novel mutational signatures across cancer types present in the VA health system
  • Discover any actionable signatures that may not have been discovered via targeted panel sequencing

2. GENCARV - GEnetics of Cancer Risk in Veterans susceptibility to cancer:

GENCARV - Genetics of Cancer Risk in Veterans susceptibility to cancer results from a combination of environmental and genetic risk factors, and several studies have identified rare, high-risk genetic variants that are associated with the development of individual or multiple cancers. The clinical utility of genetic variants comes from their use to inform personal cancer prevention and screening strategies, inform family genetic testing, and/or direct cancer treatment decisions. Genetics-driven clinical decision making is now routine for individuals with rare pathogenic germline variants (PGVs) in BRCA and Lynch Syndrome genes with substantial morbidity and mortality benefits from implementation of pre-symptomatic screening and prevention modalities and the use of targeted treatment of cancers that do develop. Rare PGVs in BRCA1, BRCA2 and the Lynch Syndrome genes together are found in approximately 1% of the population, with each variant individually being rare (<0.01% population frequency) and leading to high (>4-fold) lifetime cancer risk. Several other high-risk cancer genes have well-described associated syndromic features, such as APC, PTEN, STK11, TP53, among others, but the rarity of these syndromes impedes genotype-phenotype analyses. Finally, several PGVs are moderately rare (0.01-1% of the population) and confer moderate risks (2-4-fold) of cancer, requiring large number of individuals for well-powered studies. The vast majority of data on cancer risks comes from studies of research participants in high-risk registries or genetic testing company data on results from patients referred for high suspicion of a genetic susceptibility to cancer; therefore, both over estimate cancer risks. The emergence of biobanks allows risk estimates from unselected patient populations, but most of these resources lack detailed clinical information to yield high-level genotype-phenotype associations and outcomes-based research findings.

The VA Million Veteran Program (MVP) was established to provide genetic data representative of the United Status population for genetic association studies. MVP has performed SNP genotyping on nearly 700K and whole genome sequencing (WGS) on >130K Veterans. To perform genotype-phenotype studies and outcomes research in relation to genetic status, VA has linked research genetic data to the robust electronic health record. We use our long-standing cancer genetics expertise to accurately identify rare and moderately rare PGVs in MVP WGS and SNP genotyping data. With the VA National Precision Oncology Program, we also leverage data from our robust clinical genetic testing infrastructure in VA to validate MVP findings.

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Funding