PTEN Alterations Research Articles

Utilizing Patient-derived Xenografts to Model Precision Oncology for Biliary Tract Cancer.

Biliary tract cancers (BTCs), which are rare and aggressive malignancies, are rich in clinically actionable molecular alterations. A major challenge in the field is the paucity of clinically relevant BTC models which recapitulate the diverse molecular profiles of these tumors. The purpose of this study was to curate a collection of patient-derived xenograft (PDX) models that reflect the spectrum of genomic alterations present in BTCs to create a resource for modeling precision oncology. PDXs were derived from BTC collected from surgical resections or metastatic biopsies. Alterations present in the PDXs were identified by whole exome sequencing and RNASeq. PDXs were treated with approved and investigational agents. Efficacy was assessed by change in tumor volume from baseline. Event-free survival was defined as time to tumor doubling from baseline. Responses were categorized at day 21: >30% decrease=partial response; >20% increase in tumor volume=progressive disease, and any non-PR/PD was considered stable disease. Genomic sequencing demonstrated key actionable alterations across this cohort, including alterations in FGFR2, IDH1, ERRB2, PIK3CA, PTEN and KRAS. RNAseq demonstrated fusions and expression of antibody drug conjugate targets including TROP2, HER2 and Nectin4. Therapeutic matching revealed objective responses to approved and investigational agents that have been shown to have antitumor activity clinically. Here, we developed a catalog of BTC PDXs which underwent comprehensive molecular profiling and therapeutic modeling. To date, this is one of the largest collections of BTC PDX models and will facilitate the development of personalized treatments for patients with these aggressive malignancies.

Molecular Markers in Follicular and Oncocytic Thyroid Carcinomas: Clinical Application of Molecular Genetic Testing.

Oncocytic thyroid carcinoma (OTC) was previously considered a variant of follicular thyroid carcinoma (FTC) but has recently been reclassified as a separate form of thyroid cancer. This study aimed to demonstrate that FTC and OTC are fundamentally distinct entities that can potentially be differentiated preoperatively through cytology and/or molecular testing. A retrospective chart review of patients diagnosed with FTC and OTC operated upon at two university health centers from January 2016 to September 2023 (n = 3219) was conducted. Molecular testing results were correlated with histopathologic diagnosis. Fifty patients met the inclusion criteria. FTC was identified in 27 (54.0%) patients, and OTC in 23 (46.0%) patients. Patients with OTC were older (61.8 years) than FTC patients (51.2 years) (p = 0.013). Moreover, aggressive tumors were found in 39.1% (9/23) of OTCs compared to 11.1% (3/27) of FTCs (p = 0.021). Amongst Bethesda category III and IV nodules, 17 out of 20 (85.0%) OTC cytology reports demonstrated an oncocytic subtype compared to only 5 out of 24 FTC cytology reports (20.8%) (p = 0.002). On molecular testing, the EIF1AX alteration was exclusively present in OTCs while the PAX8/PPARy and PTEN alterations were exclusively found in FTCs. Copy number alterations (CNAs) were found to be more prevalent in OTC (66.7%) compared to FTC (33.3%), and they were not indicative of tumor aggressiveness. Within the OTC group, all three patients who had a TP53 alteration were diagnosed with aggressive cancer. Lastly, the OTCs exhibited a higher frequency of multiple alterations on molecular testing (66.7%) compared to FTCs (33.3%). To our knowledge, this is the largest study to date comparing the clinical application of abnormalities found on molecular testing for FTC and OTC. It further demonstrates the distinct clinicopathological and molecular characteristics of OTC.

1643P Impact of PTEN alterations on clinical outcomes in patients (pts) with de novo metastatic prostate cancer (mPC)

1643P Impact of PTEN alterations on clinical outcomes in patients (pts) with de novo metastatic prostate cancer (mPC)

471P PTEN alteration as a predictor of second-line efficacy in patients with recurrent IDHwt-glioblastoma

471P PTEN alteration as a predictor of second-line efficacy in patients with recurrent IDHwt-glioblastoma

Phase II study of talazoparib in advanced cancers with BRCA1/2, DNA repair, and PTEN alterations

Cancer cells with BRCA1/2 deficiencies are sensitive to poly (ADP-ribose) polymerase (PARP) inhibitors. We evaluated the efficacy of talazoparib in DNA-Damage Repair (DDR)-altered patients. In this phase II trial, patients were enrolled onto one of four cohorts based on molecular alterations: (1) somatic BRCA1/2, (2) other homologous recombination repair pathway, (3) PTEN and (4) germline BRCA1/2. The primary endpoint was a clinical benefit rate (CBR): complete response, partial response or stable disease ≥24 weeks. 79 patients with a median of 4 lines of therapy were enrolled. CBR for cohorts 1–4 were: 32.5%, 19.7%, 9.4% and 30.6%, respectively. PTEN mutations correlated with reduced survival and a trend towards shorter time to progression.Talazoparib demonstrated clinical benefit in selected DDR-altered patients. PTEN mutations/loss patients derived limited clinical benefit. Further study is needed to determine whether PTEN is prognostic or predictive of response to PARP inhibitors.

The genetic duet of concurrent RASAL1 and PTEN alterations promotes cancer aggressiveness by cooperatively activating the PI3K-AKT pathway.

The significance of the prominent tumor suppressor gene for RAS protein activator-like 1 (RASAL1) could be better understood by combined genetic, clinical, and functional studies. Here, we investigated the oncogenic and clinical impacts of genetic alterations of RASAL1, particularly when coexisting with genetic alterations of the gene for phosphatase and tensin homolog (PTEN), in 9924 cancers of 33 types in the TCGA database. We found common concurrent genetic alterations of the two genes, which were cooperatively associated with activation of the phosphatidylinositol 3-kinase (PI3K)-AKT pathway, with cancer progression and mortality rates being 46.36% and 31.72% with concurrent gene alterations, versus 29.80% and 16.93% with neither gene alteration (HR 1.64, 95% CI 1.46-1.84 and 1.77, 95% CI 1.53-2.05), respectively. This was enhanced by additional tumor protein p53 (TP53) gene alterations, with cancer progression and mortality rates being 47.65% and 34.46% with coexisting RASAL1, PTEN, and TP53 alterations versus 25.30% and 13.11% with no alteration (HR 2.21, 95% CI 1.92-2.56 and 2.76, 95% CI 2.31-3.30), respectively. In the case of breast cancer, this genetic trio was associated with a triple-negative risk of 68.75% versus 3.83% with no genetic alteration (RR 17.94, 95% CI 9.60-33.51), consistent with the aggressive nature of triple-negative breast cancer. Mice with double knockouts of Rasal1 and Pten displayed robust Pi3k pathway activation, with the development of metastasizing malignancies, while single gene knockout resulted in only benign neoplasma. These results suggest that RASAL1, like PTEN, is a critical player in negatively regulating the PI3K-AKT pathway; defect in RASAL1 causes RAS activation, thus initiating the PI3K-AKT pathway signaling, which cannot terminate with concurrent PTEN defects. Thus, the unique concurrent RASAL1 and PTEN defects drive oncogenesis and cancer aggressiveness by cooperatively activating the PI3K-AKT pathway. This represents a robust genetic mechanism to promote human cancer.

HGG-31. GENOMIC CORRELATIONS WITH HEMORRHAGE RISK IN PEDIATRIC PATIENTS WITH DIFFUSE HEMISPHERIC GLIOMA

Abstract BACKGROUND Pediatric diffuse hemispheric glioma (DHG) is a histone-mutated (H3.3G34R/V) high-grade glioma with poor prognosis. Clinical observation and previous reports have identified that a subset of patients present with tumor-associated hemorrhage. Here, we present our findings from radiological review of these patients and determine genomic risk factors predictive for hemorrhage risk in this cohort. METHODS Data was abstracted through the Children’s Brain Tumor Network (CBTN) and EGAS00001004301. Transcriptomic and genomic analyses were completed in R 4.3.1 using edgeR, msigdbr, and xCell. Presence of blood products was determined on preoperative MRI by a board certified pediatric neuroradiologist. RESULTS 48 samples were available across cohorts with transcriptomic or genomic data. 10 samples (21%) had pre-operative imaging and transcriptomic data available. Initial analysis determined that 6 samples (60%) had acute/chronic hemorrhage based on radiological review. Samples with hemorrhage had increased levels of VEGFA (LFC: 3.45, p=2E-07) and CA9 (LFC: 5.97, p=8E-05) expression on the transcriptomic level suggesting that patients with high expression of these markers (AngioHi) had increased risk of hemorrhage compared to patients with low expression of these markers (AngioLo). AngioHi patients had notably higher levels of IL8 (LFC: 3.56, p=0.01) and increased macrophage populations. Genomic information was available for 15 samples (31%, 7 AngioHi, 8 AngioLo). PTEN alterations (n=4, 57%), FBXW7 alterations (n= 2, 29%), or PDGFRa driver alterations (n= 3, 43%) were found in the AngioHi cohort while none of these alterations were identified in the AngioLo cohort. Notably, at least one of these mutations was seen in all cases in the AngioHi cohort. CONCLUSIONS These findings identify potential mutational alterations predictive for tumor-associated hemorrhage in pediatric DHG. Current work is being completed to further characterize how these mutations impact vascular remodeling for patients with DHG and may offer targeted therapeutic opportunities.

Metastatic extraneural glioblastoma diagnosed with molecular testing.

Glioblastoma, the most common malignant brain tumor in adults, is associated with a median overall survival duration of less than 2 years. Extraneural metastases occur in less than 1% of all patients with glioblastoma. The mechanism of extraneural metastasis is unclear. We present a case of extensive extraneural, extraosseous, epidural, and soft-tissue metastasis of glioblastoma. The diagnosis of metastatic glioblastoma was made only after next-generation sequencing (NGS) of the metastatic paraspinal lesions was completed. The CDK4, pTERT, PTEN, and TP53 molecular alterations seen in the initial intracranial glioblastoma were found in the paraspinal tumor, along with the addition of MYC, which is implicated in angiogenesis and epidermal-to-mesenchymal transition. Immunohistochemical stains showed that neoplastic cells were negative for GFAP. In conclusion, this case raises awareness about the role of NGS in the diagnosis of extraneural glioblastoma. This diagnosis was not possible with histology alone and only became evident after molecular profiling of the metastatic lesions and its comparison to the original tumor.

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.

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.

Testing patterns and prevalence of PIK3CA, AKT1, and PTEN alterations among patients (pts) with HR+/HER2- metastatic breast cancer (mBC) in the US.

1041 Background: Biomarkers play an essential role to inform treatment decisions in mBC. In hormone receptor positive/HER2 negative (HR+/HER2-) mBC, PIK3CA/AKT1/PTEN alterations are now actionable as pt selection biomarkers with the FDA’s Nov 2023 approval of the pan-AKT inhibitor capivasertib. Here we investigate the testing patterns and prevalence of PIK3CA/AKT1/PTEN alterations in mBC. Methods: This retrospective cohort study used two partially overlapping nationwide de-identified databases: the Flatiron Health electronic health record-derived database (FH DB) to assess next-generation sequencing (NGS) testing rates, and the Flatiron Health-Foundation Medicine clinico-genomic database (FH-FMI CGDB) to assess prevalence among tested. Pts were included if they were ≥18 years old and diagnosed with HR+/HER2- mBC from 1/1/17—6/30/22. Prevalence was assessed using any sample type or timing of testing post-mBC diagnosis and defined as pathogenic/likely pathogenic alterations as captured in FMI NGS testing. All analyses were summarized using descriptive statistics. Results: 8,049 (FH DB) and 2,912 (FH-FMI CGDB) pts were eligible. Overall, 37% of pts with mBC received NGS testing (3,002/8,049). Among pts in FH-FMI CGDB, 98% had PIK3CA tested (2,857/2,912) and 97% also had AKT1/PTEN tested (2,838/2,912) in the metastatic setting. The median time from mBC diagnosis to initial test was 148 days for PIK3CA and 188 days for AKT1/PTEN, and 92-93% of testing occurred after the start of first-line (1L) therapy (Table). 82% of PIK3CA testing was on tissue and 37% on liquid biopsy, with 19% of pts receiving both tissue and liquid biopsy. 77% of AKT1/PTEN testing was on tissue and 28% on liquid biopsy, with 5% of pts receiving both tissue and liquid biopsy. At least one PIK3CA/ AKT1/ PTEN alteration was detected in 55% of the pts (1,557/2857) tested for any of the markers, regardless of sample type. Conclusions: &lt;40% of pts with mBC received NGS testing from 2017-2021, suggesting opportunities for improving pts’ access to testing. Coupled with the high rate of PIK3CA/AKT1/PTEN alterations, NGS testing should be incorporated in routine clinical practice to identify optimal treatments. [Table: see text]

Genomic alterations (GA) in ESR1, PIK3CA, AKT1, and PTEN in HR(+)HER2(-) patients (pts) with metastatic breast cancer (MBC): Co-occurrence and prevalence along treatment course.

1060 Background: The standard of care for HR(+) HER2(-) MBC is evolving with new biomarker-guided drugs, including novel selective estrogen receptor degraders (SERDs) for pts with ESR1 mutations (mut) and PIK3CA and AKT inhibitors for pts with alterations in the PI3K pathway (PIK3CA, AKT1, and PTEN). This study aimed to evaluate the co-occurrence and the prevalence of these GA in different metastatic lines of therapy in real-world practice for MBC. Methods: This study included pts with HR(+) HER2(-) MBC who underwent genomic testing using Foundation Medicine tissue or liquid comprehensive genomic profiling assays, with specimens collected within 60 days of therapy initiation. Clinical data were obtained by the nationwide (US-based) de-identified Flatiron Health and Foundation Medicine real-world clinicogenomic breast cancer database (FH-FMI CGDB), originated from ~280 US cancer clinics (~800 sites of care) between 01/2011-06/2023. The prevalence of ESR1mut, PIK3CAmut, AKT1, PTENmut, and PTEN copy loss were calculated in tissue (TBx) and liquid biopsies (LBx) collected at time of 1st, 2nd, and 3rd lines of therapy. The co-occurrence of GA in TBx was evaluated. Results: Among patients with a TBx collected around therapy initiation (n=2,536), 81.5% (2,066) was collected at 1st, 10.2% (258) at 2nd, and 8.3% (212) at 3rd line. Among patients with a LBx collected around therapy initiation (n=545), 47% (256) was collected at 1st, 33.2% (181) at 2nd, and 19.8% (108) at 3rd line. In TBx, the prevalence of GA detected in 1st, 2nd, and 3rd lines were 58.6% (1,210), 67.4% (174), and 71.2% (151), respectively. In LBx, the prevalence was 42.7% (121), 65.7% (119), and 60.1% (65), respectively. Among LBx with tumor fraction ³1%, the prevalence was 67.9% (74/109), 89.1% (81/91), and 84.3% (43/51). Among patients with a GA detected in TBx 1st (n=1,210), 2nd (n=174), and 3rd (n=151) lines, both ESR1mut and a PIK3 pathway GA was detected in 7.4% (90), 12.1% (21), and 23.8% (36), respectively. Conclusions: In our dataset, 42-59% of patients with HR(+) HER2(-) MBC have at least one GA detected by TBx or LBx in 1st line, with prevalence increasing in later lines of therapy to 60-71% due to acquisition of an ESR1mut. PTEN copy loss is detected at much higher rates in TBx than LBx, consistent with the known inherent limitations of ctDNA testing. Serial genomic testing with LBx with reflex TBx if alterations are not identified should strongly be considered with the recent approvals of several molecularly targeted agents for HR(+) HER2(-) MBC. [Table: see text]

Abstract PS17-03: Capivasertib and fulvestrant for patients with aromatase inhibitor-resistant HR positive/HER2-negative advanced breast cancer: exploratory analysis of PFS by AKT pathway gene from the Phase 3 CAPItello-291 trial

Abstract Background: In the Phase 3 randomized, double-blind CAPItello-291 trial, the addition of capivasertib (a potent, selective pan-AKT inhibitor) to fulvestrant in patients with aromatase inhibitor-resistant, hormone receptor (HR)-positive/human epidermal growth factor receptor 2 (HER2)-negative (HER2− defined as immunohistochemistry [IHC] 0, or 1-positive or IHC2-positive/in situ hybridization-negative) advanced breast cancer (ABC) significantly improved progression-free survival (PFS) versus placebo + fulvestrant (hazard ratio [HR] 0.60, 95% confidence interval [CI] 0.51–0.71; p&amp;lt; 0.001). PFS benefit was observed in patients with detectable AKT pathway alterations (HR 0.50, 95% CI 0.38–0.65; p&amp;lt; 0.001) and without (0.70; 95% CI; 0.56–0.88). Here, we report PFS by gene within the AKT pathway-altered population. Methods: Patients were randomized 1:1 to receive fulvestrant (500 mg intramuscularly on days 1 and 15 of cycle 1, and day 1 of each subsequent 28-day cycle) with either placebo or capivasertib (400 mg twice daily; 4 days on, 3 days off). AKT pathway-alteration status (at least one qualifying alteration in the genes PIK3CA, AKT1, or PTEN) was determined post-randomization, using next-generation sequencing in tumor tissue. HRs were calculated using Cox proportional hazards models. Data cut-off Aug 15, 2022. Results: Of the 708 patients randomized to treatment, 289 (41%) had AKT pathway-altered tumors (capivasertib-fulvestrant n=155; placebo-fulvestrant n=134). In the AKT pathway-altered population, 43% had liver metastases and 40% primary endocrine therapy resistance. Prior therapy for advanced disease included: 89% of patients with ≥1 line of prior treatment, 71% with a prior cyclin-dependent kinase 4 and 6 inhibitor, and 18% with prior chemotherapy. Baseline characteristics were broadly balanced between treatment groups. Most patients with an AKT pathway-altered tumor had only one detectable alteration (272/289, 94%). Thirteen patients (capivasertib-fulvestrant n=4; placebo-fulvestrant n=9) had co-occurring PIK3CA and PTEN alterations, and four patients (capivasertib-fulvestrant n=2; placebo-fulvestrant n=2) had co-occurring PIK3CA and AKT1 alterations. Consistent PFS benefit of capivasertib-fulvestrant over placebo-fulvestrant was observed across all alterations (Table). The safety profile of capivasertib-fulvestrant in the AKT pathway-altered population was consistent with the overall population. Conclusions: Compared with fulvestrant alone, the addition of capivasertib to fulvestrant provided a consistent PFS benefit across alterations in all three key genes within the AKT pathway in patients with HR-positive/HER2-negative ABC. https://clinicaltrials.gov/: NCT04305496 Funding: CAPItello-291 is sponsored by AstraZeneca. Editorial acknowledgment: AstraZeneca-funded medical writing support was provided by Suzanne Patel, Ph.D., from BOLDSCIENCE Inc. Capivasertib was discovered by AstraZeneca subsequent to a collaboration with Astex Therapeutics (and its collaboration with the Institute of Cancer Research and Cancer Research Technology Limited). Table. Citation Format: Sacha Howell, Hope Rugo, Mafalda Oliveira, Florence Dalenc, Javier Cortés, Henry Gómez, Xichun Hu, Komal Jhaveri, Petr Krivorotko, Sibylle Loibl, Serafin Morales Murillo, Meena Okera, Yeon Park, Joo Hyuk Sohn, Masakazu Toi, Eriko Tokunaga, Lyudmila Zhukova, Andrew Lloyd, Elza de Bruin, Coumaran Egile, Celina D’Cruz, Nicholas Turner. Capivasertib and fulvestrant for patients with aromatase inhibitor-resistant HR positive/HER2-negative advanced breast cancer: exploratory analysis of PFS by AKT pathway gene from the Phase 3 CAPItello-291 trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS17-03.

Abstract PO1-06-14: Preliminary results of a phase I dose escalation study of eribulin in combination with copanlisib in patients with metastatic triple negative breast cancer (TNBC)

Abstract Background: Aberrant PI3K pathway signaling is frequently observed in TNBC. Increasing evidence indicate that PI3K pathway activation maintains the stemness and chemoresistance of breast cancer stem cells (CSCs). PI3K inhibition sensitizes CSCs to chemotherapy. Eribulin (E), a non-taxane microtubule dynamics inhibitor, showed overall survival benefit in metastatic HER2 negative BC. Preclinically, the addition of copanlisib (C), a potent pan-class I PI3K inhibitor that is highly selective against α and δ PI3K isoforms, improved anti-tumor effect in both E-sensitive and resistant TNBC patient-derived xenograft models, irrespective of PIK3CA/PTEN alteration status. Here we report the preliminary data of the phase I dose escalation trial of E+C in patients with metastatic TNBC. Methods: This trial includes a phase I study to determine the dose limiting toxicity (DLT) and recommended phase 2 dose (RP2D) of E+C, followed by a randomized phase II study of E+C (at RP2D) versus E (stratified by tumor PIK3CA/PTEN mutation status) with progression-free survival (PFS) as the primary endpoint. Key eligibility included patients with: metastatic TNBC who progressed on ≤5 lines of chemotherapy, prior anthracycline/taxane chemotherapy (unless contraindicated), ECOG 0-1, adequate organ function and known tumor PIK3CA/PTEN/AKT mutation status. Key exclusions include: prior receipt of E or any PI3K/mTOR/AKT inhibitor, grade ≥2 neuropathy, presence of tumor AKT mutation, and uncontrolled diabetes, hypertension or congenital QT prolongation. A 3+3 design was used to determine the DLT and RP2D. Dose level 1 (DL1) started with E at 1.1 mg/m2 mg IV and C at 45 mg IV on days (D) 1/8 of 21-D cycle, with the potential to escalate to 2 additional dose levels – DL2: E 1.4 mg/m2 and C 45 mg on D 1/8; DL3: E 1.4 mg/m2 and C 60 mg on D 1/8. Results: Eight patients were enrolled in the phase I study and received at least 1 dose of E+C. The median age was 48.5 (range, 26-67) years. The majority of patients identified as White (additionally 2 Hispanic, 1 Black) and had an ECOG performance status of 1 (n=6, 75.0%). Tumor PIK3CA and PTEN alterations were each observed in 1 patient. Median number of cycles received were 2 (range, 1-5). All 8 patients had at least 1 treatment-related adverse event (TRAE). The most common TRAEs across all dose levels were: lymphopenia (n=5, 62.5%), neutropenia (n=5, 62.5%), hyperglycemia (n=4, 50.0%), anemia (n=4, 50.0%) and fatigue (n=4, 50.0%); most toxicities were grade (G) 1-2. G3 hyperglycemia and hypertension were observed in 1 patient each, were medically managed and did not lead to treatment interruption. 1 patient was diagnosed with pulmonary embolism, possibly attributed to study therapy and breast cancer. DLTs were reported in 2 of 2 patients treated at DL2 - 1 patient developed G3 rash responsive to oral corticosteroids, and 1 patient developed G3 acute kidney injury in the setting of hypovolemia and discontinued study therapy. No additional patients were enrolled in DL2. No DLTs were observed in all 6 patients treated at DL1. 1 patient treated at DL1 had possible G1 non-infectious pneumonitis that resolved spontaneously on short interval imaging. Stable disease was observed in 2 of 6 response-evaluable patients (33.3%). 1 patient achieved regression of cutaneous disease following 1 cycle of therapy as assessed by the treating investigator. DL1, E at 1.1 mg/m2 plus C at 45 mg IV, was determined to be the RP2D. Tumor tissue biomarker analysis is underway. Conclusion: These data indicate that E+C at DL1 is a well-tolerated regimen warranting further investigation. The study is actively enrolling patients to the phase II study. Clinical trial information: NCT04345913. Funding Source: National Cancer Institute. Citation Format: Nusayba Bagegni, Brittney Haas, Leslie Nehring, Jingqin Luo, Laura Kennedy, Meghna Trivedi, Manali Bhave, Rabih Said, Cynthia Ma. Preliminary results of a phase I dose escalation study of eribulin in combination with copanlisib in patients with metastatic triple negative breast cancer (TNBC) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-06-14.

Abstract PO2-27-12: Genomic determinants of benefit to nivolumab (NIVO) plus low dose ipilimumab (IPI) among patients (pts) with hypermutated HER2-negative metastatic breast cancer (MBC): results of NIMBUS trial

Abstract Background: Genomic determinants of benefit to immune checkpoint inhibitors (ICI) among pts with MBC and high tumor mutational burden (TMB-H) are largely unknown. NIMBUS was an open-label, single-arm, multicenter, phase 2 study assessing the efficacy of NIVO 3 mg/kg intravenously every 14 days plus IPI 1 mg/kg IV every 6 weeks in pts with TMB-H HER2-negative MBC. The objective of this current study was to evaluate genomic profiling of pts included in NIMBUS and its association with benefit to ICI. Methods: In NIMBUS, eligible pts were required to have measurable HER2-negative MBC, TMB ≥9 Mut/Mb assessed by a cancer-gene panel evaluating &amp;gt; 300 genes and performed in a CLIA-certified laboratory, and 0-3 prior lines of chemotherapy in the advanced setting. The primary objective was overall response rate (ORR) according to RECIST 1.1. Secondary objectives include progression-free survival (PFS), and overall survival (OS). We performed genomic analyses on the pre-treatment tissue, and on baseline and on treatment plasma from pts run on the Foundation Medicine Inc (Boston, MA) FoundationOne®CDx (F1CDx®) and FoundationOne®Liquid CDx (F1LCDx®) panel, respectively. Results: A total of 30 pts (21 with estrogen receptor (ER)-positive and 9 with triple-negative breast cancer) were included in this study. After a median follow-up of 30.6 months, 6 pts (20%) achieved partial response (PR) for a confirmed ORR of 20% (95% CI: 7.7-38.6%). Exploratory analyses did not reveal differences in ORR based on ER status, PD-L1 status, or stromal TIL. The median PFS was 1.4 months (95% CI: 1.3 – 4.6), and the median OS was 16.9 months (95% CI: 7.1-not reached). Pts who achieved PR had a 3-year OS of 75% versus 25% in non-responders. A total of 29 pts (6 responders and 23 non-responders) had at least one pre-treatment genomic panel performed: 16 had both, 5 had pretreatment F1CDx only and 8 had F1LCDx panel only. Blood TMB (bTMB) was highly correlated with tissue TMB by F1CDx (r = 0.95). Using data from the most recent genomic panel available, (24 for baseline blood, and 5 for pretreatment tissue), the most frequently mutated genes ( &amp;gt; 30%) in this population were: TP53 (66%); PIK3CA (45%), ESR1 (35%), CDH1 (35%), ROS1 (31%). ESR1 and PTEN mutations were associated with absence of PR while PALB2 was associated with objective response. We observed a significant correlation between ESR1 mutations and a decrease in OS among pts with ER+ disease. Among the entire 29 pts, a negative correlation between PTEN with PFS and OS, and positive correlation between PALB2 with PFS was observed (Table). Among the 24 pts with baseline F1LCDx panel evaluable, the median TMB was 7.6 Mut/Mb ([interquartile range 0.0–113.8). Median TMB was 8.9 Mut/Mb and 41.7 Mut/mB among pts without and with PR, respectively. Baseline TMB &amp;gt; 20 mut/Mb was found in all pts with PR and in one of those without response. Paired serial blood samples demonstrated that among responding pts, TMB was significantly reduced from baseline to end of treatment (EOT) compared to those with no clinical benefit [n=16, p=0.0015]. Lastly, the tumor fraction (TF) estimates from 7 paired baseline to EOT samples demonstrated that all responders had a decrease or no change in TF compared to an increase in non-responders [n=7, p=0.017]. Conclusion: In the NIMBUS trial, all pts with PR to NIVO plus low dose IPI had a TMB &amp;gt; 20 Mut/Mb and had a decrease in TMB during treatment. ESR1 and PTEN mutations were associated with lack of benefit, while PALB2 mutation was associated with increased benefit. If validated, these results could help tailor the use of ICI among pts with TMB-H MBC. Association between genomic alterations in ESR1, PTEN and PALB2 and benefit to nivolumab and low dose ipilimumab among patients with HER2-negative MBC and TMB-H HR: hazard ratio; MBC: metastatic breast cancer; mOS: median overall survival (months); mPFS: median progression-free survival (months); TMB-H: tumor mutational burden above &amp;gt; 9 mutations/megabase. Citation Format: Romualdo Barroso-Sousa, Tyler Chinsky, Tianyu Li, Sangeetha Reddy, Leisha Emens, Beth Overmoyer, Saud AlDubayan, Hoyin Chu, Paulina Lange, Julie Kasparian, Ameer Basta, Molly DiLullo, Victoria Attaya, Melissa Hughes, Nancy Lin, Nabihah Tayob, Rinath Jeselsohn, Elizabeth Mittendorf, Sara Tolaney. Genomic determinants of benefit to nivolumab (NIVO) plus low dose ipilimumab (IPI) among patients (pts) with hypermutated HER2-negative metastatic breast cancer (MBC): results of NIMBUS trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-27-12.

Abstract PO5-13-06: Classifying HER2-low breast cancer using a combination of ERBB2 mRNA expression and altered genes

Abstract Background It has been recently demonstrated that some HER2-low, defined as immunohistochemistry (IHC) 1+ or 2+ with no gene amplification by FISH, breast cancer (BC) patients respond to trastuzumab deruxtecan (T-DXd). Results of the DAISY trial also suggested clinical activity with T-DXd in some of those classified as HER2-0 cancers by IHC, indicating an unmet need to better identify cancers that may truly respond to T-DXd. We compared ERBB2 (HER2) mRNA expression obtained from whole-transcriptome sequencing with IHC/FISH assignment of HER2 status. Furthermore, we constructed a classifier using ERBB2 expression and alterations in other genes as a proof of concept to illustrate classification of BC samples into HER2-0 versus HER2-low status. Methods We chose consecutive HR positive/HER2-negative BC samples that were submitted for OncoExTraTM testing between April 2020 and October 2022. The OncoExTra test uses tumor-normal whole exome and whole transcriptome sequencing to detect somatic single base substitutions, indels, copy number alterations, gene fusions and alternative transcripts. In addition, we investigated the expression profile of select genes for this analysis. We utilized OncoExTra data to examine the relationship between HER2 IHC status and gene expression, mutation and copy number profiles of selected genes, and built a classifier to predict HER2 status. Results A total of 279 BC samples were analyzed, including 106 HER2-IHC 0, 120 HER2-IHC 1+ and 53 HER2-IHC 2+. Of these, 273 were primary and 6 were metastatic samples. The distributions of patient age and tumor fraction were similar among these categories (p=0.54 and p=0.41, respectively; one-way ANOVA). ERBB2 expression differed among all three HER2-IHC categories and was lowest in HER2-0 and highest in HER2-2+ (HER2-0 vs HER2-1+, p&amp;lt; 0.001; HER2-0 vs HER2-2+, p&amp;lt; 0.0001; HER2-1+ vs HER2-2+, p=0.011; HER2-0 vs HER2-low, p&amp;lt; 0.0001; Mann-Whitney U test in all comparisons). Five genes were mutated in more than 10% of samples: PIK3CA (45.5%), GATA3 (15.4%), CDH1 (12.5%), MAP3K1 (11.1%) and TP53 (10.8%). Mutations in two genes, GATA3 and PTEN, appeared to differ in frequency between the HER2-0 and HER2-low categories (Table 1). We selected 4 genes with mutations, 2 genes with copy number changes, and expression from ERBB2, ESR1, CD274, HLA-A, HLA-B and HLA-C to train a logistic regression classifier to predict HER2 IHC status. The classifier was trained on 163 samples, hyperparameter tuning was done using 55 samples, and testing was performed on 61 samples. Preliminary results indicated that of the 28 HER2-IHC 0 samples in the test set, 17 (65%) were classified as HER2-low. Further development and evaluation of the classifier using a blinded testing set is currently in progress. Conclusion In this cohort of BC samples, ERBB2 mRNA expression was positively associated with HER2 protein IHC status. Additional studies are required to determine whether a classifier including ERBB2 mRNA expression could be used to identify patients for T-DXd therapy. The relative enrichment of PTEN alterations in HER2-IHC 0 samples suggests that mTOR/AKT inhibitors may sometimes be appropriate in patients who are not eligible for T-DXd therapy, and further investigation should be considered. Table 1. Number (frequency) of altered genes by HER2 IHC status. Citation Format: Gargi Basu, Niru Chennagiri, Turgut Dogruluk, David Hall, Jess Hoag, Cynthia Flannery, Snehal Thakkar, Melanie Palomares, Frederick Baehner, Lajos Pusztai. Classifying HER2-low breast cancer using a combination of ERBB2 mRNA expression and altered genes [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-13-06.

Thyroid nodules with DICER1 mutation or PTEN alteration: A comparative cytologic, clinical, and molecular study of 117 FNA cases.

DICER1 mutations and PTEN alterations are increasingly detected by thyroid fine-needle aspiration (FNA). Both are associated with nodular thyroid disease and cancer. The authors analyzed a large comparative thyroid FNA cohort with DICER1 mutation or PTEN alteration. A total of 117 thyroid FNAs with DICER1 or PTEN alterations were retrieved from the databases of two academic medical institutions. Demographic, clinical, and radiologic data were collected; FNA slides were analyzed for 29 cytomorphologic features. Of 117 thyroid FNAs, 36 (30.8%) had DICER1 mutation and 81 (69.2%) showed PTEN alteration. The DICER1 cohort had 33 (91.7%) females and three (8.3%) males (mean, 40.9 years); 61.8% had multinodular disease. FNAs were classified as atypia of undetermined significance (AUS), 23 (63.9%); follicular neoplasm (FN), 12 (33.3%); and malignant, 1 (2.8%). The PTEN subgroup had 66 (81.5%) females and 15 (18.5%) males (mean, 55.2 years) with increased multinodular disease (93.8%, p=.0016). PTEN FNAs had greater cytologic diversity: non-diagnostic, 2 (2.5%); benign, 5 (6.2%); AUS, 44 (54.3%); FN, 24 (29.6%); and malignant, 6 (7.4%). Both DICER1 and PTEN cases showed a range of resected tumor subtypes. The DICER1 cohort included thyroblastoma, and the PTEN group included anaplastic carcinoma. The cytomorphology of DICER1 and PTEN cases showed overlapping features, especially microfollicular patterns. Minor cytomorphologic differences included papillary patterns in DICER1 (p=.039), and oncocytic changes (p<.0001) in PTEN. DICER1 and PTEN FNAs reveal many cytologic similarities. DICER1 patients are younger, and PTEN patients had multinodular disease. Awareness of these genetic cohorts can identify patients at risk for thyroid cancer.

SMARCA4 Mutations in Gastroesophageal Adenocarcinoma: An Observational Study via a Next-Generation Sequencing Panel.

The clinical impact of SMARCA4 mutations (SMARCA4ms) in gastroesophageal adenocarcinoma (GEA) remains underexplored. This study aimed to examine the association of SMARCA4ms with clinical outcomes and co-occurrence with other gene mutations identified through a next-generation sequencing (NGS) panel in GEA patients. A total of 256 patients with metastatic or recurrent GEA who underwent NGS panel profiling at the MD Anderson Cancer Center between 2016 and 2022 were included. Comparative analyses were performed to assess clinical outcomes related to SMARCA4ms. The frequency and types of SMARCA4ms and their co-occurrence with other gene mutations were also examined. SMARCA4ms were identified in 19 patients (7.4%). These SMARCA4ms were significantly associated with non-signet ring cell subtype (p = 0.044) and PD-L1 positive expression (p = 0.046). No difference in survival between the SMARCA4m and SMARCA4-normal group was observed (p = 0.84). There were significant associations between SMARCA4ms and FANCA, IGF1R, KRAS, FANCL, and PTEN alterations. Notably, 15 of the 19 SMARCA4m cases involved SNV missense mutations, with frequent co-occurrences noted with TP53, KRAS, ARID1A, and ERBB2 mutations. These results serve as the first comprehensive examination of the relationship between SMARCA4ms and clinical outcomes in GEA.

Abstract 1620: Multiregion spatial sequencing of early-stage head and neck squamous cell carcinoma revealed theranostic intratumor heterogeneity

Abstract Introduction: Head and neck squamous cell carcinoma (HNSCC) is characterized by substantial intratumor heterogeneity (ITH) at the molecular level that may foster tumor evolution, adaptation, and therapeutic failure. Personalized-medicine strategies mostly rely on single tumor-biopsy samples which might not capture the mutational burden of heterogeneous tumors. We investigated genetic ITH portrayed from multiregion spatial sequencing in early-stage HNSCC patients. Patients and Method: Seventy-eight early-stage HNSCC patients treated with upfront curative-intent surgery in the prospective biobanking SCANDARE study (NCT03017573) were included. We developed a pathological measure of ITH, based on the relative contribution of multiple poorly and well-differentiated intratumor regions (n = 199) in resected tumor samples. To characterize genetic ITH, we performed targeted next-generation sequencing on multiple spatially separated intratumor regions by using a custom panel of 571 genes (named DRAGON), in combination with 3’-Tag RNA sequencing and pathway enrichment analysis. Results: Fifty-nine patients displaying a minimum of two sequenced intratumor regions (total, n = 180) were analyzed. We observed genetic ITH in 22/59 patients (37%), of whom 6/22 (27%) based on theranostic molecular alterations. Three of the six patients exhibited molecular alterations in PIK3CA (n = 2) and TSC2 (n = 1) that have supporting clinical evidence of actionability in some cancer types (OncoKB level 1) and three patients exhibited actionable molecular alterations in PTEN (n = 2) and CDKN2A/B (n = 1) supported by substantial research evidence (OncoKB level 4). Two patients exhibited molecular alterations in SMARCA4 that is likely oncogenic, albeit not yet annotated by OncoKB. The pathological measure of ITH accounted for a significant source of variation in gene expression. Notably, we identified 346 differentially expressed genes imposing p-values ≤ 0.05 and |log fold-change| ≥ 1 involved in several Reactome’s signaling pathways, including the cell junction organization (R-HSA-446728) and the tight junction interaction (R-HSA-420029) pathways. Conclusions: We reported clinically relevant genetic ITH in early-stage HNSCC, which can lead to underestimation of the mutational landscape portrayed from single tumor biopsy and may present major challenges to precision medicine. Our findings uncovered the biology behind ITH and may have translational implications for biomarkers discovery in HNSCC. Citation Format: Grégoire Marret, Sophie Vacher, Constance Lamy, Ladidi Ahmanache, Laura Courtois, Choumouss Kamoun, Maral Halladjian, Zakhia El Beaino, Jerzy Klijanienko, Charlotte Martinat, Emmanuelle Jeannot, Joey Martin, Guillaume Rougier, Nathalie Badois, Maria Lesnik, Antoine Dubray-Vautrin, Olivier Choussy, Wahib Ghanem, Rabah Taouachi, Julien Masliah Planchon, Nicolas Servant, Maud Kamal, Ivan Bièche, Christophe Le Tourneau. Multiregion spatial sequencing of early-stage head and neck squamous cell carcinoma revealed theranostic intratumor heterogeneity [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 1620.