Alterations In Tumor Suppressor Genes Research Articles

HIPK2 in Colon Cancer: A Potential Biomarker for Tumor Progression and Response to Therapies.

Colon cancer, one of the most common and fatal cancers worldwide, is characterized by stepwise accumulation of specific genetic alterations in tumor suppressor genes or oncogenes, leading to tumor growth and metastasis. HIPK2 (homeodomain-interacting protein kinase 2) is a serine/threonine protein kinase and a "bona fide" oncosuppressor protein. Its activation inhibits tumor growth mainly by promoting apoptosis, while its inactivation increases tumorigenicity and resistance to therapies of many different cancer types, including colon cancer. HIPK2 interacts with many molecular pathways by means of its kinase activity or transcriptional co-repressor function modulating cell growth and apoptosis, invasion, angiogenesis, inflammation and hypoxia. HIPK2 has been shown to participate in several molecular pathways involved in colon cancer including p53, Wnt/β-catenin and the newly identified nuclear factor erythroid 2 (NF-E2) p45-related factor 2 (NRF2). HIPK2 also plays a role in tumor-host interaction in the tumor microenvironment (TME) by inducing angiogenesis and cancer-associated fibroblast (CAF) differentiation. The aim of this review is to assess the role of HIPK2 in colon cancer and the underlying molecular pathways for a better understanding of its involvement in colon cancer carcinogenesis and response to therapies, which will likely pave the way for novel colon cancer therapies.

NF1-Driven Rhabdomyosarcoma Phenotypes: A Comparative Clinical and Molecular Study of NF1-Mutant Rhabdomyosarcoma and NF1-Associated Malignant Triton Tumor.

Alterations of the NF1 tumor suppressor gene is the second most frequent genetic event in embryonal rhabdomyosarcoma (ERMS), but its associations with clinicopathologic features, outcome, or coexisting molecular events are not well defined. Additionally, NF1 alterations, mostly in the setting of neurofibromatosis type I (NF1), drive the pathogenesis of most malignant peripheral nerve sheath tumor with divergent RMS differentiation (also known as malignant triton tumor [MTT]). Distinguishing between these entities can be challenging because of their pathologic overlap. This study aims to comprehensively analyze the clinicopathologic and molecular spectrum of NF1-mutant RMS compared with NF1-associated MTT for a better understanding of their pathogenesis. We investigated the clinicopathologic and molecular landscape of a cohort of 22 NF1-mutant RMS and a control group of 13 NF1-associated MTT. Cases were tested on a matched tumor-normal hybridization capture-based targeted DNA next-generation sequencing. Among the RMS group, all except one were ERMS, with a median age of 17 years while for MTT the mean age was 39 years. Three MTTs were misdiagnosed as ERMS, having clinical impact in one. The most frequent coexisting alteration in ERMS was TP53 abnormality (36%), being mutually exclusive from NRAS mutations (14%). MTT showed coexisting CDKN2A/B and PRC2 complex alterations in 38% cases and loss of H3K27me3 expression. Patients with NF1-mutant RMS exhibited a 70% 5-year survival rate, in contrast to MTT with a 33% 5-year survival. All metastatic NF1-mutant ERMS were associated with TP53 alterations. Patients with NF1-mutant ERMS lacking TP53 alterations may benefit from dose-reduction chemotherapy. On the basis of the diagnostic challenges and significant treatment and prognostic differences, molecular profiling of challenging tumors with rhabdomyoblastic differentiation is recommended.

Emerging New Targets in Systemic Therapy for Malignant Pleural Mesothelioma.

Malignant pleural mesothelioma (MPM) is a heterogeneous cancer composed of distinct molecular and pathologic subtypes. Unfortunately, MPM is aggressive, and current therapies for advanced, unresectable disease remain limited to cytotoxic chemotherapy and immunotherapy. Our understanding of the genomic landscape of MPM is steadily growing, while the discovery of effective targeted therapies in MPM has advanced more slowly than in other solid tumors. Given the prevalence of alterations in tumor suppressor genes in MPM, it has been challenging to identify actionable targets. However, efforts to characterize the genetic signatures in MPM over the last decade have led to a range of novel targeted therapeutics entering early-phase clinical trials. In this review, we discuss the advancements made thus far in targeted systemic therapies in MPM and the future direction of targeted strategies in patients with advanced MPM.

Abstract 6568: PTCH1 loss-of-function alterations: Mutational landscape and therapeutic outcomes to hedgehog pathway inhibitors in non-canonical histologies

Abstract Background: Loss-of-function (LOF) alterations of the PTCH1 tumor suppressor gene represent canonical events involved in basal cell carcinoma (BCC) oncogenesis. Hedgehog pathway inhibitors (HPIs) are approved for the treatment of advanced BCC, but genomic characterization and therapeutic outcomes remain limited in tumor agnostic cohorts. Methods: Using the MSK Clinical Sequencing Cohort, we performed a mutational landscape analysis of PTCH1 LOF alterations in non-BCC histologies, including Fraction and Allele-Specific Copy Number Estimates from Tumor Sequencing (FACETS) annotation, and analyzed response and progression-free survival (PFS) with HPIs. Results: Out of 115,619 samples, 1972 samples from advanced solid tumors with PTCH1 LOF alterations were identified. Excluding 1306 variants of unknown significance (VUSs) using OncoKB knowledge base as well as 44 BCC samples, a total of 622 samples from 570 patients with oncogenic PTCH1 alterations were identified across histologies. In addition, 124 patients received ≥1 line(s) of HPI at our institution between 2013 and 2023, including 16 patients with non-BCC histologies presenting PTCH1 alterations. At the patient level (n=570), PTCH1 LOF alterations were present across 33 primary sites, including colorectal cancer (CRC; 34%; n=192), endometrial cancer (17%; n=98), gastroesophageal adenocarcinomas (ADC) (8%; n=46), non-small cell lung cancer (NSCLC; 6%; n=34) and soft tissue sarcomas (STS; 4%; n=25). At the sample level (n=622), TMB ≥10 was seen in 83% (n=516), including 324 samples with MSI-high status. PTCH1 alterations most frequently involved frameshift (fs) indels (69%; n=429), followed by nonsense mutations (18%; n=114), splice site mutations (8%; n=47) and fusions (5%; n=32). Recurrent fs events involved S1203Afs*52 (24%; n=150) and R1308Efs*64 (14%; n=89), among others. When comparing monoallelic (monoall) vs biallelic (biall) PTCH1 alterations through FACETS annotation, monoall showed a strong association with higher TMB (p<0.01) whereas biall were associated with higher loss of heterozygosity (LOH; p<0.01). In the 16 non-BCC patients treated with HPIs, histologies included lung squamous cell cancer (SCC; n=4), cutaneous SCC (n=2), medulloblastoma (n=2), carcinosarcoma (n=2), osteosarcoma, chondrosarcoma, chordoma, sebaceous ADC, anal SCC and CRC (each n=1). TMB ≥10 was seen in 31% (n=5), all of which were MSI-low. Tumor regressions on HPI were seen in 50% (n=8), including lung SCC, cutaneous SCC, sebaceous ADC and medulloblastoma, with median time to response of 3.6 months (mo). Median PFS on HPI was 3.9 mo and median OS was 22.7 mo. Median PFS was higher in patients with TMB ≥10 vs TMB <10 (11.8 vs. 1.8 mo, p=0.02). Conclusion: PTCH1 LOF alterations appear actionable in non-BCC cutaneous/adnexal primaries, lung SCC and medulloblastoma, including TMB-high tumors. Citation Format: Antoine Desilets, Matteo Repetto, Soo Ryum Yang, Monica Chen, Dazhi Liu, Nikhil Rizvi, Ezra Rosen, Alan L. Ho, Alexander Drilon, Yonina R. Murciano-Goroff. PTCH1 loss-of-function alterations: Mutational landscape and therapeutic outcomes to hedgehog pathway inhibitors in non-canonical histologies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6568.

Abstract 1202: Signatures of selection for biallelic inactivation in tumor suppressor genes across cancer types

Abstract Background: The canonical model of tumor suppressor gene (TSG)-mediated oncogenesis, Knudson’s “two hit hypothesis,” posits that loss of both alleles is necessary for TSG inactivation. Many key TSGs accordingly exhibit near universal biallelic loss in associated cancers (e.g., APC in colorectal cancer and RB1 in retinoblastoma). However, because the majority of large-scale cancer genomics studies have utilized non-allele-specific copy number analysis methods, neither the extent of biallelic inactivation across TSGs, nor patterns of selective pressure for biallelic inactivation of TSGs, nor the functional and translational consequences of biallelic inactivation, are well-understood. Methods: Here, through allele-specific analysis of sequencing data from 48,179 cancer patients, we define the prevalence, selective pressure for, and functional consequences of biallelic inactivation across TSGs. We classify TSGs according to their patterns of selection for or against biallelic inactivation across cancer types. Finally, we investigate in detail the association of biallelic status with gene expression and clinical outcome for two TSGs, KEAP1 (in lung adenocarcinoma) and APC (in lung and prostate adenocarcinoma). Results: TSGs largely assort into distinct classes associated with either pan-cancer (Class I) or lineage-specific (Class II) patterns of biallelic loss, while some TSGs, mostly transcription factors, were predominantly monoallelic (Class III/IV). We discover that gene expression and co-mutation patterns indicate selection for biallelic losses in noncanonical contexts, including APC in lung and prostate adenocarcinomas. Selection for biallelic inactivation is also significantly elevated among variants of unknown significance (VUS) of several TSGs, including in KEAP1 in lung adenocarcinoma, nominating these VUS as worthy of in depth functional characterization. Patients with KEAP1 VUS also show similarly poor survival and response to chemoimmunotherapy to patients with known KEAP1 driver mutations, indicating that KEAP1 VUSs phenocopy established KEAP1 oncogenic alleles, and that zygosity, rather than variant classification, is prognostic of therapeutic response. Conclusion: TSGs vary significantly in their patterns of biallelic inactivation, and can be classified according to whether and how consistently, across cancer types, they are selected for loss of both copies. Additionally, taking into account the zygosity of TSG alteration events is often necessary to understand their phenotypic and clinical consequences, and can be useful in identifying novel driver events. Citation Format: Mark R. Zucker, Maria A. Perry, Arielle Elkrief, Anton Safonov, Rohit Thummalapalli, Samuel I. Gould, Debyani Chakravarty, Rose Brannon, Marc Ladanyi, Pedram Razavi, Mark T. Donoghue, Yonina R. Murciano-Goroff, Francisco J. Sánchez-Rivera, Yuan Chen, Ronglai Shen, Sarat Chandarlapaty, David B. Solit, Nikolaus Schultz, Michael F. Berger, Adam J. Schoenfeld, Jason Chang, Ed Reznik, Chaitanya Bandlamudi. Signatures of selection for biallelic inactivation in tumor suppressor genes across cancer types [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1202.

Abstract 4631: Evaluation of the impact of homozygous MTAP truncations on the activity and selectivity of MTA-cooperative PRMT5 inhibitors

Abstract Homozygous deletion of the MTAP gene occurs in 10-15% of all human cancers. To benefit this large and diverse patient population, MTA-cooperative PRMT5 inhibitors, including TNG908 and TNG462, have been developed to leverage the synthetic lethal relationship between MTAP deletion and PRMT5 inhibition. MTA-cooperative PRMT5 inhibitors selectively bind the PRMT5-MTA complex driving selective inhibition of PRMT5 in MTAP-deleted cancers while sparing normal, MTAP-proficient cells. Our PRMT5 inhibitors are currently in Phase I/II clinical trials (NCT05275478 and NCT05732831), and eligibility is restricted to patients with tumors with confirmed MTAP loss either detected by next-generation sequencing (NGS) or immunohistochemistry. MTAP gene loss occurs in cancers because of its chromosomal proximity to one of the most common genetically altered tumor suppressor genes, CDKN2A, but the chromosomal 9p breakpoints for the co-deletion are not uniform. Indeed, while clinical NGS testing and preclinical data confirm that homozygous intragenic MTAP breakpoints occur, the functional consequence of any given breakpoint on MTAP enzymatic activity and protein function remains unknown. Given the potential implications for homozygous intragenic MTAP deletions to impact the clinical response to MTA-cooperative PRMT5 inhibitors, we have started to evaluate the loss-of-function phenotype of various MTAP truncations to determine whether they retain MTAP activity. Here, we present our initial functional genomics analysis of this important diagnostic biomarker using in vitro cDNA reconstitution approaches for MTAP activity combined with analysis of PRMT5 inhibitor sensitivity. Ultimately, these data may help refine patient enrollment on clinical trials to drive the maximum benefit for patients with MTAP-deleted cancers. Citation Format: Matthew R. Tonini, Andre A. Mignault, Douglas A. Whittington, Steven A. Lombardo, Binzhang Shen, Hannah Stowe, Satoshi Yoda, Shangtao Liu, Minjie Zhang, Kevin M. Cottrell, Samuel R. Meier, Heidi Rego, Jennifer Morawiak, Ellen Hooper, Yi Yu, Heather DiBenedetto, Adam S. Crystal, Teng, Kimberly Briggs. Evaluation of the impact of homozygous MTAP truncations on the activity and selectivity of MTA-cooperative PRMT5 inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4631.

Tumor suppressor genes (TSG), treatment, and survival in de novo metastatic hormone-sensitive prostate cancer.

187 Background: The standard of care for metastatic hormone sensitive prostate cancer (mHSPC) has changed substantially in less than a decade. For patients with suitable functional status, combination therapy is recommended. A diverse molecular landscape of mHSPC exists, however molecular findings have yet to inform treatment in mHSPC. Outcomes for patients vary according to baseline molecular features and patients with mutations of tumor suppressor genes (TSG) have worse survival. We sought to examine the impact of TSG alterations on overall survival (OS) for men with de novo mHSPC. Methods: We identified patients diagnosed with de-novo mHSPC from July 2017-December 2021 within the Veterans Health Affairs. Alterations in TSG including TP53, PTEN, and RB1 were determined from National Precision Oncology Program (NPOP) testing collected at any time and all alterations were included. Only patients who survived 4 months and treated within 4 months of diagnosis were included. Treatments included androgen deprivation therapy (ADT) alone, docetaxel, or androgen receptor pathway inhibitors (ARPI). Kaplan-Meier and Cox proportional hazard models to assess the association between TSG and age, BMI, weight change in the year prior to mHSPC, Charlson comorbidity index, PSA at diagnosis, Black race, treatments for mHSPC, and OS. Results: We identified 2808 veterans with de novo mHSPC and 915 (32.6%) had NPOP data available. Patients with NPOP data were younger (mean age 71.4 vs. 75.2 years, p<0.001) and had longer OS than veterans without testing (43.7 months vs. 31.0, p<0.001). TSG alterations were found in 475 of 915 veterans (51.9%) and included 344 TP53 (37.6%), 206 PTEN (22.5%) and 50 RB1 (5.5%). Veterans with TSG had shorter OS compared to no TSG, median OS 48.5 months without TSG vs 37.7 with 1 or more TSG (p<0.001). Each TSG alteration was associated with mortality, TP53 adjusted Hazard Ratio (aHR) and 95% confidence interval 1.68 (1.35-2.09), PTEN aHR 1.33 (1.02-1.73), RB1 aHR 1.51 (1.01-2.26). ADT alone was given in 354 veterans (38.7%), docetaxel in 139 (15.2%), and ARPI in 422 (46.1%). In veterans with one or more TSG neither docetaxel nor ARPI therapy was associated with OS in unadjusted or adjusted analyses compared to ADT alone (Table). Conclusions: TSG alterations were associated with shorter survival in de novo mHSPC in veterans with molecular testing. In this real-world observational study, treatment intensification with docetaxel or ARPI in veterans with TSG was not associated with improved overall survival. [Table: see text]

Applications of CRISPR screening to lung cancer treatment.

Lung cancer is an extremely aggressive and highly prevalent disease worldwide, and it is one of the leading causes of cancer death. Deciphering intrinsic genetic mechanism, finding new targets, and overcoming drug resistance are the key to lung cancer treatment. High-throughput CRISPR screening has been extensively used to obtain the genes related to cancers including lung cancer. This review describes CRISPR/Cas9 or CRISPR/dCas9-based technologies for high-throughput screening. We summarize the applications of CRISPR screening technology in exploring the mechanism of lung cancer development in vivo or in vitro, overcoming drug resistance, improving the effect of immunotherapy, and discovering new therapeutic targets. This review highlights the potential of CRISPR screening in combination with tumor barcoding and high-throughput sequencing (Tuba-seq) to precisely quantify the impact of alterations in many tumor suppressor genes on lung cancer.

Oncogenic context shapes the fitness landscape of tumor suppression

Tumors acquire alterations in oncogenes and tumor suppressor genes in an adaptive walk through the fitness landscape of tumorigenesis. However, the interactions between oncogenes and tumor suppressor genes that shape this landscape remain poorly resolved and cannot be revealed by human cancer genomics alone. Here, we use a multiplexed, autochthonous mouse platform to model and quantify the initiation and growth of more than one hundred genotypes of lung tumors across four oncogenic contexts: KRAS G12D, KRAS G12C, BRAF V600E, and EGFR L858R. We show that the fitness landscape is rugged—the effect of tumor suppressor inactivation often switches between beneficial and deleterious depending on the oncogenic context—and shows no evidence of diminishing-returns epistasis within variants of the same oncogene. These findings argue against a simple linear signaling relationship amongst these three oncogenes and imply a critical role for off-axis signaling in determining the fitness effects of inactivating tumor suppressors.

Co-Occurring Alterations in Multiple Tumor Suppressor Genes Are Associated With Worse Outcomes in Patients With EGFR-Mutant Lung Cancer

IntroductionPatients with metastatic EGFR-mutant NSCLC inevitably have disease progression while on tyrosine kinase inhibitor (TKI) therapy. Co-occurring tumor suppressor gene (TSG) alterations have been associated with poor outcomes, however, detailed analyses of their impact on patient outcomes are limited. MethodsPatients with EGFR-mutant NSCLC treated with EGFR TKIs who had tumor genomic profiling were included. Alterations in TP53 and five additional TSGs (RB1, NF1, ARID1A, BRCA1, and PTEN) were used to stratify the cohort into the following three subgroups: patients with tumors harboring a TP53 mutation plus a mutation in at least one additional TSG (TP53mut/TSGmut), those having a TP53 mutation without additional TSG mutations (TP53mut/TSGwt), and those with TP53wt. Patient characteristics and clinical outcomes were assessed in two independent cohorts. ResultsA total of 101 patients from the Yale Cancer Center and 182 patients from the American Association for Cancer Research Project GENIE database were included. In the Yale cohort, TP53 mutations were identified in 65 cases (64%), of which 23 were TP53mut/TSGmut and 42 were TP53mut/TSGwt. Although the presence of a TP53 mutation was associated with worse outcomes, the additional TSG alteration in TP53mut tumors identified a subset of patients associated with particularly aggressive disease and inferior clinical outcome in both the Yale and the GENIE cohorts. Specifically, in the Yale cohort for patients receiving first-line TKIs, those with TP53mut/TSGmut tumors had shorter progression-free survival (PFS) and overall survival (OS) than TP53mut/TSGwt (PFS: hazard ratio [HR] = 2.03, confidence interval [CI]: 1.12–3.69, p < 0.01, OS: HR = 1.58, CI: 0.82–3.04, p = 0.12) or TP53wt cases (PFS: HR 2.4, CI: 1.28–4.47, p < 0.001, OS: HR = 2.54, CI: 1.21–5.34, p < 0.005). Inferior outcomes in patients with TP53mut/TSGmut tumors were also found in those receiving osimertinib as second-line therapy. Similar findings were seen in patients in the GENIE cohort. ConclusionsPatients with TP53mut/TSGmut tumors represent a patient subgroup characterized by an aggressive disease phenotype and inferior outcomes on EGFR TKIs. This information is important for understanding the biological underpinnings of differential outcomes with TKI treatment and has implications for identifying patients who may benefit from additional therapeutic interventions beyond osimertinib monotherapy.

Targeted Approaches to Treatment of Pleural Mesothelioma: A Review.

Pleural mesothelioma is an aggressive disease that is enriched for inactivating alterations in tumor suppressor genes. Systemic therapeutic strategies for pleural mesothelioma generally involve chemotherapies and immunotherapies that are chosen without consideration of the tumor's molecular profile. As this generalized approach to treatment rarely yields durable responses, alternative therapeutic regimens are urgently indicated. Preclinical studies have identified synthetic lethal protein and metabolic interactions, recurrently overexpressed proteins, and frequent pathway perturbations that may be therapeutically exploited in mesothelioma. This review discusses the mechanism of action of emerging investigational therapies and summarizes findings from phase I-II clinical trials exploring selective, biomarker-driven therapeutic strategies for mesothelioma, with a focus on five common targets. Finally, using lessons learned from these clinical trials, imperatives for successful implementation of targeted therapy in mesothelioma are discussed.

Metabolic barriers in non-small cell lung cancer with LKB1 and/or KEAP1 mutations for immunotherapeutic strategies.

Currently, immune checkpoint inhibitors (ICIs) are widely considered the standard initial treatment for advanced non-small cell lung cancer (NSCLC) when there are no targetable driver oncogenic alternations. NSCLC tumors that have two alterations in tumor suppressor genes, such as liver kinase B1 (LKB1) and/or Kelch-like ECH-associated protein 1 (KEAP1), have been found to exhibit reduced responsiveness to these therapeutic strategies, as revealed by multiomics analyses identifying immunosuppressed phenotypes. Recent advancements in various biological approaches have gradually unveiled the molecular mechanisms underlying intrinsic reprogrammed metabolism in tumor cells, which contribute to the evasion of immune responses by the tumor. Notably, metabolic alterations in glycolysis and glutaminolysis have a significant impact on tumor aggressiveness and the remodeling of the tumor microenvironment. Since glucose and glutamine are essential for the proliferation and activation of effector T cells, heightened consumption of these nutrients by tumor cells results in immunosuppression and resistance to ICI therapies. This review provides a comprehensive summary of the clinical efficacies of current therapeutic strategies against NSCLC harboring LKB1 and/or KEAP1 mutations, along with the metabolic alterations in glycolysis and glutaminolysis observed in these cancer cells. Furthermore, ongoing trials targeting these metabolic alterations are discussed as potential approaches to overcome the extremely poor prognosis associated with this type of cancer.

Impact of tumor suppressor genes inactivation on the multidrug resistance phenotype of hepatocellular carcinoma cells

The response of advanced hepatocellular carcinoma (HCC) to pharmacological treatments is unsatisfactory and heterogeneous. Inactivation of tumor suppressor genes (TSGs) by genetic and epigenetic events is frequent in HCC. This study aimed at investigating the impact of frequently altered TSGs on HCC chemoresistance. TSG alterations were screened by in silico analysis of TCGA-LIHC database, and their relationship with survival was investigated. These TSGs were silenced in HCC-derived cell lines using CRISPR/Cas9. TLDA was used to determine the expression of a panel of 94 genes involved in the resistome. Drug sensitivity, cell proliferation, colony formation and cell migration were assessed. The in silico study revealed the down-regulation of frequently inactivated TSGs in HCC (ARID1A, PTEN, CDH1, and the target of p53, CDKN1A). The presence of TP53 and ARID1A variants and the low expression of PTEN and CDH1 correlated with a worse prognosis of HCC patients. In PLC/PRF/5 cells, ARID1A knockout (ARID1AKO) induced increased sensitivity to cisplatin, doxorubicin, and cabozantinib, without affecting other characteristics of malignancy. PTENKO and E-CadKO showed minimal changes in malignancy, resistome, and drug response. In p53KO HepG2 cells, enhanced malignant properties and higher resistance to cisplatin, doxorubicin, sorafenib, and regorafenib were found. This was associated with changes in the resistome. In conclusion, the altered expression and function of several TSGs are involved in the heterogeneity of HCC chemoresistance and other features of malignancy, contributing to the poor prognosis of these patients. Individual identification of pharmacological vulnerabilities is required to select the most appropriate treatment for each patient.

Epigenetic Silencing of p16INK4a gene in Sporadic Breast Cancer.

Epigenetic alterations of tumor suppressor genes (TSG) involved in the onset and progression of Breast Cancer (BC) may serve as biomarkers for early detection and prediction of disease prognosis. We have herein tried to determine the methylation status of TSG, p16INK4a, in our 50 BC patients and their association with clinicopathological parameters. The methylation status of the p16INK4a gene in fresh tissue samples from 50 patients with BC was assessed by methylation-specific polymerase chain reaction (MS-PCR). The mean age of BC patients was 49.30 ± 9.75years. Of 50 BC samples tested, 21 (42%) had methylated p16INK4a gene. p16INK4a gene hypermethylation was significantly associated with age ≤ 50years, premenopausal status and advanced BC stage. Multivariate analysis revealed a strong association between advanced BC stage (Stage III and Stage IV) and p16INK4a hypermethylation (P = 0.008, RR = 5.996, 95% CI = 1.581-22.739). p16INK4a methylation was significantly associated with Triple Negative BC (TNBC) (P = 0.045, OR = 4.181, 95% CI = 1.030-16.981). These findings indicate that p16INK4a hypermethylation frequently occurs in BC. Hypermethylation of p16INK4a in young, premenopausal, TNBC and with advance stage in BC patients suggests its association with aggressive BC.

Association of alterations in multiple tumor suppressor genes with poor outcomes in patients with EGFR-mutant lung cancer.

9104 Background: Co-occurring alterations in tumor suppressor genes (TSG) have been described as determinants of disease heterogeneity in EGFR-mutant non-small cell lung cancer (NSCLC), however, detailed analyses of their impact on patient outcomes are limited. Methods: Patients with EGFR-mutant NSCLC treated with EGFR tyrosine kinase inhibitors (TKIs) at the Yale Cancer Center who had tumor genomic profiling either before EGFR-TKI therapy (pre-TKI) or at disease progression (post-TKI) were included. Whole exome sequencing (WES) of paired pre- and post-TKI tumor specimens were performed for a subset of patients. The cohort was divided into three subgroups based on alterations in TP53 and five additional TSGs ( RB1, NF1, ARID1A, BRCA1 and PTEN): patients with tumors harboring a TP53 mutation plus a mutation in at least 1 additional TSG ( TP53mut/TSGmut), patients with tumors harboring a TP53 mutation without an additional TSG mutation ( TP53mut/TSGwt), and patients with TP53wt tumors. Clinical characteristics including progression-free (PFS) and overall survival (OS) were assessed. Results: One-hundred-one patients were included in this retrospective study. TP53 mutations were identified in 65 (64%) tumors, of which 23 (35%) and 42 (65%) were classified as TP53mut/TSGmut and TP53mut/TSGwt, respectively. Among those cases with paired WES available (n = 34), frequencies of alterations in the six included TSGs did not significantly differ between pre- and post-TKI tumor specimens. In the full study cohort, the presence of a TP53 mutation was associated with numerically worse PFS (HR 1.46, CI 0.97 – 2.21, p = 0.09) and OS (HR 1.68, CI 1.05 – 2.70, p = 0.04) on first-line EGFR-TKI. Strikingly, after dividing the TP53mut cohort into TP53mut /TSGmut and TP53mut /TSGwt cases, alterations in additional TSG were found to drive the poor outcomes: TP53mut /TSGmut cases had significantly worse PFS and OS on first-line EGFR TKI than TP53mut /TSGwt(mPFS 8.0 vs 10.6 months, p = 0.006; mOS 30.0 vs 33.3 months, p = 0.12) or TP53wt cases (mPFS 8.0 vs 12.6 months, p &lt; 0.0001; mOS 30.0 vs 48.8 months, p = 0.001). There was no significant difference in PFS or OS between patients with TP53mut /TSGwt and TP53wt tumors. Similar outcome differences between the three groups were found in patients who received osimertinib in the second-line setting. Conclusions: The inferior outcomes associated with EGFR/TP53-mutant NSCLC tumors may be due to additional TSG alterations rather than TP53 mutational status alone. Alterations in the investigated TSGs did not appear to emerge under EGFR-TKI therapy, which suggests that they are truncal. Our findings may have implications for understanding the biologic underpinnings of differential outcomes to EGFR TKIs.

Abstract 230: Characterizing pathogenic germline variants in a large Chinese lung cancer cohort

Abstract Objective: To characterize germline pathogenic/likely pathogenic variants (GPVs) in Chinese lung cancer patients and analyze the prevalence of the GPVs and their association with lung cancer risk as well as various clinical and genomic features. Methods: A total of 17,904 Chinese lung cancer patients were included in the study. Samples of all 17,904 patients were subjected to next-generation targeted sequencing by using a panel covering more than 400 cancer genes, of which 146 are known cancer predisposition genes. Baseline somatic mutations of 5,460 patients were also obtained by targeted sequencing. Germline variants were called from white blood cells, and the GPVs were determined according to the 2015 ACMG Guideline. The prevalence of the GPVs as well as corresponding pathways in the whole cohort and in various subgroups was compared. The odds ratios of the GPVs for lung cancer risk were estimated by case-control analysis based on the local cohort and the gnomAD East Asia database. Interaction between GPVs and somatic mutations was investigated. Results: Among 17,904 patients, 1,738 patients (9.7%) carried 1,840 GPVs from 87 cancer susceptibility genes. SBDS (1.37%), TSHR (1.20%), BLM (0.62%), BRCA2 (0.62%) and ATM (0.45%) were the top 5 genes with the highest prevalence in the whole cohort. GPVs in thirteen genes including FANCF, KIT, LHCGR, MAP2K2, NRAS, etc. have not been reported in lung cancer patients previously. GPVs in certain genes showed histological variation. GPVs in PTEN, BRCA2, WRN, RAD51C, JAK2 and ERBB2 were more enriched in lung squamous cell carcinoma compared with lung adenocarcinoma (LUAD). At pathway level, homologous recombination repair (HRR, 2.09%), TP53 pathway (0.83%), nucleotide excision repair (NER, 0.7%), Fanconi anemia (FA, 0.6%) and DNA damage sensor (0.53%) ranked the first 5 pathways with the highest overall prevalence. Case-control analysis showed SBDS c.184A&amp;gt;T(p.K62*) had the highest odds ratio for lung cancer risk (OR=14.42, P&amp;lt;0.001), followed by JAK2 c.1849G&amp;gt;T(p.V617F)(OR=10.8, P=0.002) and MUTYH c.55C&amp;gt;T(p.R19*)(OR=9.77, P=0.004). The rate of second hit was 4.74% in 464 patients with germline alterations in tumor suppressor genes. Patients with second hit had a higher percentage with family cancer history (68.2% vs. 35.2%, P=0.011), and tumors with second hit showed higher tumor mutation burden (P=3.2e-05) and chromosome instability score (CIS, P=0.12). PGVs in certain genes were associated with early onset of lung cancer, and co-occurrence analysis showed there existed germline-somatic mutation interaction in lung cancer. Conclusion: Chinese lung cancer patients have a relatively high carry rate of PGVs. PGVs are distributed in multiple pathways and dominated by DNA damage repair associated pathways. The association between identified PGVs and lung cancer risk requires further studies to verify. Citation Format: Xiaoying Wu, Xiaojun Fan, Jiaohui Pang, Hua Bao, Jiani Yin, Haimeng Tang, Xue Wu, Yang Shao. Characterizing pathogenic germline variants in a large Chinese lung cancer cohort [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 230.

Gastric Tumor Suppressor Genes Alterations Associated with cag A Positive H pylori among Patients with Gastric Cancer Systemic Review and Meta-Analysis

Introduction: Gastric cancer is the fifth most frequent cancer worldwide After lung, breast, colorectal, and prostate cancers. Helicobacter pylori (H. pylori) is considered the most important causative agent of gastrointestinal diseases such as peptic ulcer, gastritis, gastric adenocarcinoma, and mucosa-associated lymphoid tissue (MALT) lymphoma. Objective: to identify the tumor suppressor genes alterations associated with CagA in patients with gastric cancer. Methods: All the available papers published before 2022 were collected by searching in PubMed and Scopus. The keywords included in the research were “H.pylori”, “gastric cancer”, “virulence factors”, “tumor suppressor genes” “ gene mutations” “cagA+” used by Boolean operators to obtain the articles with the keywords in their titles or abstracts. Result: Initial searches yielded 111 articles, four articles were excluded as a duplication using the computer program Zotero (v5), then one hundred and seven articles were screened for the title and abstract evaluation using the Rayyan website, among them seventy-one articles were excluded. Thirty-six articles were scanned for full-text review and eligibility, furthermore, twenty-five articles were excluded because there were either Reviews and case reports, Not relevant studies, Insufficient data, and Unclear methods and results. Eleven articles were included for the literature review. In addition, the studies were in different regions of the world including Asia, Europe, North America, and Latin America. However, most of the studies were related to the USA. Conclusion: Cag A can cause alterations on gastric tumor suppressor genes by either decreased expression by increasing the methylation, inducing point mutation as mentioned, inactivation by increasing the methylation levels, increasing the levels of degradation and methylation the promotor of the tumor suppressor gene as mentioned.

Comprehensive Genomic Profiling of NF2-Mutated Kidney Tumors Reveals Potential Targets for Therapy.

Genomic alterations (GA) in NF2 tumor-suppressor gene have been associated with aggressive behavior in kidney tumors. We used comprehensive genomic profiling (CGP) to evaluate the frequencies of NF2 GA in histologic subtypes of kidney tumors and co-occurring GA in other genes and biomarkers. Advanced kidney tumors included 1875 clear cell (ccRCC), 405 papillary (pRCC), 108 chromophobe (chRCC), 171 sarcomatoid (sRCC), 61 collecting duct (cdRCC), 49 medullary (mRCC), 134 unclassified (uRCC), 906 urothelial carcinoma of renal pelvis (UC), and 147 Wilms tumors underwent hybrid-capture based CGP to evaluate all classes of GA. 192 (4.9%) of kidney tumors featured NF2 GA which were predominantly structural variant mutations (89%), followed by copy number alterations (9%). Gender and age were similar between NF2-mutant (NF2mut) and NF2-wild type (NF2wt) cohorts with male preponderance. NF2 GA frequency was highest in cdRCC (30%), sRCC (21%), uRCC (15%), and pRCC (12%) while lowest in ccRCC (3%), UC (3%) Wilms tumor (1%), and chRCC (0%). NF2 mutational status was associated with loss of Ch 22 (P < .001). NF2mut RCC harbored co-occurring GA including CDKN2A, CDKN2B, SETD2, and BAP1. VHL, PBRM1, PTEN, and FGFR3 GA were significantly more frequent in NF2wt than in NF2mut tumors. MTOR pathway GAs were uncommon in NF2mut tumors. No NF2 mutated RCC featured MSI-high or high TMB. sRCC was associated with high PD-L1 expression. PD-L1 SP142 tumoral (P = .04) and immune cells (P = .013) were more frequent in NF2mut as compared to NF2wt group. Among histologic subtypes of RCC, cdRCC, sRCC, pRCC, and uRCC are enriched in NF2 GA. Co-occurrent GA in CDKN2A/B, SETD2, and BAP1 may represent potential therapeutic targets. Higher level of PD-L1 expression in NF2mut cohort suggests that these tumors might be sensitive to immune checkpoint inhibitor therapies.

Discovering cryptic splice mutations in cancers via a deep neural network framework.

Somatic mutations can disrupt splicing regulatory elements and have dramatic effects on cancer genes, yet the functional consequences of mutations located in extended splice regions is difficult to predict. Here, we use a deep neural network (SpliceAI) to characterize the landscape of splice-altering mutations in cancer. In our in-house series of 401 liver cancers, SpliceAI uncovers 1244 cryptic splice mutations, located outside essential splice sites, that validate at a high rate (66%) in matched RNA-seq data. We then extend the analysis to a large pan-cancer cohort of 17714 tumors, revealing>100 000 cryptic splice mutations. Taking into account these mutations increases the power of driver gene discovery, revealing 126 new candidate driver genes. It also reveals new driver mutations in known cancer genes, doubling the frequency of splice alterations in tumor suppressor genes. Mutational signature analysis suggests mutational processes that could give rise preferentially to splice mutations in each cancer type, with an enrichment of signatures related to clock-like processes and DNA repair deficiency. Altogether, this work sheds light on the causes and impact of cryptic splice mutations in cancer, and highlights the power of deep learning approaches to better annotate the functional consequences of mutations in oncology.

Tumoral Intraductal Neoplasms of the Bile Ducts Comprise Morphologically and Genetically Distinct Entities.

Tumoral (grossly visible) intraductal neoplasms of the bile ducts are still being characterized. To investigate their morphologic, immunohistochemical, and molecular features. Forty-one cases were classified as gastric-, intestinal-, pancreatobiliary-type intraductal papillary neoplasm (IPN), intraductal oncocytic papillary neoplasm (IOPN), or intraductal tubulopapillary neoplasm (ITPN) on the basis of histology. All neoplasms were subjected to targeted next-generation sequencing. The mean age at diagnosis was 69 years (42-81 years); male to female ratio was 1.3. Most neoplasms (n = 23, 56%) were extrahepatic/large (mean size, 4.6 cm). The majority (n = 32, 78%) contained high-grade dysplasia, and 68% (n = 28) revealed invasion. All gastric-type IPNs (n = 9) and most ITPNs/IOPNs showed consistent colabeling for CK7/MUC6, which was less common among others (P = .004). Intestinal-type IPNs (n = 5) showed higher rates of CK20 expression than others (P < .001). Overall, the most commonly mutated genes included TP53 and APC, while copy number variants affected ELF3 and CDKN2A/B. All gastric-type IPNs contained an alteration affecting the Wnt signaling pathway; 7 of 9 (78%) showed aberrations in the MAPK pathway. Mutations in APC and KRAS were common in gastric-type IPNs as compared with others (P = .01 for both). SMAD4 was more frequently mutated in intestinal-type IPNs (P = .02). Pancreatobiliary-type IPNs (n = 14) exhibited frequent alterations in tumor suppressor genes including TP53, CDKN2A/B, and ARID2 (P = .04, P = .01 and P = .002, respectively). Of 6 IOPNs analyzed, 3 (50%) revealed ATP1B1-PRKACB fusion. ITPNs (n = 6) showed relatively few recurrent genetic aberrations. Follow-up information was available for 38 patients (median, 58.5 months). The ratio of disease-related deaths was higher for the cases with invasion (56% versus 10%). Tumoral intraductal neoplasms of the bile ducts, similar to their counterparts in the pancreas, are morphologically and genetically heterogeneous.