TP53 Alterations Research Articles

DIPG-78. OSSEOUS METASTASIS IN DIFFUSE MIDLINE GLIOMA: A CASE REPORT AND REVIEW OF LITERATURE

Abstract BACKGROUND Diffuse midline glioma (DMG), H3K 27M-altered are uniformly fatal CNS tumors with 2-year overall survival of less than 10%. Neuroaxis dissemination with disease progression is well known. However, osseous metastasis is a rare occurrence. METHODS We report a patient with localized DMG at diagnosis with subsequent craniospinal and osseous metastasis. We also performed an updated literature review on DMG with extra-neural metastasis (ENM). RESULTS At 11 years of age, the patient presented with progressively worsening headache and vomiting and was found to have a tumor within the right thalamus with mild mass effect on the right lateral ventricle without hydrocephalus. The patient underwent stereotactic tumor biopsy with pathology consistent with DMG, H3K 27-altered. Whole exome sequencing revealed alterations in H3-3A, TP53 and PTPN11. Treatment consisted of upfront focal radiation therapy with subsequent stabilization of the thalamic lesion. However, within 1.5 months of completing radiotherapy, the patient developed drop metastasis, intracranial leptomeningeal dissemination, and diffuse osseous metastasis within the spine, bilateral iliac wings, and ribs. The patient died from disease progression within 2 months of metastatic progression, 6 months from initial diagnosis. Review of literature yielded 11 cases of ENM in pediatric patients with DMG, H3K 27M-altered. Four patients had ENM at diagnosis. The remaining seven patients developed extra CNS metastasis within 1-6 months of diagnosis. Eight patients received upfront radiotherapy with variation in adjuvant chemotherapy. All patients died with median overall survival of 9 months (range, 0.5-24) from diagnosis and 6 months (range, 0.5-22) from ENM occurrence. CONCLUSION ENM in DMG, H3K 27-altered is uncommon. Mechanisms for development of ENM in DMG is unclear. Biopsy and molecular testing for both primary site and ENM should be considered to help elucidate biologic mechanisms of ENM and potentially guide management.

DIPG-77. SOMATICTP53,PIK3R1, NF1 ORTERT MUTATIONS ARE ASSOCIATED WITH INFERIOR CLINICAL OUTCOME IN H3K27M-ALTERED DIFFUSE MIDLINE GLIOMA

Abstract BACKGROUND H3K27-altered diffuse midline gliomas (DMGs) have poor prognosis with no standard of care therapy beyond radiation (RT). While RT prologues survival, not all patients respond. Prior work has shown somatic TP53 mutations predict radiation resistance, and poorer outcomes. We report on a multi-center retrospective cohort study to identify molecular biomarkers of RT response in DMG patients. METHODS We performed retrospective chart reviews of patients with biopsy-proven H3K27M-altered DMG between 2013-2023 from six US medical centers and the DMG Center in Zurich. Patients with tumor genomic sequencing and completion of RT were eligible. Genomic alterations were evaluated for somatic mutations, fusions, and chromosomal instability. Cox proportional hazard models were used to evaluate associations between genomic alterations and survival. Multivariate analysis included all significant variables at P <0.1 in initial analysis, including age at diagnosis, tumor location, TP53 and PIK3R1 alterations. RESULTS 297 patients (135 female; median age 8.3years; range 0.2-71.4 years) were included. Median progression-free survival (PFS) and overall survival (OS) was 7.6months (95%-CI 6.9-8.6) and 14.0months (95%-CI 12.9-15.7). Univariate analysis identified TP53 or PIK3R1 status to be associated with shorter OS (TP53-mutant 12.5months vs. TP53-wildtype 17.5months; HR=1.5; 95%-CI (1.1-2.0), P=0.005); (PIK3R1-mutant 12.6months vs. PIK3R1-wildtype 14.5months; HR=1.9, 95%-CI (1.1-3.4), P=0.03). Multivariate analysis corroborated TP53 status association with shorter OS (HR=1.5; 95%-CI (1.2-2.0), P=0.003). Subgroup univariate analysis of pontine DMG patients showed reduced OS with TP53, NF1, or TERT mutations (TP53-mutant 11.9months vs. TP53-wildtype 15.2months; HR=1.6; 95%-CI (1.1-2.3), P=0.02); (NF1-mutant 8.4months vs. NF1-wildtype 13.6months; HR=2.3; 95%-CI (1.1-5.0), P=0.03); (TERT-mutant 8.5months vs. TERT-wildtype 13.6months; HR=2.5; 95%-CI (1.1-5.7), P=0.03). Subsequent multivariate analysis corroborated the association of TERT mutations with shorter OS in pontine DMG (HR=2.5; 95%-CI (1.05, 5.9), P=0.03). CONCLUSIONS This is one of the largest molecular characterized cohorts of H3K27M-altered DMG patients. DMGs with somatic TP53 or PIK3R1 mutations demonstrate inferior OS. Pontine DMG patients with somatic TP53, NF1 or TERT mutations demonstrate inferior OS.

Molecular profiling of visible polypoid and invisible conventional intestinal-type low-grade dysplasia in patients with idiopathic inflammatory bowel disease

AimsLittle is known about the molecular features of visible polyps with low-grade intestinal-type dysplasia in patients with inflammatory bowel disease (IBD). To better understand their origins and biological potential, we...

Surgical Outcomes and Genomic Insights of Nonclear Cell Renal Cell Carcinoma With Synchronous and Metachronous Nodal Disease.

Oncological outcomes in patients with nonclear cell renal cell carcinoma (non-ccRCC) treated with surgery for locoregional nodal disease (ND) remain incompletely characterized. The objective was to investigate the characteristics and outcomes of non-ccRCC patients treated with lymph node dissection (LND) and salvage-LND (S-LND). A total of 1627 patients underwent nephrectomy for nonmetastatic non-ccRCC at Memorial Sloan Kettering Cancer Center between 2007 and 2023. Histology was grouped as papillary, chromophobe, unclassified, and rare subtypes. Retrospective evaluation identified 2.5% (n = 40) of patients with nodal disease at time of nephrectomy (synchronous-ND) and 1.1% (n = 18) with metachronous nodal disease limited to the retroperitoneum (metachronous-ND). Patients' demographics and tumor characteristics were recorded and evaluated by univariate and multivariate cox regression models. Recurrence-free survival (RFS) and overall survival (OS) were estimated by the Kaplan-Meier method. Patients who underwent tumor DNA sequencing during their clinical course were considered for genomic analysis. OS trended toward longer in metachronous-ND (51 vs 105 months; P = .2), though 23% of patients with synchronous-ND were recurrence-free at 45 months median follow-up. In multivariate analysis, rare histologies were associated with decreased OS (P = .030) and metachronous-ND with improved OS (P = .036). RFS and OS after S-LND was 15 and 96 months, respectively. Late onset of metachronous-ND/recurrence was associated with improved OS (P = .008). Genetic alterations in SETD2, TP53, B2M, and FGFR3 were exclusively seen in synchronous-ND, and tumor mutation burden (TMB) was also higher in patients with synchronous-ND (P = .016). Patients with metachronous-ND tend to have prolonged OS compared to synchronous-ND, but a substantial portion of patients with synchronous-ND still enter a durable disease-free state following LND. S-LND can likewise provide long-term survival, particularly in patients with longer time to metachronous nodal recurrence. Synchronous-ND was associated with SETD2, TP53, and NF2 alteration as well as higher TMB.

NGS-based stratification refines the risk stratification in T-ALL and identifies a very-high-risk subgroup of patients

AbstractWe previously reported a better outcome in adult and pediatric T-cell acute lymphoblastic leukemia (T-ALL) harboring NOTCH1 and/or FBXW7 mutations without alterations of K-N-RAS and PTEN genes. Availability of high-throughput next-generation sequencing (NGS) strategies led us to refine the outcome prediction in T-ALL. Targeted whole-exome sequencing of 72 T-ALL–related oncogenes was performed in 198 adults with T-ALLs in first remission from the GRAALL-2003/2005 protocols and 242 pediatric patients with T-ALLs from the FRALLE2000T. This approach enabled the identification of, to our knowledge, the first NGS-based classifier in T-ALL, categorizing low-risk patients as those with N/F, PHF6, or EP300 mutations, excluding N-K-RAS, PI3K pathway (PTEN, PIK3CA, and PIK3R1), TP53, DNMT3A, IDH1/2, and IKZF1 alterations, with a 5-year cumulative incidence of relapse (CIR) estimated at 21%. Conversely, the remaining patients were classified as high risk, exhibiting a 5-year CIR estimated at 47%. We externally validated this stratification in the pediatric cohort. NGS-based classifier was highly prognostic independently of minimal residual disease (MRD) and white blood cell (WBC) counts, in both adult and pediatric cohorts. Integration of the NGS-based classifier into a comprehensive risk-stratification model, including WBC count at diagnosis and MRD at the end of induction, enabled the identification of an adverse-risk subgroup (25%) with a 5-year CIR estimated at 51%, and a favorable-risk group (32%) with a 5-year CIR estimated at 12%. NGS-based stratification combined with WBC and MRD sharpens the prognostic classification in T-ALL and identifies a new subgroup of patients who may benefit from innovative therapeutic approaches. The GRAALL-2003/2005 studies were registered at www.ClinicalTrials.gov as #NCT00222027 and #NCT00327678.

Abstract A009: Synthetic lethality in the context of STAG2-mutant Ewing sarcoma

Abstract Introduction: Ewing sarcoma is an aggressive type of pediatric bone cancer, characterized by a fusion transcription factor (EWSR1-FLI1 in 85% of cases1,2). Recurrent secondary mutations are limited to alterations in TP53, CDKN2A and most frequently STAG2 (15-22%3-6). STAG2-loss-of-function (LOF) mutations correlate with poor prognosis, metastasis and relapse3,5,6, and are shown to induce a metastatic phenotype in preclinical studies7,8. Despite intensive multimodal treatment (surgery, radiation and chemotherapy), the estimated 5-year survival is a mere 65-75% for patients with localized disease, and below 30% for those presenting with metastasis or relapse9. Targeted treatment is currently not available. Our current work aims at identifying synthetic lethal (SL) interactions with STAG2-LOF mutations in the context of Ewing sarcoma, which could represent potential avenues for targeted therapy. Methods: In order to identify STAG2-SL interactions, we have performed genome-wide and subsequent targeted CRISPR knockout (KO) screening in Ewing sarcoma cell lines and their isogenic CRISPR-engineered STAG2-KO clones. To capture Ewing sarcoma inter- and intra-tumoral heterogeneity, we engineered additional bulk STAG2-KO populations across a wide range of cell lines and primary samples, using a lentiviral approach. These isogenic (STAG2 +/-) models are used for high throughput drug screening, validation of screening results and investigation of underlying SL mechanisms. Results: The quality of our genetic screening is validated by the recurrent emergence of (untargetable) paralog STAG1 as the top STAG2-SL hit. Additional preliminary results include a targetable candidate that emerged from both CRISPR- and drug-based screens in most of our tested models. Validation experiments across a broader panel of isogenic cell lines and primary models are currently ongoing. Conclusion: In this work, we employ genetic and drug-based screening approaches across a panel of isogenic (STAG2+/-) Ewing sarcoma cell lines, aiming to identify STAG2-SL interactions. These approaches have yielded potential hits that are currently being investigated across a broader panel of Ewing cell lines and primary models. Ultimately, this work aims to identify novel targeted therapeutic strategies for this aggressive subtype of Ewing sarcoma.

Racial Disparities in Glioblastoma Genomic Alterations: A Comprehensive Analysis of a Multi-Institution Cohort of 2390 Patients

Although molecular biomarkers have significantly advanced precision oncology in glioblastoma, the prevalence of these biomarkers by race remains underexplored. This study aims to characterize the genomic alterations in glioblastoma across Asian, Black, and White patients, offering insights into racial disparities that may influence treatment outcomes and disease progression. Analyzing data from the American Association for Cancer Research Project Genomics Evidence Neoplasia Information Exchange database V13.0, this study examined 2390 primary glioblastoma samples from unique patients. Genomic alterations in 566 cancer-related genes were assessed using targeted next-generation sequencing panels from 3 large cancer institutes. The patient cohort included 112 Asians, 67 Blacks, and 2211 Whites. Statistical significance of associations between genomic alterations and race was evaluated using the chi-squared test, with the Benjamini-Hochberg method applied to control for multiple testing adjustments. Significant racial differences were observed in the frequency of genomic alterations. Asians exhibited a higher frequency of TP53 alterations (52.68%, P < 0.001), Blacks showed more frequent alterations in NRAS (7.46%, P < 0.001), MTOR (10.45%, P= 0.039), and TET2 (8.96%, P=0.039), and Whites had a higher occurrence of PTEN alterations (48.67%, P= 0.045). Additionally, Black patientshad an elevated rate of RET deletions (14.29%, P<0.001). This study identifies significant racial disparities in the alteration frequencies of 6 key glioblastoma genes: NRAS, TP53, MTOR, TET2, PTEN, and RET. These findings underscore the need for racial considerations in glioblastoma treatment strategies and highlight potential avenues for targeted therapeutic interventions. Further research is needed to explore the clinical implications of these genomic disparities.

A prostate cancer gastrointestinal transcriptional phenotype may be associated with diminished response to AR-targeted therapy.

Prostate cancer is a heterogenous disease, but once it becomes metastatic it eventually becomes treatment resistant. One mechanism of resistance to AR-targeting therapy is lineage plasticity, where the tumor undergoes a transformation to an AR-indifferent phenotype, most studied in the context of neuroendocrine prostate cancer (NEPC). However, activation of additional de- or trans-differentiation programs, including a gastrointestinal (GI) gene expression program, has been suggested as an alternative method of resistance. In this study, we explored the previously identified GI prostate cancer phenotype (PCa-GI) in a large cohort of metastatic castration-resistant prostate cancer (mCRPC) patient biopsy samples. We analyzed a dataset of 634 mCRPC samples with batch effect corrected gene expression data from the West Coast Dream Team (WCDT), the East Coast Dream Team (ECDT), the Fred Hutchinson Cancer Research Center (FHCRC) and the Weill Cornell Medical center (WCM). Survival data was available from the WCDT and ECDT cohorts. We calculated a gene expression GI score using the sum of z-scores of genes from a published set of PCa-GI-defining genes (N=38). Survival analysis was performed using the Kaplan-Meier method and Cox proportional hazards regression with endpoint overall survival from time of biopsy to death of any cause. We found that the PCa-GI score had a bimodal distribution, identifying a distinct set of tumors with an activated GI expression pattern. Approximately 35% of samples were classified as PCa-GI high, which was concordant with prior reports. Liver metastases had the highest median score but after excluding liver samples, 29% of the remaining samples were still classified as PCa-GI high, suggesting a distinct phenotype not exclusive to liver metastases. No correlation was observed between GI score and proliferation, AR signaling, or NEPC scores. Furthermore, the PCa-GI score was not associated with genomic alterations in AR, FOXA1, RB1, TP53 or PTEN. However, tumors with MYC amplifications showed significantly higher GI scores (p=0.0001). Patients with PCa-GI tumors had a shorter survival (HR=1.5 [1.1-2.1], p=0.02), but this result was not significant after adjusting for the liver as metastatic site (HR=1.2 [0.82-1.7], p=0.35). Patients with PCa-GI low samples had a better outcome after androgen receptor signaling inhibitors (ASI, abiraterone or enzalutamide) than other therapies (HR=0.37 [0.22-0.61], p=0.0001) while the benefit of ASI was smaller and non-significant for PCa-GI high samples (HR=0.55 [0.29-1.1], p=0.07). A differential pathway analysis identified FOXA2 signaling to be upregulated PCa-GI high tumors (FDR = 3.7 × 10-13). The PCa-GI phenotype is prevalent in clinical mCRPC samples and may represent a distinct biological entity. PCa-GI tumors may respond less to ASI and could offer a strategy to study novel therapeutic targets.

Comprehensive genomic and immune profiling of non-V600 BRAF-mutated melanomas: Implications for therapeutic strategies beyond BRAF/MEK inhibitors.

e21544 Background: The development of targeted therapies ( BRAF plus MEK inhibitors) and immune checkpoint inhibitors (ICIs) has significantly improved the outcomes of patients with advanced melanoma, particularly those harboring BRAF V600 mutations (Class 1). However, melanomas with non-V600 BRAF mutations (classes 2 and 3) remain a subset where effective therapeutic options beyond ICIs are lacking. In this study, we investigated the distinct genomic and immune profile of non-V600 BRAF melanomas to further understand the mechanism of resistance to BRAF/MEK inhibitors and explore other pathways of potential therapeutic targets. Methods: A retrospective cohort of 175 melanoma samples was evaluated by comprehensive genomic and immune profiling, including tumor mutational burden (TMB) and the expression of 395 immune genes. Tumor specimens were classified as BRAFClass1, BRAFClass2, and BRAFClass3. TMB (high ≥10 Mut/Mb) was obtained by DNA sequencing. Gene expression signatures of tumor inflammation and cell proliferation were determined by targeted RNA-sequencing. We used chi-square and Wilcoxon rank-sum tests to determine the association of BRAF mutational status to genomic and immune correlates. Results: The median age of diagnosis was 69 years, with 57% of patients being male. A total of 152 (86.9%), 13 (7.4%), and 10 (5.7%) patients had BRAFClass1, BRAFClass2, and BRAFClass3 alterations, respectively. Genomic alterations in TP53, NF1, MET and ROS1 were more prevalent in BRAFClass2 and BRAFClass3 as opposed to BRAFClass1. BRAFClass2 and BRAFClass3 tumors combined had significantly higher tumor inflammation (median = 55.38; p &lt; 0.05) compared to BRAFClass1 (median = 41.79) as well as higher TMB (median = 54.8; p &lt; 0.001) compared to BRAFClass1 (median = 9.75). BRAFClass3 tumors showed significantly higher cellular proliferation (median = 47.5; p &lt; 0.05) compared to BRAFClass1 (median = 33.5) and BRAFClass2 (median = 31.8) tumors, as well as higher TMB (median = 71.5; p &lt; 0.01) compared to BRAFClass1 (median = 10.9). When looking at differentially expressed genes, CDK1 was upregulated in BRAFClass3 compared to BRAFClass1 and BRAFClass2, and IL12B was upregulated in BRAFClass2 compared to BRAFClass1. Conclusions: Our study showed that the non-Class 1 BRAF (V600E or K) mutated tumors have higher immune response suggesting a favorable response to ICIs. Additionally, we found that BRAFClass2 and BRAFClass3 tumors more commonly harbor alterations in NF1 and TP53, supporting the prior understanding that BRAF non-V600 tumors have a more aggressive clinical course. Comprehensive molecular analysis offers insight into the immunogenicity and distinct oncogenic drivers associated with unfavorable clinical outcomes, which could guide therapeutic development beyond BRAF and MEK inhibitors in BRAF non-V600 melanomas.

Genomic biomarkers of CNS-specific outcomes in patients with breast cancer brain metastases.

2028 Background: The diagnosis of brain metastasis (BM) is a life- and prognosis-altering event for patients with breast cancer (BC). The standard of care for the management of limited BM remains ablation with stereotactic radiosurgery (SRS). There are no established genomic biomarkers to guide CNS-directed treatment among patients with breast cancer brain metastases (BCBM). Methods: We retrospectively reviewed clinical and genomic features of a cohort of patients who received SRS for newly diagnosed BCBM between 2010 and 2021 at Memorial Sloan Kettering Cancer Center (MSK). Next-generation genomic sequencing (NGS) was performed using the MSK-IMPACT assay, covering all exonic and selected intronic regions for up to 505 genes. Time-to-CNS progression (TTCP) and overall survival (OS) were analyzed using multivariable Fine-Gray and Cox proportional hazards models, respectively, adjusted for clinically relevant factors. Relevant clinical factors and genomic alterations present at an instance at 5% or greater were included in the univariate and multivariate analyses, completed for the overall cohort and BC receptor subtypes. Results: From 2010-2021, 260 patients were identified; 123 (47%), 74 (28%), and 63 (24%) patients had hormone receptor-positive (HR+), HER2+, and triple-negative (TN) disease, respectively. 116 (45%) patients had 1 BM and 184 (71%) patients had at least 1 BM greater than 1 cm in axial diameter. Median follow up for the overall cohort was 18 months (Interquartile range (IQR) 9, 36). Among patients who progressed, median TTCP was 6 (IQR 4, 13) and 6 (4, 13), 11 (5, 18), and 4(3, 8) months for the overall cohort, and HR+, HER2+, and TN subgroups, respectively. Median OS was 19 (95% CI 16, 23) and 15 (13, 21), 58 (39, --), and 13 (11, 20), for the overall cohort, and HR+, HER2+, and TN subgroups, respectively. In the overall cohort, shorter TTCP was associated with TN subtype, &gt;5 BMs, and pathogenic alterations in MYC, AGO2, PTEN, AURKA, and NF1 (p &lt; 0.05); decreased OS was associated with non-HER2+ subtype, presence of extracranial metastatic disease, and oncogenic alterations in TP53, CDH1 and NF1 (p &lt; 0.05). In stratified analyses by receptor subtype, shorter TTCP was associated with oncogenic alterations in GAB2 and PTEN in the HR+ subgroup, AURKA, DDR2, and NF1 in the HER2+ subgroup, and with MYC and RB1 in the TN subgroup (p &lt; 0.05). Conclusions: In this large clinico-genomic analysis of CNS-specific outcomes in BC, we identified clinical and putative genomic biomarkers associated with BCBM receiving SRS that could be used to risk stratify patients and optimize treatment strategies.

Comprehensive germline and somatic DDR genomic profiles of Chinese patients with multiple primary lung cancer.

e20019 Background: The number of multiple primary lung cancer (MPLC) patients has rapidly increased in recent years. Disruption of the DNA damage repair (DDR) gene is related to cancer risk, progression, and treatment selection. However, the characteristics and significance of DDR alterations remain undefined in MPLC. Methods: A total of 133 Chinese MPLC patients with 308 lesions were enrolled and 784 single-focus lung cancer (SFLC) patients were analyzed as a control in the present study. Genetic testing was performed by utilizing the next-generation sequencing (NGS) of 808 cancer-related genes including 276 DDR genes. The patients were divided into 3 groups: DDR-germline (germline variants, N = 11), DDR-somatic group (somatic variants, N = 98), and the non-DDR group (no DDR variants, N = 24). Results: The overall DDR variants were identified in 82.0% (109/133) of the patients, and a total of 283 variants were found, with almost all being somatic (254/283). The most commonly germline alterations were found in RAD50 (1.5%), WRN (1.5%) and BRCA2 (0.8%), while in the somatic mutations, TP53 (26.3%) showed the highest frequency. DDR-somatic group was more likely to have alterations in TP53, LRP1B, ERBB2 and MYC compared with DDR-germline group. Taken non-DDR group as reference, mutations in TP53 (P&lt;0.05), ALK and PIK3CA were more common in DDR-germline group, but EGFR, RBM10, TP53, LRP1B, TERT and MYC (all P&lt;0.05) in DDR-somatic group. Furthermore, the DDR-somatic group exhibits the highest median tumor mutational burden (6.56) compared with 3.55 (P&lt;0.01) in DDR-germline and 3.69 (P&lt;0.01) in non-DDR group. In addition, the copy number variation (CNV) was statistically different among the three groups (P&lt;0.001). The proportion of copy number gain patients in DDR-somatic group is the highest (28.3%) compared with 12.0% in DDR-germline group and 10.9% in non-DDR group (P&lt;0.01). DDR-somatic MPLC and SFLC patients possessed distinct somatic genomic characteristics, especially with significantly different prevalence in TP53, RBM10, KRAS, BRAF, and ALK (all P&lt;0.05). Moreover, we also found notable differences in the prevalence of TP53 and CNV (both P&lt;0.05) between DDR-germline MPLC and SFLC patients. Conclusions: MPLC patients exhibit a distinctive DDR mutations landscape. The differences in the mutation profile between the DDR-germline and DDR-somatic groups and the distinct actionable genes may indicate the different target-therapy choices for MPLC patients.

Genomic profiling of lung adenocarcinoma and squamous carcinoma in Indian patients: Implications for precision oncology.

e20599 Background: Lung cancer, the leading cause of cancer related mortalities, remains a significant global health burden. Despite advances in next generation sequencing technologies, the genomic landscape of non-small cell lung cancer (NSCLC) in the Indian patients remains underexplored. In this study, we aimed to elucidate the genomic landscape of lung adenocarcinoma and squamous carcinoma among Indian patients using targeted sequencing data of over 400 lung cancer patients. Methods: Utilizing targeted sequencing of 50 established cancer-related genes, we conducted a comprehensive analysis of the genomic alteration profiles in 325 lung adenocarcinoma and 81 lung squamous carcinoma samples from Indian patients. Further, we employed robust statistical methodologies to delineate correlations between genomic alterations and clinical characteristics including age, gender, extent of disease, habits and PD-L1 status. Furthermore, our investigation revealed distinctive genomic signatures of NSCLC specific to the Indian population through comparative analyses across various ethnicities. Results: EGFR mutations were detected in 45.8% of lung adenocarcinoma and a notable 18% of lung squamous carcinoma cases among Indian patients, showing significant prevalence among females and non-smokers. Alongside EGFR, lung adenocarcinoma patients exhibited frequent alterations in TP53 (27.4%), ALK (11.4%), and KRAS (10.2%), while lung squamous carcinoma cases predominantly showed mutations in TP53 (40.7%), PIK3CA (17.3%), and CDKN2A (8.6%). Remarkably, 68% of lung adenocarcinoma cases deemed eligible for NCCN-approved targeted therapies. Additionally, it was observed that 60% of PD-L1-negative squamous carcinoma patients harbored TP53 alterations, indicating intriguing therapeutic implications. Overall, we observed alterations in 88% lung adenocarcinoma and 70% lung squamous carcinoma patients. Conclusions: Our study unveiled distinct genomic variations present in both adenocarcinoma and squamous carcinoma patients from India, revealing crucial insights for precision medicine and clinical management of lung cancer. Our findings provide a significant lead for designing clinical trials aimed at evaluating EGFR inhibitors in lung squamous carcinoma patients, potentially expanding therapeutic options for Indian lung cancer patients.

Genetic ancestry-associated differences in genomic profiling and treatment patterns in pancreatic ductal adenocarcinoma (PDAC).

4138 Background: Prior epidemiological studies have explored race both as a population-level risk factor and as a factor influencing outcomes in PDAC. However, the impact of genetic ancestry on PDAC remains unclear. We assessed the genomic landscape, clinical care patterns, and outcomes across ancestries in a large and diverse PDAC cohort to determine the potential impact of genomics on ancestral disparities. Methods: A total of 24,948 patients diagnosed with PDAC who received tissue-biopsy based comprehensive genomic profiling (CGP; Foundation Medicine, Inc. (FMI)) as part of routine clinical care from Aug 2014-Sep 2023 were examined for genomic differences based on genetic ancestry which was determined using a SNP-based approach. The US-based de-identified Flatiron Health (FH)-FMI clinico-genomic database (CGDB) was used to assess testing and treatment patterns based on ancestry in 3,863 patients of European (EUR) and 414 patients of African (AFR) ancestry diagnosed with PDAC. Results: The genetic ancestry distribution in the overall cohort was 80.9% EUR, 10.0% AFR, 4.9% Admixed American, 3.5% East Asian, and 0.7% South Asian. AFR patients showed a slightly younger median age at biopsy than EUR (64 vs 67, p &lt; .001). KRAS, TP53, CDKN2A/ 2B and SMAD4 were the most commonly altered genes in the cohort, as expected. Compared to EUR, AFR patients were enriched in alterations in TP53 (85% vs 78%, p &lt; .001) and KRAS (95% vs 93%, p &lt; .001). While rare, alterations in GNAS, CHEK2, and BRAF were observed less frequently in AFR (1.7% vs 3.3%, 0.3% vs 1.7%, 0.9% vs 2.2%, respectively, all p &lt; .001). In the CGDB cohort, AFR patients were likely to have lower socioeconomic status (SES) compared to EUR patients (33% of AFR in the lowest SES quintile compared to 9% of EUR; p &lt; .001 across all quintiles). AFR patients were less likely to receive care at academic centers (AFR 14.8% vs EUR 22.2%, p &lt; .001) and less likely to receive an investigational drug during their cancer treatment course (5.9% vs 14.1%, p = .001). Rates of surgery (AFR 30.2% vs EUR 34.6%, p = .08), immunotherapy (3.2% vs 3.3%, p &gt; .99), and targeted therapy (4.1% vs 6.2%, p = .28) were not significantly different. While AFR patients comprised a significantly larger proportion of AFR+EUR CGP volume following the decision to implement Medicare coverage for NGS testing (10.6% after Mar 16, 2018 vs 6.6% prior, p &lt; .001), this remained below the expected based on share of the overall US population. Conclusions: To our knowledge, this is the largest study of genetic ancestry in PDAC to date. There were largely similar patterns of gene alterations across AFR and EUR ancestry groups suggesting non-genomic factors may underlie clinical outcomes disparities which have been reported in other studies. Notably, there were ancestry-based differences associated with SES and both CGP testing and clinical trial enrollment patterns, which may impact patient outcomes.

CCNE1 amplification as marker of poor prognosis and novel therapeutic target in advanced breast cancer.

1040 Background: Cyclin E1 (CCNE1) overexpression or gene amplification is linked to dismal outcomes in various tumors, including breast cancer (BC). Recent studies suggest that CCNE1 amplification is a potential target for new synthetic lethality-based treatments, including CDK2-selective and PKMYT1 inhibition. This study explores the clinical and genomic characteristics of CCNE1-amplified BCs. Methods: We analyzed genomic and clinical data from consecutive BCs that underwent clinical tumor-normal targeted panel sequencing (MSK-IMPACT) between April 2014 and December 2021. Allele-specific copy number, including CCNE1 amplification, and fraction genome altered (FGA) were calculated by FACETS. Mutual exclusivity and co-occurrence analyses were performed using CoMEt. Benjamini–Hochberg method for multiple testing correction (q) was applied. Real-world progression-free survival (rwPFS) was assessed by the Kaplan Meier method and Cox models, focusing on patients with available pre-treatment samples. Results: Out of 3,753 BCs, 125 (3.3%) had CCNE1 amplification. A higher occurrence of CCNE1 amplification was noted in post-treatment (n=2,368) compared to treatment-naïve tumors (n=1,385; 4% vs 2.4%, p=0.007). CCNE1 amplification was less common in hormone receptor (HR)+/HER2- BCs (2%) compared to HER2+ (7.6%) and triple-negative (7.2%) tumors (p&lt;0.001). BCs with CCNE1 amplification showed higher median FGA, indicating increased genomic instability. In primary BC, TP53 alterations were more frequent in CCNE1-amplified tumors (q&lt;0.001), while CDH1 alterations were mutually exclusive (q&lt;0.001) suggesting that lobular BCs rarely harbor CCNE1 amplification. Similar trends were seen in post-treatment samples. CCNE1 amplification detected at baseline was linked to shorter median rwPFS in HR+/HER2- metastatic BCs treated with CDK4/6 inhibitors plus endocrine therapy (8.8 vs 15.2 months in CCNE1-amplified [n=9] vs CCNE1-non amplified [n=402]; hazard ratio [HR] 2.82, 95% CI 1.38-5.75), and in HER2+ metastatic BCs treated with first-line THP regimen (7.3 vs 20.8 months in CCNE1-amplified [n=5] vs CCNE1-non amplified [n=106]; HR 3.1, 95% CI 1.24-7.87). In a CCNE1-amplified triple-negative cell model (HCC1569), treatment with the PKMYT1 inhibitor RP-6306 significantly reduced tumor growth. Conclusions: CCNE1 amplification in BCs is associated with greater genomic instability and characterizes a group of metastatic BCs with poor response to standard of care therapies. Novel therapies targeting CCNE1-amplified tumors are under evaluation in preclinical and clinical studies.

Comparing Genomic Profiles of ALK Fusion-Positive and ALK Fusion-Negative Nonsmall Cell Lung Cancer Patients.

Background Anaplastic lymphoma kinase ( ALK ) fusion events account for 3 to 7% of genetic alterations in patients with nonsmall cell lung cancer (NSCLC). This study aimed to explore the landscape of ALK fusion-positive and ALK fusion-negative in a large cohort of NSCLC patients. Methods The formalin-fixed paraffin-embedded specimens of NSCLC patients who underwent next-generation sequencing from 2020 to 2023 in Yinfeng Gene Technology Co., Ltd. Clinical laboratory were included in this study. Results In the current study, a total of 180 (3.20%) patients tested positive for ALK fusions in 5,622 NSCLC samples. Within the ALK -positive cohort, a total of 228 ALK fusions were identified. Furthermore, five novel ALK fusion partners, including DAB1-ALK , KCMF1-ALK , KIF13A-ALK , LOC643770-ALK , and XDH-ALK were identified. In cases with ALK fusion-positive, TP53 alterations were the most prevalent (26.3%), followed by CDKN2A (8.4%), epidermal growth factor receptor ( EGFR , 5.6%), and ALK (5.6%). By contrast, EGFR alterations were most prevalent (51%) in patients with ALK fusion-negative NSCLC, followed by TP53 (42.7%), KRAS (11.6%), and CDKN2A (11.3%). A total of 10 cases where ALK fusion co-occurred with EGFR mutations were also identified. Notably, the ALK fusion positivity rate was higher in younger patients ( p < 0.0001) and in female patients ( p = 0.0429). Additionally, positive ALK test results were more prevalent in patients with high programmed death-ligand 1 expression, especially when applying a 50% cutoff. Conclusions Collectively, these findings offer valuable genomic insights that could inform the personalized clinical care of patients with NSCLC harboring ALK fusions within the context of precision medicine.

Clinical outcomes of NOTCH pathway-activated adenoid cystic carcinoma with and without co-occurrent TP53 mutation.

6101 Background: Adenoid cystic carcinoma (ACC) is a salivary cancer with a variable disease course. Inoperable locally advanced, recurrent or metastatic ACC (LA-R/M-ACC) are its commonest manifestations. The disease course is typically indolent with patients remaining on surveillance for many years before initiating relatively ineffective systemic therapies. A sub-group of patients experience rapid disease progression and surveillance is not indicated. NOTCH gain-of-function mutation (NOTCHgof) is well characterised, defining a genomic sub-group with shorter survival times. However, there remains a need to identify additional prognostic biomarkers to inform the clinical management of these patients. TP53 mutation (TP53mt) has been proposed as a prognostic biomarker warranting further evaluation. Methods: Clinical-genomic data from 349 ACC patients with LA-R/M-ACC were included. 198 patients provided written informed consent to an ethically approved study, and a further 151 were identified through cBioPortal (MetTropsim, Cell, 2021). ACC tumour samples were evaluated by next-generation sequencing (NGS) for the presence of NOTCHgof and TP53mt. Kaplan-Meier analysis was performed, and p values calculated with the log-rank test, to calculate overall survival (OS) from inoperable LA or R/M disease for patients with either NOTCHgof alone, TP53mt alone or both NOTCHgof/TP53mt combined. Results: ACC tumours harboured NOTCHgof in 14.0% (49/349) and TP53mt in 13.8% (48/349) of cases, respectively. Co-occurrence of TP53mt was identified in 16% of ACC tumours with NOTCHgof (8/49), being in 2.3 % of all LA-R/M-ACC cases. Median OS from inoperable LA-R/M disease was significantly reduced for patients with co-occurrent NOTCHgof/TP53mt (n=8, 10.0 mo; 95% CI 8–12), followed by NOTCHgof/TP53wt (n=41, 15.2 mo; 95% CI 8–22), NOTCHwt/TP53wt (n=262, 58.6 mo; 95% CI 52–65) and NOTCHwt/TP53mt (n=40, 69 mo; 95% CI 13–125) (p=&lt;0.001). TP53 alterations were available for functional classification in 33 patients and were classed as loss-of-function point mutations in 81.8% or truncating frameshift mutations in 18.2%. Truncating TP53mt were mutually exclusive with NOTCHgof. Although the smaller number in this group limits statistical power, patients with truncating TP53mt had shorter OS from recurrence (15.9 mo; 95% CI 0-39) compared with other TP53mt (51.0 mo; 95% CI 0-108; p=0.41). Conclusions: A subgroup of LA-R/M ACC patients with co-occurrent TP53mt and NOTCHgof were identified with significantly shorter OS from recurrence than either NOTCHgof or TP53mt alone. Truncating TP53mt may have greater prognostic value than other TP53mt in NOTCHwt ACC. This study provides further evidence of the potential prognostic utility of some TP53 mutations in ACC and highlights groups of patients with aggressive disease who are potentially suitable for inclusion in therapeutic research programmes.

Young lung adenocarcinoma: Molecular analysis pointing to early acquisition of aggressive mutations.

e20089 Background: Lung adenocarcinoma in young adults (≤50) has an aggressive course even in the presence of targetable mutations. Prior US studies were conducted predominantly in the Northeast where smoking prevalence is low compared to Kentucky. To assess the mutation spectrum in this high-risk population we performed a descriptive analysis of mutations in patients 50 years old and younger (YA), compared the survival of YA patients with and without NGS, and compared the survival of YA to patients older than 50 seen at an NCI designated Comprehensive Cancer Center situated in the tobacco belt. Methods: Using the Kentucky Cancer Registry (KCR) we identified 139 YA patients with lung adenocarcinoma seen at our institution between 01/2015 to 11/2023, 54 had NGS testing. 933 patients over age 50 were identified for comparison. Adenocarcinoma specimens were analyzed using next-generation sequencing of DNA (DNA-592-gene panel or whole exome) or RNA (whole transcriptome or +1,500-gene panel) at Caris Life Sciences or Foundation Medicine. Hazard ratios (HR) for survival were calculated using the Cox proportional hazards model. The cutoff for statistical significance was a p-value &lt; 0.05 as calculated by the Chi-square test. Results: Of the 54 YA patients with NGS, 28 (52%) were female, 48 were white (88%), 3 Black (6%), and 3 Asian (6%). Only 18% were never smokers (median 28 pack years), and 44% had a family history of lung cancer. Targetable mutations were noted in 27 (50%). EGFR was mutated in 9 patients (17%), 2 with exon 19 deletion, 2 with exon 20 insertion, and 5 with exon 21 L858R. ALK rearrangement was seen in 3 patients (5%), while 3 (5%) had ERBB2 mutations. There was a mutation of KRAS in 17 patients (31%), and 16 of the 17 were transversion mutations. (13/17 G12C, 1/17 G12R, 1/17 G12V and 1/17 G13C). TP53 was mutated in 44 patients (81%), and 16 patients (30%) had mutations in homologous recombination repair (HHR) genes. The tumor mutation burden was high in 28 (52%) of patients. PD-L1 expression was &gt; 50% in 22/54, 1-49% in 18/54, and 0% in 10/54 (4/54 NA). Survival was not significantly different between YA patients with and without NGS nor between YA and older patients. Conclusions: We identified a high prevalence of KRAS G12C, previously associated with increasing age and environmental insults, and EGFR exon 21 L858R in a YA population. Alterations in HHR genes and TP53 were present at high rates. Acquisition of aggressive forms of KRAS and EGFR mutation, as well as the high prevalence of TP53 and HHR defects, may contribute to early disease development. This highlights the importance of early tobacco cessation efforts and the need to assess germline mutations. Lung cancer remains a therapeutic challenge even in young patients with actionable mutations, as outcomes remain poor, equivalent to older patients.

Genomic profile of Japanese patients with rare ductal adenocarcinoma of the prostate has high grade and poor prognostic aspects.

e17046 Background: Ductal adenocarcinoma (DAC) is a rare subtype of prostate cancer with unknown etiology and poor prognosis. Genomic profile analysis using next-generation sequencing has the potential to elucidate the pathogenesis of rare cancers. To date, some studies have investigated the genetic profile of the DAC genotype; however, no definitive findings have been obtained due to the small number of patients and limited geographic coverage. The discovery of novel prognostic biomarkers and identification of new pharmacotherapeutic targets for DAC are currently of research interest. The aim of this study is to analyze the genomic profile of DAC in a Japanese population and to summarize its features. Methods: Clinical and histopathological records from our three hospitals were searched for radical prostatectomy, transurethral resection of the prostate, and prostate needle biopsy specimens containing a DAC component in the past 10 years. Twenty-eight patients diagnosed with DAC of the prostate were identified, of whom 15 were eligible for genomic profile analysis and 10 patients (66.7%) had the pure type. Genomic evaluation was performed using either the FoundationOne CDx or PleSSision testing platform. Results: At least one genomic alteration occurred in 14 patients (93.3%), and the most frequently mutated gene was tumor suppressor protein p53 ( TP53), which was found in seven cases (46.7%). Additionally, five cases (33.3%) had mutations in retinoblastoma transcriptional corepressor 1 ( RB1), and all these patients had the pure type. Four cases had both TP53 and RB1 alterations, and most of these possessed the pure type and had PSA levels of less than 4.0. Compared to previous study results, the frequency of genetic alterations in the phosphoinositide 3-kinase (PI3K) pathway (n = 3; 20.0%) and Wnt-β/catenin pathway (n = 6; 40.0%) was comparable, but alterations in the DNA damage repair (DDR) pathway were few (n=1; 6.7%). None of the patients presented high tumor mutation burden or microsatellite instability. Conclusions: We found that the Japanese cohort with DAC has unique features such as high frequency of alterations in tumor suppressor gene such as TP53 and RB1, and few DDR pathway alterations, suggesting the existence of regional and racial differences in genomic profiles. Genomic analysis using next-generation sequencing is expected to be useful in elucidating the pathogenesis of DAC, and further accumulation of cases is considered important.

Pre-clinical modeling of navtemadlin pharmacokinetics (PK), pharmacodynamics (PD), and efficacy in glioblastoma, IDH-wildtype.

2012 Background: Navtemadlin (nvtm) is a potent, selective, orally available small molecule inhibitor of murine double minute 2 homologue (MDM2), which has completed a Ph 0 surgical window of opportunity study in glioblastoma, IDH-wildtype (GBM; Lee SNO 2022). To optimally interpret the clinical data, an extensive evaluation of nvtm PK, PD, and efficacy was performed in GBM patient derived xenografts (PDXs). Methods: Response to nvtm +/- radiotherapy (RT) was characterized in vitro and in vivo across a panel of GBM PDXs with various TP53 and MDM2 alterations. Efficacy, PK, and PD were evaluated in flank or orthotopic PDX models. p53 pathway activation was evaluated by qPCR, Western blot, and IHC. Nvtm brain penetration was evaluated in immunodeficient abcb1a/b-/- (P-glycoprotein; P-gp), abcg2-/- (BCRP), or double-knockout mice. Binding was assessed by rapid equilibrium dialysis, and nvtm concentrations by LC/MS-MS and MALDI-MSI. Results: Nvtm decreased in vitro viability of TP53WT GBM PDX explant cultures with IC50s &lt;100 nM, but was ineffective in TP53-mutant GBM. In flank PDX models, nvtm (100 mg/kg daily for 4 weeks) delayed tumor regrowth by 1.9 to 4.8-fold in TP53 WT PDXs without MDM2amplification (n=9 models), and more markedly by 20.1 to 29.4-fold in MDM2amplified models (n=4; 2 PDXs did not recur). In a flank PDX dose-response study, nvtm delayed the growth of MDM2amplified GBM108 at 25 mg/kg vs. placebo (61 vs. 11 days; p=0.014; mean intra-tumoral nvtm = 590 nM). In contrast, growth of MDM2 non-amplified GBM14 was delayed only at 100 mg/kg (37 vs. 15 days; p=0.082; mean intra-tumoral nvtm = 2990 nM). In both flank models, p53 target gene upregulation was similar between 25 and 100 mg/kg and did not correlate with regrowth. Nvtm was highly bound (fraction unbound = 0.090 in culture media and 0.014 in GBM tissue homogenate). In mice after oral dosing, distribution to CNS was limited by P-gp, with a brain to plasma ratio of 0.009 and unbound ratio of 0.005, whereas BCRP deficiency did not affect CNS distribution. With orthotopic GBM108, nvtm efficacy was evaluated in nude vs. efflux-deficient mice. In nude mice, nvtm was ineffective at doses up to 100 mg/kg; however, in efflux deficient mice, the mean intra-tumoral nvtm level at 25 mg/kg was similar to flank tumors (690 nM) and survival was significantly extended (134 vs. 62 days, p=0.0002) with additional benefit at 100 mg/kg (&gt;300 vs. 62 days, p=0.002). Conclusions: Nvtm showed significant efficacy in TP53 WT GBM flank PDX models but efficacy in orthotopic PDX was highly dependent on CNS penetration. Targeting nvtm levels based on the in vitroIC50 may significantly underestimate the in vivo effective concentration . In vivo studies showed lower intra-tumoral nvtm levels are required to significantly inhibit growth of MDM2amplified compared with non-amplified tumors.

High PD-L1 expression among patients with ALK rearranged non–small cell lung cancer and response to first line ALK tyrosine kinase inhibitors.

8626 Background: Patients with ALK rearrangements are highly responsive to ALK tyrosine kinase inhibitors (TKIs). Negative prognostic markers to ALK TKIs include TP53 alterations and ALK Variants 3a/b. The prognostic role that PD-L1 expression plays in first line ALK TKI outcomes is less clear. Methods: Retrospective review of patients with metastatic ALK-rearranged NSCLC seen between 2015 - 2022 at the University of Colorado who received first line ALK TKIs was performed. PD-L1 expression was denoted as high (≥50%) or low (&lt; 1% or 1-49%) based on pre-treatment biopsies. Clinicopathologic features, treatment outcomes, and genomic data were collected. Survival outcomes were assessed using Kaplan Meier methods. Multivariate modeling adjusted for brain metastases, TP53 mutations, ALK variant status, and ALK TKI. PD-L1 was overexpressed in EML4-ALK murine cell lines with lentiviruses. In vitro and in vivo response to ALK TKIs were compared to non-infected cell lines. Neutrophils and lymphocytes were quantified on pre-treatment H&amp;E slides. Results: Baseline characteristics of 130 patients are shown. Median PFS was significantly (p &lt; 0.001) longer in the PD-L1 low cohort than the PD-L1 high cohort (21 vs 11 mos, HR 0.44, 95% CI 0.29 – 0.66). There was no difference in OS between the PD-L1 low and PD-L1 high cohort (p = 0.4, 107 vs 91 mos, HR 0.86, 95% CI 0.46 – 1.61). In an alectinib sub-group analysis (n = 89), PD-L1 high expressers had a higher rate of CNS progression (93.0 vs 73.9%, p = 0.022). Forced overexpression of PD-L1 in murine EML4-ALK cell lines exerted no difference to in vitroor in vivo response to alectinib. Pre-treatment biopsies from the PD-L1 high cohort (n = 19) revealed significantly more neutrophils (p = 0.021), but not lymphocytes (p = 0.904). Conclusions: High PD-L1 expression (≥50%) is associated with worse PFS and higher intracranial progression rate among those receiving ALK TKIs. Induced PD-L1 expression in ALK cell lines did not alter in vitro or in vivo ALK TKI sensitivity. Higher neutrophil counts track with high PD-L1 expression (≥50%), suggesting that differences within the immune tumor microenvironment may be relevant. [Table: see text]