Publications by authors named "Sanjeevani Arora"

19 Publications

  • Page 1 of 1

Patterns of germline and somatic testing after universal tumor screening for Lynch syndrome: A clinical practice survey of active members of the Collaborative Group of the Americas on Inherited Gastrointestinal Cancer.

J Genet Couns 2022 Feb 26. Epub 2022 Feb 26.

Department of Medical Oncology, Dana-Farber Cancer Institute/ Harvard Medical School, Boston, Massachusetts, USA.

Clinical guidelines recommend universal tumor screening (UTS) of colorectal and endometrial cancers for Lynch syndrome (LS). There are limited guidelines for how to integrate germline testing and somatic tumor testing after a mismatch repair deficient (dMMR) tumor is identified. We sought to characterize current practice patterns and barriers to preferred practice among clinical providers in high-risk cancer programs. A clinical practice survey was sent to 423 active members of the Collaborative Group of the Americas on Inherited Gastrointestinal Cancer (CGA-IGC) with a follow-up survey sent to 103 clinician responders. The survey outlined clinical vignettes and asked respondents their preferred next test. The survey intended to assess: (1) the role of patient age and family history in risk assessment and (2) barriers to preferred genetic testing. Genetic test options included targeted germline testing based on dMMR expression, germline testing for LS, germline testing with a multigene cancer panel including LS, or paired tumor/germline testing including LS. In October 2020, 117 of 423 (28%) members completed the initial survey including 103 (88%) currently active clinicians. In April 2021, a follow-up survey was sent to active clinicians, with 45 (44%) completing this second survey. After selecting their preferred next germline or paired tumor/germline tumor test based on the clinical vignette, 39% of respondents reported wanting to make a different choice for the initial genetic test without any testing barriers. The proportion of respondents choosing a different initial genetic test was dependent on the proband's age at diagnosis and specified family history. The reported barriers included patient's lack of insurance coverage, patient unable/unwilling to self-pay for proposed testing, and inadequate tumor tissue. Responders reported insurance, financial constraints, and limited tumor tissue as influencing preferred genetic testing in high-risk clinics, thus resulting in possible under-diagnosis of LS and impacting potential surveillance and cascade testing of at-risk relatives.
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http://dx.doi.org/10.1002/jgc4.1567DOI Listing
February 2022

MetaMutationalSigs: Comparison of mutational signature refitting results made easy.

Bioinformatics 2022 02 14. Epub 2022 Feb 14.

Ecological and Evolutionary Signal-processing and Informatics Laboratory, Department of Electrical and Computer Engineering, College of Engineering, Drexel University, Philadelphia, PA, USA.

Motivation: The analysis of mutational signatures is becoming increasingly common in cancer genetics, with emerging implications in cancer evolution, classification, treatment decision and prognosis. Recently, several packages have been developed for mutational signature analysis, with each using different methodology and yielding significantly different results. Because of the nontrivial differences in tools' refitting results, researchers may desire to survey and compare the available tools, in order to objectively evaluate the results for their specific research question, such as which mutational signatures are prevalent in different cancer types.

Results: Due to the need for effective comparison of refitting mutational signatures, we introduce a user-friendly software that can aggregate and visually present results from different refitting packages.

Availability: MetaMutationalSigs is implemented using R and python and is available for installation using Docker and available at: https://github.com/EESI/MetaMutationalSigs.

Supplementary Information: More information about the package including test data and results are available at https://github.com/EESI/MetaMutationalSigs.
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http://dx.doi.org/10.1093/bioinformatics/btac091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9004636PMC
February 2022

Therapeutic implications of germline vulnerabilities in DNA repair for precision oncology.

Cancer Treat Rev 2022 Mar 5;104:102337. Epub 2022 Jan 5.

Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, PA, United States; Department of Radiation Oncology, Fox Chase Cancer Center, Philadelphia, PA, United States. Electronic address:

DNA repair vulnerabilities are present in a significant proportion of cancers. Specifically, germline alterations in DNA repair not only increase cancer risk but are associated with treatment response and clinical outcomes. The therapeutic landscape of cancer has rapidly evolved with the FDA approval of therapies that specifically target DNA repair vulnerabilities. The clinical success of synthetic lethality between BRCA deficiency and poly(ADP-ribose) polymerase (PARP) inhibition has been truly revolutionary. Defective mismatch repair has been validated as a predictor of response to immune checkpoint blockade associated with durable responses and long-term benefit in many cancer patients. Advances in next generation sequencing technologies and their decreasing cost have supported increased genetic profiling of tumors coupled with germline testing of cancer risk genes in patients. The clinical adoption of panel testing for germline assessment in high-risk individuals has generated a plethora of genetic data, particularly on DNA repair genes. Here, we highlight the therapeutic relevance of germline aberrations in DNA repair to identify patients eligible for precision treatments such as PARP inhibitors (PARPis), immune checkpoint blockade, chemotherapy, radiation therapy and combined treatment. We also discuss emerging mechanisms that regulate DNA repair.
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http://dx.doi.org/10.1016/j.ctrv.2021.102337DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9016579PMC
March 2022

Combination Strategies for Immune Checkpoint Inhibitors in PBRM1-mutant Renal Cell Carcinoma: To PARP or Not To PARP?

Eur Urol 2022 02 6;81(2):149-150. Epub 2021 Nov 6.

Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, PA, USA; Department of Radiation Oncology, Fox Chase Cancer Center, Philadelphia, PA, USA. Electronic address:

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http://dx.doi.org/10.1016/j.eururo.2021.10.028DOI Listing
February 2022

RRM2B Is Frequently Amplified Across Multiple Tumor Types: Implications for DNA Repair, Cellular Survival, and Cancer Therapy.

Front Genet 2021 12;12:628758. Epub 2021 Mar 12.

Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, PA, United States.

plays a crucial role in DNA replication, repair and oxidative stress. While germline mutations have been implicated in mitochondrial disorders, its relevance to cancer has not been established. Here, using TCGA studies, we investigated alterations in cancer. We found that is highly amplified in multiple tumor types, particularly in -amplified tumors, and is associated with increased mRNA expression. We also observed that the chromosomal region 8q22.3-8q24, is amplified in multiple tumors, and includes , along with several other cancer-associated genes. An analysis of genes within this 8q-amplicon showed that cancers that have both -amplified along with have a distinct pattern of amplification compared to cancers that are unaltered or those that have amplifications in or only. Investigation of curated biological interactions revealed that gene products of the amplified 8q22.3-8q24 region have important roles in DNA repair, DNA damage response, oxygen sensing, and apoptosis pathways and interact functionally. Notably, -amplified cancers are characterized by mutation signatures of defective DNA repair and oxidative stress, and at least -amplified breast cancers are associated with poor clinical outcome. These data suggest alterations in RR2MB and possibly the interacting 8q-proteins could have a profound effect on regulatory pathways such as DNA repair and cellular survival, highlighting therapeutic opportunities in these cancers.
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http://dx.doi.org/10.3389/fgene.2021.628758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8045241PMC
March 2021

Prevalence of pathogenic variants in DNA damage response and repair genes in patients undergoing cancer risk assessment and reporting a personal history of early-onset renal cancer.

Sci Rep 2020 08 11;10(1):13518. Epub 2020 Aug 11.

Cancer Prevention and Control Program, Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, PA, 19111-2497, USA.

Pathogenic variants (PVs) in multiple genes are known to increase the risk of early-onset renal cancer (eoRC). However, many eoRC patients lack PVs in RC-specific genes; thus, their genetic risk remains undefined. Here, we determine if PVs in DNA damage response and repair (DDRR) genes are enriched in eoRC patients undergoing cancer risk assessment. Retrospective review of de-identified results from 844 eoRC patients, undergoing testing with a multi-gene panel, for a variety of indications, by Ambry Genetics. PVs in cancer-risk genes were identified in 12.8% of patients-with 3.7% in RC-specific, and 8.55% in DDRR genes. DDRR gene PVs were most commonly identified in CHEK2, BRCA1, BRCA2, and ATM. Among the 2.1% of patients with a BRCA1 or BRCA2 PV, < 50% reported a personal history of hereditary breast or ovarian-associated cancer. No association between age of RC diagnosis and prevalence of PVs in RC-specific or DDRR genes was observed. Additionally, 57.9% patients reported at least one additional cancer; breast cancer being the most common (40.1% of females, 2.5% of males). Multi-gene testing including DDRR genes may provide a more comprehensive risk assessment in eoRC patients. Further validation is needed to characterize the association with eoRC.
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http://dx.doi.org/10.1038/s41598-020-70449-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7419503PMC
August 2020

Comprehensive characterization of RAS mutations in colon and rectal cancers in old and young patients.

Nat Commun 2019 08 19;10(1):3722. Epub 2019 Aug 19.

Program in Molecular Therapeutics, Fox Chase Cancer Center, Philadelphia, PA, 19111, USA.

Colorectal cancer (CRC) is increasingly appreciated as a heterogeneous disease, with factors such as microsatellite instability (MSI), cancer subsite within the colon versus rectum, and age of diagnosis associated with specific disease course and therapeutic response. Activating oncogenic mutations in KRAS and NRAS are common in CRC, driving tumor progression and influencing efficacy of both cytotoxic and targeted therapies. The RAS mutational spectrum differs substantially between tumors arising from distinct tissues. Structure-function analysis of relatively common somatic RAS mutations in G12, Q61, and other codons is characterized by differing potency and modes of action. Here we show the mutational profile of KRAS, NRAS, and the less common HRAS in 13,336 CRC tumors, comparing the frequency of specific mutations based on age of diagnosis, MSI status, and colon versus rectum subsite. We identify mutation hotspots, and unexpected differences in mutation spectrum, based on these clinical parameters.
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http://dx.doi.org/10.1038/s41467-019-11530-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6700103PMC
August 2019

Existing and Emerging Biomarkers for Immune Checkpoint Immunotherapy in Solid Tumors.

Adv Ther 2019 10 13;36(10):2638-2678. Epub 2019 Aug 13.

Department of Hematology/Oncology, Fox Chase Cancer Center, Philadelphia, PA, USA.

In the last few years, immunotherapy has transformed the way we treat solid tumors, including melanoma, lung, head neck, breast, renal, and bladder cancers. Durable responses and long-term survival benefit has been experienced by many cancer patients, with favorable toxicity profiles of immunotherapeutic agents relative to chemotherapy. Cures have become possible in some patients with metastatic disease. Additional approvals of immunotherapy drugs and in combination with other agents are anticipated in the near future. Multiple additional immunotherapy drugs are in earlier stages of clinical development, and their testing in additional tumor types is under way. Despite considerable early success and relatively fewer side effects, the majority of cancer patients do not respond to checkpoint inhibitors. Additionally, while the drugs are generally well tolerated, there is still the potential for significant, unpredictable and even fatal toxicity with these agents. Improved biomarkers may help to better select patients who are more likely to respond to these drugs. Two key biologically important predictive tissue biomarkers, specifically, PD-L1 and mismatch repair deficiency, have been FDA-approved in conjunction with the checkpoint inhibitor, pembrolizumab. Tumor mutation burden, another promising biomarker, is emerging in several tumor types, and may also soon receive approval. Finally, several other tissue and liquid biomarkers are emerging that could help guide single-agent immunotherapy and in combination with other agents. Of these, one promising investigational biomarker is alteration or deficiency in DNA damage response (DDR) pathways, with altered DDR observed in a broad spectrum of tumors. Here, we provide a critical overview of current, emerging, and investigational biomarkers in the context of response to immunotherapy in solid tumors.
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http://dx.doi.org/10.1007/s12325-019-01051-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778545PMC
October 2019

Interaction of germline variants in a family with a history of early-onset clear cell renal cell carcinoma.

Mol Genet Genomic Med 2019 03 24;7(3):e556. Epub 2019 Jan 24.

Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania.

Background: Identification of genetic factors causing predisposition to renal cell carcinoma has helped improve screening, early detection, and patient survival.

Methods: We report the characterization of a proband with renal and thyroid cancers and a family history of renal and other cancers by whole-exome sequencing (WES), coupled with WES analysis of germline DNA from additional affected and unaffected family members.

Results: This work identified multiple predicted protein-damaging variants relevant to the pattern of inherited cancer risk. Among these, the proband and an affected brother each had a heterozygous Ala45Thr variant in SDHA, a component of the succinate dehydrogenase (SDH) complex. SDH defects are associated with mitochondrial disorders and risk for various cancers; immunochemical analysis indicated loss of SDHB protein expression in the patient's tumor, compatible with SDH deficiency. Integrated analysis of public databases and structural predictions indicated that the two affected individuals also had additional variants in genes including TGFB2, TRAP1, PARP1, and EGF, each potentially relevant to cancer risk alone or in conjunction with the SDHA variant. In addition, allelic imbalances of PARP1 and TGFB2 were detected in the tumor of the proband.

Conclusion: Together, these data suggest the possibility of risk associated with interaction of two or more variants.
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http://dx.doi.org/10.1002/mgg3.556DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6418363PMC
March 2019

Targeting the DNA Repair Endonuclease ERCC1-XPF with Green Tea Polyphenol Epigallocatechin-3-Gallate (EGCG) and Its Prodrug to Enhance Cisplatin Efficacy in Human Cancer Cells.

Nutrients 2018 Nov 3;10(11). Epub 2018 Nov 3.

Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI 48201, USA.

The 5'-3' structure-specific endonuclease ERCC1/XPF (Excision Repair Cross-Complementation Group 1/Xeroderma Pigmentosum group F) plays critical roles in the repair of cisplatin-induced DNA damage. As such, it has been identified as a potential pharmacological target for enhancing clinical response to platinum-based chemotherapy. The goal of this study was to follow up on our previous identification of the compound NSC143099 as a potent inhibitor of ERCC1/XPF activity by performing an in silico screen to identify structural analogues that could inhibit ERCC1/XPF activity in vitro and in vivo. Using a fluorescence-based DNA-endonuclease incision assay, we identified the green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) as a potent inhibitor of ERCC1/XPF activity with an IC (half maximal inhibitory concentration) in the nanomolar range in biochemical assays. Using DNA repair assays and clonogenic survival assays, we show that EGCG can inhibit DNA repair and enhance cisplatin sensitivity in human cancer cells. Finally, we show that a prodrug of EGCG, Pro-EGCG (EGCG octaacetate), can enhance response to platinum-based chemotherapy in vivo. Together these data support a novel target of EGCG in cancer cells, namely ERCC1/XPF. Our studies also corroborate previous observations that EGCG enhances sensitivity to cisplatin in multiple cancer types. Thus, EGCG or its prodrug makes an ideal candidate for further pharmacological development with the goal of enhancing cisplatin response in human tumors.
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http://dx.doi.org/10.3390/nu10111644DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6267282PMC
November 2018

Gap Junction Intercellular Communication Positively Regulates Cisplatin Toxicity by Inducing DNA Damage through Bystander Signaling.

Cancers (Basel) 2018 Oct 2;10(10). Epub 2018 Oct 2.

Department of Oncology, Karmanos Cancer Institute and Wayne State University, Detroit, MI 48201, USA.

The radiation-induced bystander effect (RIBE) can increase cellular toxicity in a gap junction dependent manner in unirradiated bystander cells. Recent reports have suggested that cisplatin toxicity can also be mediated by functional gap junction intercellular communication (GJIC). In this study using lung and ovarian cancer cell lines, we showed that cisplatin cytotoxicity is mediated by cellular density. This effect is ablated when or Connexin 43 (Cx43) is targeted, a gap junction gene and protein, respectively, leading to cisplatin resistance but only at high or gap junction forming density. We also observed that the cisplatin-mediated bystander effect was elicited as DNA Double Strand Breaks (DSBs) with positive H2AX Ser139 phosphorylation (γH2AX) formation, an indicator of DNA DSBs. These DSBs are not observed when gap junction formation is prevented. We next showed that cisplatin is not the "death" signal traversing the gap junctions by utilizing the cisplatin-GG intrastrand adduct specific antibody. Finally, we also showed that cells deficient in the structure-specific DNA endonuclease - (ERCC1-XPF), an important mediator of cisplatin resistance, further sensitized when treated with cisplatin in the presence of gap junction forming density. Taken together, these results demonstrate the positive effect of GJIC on increasing cisplatin cytotoxicity.
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http://dx.doi.org/10.3390/cancers10100368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6210410PMC
October 2018

Functional analysis of rare variants in mismatch repair proteins augments results from computation-based predictive methods.

Cancer Biol Ther 2017 Jul 11;18(7):519-533. Epub 2017 May 11.

a Molecular Therapeutics Program , Fox Chase Cancer Center , Philadelphia , PA , USA.

The cancer-predisposing Lynch Syndrome (LS) arises from germline mutations in DNA mismatch repair (MMR) genes, predominantly MLH1, MSH2, MSH6, and PMS2. A major challenge for clinical diagnosis of LS is the frequent identification of variants of uncertain significance (VUS) in these genes, as it is often difficult to determine variant pathogenicity, particularly for missense variants. Generic programs such as SIFT and PolyPhen-2, and MMR gene-specific programs such as PON-MMR and MAPP-MMR, are often used to predict deleterious or neutral effects of VUS in MMR genes. We evaluated the performance of multiple predictive programs in the context of functional biologic data for 15 VUS in MLH1, MSH2, and PMS2. Using cell line models, we characterized VUS predicted to range from neutral to pathogenic on mRNA and protein expression, basal cellular viability, viability following treatment with a panel of DNA-damaging agents, and functionality in DNA damage response (DDR) signaling, benchmarking to wild-type MMR proteins. Our results suggest that the MMR gene-specific classifiers do not always align with the experimental phenotypes related to DDR. Our study highlights the importance of complementary experimental and computational assessment to develop future predictors for the assessment of VUS.
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http://dx.doi.org/10.1080/15384047.2017.1326439DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5639829PMC
July 2017

Identification of small molecule inhibitors of ERCC1-XPF that inhibit DNA repair and potentiate cisplatin efficacy in cancer cells.

Oncotarget 2016 Nov;7(46):75104-75117

Department of Oncology, Karmanos Cancer Institute, Wayne State University, Detroit, MI, USA.

ERCC1-XPF heterodimer is a 5'-3' structure-specific endonuclease which is essential in multiple DNA repair pathways in mammalian cells. ERCC1-XPF (ERCC1-ERCC4) repairs cisplatin-DNA intrastrand adducts and interstrand crosslinks and its specific inhibition has been shown to enhance cisplatin cytotoxicity in cancer cells. In this study, we describe a high throughput screen (HTS) used to identify small molecules that inhibit the endonuclease activity of ERCC1-XPF. Primary screens identified two compounds that inhibit ERCC1-XPF activity in the nanomolar range. These compounds were validated in secondary screens against two other non-related endonucleases to ensure specificity. Results from these screens were validated using an in vitro gel-based nuclease assay. Electrophoretic mobility shift assays (EMSAs) further show that these compounds do not inhibit the binding of purified ERCC1-XPF to DNA. Next, in lung cancer cells these compounds potentiated cisplatin cytotoxicity and inhibited DNA repair. Structure activity relationship (SAR) studies identified related compounds for one of the original Hits, which also potentiated cisplatin cytotoxicity in cancer cells. Excitingly, dosing with NSC16168 compound potentiated cisplatin antitumor activity in a lung cancer xenograft model. Further development of ERCC1-XPF DNA repair inhibitors is expected to sensitize cancer cells to DNA damage-based chemotherapy.
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http://dx.doi.org/10.18632/oncotarget.12072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5342726PMC
November 2016

POLD1: Central mediator of DNA replication and repair, and implication in cancer and other pathologies.

Gene 2016 Sep 16;590(1):128-41. Epub 2016 Jun 16.

Program in Molecular Therapeutics, Fox Chase Cancer Center, Philadelphia, PA 19111, USA. Electronic address:

The evolutionarily conserved human polymerase delta (POLD1) gene encodes the large p125 subunit which provides the essential catalytic activities of polymerase δ (Polδ), mediated by 5'-3' DNA polymerase and 3'-5' exonuclease moieties. POLD1 associates with three smaller subunits (POLD2, POLD3, POLD4), which together with Replication Factor C and Proliferating Nuclear Cell Antigen constitute the polymerase holoenzyme. Polδ function is essential for replication, with a primary role as the replicase for the lagging strand. Polδ also has an important proofreading ability conferred by the exonuclease activity, which is critical for ensuring replicative fidelity, but also serves to repair DNA lesions arising as a result of exposure to mutagens. Polδ has been shown to be important for multiple forms of DNA repair, including nucleotide excision repair, double strand break repair, base excision repair, and mismatch repair. A growing number of studies in the past decade have linked germline and sporadic mutations in POLD1 and the other subunits of Polδ with human pathologies. Mutations in Polδ in mice and humans lead to genomic instability, mutator phenotype and tumorigenesis. The advent of genome sequencing techniques has identified damaging mutations in the proofreading domain of POLD1 as the underlying cause of some inherited cancers, and suggested that mutations in POLD1 may influence therapeutic management. In addition, mutations in POLD1 have been identified in the developmental disorders of mandibular hypoplasia, deafness, progeroid features and lipodystrophy and atypical Werner syndrome, while changes in expression or activity of POLD1 have been linked to senescence and aging. Intriguingly, some recent evidence suggests that POLD1 function may also be altered in diabetes. We provide an overview of critical Polδ activities in the context of these pathologic conditions.
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http://dx.doi.org/10.1016/j.gene.2016.06.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4969162PMC
September 2016

Systematic evaluation of underlying defects in DNA repair as an approach to case-only assessment of familial prostate cancer.

Oncotarget 2015 Nov;6(37):39614-33

Cancer Prevention and Control, Fox Chase Cancer Center, Philadelphia, PA, USA.

Risk assessment for prostate cancer is challenging due to its genetic heterogeneity. In this study, our goal was to develop an operational framework to select and evaluate gene variants that may contribute to familial prostate cancer risk. Drawing on orthogonal sources, we developed a candidate list of genes relevant to prostate cancer, then analyzed germline exomes from 12 case-only prostate cancer patients from high-risk families to identify patterns of protein-damaging gene variants. We described an average of 5 potentially disruptive variants in each individual and annotated them in the context of public databases representing human variation. Novel damaging variants were found in several genes of relevance to prostate cancer. Almost all patients had variants associated with defects in DNA damage response. Many also had variants linked to androgen signaling. Treatment of primary T-lymphocytes from these prostate cancer patients versus controls with DNA damaging agents showed elevated levels of the DNA double strand break (DSB) marker γH2AX (p < 0.05), supporting the idea of an underlying defect in DNA repair. This work suggests the value of focusing on underlying defects in DNA damage in familial prostate cancer risk assessment and demonstrates an operational framework for exome sequencing in case-only prostate cancer genetic evaluation.
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http://dx.doi.org/10.18632/oncotarget.5554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4741850PMC
November 2015

Genetic Variants That Predispose to DNA Double-Strand Breaks in Lymphocytes From a Subset of Patients With Familial Colorectal Carcinomas.

Gastroenterology 2015 Dec 5;149(7):1872-1883.e9. Epub 2015 Sep 5.

Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania; Medicine, Fox Chase Cancer Center, Philadelphia, Pennsylvania. Electronic address:

Background & Aims: DNA structural lesions are prevalent in sporadic colorectal cancer. Therefore, we proposed that gene variants that predispose to DNA double-strand breaks (DSBs) would be found in patients with familial colorectal carcinomas of an undefined genetic basis (UFCRC).

Methods: We collected primary T cells from 25 patients with UFCRC and matched patients without colorectal cancer (controls) and assayed for DSBs. We performed exome sequence analyses of germline DNA from 20 patients with UFCRC and 5 undiagnosed patients with polyposis. The prevalence of identified variants in genes linked to DNA integrity was compared with that of individuals without a family history of cancer. The effects of representative variants found to be associated with UFCRC was confirmed in functional assays with HCT116 cells.

Results: Primary T cells from most patients with UFCRC had increased levels of the DSB marker γ(phosphorylated)histone2AX (γH2AX) after treatment with DNA damaging agents, compared with T cells from controls (P < .001). Exome sequence analysis identified a mean 1.4 rare variants per patient that were predicted to disrupt functions of genes relevant to DSBs. Controls (from public databases) had a much lower frequency of variants in the same genes (P < .001). Knockdown of representative variant genes in HCT116 CRC cells increased γH2AX. A detailed analysis of immortalized patient-derived B cells that contained variants in the Werner syndrome, RecQ helicase-like gene (WRN, encoding T705I), and excision repair cross-complementation group 6 (ERCC6, encoding N180Y) showed reduced levels of these proteins and increased DSBs, compared with B cells from controls. This phenotype was rescued by exogenous expression of WRN or ERCC6. Direct analysis of the recombinant variant proteins confirmed defective enzymatic activities.

Conclusions: These results provide evidence that defects in suppression of DSBs underlie some cases of UFCRC; these can be identified by assays of circulating lymphocytes. We specifically associated UFCRC with variants in WRN and ERCC6 that reduce the capacity for repair of DNA DSBs. These observations could lead to a simple screening strategy for UFCRC, and provide insight into the pathogenic mechanisms of colorectal carcinogenesis.
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http://dx.doi.org/10.1053/j.gastro.2015.08.052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4663158PMC
December 2015

A new strategy to ERADicate HER2-positive breast tumors?

Sci Signal 2015 May 26;8(378):fs11. Epub 2015 May 26.

Program in Molecular Therapeutics, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.

HER2-positive breast cancers that have become resistant to HER2-targeting agents, such as trastuzumab (also known as Herceptin), have limited treatment options. In this issue of Science Signaling, Singh et al. have identified a characteristic increase in the endoplasmic reticulum (ER)-associated degradation (ERAD) system in HER2-positive tumors as a mechanism of relieving proteotoxic stress. Synthetic lethality arising from targeted disruption of ERAD signaling in conjunction with other HER2-dependent signaling may improve therapeutic management of this difficult class of breast tumors.
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http://dx.doi.org/10.1126/scisignal.aac4746DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4483198PMC
May 2015

Reversible cell cycle inhibition and premature aging features imposed by conditional expression of p16Ink4a.

Aging Cell 2015 Feb 6;14(1):139-47. Epub 2014 Dec 6.

Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, PA, USA; Department of Medicine, Fox Chase Cancer Center, Philadelphia, PA, USA.

The cyclin-dependent kinase (Cdk) inhibitor p16(Ink4a) (p16) is a canonical mediator of cellular senescence and accumulates in aging tissues, where it constrains proliferation of some progenitor cells. However, whether p16 induction in tissues is sufficient to inhibit cell proliferation, mediate senescence, and/or impose aging features has remained unclear. To address these issues, we generated transgenic mice that permit conditional p16 expression. Broad induction at weaning inhibited proliferation of intestinal transit-amplifying and Lgr5+ stem cells and rapidly imposed features of aging, including hair loss, skin wrinkling, reduced body weight and subcutaneous fat, an increased myeloid fraction in peripheral blood, poor dentition, and cataracts. Aging features were observed with multiple combinations of p16 transgenes and transactivators and were largely abrogated by a germline Cdk4 R24C mutation, confirming that they reflect Cdk inhibition. Senescence markers were not found, and de-induction of p16, even after weeks of sustained expression, allowed rapid recovery of intestinal cell proliferation and reversal of aging features in most mice. These results suggest that p16-mediated inhibition of Cdk activity is sufficient to inhibit cell proliferation and impose aging features in somatic tissues of mammals and that at least some of these aging features are reversible.
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http://dx.doi.org/10.1111/acel.12279DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4326901PMC
February 2015

Downregulation of XPF-ERCC1 enhances cisplatin efficacy in cancer cells.

DNA Repair (Amst) 2010 Jul 24;9(7):745-53. Epub 2010 Apr 24.

Department of Biochemistry & Cancer Biology, University of Toledo Health Science Campus, OH, United States.

Bulky cisplatin lesions are repaired primarily by nucleotide excision repair (NER), in which the structure specific endonuclease XPF-ERCC1 is a critical component. It is now known that the XPF-ERCC1 complex has repair functions beyond NER and plays a role in homologous recombination (HR). It has been suggested that expression of ERCC1 correlates with cisplatin drug resistance in non-small cell lung cancer (NSCLC). In our study, using NSCLC, ovarian, and breast cancer cells, we show that the XPF-ERCC1 complex is a valid target to increase cisplatin cytotoxicity and efficacy. We targeted XPF-ERCC1 complex by RNA interference and assessed the repair capacity of cisplatin intrastrand and interstrand crosslinks by ELISA and alkaline comet assay, respectively. We also assessed the repair of cisplatin-ICL-induced double-strand breaks (DSBs) by monitoring gamma-H2AX focus formation. Interestingly, XPF protein levels were significantly reduced following ERCC1 downregulation, but the converse was not observed. The transcript levels were unaffected suggesting that XPF protein stability is likely affected. The repair of both types of cisplatin-DNA lesions was decreased with downregulation of XPF, ERCC1 or both XPF-ERCC1. The ICL-induced DSBs persist in the absence of XPF-ERCC1. The suppression of the XPF-ERCC1 complex significantly decreases the cellular viability which correlates well with the decrease in DNA repair capacity. A double knockdown of XPF-ERCC1 displays the greatest level of cellular cytotoxicity when compared with XPF or ERCC1 alone. The difference in cytotoxicity observed is likely due to the level of total protein complex remaining. These data demonstrate that XPF-ERCC1 is a valid target to enhance cisplatin efficacy in cancer cells by affecting cisplatin-DNA repair pathways.
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http://dx.doi.org/10.1016/j.dnarep.2010.03.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4331052PMC
July 2010
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