Genomic Landscape Research Articles

Molecular alterations in claudin 18 suppressed and non-suppressed gastric adenocarcinomas to guide targeted therapies.

Gastric adenocarcinoma represents an aggressive type of cancer and an important cause of cancer mortality. Progress in gastric cancer therapeutics has resulted from a better understanding of the molecular pathogenesis of the disease and introduction of targeted therapies, but most gastric cancer patients still rely on non-targeted chemotherapy as the mainstay of treatment for advanced disease. An analysis of publicly available series from The Cancer Genome Atlas (TCGA) gastric cancer cohort was undertaken to delineate the clinical and genomic landscape of gastric cancers with suppressed expression of claudin 18 compared with cancers with non-suppressed claudin 18. Claudin 18 suppressed cancers were defined as having an mRNA expression z-score relative to normal samples (log RNA Seq V2) of less than -1. Claudin 18 non-suppressed cancers were defined as having an mRNA expression z-score relative to normal samples (log RNA Seq V2) above 0.5. Gastric cancers with claudin 18 mRNA suppression represented 7.7% of the gastric adenocarcinomas of TCGA cohort, while non-suppressed cancers represented 46.6% of the cases. The two groups did not differ in clinical and genomic characteristics, such as mean age, histology, grade, and stage. The mutation landscape of claudin 18 suppressed cases included high mutation rates of TP53, of genes of the WNT/β-catenin pathway and of ubiquitin ligase FBXW7. Moreover, a subset of both claudin 18 suppressed and non-suppressed cancers displayed mutations in Mismatch Repair (MMR) associated genes or a high tumor mutation burden (TMB). At the mRNA expression level, claudin 18 suppressed gastric cancers showed up-regulation of EMT core transcription factor Snail 2 and down-regulation of genes of HLA cluster. The survival of gastric cancer patients with claudin 18 mRNA suppression was not significantly different compared with patients with non-suppressed claudin 18. Sub-sets of gastric cancers with claudin 18 mRNA suppression displayed characteristics of potential therapeutic interest, such as mutations in WNT and PI3K pathways and MMR defects. These may guide the development of alternative targeted therapies, in this sub-set of gastric cancers which are not candidates for claudin 18 targeting therapies.

Abstract PO5-06-14: Genomic Landscape and Clinical Outcomes of Triple-Negative Invasive Lobular Carcinoma

Abstract Background: Triple-negative (TN) invasive lobular carcinoma (ILC) of the breast is a rare entity, comprising roughly 2% of primary ILC. Clinicopathologic studies have shown that TN ILC is associated with more pleomorphism, higher nuclear grade, older patient age, and worse disease-related outcomes compared to estrogen receptor-positive (ER+) ILC. Intrinsic subtyping indicates that TN ILC less frequently expresses basal markers than TN invasive ductal carcinoma (IDC). This prompted investigation of the genomic landscape and clinical management of TN ILC to provide insight into potential therapeutic approaches. Methods: Twenty patients with TN ILC were included in this study. Three patients were treated at a single academic center and underwent comprehensive genomic profiling (CGP) of breast or metastatic tissue; clinicopathologic details were obtained from their electronic health records. Seventeen patients with TN ILC who underwent CGP of breast or metastatic biopsies were included from the Foundation Medicine, Inc. database. PD-L1 testing for all patients was determined by immunohistochemistry (IHC) using the VENTANA SP142 or Dako 22C3 commercial assays. HER2 expression was determined by IHC. Negative HER2 expression was defined as an IHC score of 0, or IHC 1+ or 2+ with non-amplified fluorescence in situ hybridization (FISH). Results: In the Foundation Medicine database of patients with TN ILC, the most frequent genomic alterations were in CDH1 (15/17 patients [88.2%]), TP53 (10/17 patients [58.8%]), and PIK3CA (10/17 patients [58.8%]). Short variant mutations in ERBB2 were found in 3/17 patients (17.6%). Fourteen patients were HER2-low (HER2 IHC 1+ or 2+ with non-amplified FISH). No cases were MSI high or expressed PD-L1 (8 of 17 cases underwent PD-L1 assessment by SP142). Median tumor mutational burden (TMB) was 5 muts/Mb. Among the three patients with TN ILC treated at a single center, the most common somatic mutations were in CDH1 and TP53, and no cases were MSI high or expressed PD-L1 (by SP142 or 22C3). All three patients had pleomorphic ILC, histologic grade 2 or 3, and HER2 IHC 0. One patient with mpT1cN1a(sn) ER+/HER2- ILC of the right breast in 2011 received adjuvant TAC (docetaxel, doxorubicin, cyclophosphamide), breast/axillary radiation, and ten years of adjuvant endocrine therapy. She was diagnosed with TN ILC of the left supraclavicular nodes in 2022; she did not exhibit a response to docetaxel and carboplatin with pembrolizumab, and she subsequently received palliative radiation followed by capecitabine. Another patient with locally advanced TN ILC progressed on the KEYNOTE-522 regimen, immediately developed metastases, and is currently receiving palliative sacituzumab govitecan. The third patient with cT3N1 TN ILC progressed on neoadjuvant therapy (through the I-SPY 2 trial) with oral paclitaxel, encequidar, and dostarlimab, followed by dostarlimab with dose-dense doxorubicin and cyclophosphamide. Her surgical pathology demonstrated ypT3N3a disease with 10 × 7 cm of tumor, and she developed skin metastases on the chest wall within weeks post-operatively. She received comprehensive chest wall and nodal radiation and is currently receiving palliative capecitabine with neratinib given a high variant allele frequency driver somatic ERBB2 mutation. Conclusions: The most common somatic mutations among TN ILC patients included in this study were in CDH1, TP53, and PIK3CA. Currently the management of TN ILC is extrapolated from that of TN IDC, however our patients did not respond to chemoimmunotherapy. The presence of HER2-low status or somatic ERBB2 mutations may provide opportunities for treatment with an anti-HER2 antibody-drug conjugate or tyrosine kinase inhibitor. Studies with larger sample sizes of TN ILC are needed for molecular profiling and assessment of outcomes with specific therapeutic approaches. Citation Format: Hemali Batra-Sharma, Smruthy Sivakumar, Prashanth Ashok Kumar, Ethan Sokol, Rebecca Shatsky, Jeffrey Ross. Genomic Landscape and Clinical Outcomes of Triple-Negative Invasive Lobular Carcinoma [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 PO5-06-14.

Abstract PO3-16-01: Personalized circulating tumor DNA assay precisely predicts the response of neoadjuvant chemotherapy in breast cancer patients

Abstract Introduction Circulating tumor DNA (ctDNA), which is detected in the blood as cell free DNA fragment from tumor cells, could indicates genomic landscape, tumor burden and treatment response during chemotherapy as a non-invasive method. Especially, personized ctDNA assay by next-generation sequencing(NGS) is able to detect a trace amount of ctDNA and precisely figure out the change of the amount of ctDNA during NAC and follow up period after curative surgery. Here, we performed serial ctDNA evaluations in EBC patients who diagnosed as TNBC or HER2-positive BC and received NAC followed by curative surgery. We aimed to predict NAC response and detect minimal residual disease (MRD) using personalized ctDNA assay. Methods CtDNA was detected by AlphaLiquid®Detect, a tumor-informed personalized MRD detection assay exploring most of the mutations in tumor. Whole exome sequencing (WES) of tumor tissue and peripheral blood mononuclear cells (PBMCs) was performed. Patient-specific somatic mutations were selected using a proprietary algorithm. In brief, clonal variants were prioritized using integrated information including variant allele frequency, population allele frequency databases, somatic variant databases, variant pathogenicity, and genomic context. Up to 100 variants were selected for patient monitoring. A hybridization capture panel consisting of a pool of 4 patients’ selected variants was synthesized. These bespoke panels (BSPs) were used for ctDNA detection. Patients with more than 2 tumor-derived mutations detected in plasma were considered as ctDNA positive. Inclusion criteria included patient who diagnosed as stage IIA-IIIC BC planned to NAC followed by curative surgery. Among BC subtypes, TNBC and HER2-positive BC were allowed. Collection of specimens and associated clinical data used in this study was approved by the Institutional Review Board of Samsung Medical Center (IRB File No.2021-02-033), and we received informed consents for this study. Results From May 2021 to Sep 2022, 158 patients has been enrolled. Archival tissues were not available in 47 patients and tissue WES had failed in six patients. Therefore, 105 patients were enrolled and analyzed their ctDNA at diagnosis based on tissue WES data. Of 105 patients, Median age of BC patients was 49.3 years of age (range: 26.1, 67.8). Thirty-six patients (34.3%) were post-menopausal status and others (65.7%) were pre-menopausal status. In BC pathology at diagnosis, 56 (53.3%) were TNBC and other 49 (46.7%) were HER2-positive BC. Among HER2-positive BC, hormone receptor (HR)-positive were in 21 (20.0%). In clinical stage at diagnosis, stage II were 46 (43.8%) and stage III in 59 (56.2%). In 105 patients, median number of somatic mutations was 60 (interquartile range [IQR]: 38.5, 91). CtDNA detection rate at BC diagnosis was 90.5% and all of BC which not be detected ctDNA was HER2-positive BC with clinical stage II. Median amount of ctDNA at diagnosis was 20 (IQR: 6, 44). In the number of somatic mutations, there was no difference according to BC subtypes (P=0.121) and clinical stage (P=0.700) but the amounts of ctDNA at diagnosis was different. CtDNA was much more detected in TNBC compared to HER2-positive BC (Median: 34.5 vs. 15.9; P< 0.001) and clinical stage III compared to clinical stage II (Median 33.2 vs. 19.5; P=0.002). After NAC, ctDNA at curative surgery was tested in 70 patients. In 70 patients, pathologic complete response(pCR) was observed in 44 patients. CtDNA was detected in ten patients (14.3%) and eight did not achieve pCR and two with pCR (Specificity of assay: 95.5%, positive predictive value: 0.80, P=0.032). Conclusions Personalized ctDNA assay can precisely detected ctDNA at diagnosis and their detection rate was associated with BC subtypes and clinical stage. In addition, ctDNA at curative surgery also can predict NAC response in EBC patients. Long term ctDNA follow up of MRD would be warranted. Citation Format: Ji-Yeon Kim, Hae Hyun Jung, Jeong Eon Lee, Seok Won Kim, Hwang-Phill Kim, Sunghoon Heo, Jongseong Ahn, Tae-You Kim, Young-Hyuck Im. Personalized circulating tumor DNA assay precisely predicts the response of neoadjuvant chemotherapy in breast cancer patients [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 PO3-16-01.

Abstract PO4-15-06: Genomic landscape characterization after exposure to cyclin dependent kinase 4/6 inhibitors: a retrospective multi-institutional consortium analysis

Abstract Background: Limited data are available to determine the best therapeutic strategy for hormone-receptor positive (HR+) HER2 negative (HER2-) advanced breast cancer (ABC) after progression on cyclin dependent kinase 4/6 inhibitors (CDK4/6i). The aim of this study was to characterize the genomic and prognostic profile of patients (pts) that experienced disease progression after first-line therapy with a CDK4/6i. Methods: The study retrospectively analyzed a multi-institutional cohort of 75 patients (pts) with HR+/HER2- ABC after experiencing progression on first-line endocrine therapy (ET) with CDK4/6i and characterized by circulating tumor DNA (ctDNA) next-generation sequencing (Guardant 360). Oncogenic pathways (i.e., RTK, RAS, RAF, MEK, NRF2, ER, WNT, MYC, p53, cell cycle, notch, and PI3K) were defined based on previous research (Sanchez-Vega et al. Cell, 2018). Associations across single nucleotide variations (SNVs), copy number variations (CNVs), and pathway classification were tested by uni- and multivariable logistic regression with respect to a CDK4/6i naïve group comprising 247 patients with HR+/HER2- ABC (control). Prognosis was analyzed through Cox regression for overall survival (OS). Result: Our study cohort included 75 ABC pts who experienced progression after first line CDK4/6i plus ET: 43 pts (57.3%) were treated with ET while 31 (41.3%) with a non-ET- second line therapy. The most common histology was invasive ductal carcinoma (70.6%), 37.3% of pts were negative for progesterone expression and 26.7% had de novo metastatic disease. The PI3K (33.3%), p53 (28%) and ER (25.3%) pathways were the most mutated in the CDK4/6i-treated cohort. Comparing our population with a CDK4/6i naïve cohort (N=247), the multivariate analysis showed a higher prevalence in the study cohort of SNVs mutations in: i) ESR1 gene [Odds ratio (OR) 2.93; p=0.005]; ii) cell-cycle pathway (OR 4.24; p=0.033); iii) ER pathway (OR 2.04; p= 0.034). Moreover, in the study cohort a numerical but not significant increase of RB1 SNVs was also observed. Multivariable analyses of the 43 pts that received ET second line therapy showed a negative prognostic impact with TP53 SNVs in both progression-free survival (PFS) [Hazard ratio (HR) = 3.34; 95% CI: 1.15-9.66, p=0.026] and OS (HR = 3.85; 95% CI: 1.52-9.77, p=0.005). The prognostic role of p53 pathway mutations was also confirmed for both PFS (HR = 4.71, 95% CI: 1.64-13.50; p=0.004) and OS (HR = 3.25; 95% CI: 1.27-8.3; p=0.014). The potential interaction between the prognostic oncogenic pathways and the outcome was investigated according to the treatment strategy (ET vs non-ET). A consistent impact was observed across the above prognostic pathways both for PFS and OS. A numerical difference was observed in terms of PFS in pts with p53 pathway mutations that underwent non-ET second line. Conclusions: The genomic landscape of progressive disease after CDK4/6i exposure is significantly different from the genomic alterations detectable in treatment-naïve pts, which could potential impact future treatment strategies for patients with disease progression after adjuvant CDK4/6i exposure. Our data suggest that the choice of second-line treatment could potentially be guided by the identification of actionable mutation by ctDNA, although further prospective studies are needed to validate the clinical utility of this approach. Citation Format: Letizia Pontolillo, Carolina Reduzzi, Andrew Davis, Lorenzo Gerratana, Arielle Medford, Katherine Clifton, Whitney L. Hensing, Marko Velimirovic, Ami N. Shah, Jeannine Donahue, Laura Munoz Arcos, Charles S. Dai, Jennifer Keenan, Amir Behdad, William Gradishar, Emilio Bria, Cynthia Ma, Aditya Bardia, Massimo Cristofanilli. Genomic landscape characterization after exposure to cyclin dependent kinase 4/6 inhibitors: a retrospective multi-institutional consortium analysis [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 PO4-15-06.

Abstract ED02-01: Biological peculiarities of ILC

Abstract One in 8 women will be diagnosed with breast cancer in her lifetime and approximately 15% of them have invasive lobular breast cancer (ILC). ILC differs in many ways from the most common BC subtype, called non-special type (NST) (Van Baelen et al. Ann Oncol 2022). The microscopic hallmark of ILC is the presence of single files of relatively small discohesive tumor cells, which stems from a dysfunctional E-cadherin-catenins complex at the intercellular junctions, a feature that is uncommon in NST. More than 90% of ILC express the estrogen (ER) and/or progesterone receptor (PR) and lack HER2 amplification, while this is only the case in ~65% of NST. Patients with ILC also show a distinct clinical behavior as compared to NST: generally, a poor response to chemotherapy, more late recurrences, and a peculiar metastatic pattern. The first part of the presentation will focus on presenting the biological knowledge we have accumulated over the years about primary ILC. We will review the studies which compared ILC and NST at the genomic, transcriptomic and immune level. The second part of the presentation will be focused on metastatic breast cancer. Here we will review the published evidence regarding the metastatic dissemination of ILC, its genomic landscape and the contribution of the local tumor microenvironment. We will also dedicate attention to the important biological and clinical knowledge which can be gained by studying samples collected from patients with metastatic ILC who consented to post-mortem tissue donation. Finally, the last part of the presentation will be dedicated to mixed NST/ILC tumors. These tumors account for up to 18% of all invasive breast cancers and represent an even more under investigated area of research as pure ILC. Citation Format: C. Desmedt. Biological peculiarities of ILC [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 ED02-01.

Abstract PO2-03-07: Immune Landscape in PD-L1-positive and TROP2-positive Triple-Negative Breast Cancer

Abstract Background: Triple-negative breast cancer (TNBC) tends to be more aggressive, and more likely to recur than other subtypes of breast cancer, and patients with metastatic TNBC have a poor prognosis. The recent approvals and data of pembrolizumab (anti-PD-1 inhibitor) in combination with chemotherapy in advanced-metastatic and early high-risk TNBC, and sacituzumab govitecan (an ADC that targets the tumor cell surface antigen TROP2 coupled with the irinotecan metabolite SN-38) in metastatic TNBC, has shed light on some of the immune and cancer signaling associated with TNBC. Although exciting progress has been made in the treatment of TNBC, there are still important gaps to fill to understand the biology of TNBC and identify additional therapies. Materials and Methods: We identified a total of 1500 patients whose tumors were negative for the expression of ER and HER2 receptors by gene array. PD-L1 and TROP2 expression data were available for 802 early TNBC patients, of whom 657 were treated with adjuvant chemotherapy. A comprehensive identification and analysis of immune-related genes (n= 1169) was conducted. Mann-Whitney test comparing the gene expression for all immune related-genes was evaluated in patients expressing PD-L1 and TROP2 at low and high expression levels. Immune gene modules resembling key immune gene signaling were analyzed in all subgroups identified. Fold changes and Mann-Whitney P values were calculated in each subgroup. Results: Patients were stratified based on high and low expression levels of PD-L1 and TROP-2 mRNA. PD-L1 highly expressing (PD-L1+) tumors showed a marked difference in immune-related genes as compared to PD-L1 low-level tumors (PD-L1-) with PD-L1+ tumors demonstrating a significantly higher proportion of correlated immune genes (CXCL9/10/11, IFN-γ, CCL5 and STAT1). TROP2 highly and low expressing tumors (TROP2+ and TROP2-, respectively) showed a relatively limited difference in terms of immune-related gene expression with key immune-related genes having statistical differential expression in the two subgroups. TROP2+ tumors showed a high presence of several Mucin family gene members (MUC1, -5AC, -4). In addition, CLDN 4, VTCN1, and MET were among the key immune genes significantly expressed in TROP2+ tumors (P = 5.73E-20; P = 6.2E-13, P = 2.1E-11). Overall, PD-L1+ tumors and TROP2+ tumors showed an immune suppressive genomic landscape but were potentially regulated by different signaling. Of note, tumors with very high expression of PD-L1 showed an inverse correlation with VTCN1 gene expression. Conclusions: This preliminary analysis confirmed that only a limited number of early TNBC highly expressed PD-L1 and that a different immune suppressive landscape for PD-L1± and TROP2± tumors exists. In addition, our data suggest that early TNBC cases lacking PD-L1 might escape immune surveillance by virtue of the upregulation of key alternate signaling. Ongoing efforts investigating the expression of such markers on stroma/epithelial tumor components will help to clarify their biological, clinical, and potential therapeutic role in TNBC. Citation Format: Balazs Gyorffy, Libero Santarpia. Immune Landscape in PD-L1-positive and TROP2-positive Triple-Negative Breast Cancer [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 PO2-03-07.

Abstract PO2-16-03: ESR1 genomic alterations (GAs) and coexistent putative resistance alterations in comprehensive genomic profiling (CGP) of metastatic breast cancer (MBC)

Abstract Background: ESR1 mutations (ESR1mut) are an established biomarker of endocrine therapy (ET) resistance in patients (pts) with hormone receptor positive (HR+) MBC. Moreover, ESR1mut acquired in response to standard-of-care ET now confer access to novel ET recently approved by health authorities. However, the nuanced genomic landscape beyond the simple presence or absence of ESR1mut may be critically important. We report on detailed ESR1 GAs detected by tissue and liquid biopsies (TBx, LBx) and examine the prevalence of co-occurring GAs in other genes reported to confer intrinsic or acquired resistance to ET. Methods: CGP results from MBC biopsies were retrospectively analyzed (TBx 33,653 pts; LBx 7,134 pts) using FoundationOne® CDx and FoundationOne® Liquid CDx, hybrid-capture based sequencing platforms profiling 324 cancer-related genes. Foundation Medicine’s ctDNA tumor fraction (TF) is a composite algorithm prioritizing aneuploidy at higher levels to avoid germline signal and prioritizing variant allele frequency (VAF) of canonical alterations at lower levels to maximize dynamic range. Results: ESR1mut were detected in 3,755 (11.2%) TBx and 1,513 (21.2%) LBx. Among samples with ESR1mut, multiple ESR1mut were more common in LBx (19% 2, 12% 3+) than in TBx (6% 2, 0.5% 3+), with up to 10 different ESR1mut seen in one LBx. The most common mutations were D538G (39% and 52% of TBx and LBx with ESR1mut), Y537S (27%, 38%), Y537N (10%, 15%), and E380Q (10%, 14%). Activating ESR1 insertion-deletion mutations (indels) were detected in 81 TBx and 62 LBx (2% and 4%), representing 23 distinct variants at the protein level, including V422del (107), indels affecting codons 532-538 (29), G344_L345insC (7), and Y328_S329del (6). 28 fusions of the ESR1 DNA binding domain to 13 unique gene partners were detected; 10/28 (36%) were ESR1-CCDC170 fusions. ESR1 amplifications (ESR1amp) were detected in 704 (2.1%) TBx; 62 (8.8%) also harbored ESR1mut. ESR1amp were detected among 34 (0.5%) LBx; 8 (24%) also had ESR1mut. In TBx with ESR1mut and LBx collected from the same pt within 8 months (43 pairs), LBx sensitivity of detection of ESR1mut was 96% (26/27, 95% confidence interval CI [95%CI]: 79%-99%) when TF was ≥1%, but only 6% (1/16, 95%CI: 0.3%-32%) when LBx had TF < 1%. In 988/1,513 (65%) of LBx with ESR1mut, ≥1 mutations or fusions in genes associated with alternative ET resistance pathways were detected, including PIK3CA (46% of ESR1mut LBx), RB1 (12%), PTEN (10%), NF1 (7%), ARID1A (7%), AKT1 (5%), FGFR2 (4%), and KRAS (4%) and ERBB2 (3%). Prevalence of co-occurring GA associated with ET resistance was higher in LBx than in TBx (RB1, NF1, FGFR2, PTEN, EGFR, KRAS, PIK3CA, FDR < 0.001). Moreover, prevalence was higher in 240 LBx where ESR1 was a minor allele (VAF/TF < 10%) compared to 1,273 LBx were ESR1 was a major allele (FGFR2, PIK3CA, FGFR3, BRAF, ERBB2, RB1, FDR < 0.05). In pts with ESR1mut LBx and a historical TBx collected < 5 years earlier (182 pairs), GAs in RB1, FGFR2, KRAS, EGFR, and BRAF were detected primarily on the LBx but not the preceding TBx. Conclusions: CGP detects a wide spectrum of mutations in ESR1, including missense and indel mutations and fusions. When LBx TF is < 1%, sensitivity for ESR1mut is reduced and repeat testing should be considered. ESR1mut can coexist with complementary or competing resistance mechanisms, particularly when ESR1 is a minor allele, which could impact benefit from novel ET approved for patients with ESR1 mutations. Currently, CGP of LBx informs therapeutic recommendations for pts with HR+ MBC based on the binary presence or absence of ESR1mut; however, the clinical implications of the ESR1 mutation spectrum described herein and the potential impact of co-expression of other key pathway signals warrant further investigation. Citation Format: Heather McArthur, Hanna Tukachinsky, Alexa Schrock, Russell Madison, Oliver Holmes, Smruthy Sivakumar, Ethan Sokol, Ryon Graf, Julia Quintanilha, Kali Dougherty, Lincoln Pasquina, Geoffrey Oxnard, Mia Levy, Eric Winer. ESR1 genomic alterations (GAs) and coexistent putative resistance alterations in comprehensive genomic profiling (CGP) of metastatic breast cancer (MBC) [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 PO2-16-03.

Abstract PO2-20-05: Onco-Genome Brazil: Mapping Breast and Prostate Cancer in the Brazilian Public Health System

Abstract Background: Breast and prostate cancers are the most common malignancies diagnosed in women and men respectively, and present with great clinical heterogeneity, even in tumors with the same phenotypic characteristics. Somatic and germline molecular alterations in DNA may have prognostic and predictive impact, influencing response to therapies and overall survival. In addition, the evaluation of germline mutations can help identify individuals and families at higher risk for developing cancer. Aims: Our aim is to characterize the somatic and germline genomic landscape of women with locally advanced HER2-positive breast cancer and men with metastatic prostate cancer from Brazil. Secondarily, we aim to identify genetic variants associated with tumor prognosis and treatment response, identify patients carrying pathogenic alterations in cancer-predisposing genes, and characterize the genetic ancestry of the population included in the study. Trial design: This observational multicenter cohort study (NCT05306600) will include 550 adult patients from the five macro-regions of Brazil, divided into two arms: 1) breast cancer and 2) prostate cancer. Clinical and pathological data will be collected, as well as DNA samples from peripheral blood and tumor tissue. Whole-exome sequencing (WES) will be performed to identify variants that may be drivers and/or actionable in a specific patient or tumor. These variants will be interpreted and classified according to their population frequencies, in silico predictors, functional studies, and literature data, following international guidelines proposed by expert societies. This trial is supported by the Ministry of Health in Brazil and includes patients of the Public Health System. Eligibility criteria: In arm 1, the inclusion criteria are patients with a histological diagnosis of breast carcinoma with overexpression of HER-2, clinical stage II or III, and undergoing neoadjuvant treatment with chemotherapy plus trastuzumab. In arm 2, the criteria are patient with histological diagnosis of prostate adenocarcinoma, currently in clinical stage IV. Patients without availability of biological material collection are excluded from the study, as well as patients in whom there was no central confirmation of required histology. Statistical methods: Univariate and multivariate analyses will be conducted to evaluate the possible effect of specific genes or mutations on the investigated outcomes, including tumor prognosis and treatment response. Discussion: Currently, 265 participants have already been included in the study, 150 in arm 1 and 115 in arm 2. This trial will contribute to the construction of a robust database that should provide a better understanding of the genomic profile of patients with breast and prostate cancer in Brazil. For more information about the trial, please contact Daniela Rosa (daniela.rosa@hmv.org.br). Citation Format: Jaqueline Schuch, Cláudia Bordignon, Mahira Rosa, Angélica de Baumont, Marina Bessel, Gabriel Macedo, Daniela Rosa. Onco-Genome Brazil: Mapping Breast and Prostate Cancer in the Brazilian Public Health System [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 PO2-20-05.

Abstract PO4-14-02: Enhancing Informative Outcomes with Liquid Biopsy in a Real-World Population of Patients with Advanced Breast Cancer: Analysis of the SOLTI-1903 HOPE Study

Abstract Background: The integration of next-generation sequencing (NGS) into routine clinical practice is often hindered by financial and logistical challenges, resulting in diagnostic and therapeutic disparities among patients. Addressing these issues, the SOLTI-1903 HOPE study (NCT04497285) seeks to evaluate the feasibility of implementing a molecular screening program that actively involves patients with advanced breast cancer (ABC) in the management of their disease. By empowering patients, this study aims to gain a comprehensive understanding of the genomic landscape of ABC and to facilitate patient access to matched targeted therapies in Spain Methods: In the SOLTI-1903 HOPE study, a patient-centric approach was adopted, allowing patients living anywhere in Spain, and diagnosed with ABC, to actively lead their inclusion, participation, and follow-up using a digital tool (DT). Patients signed an informant consent, and then provided clinical information and underwent a liquid biopsy (LBx) using the Guardant360 panel at a local laboratory. LBx samples could be obtained at the moment of progressive disease (PD), when a new line of treatment was started < 8 weeks before extraction (PT), or during treatment with stable disease o partial response lasting more than 8 weeks (SD/PR). Clinical and molecular findings were evaluated by a Molecular Advisory Board (MAB), which generated a comprehensive report explaining the observed alterations and listing potentially beneficial matched targeted treatments. The primary endpoint of this subanalysis was to compare the genomic outcomes between LBx samples obtained at SD and those obtained at PD or PT. In the study, tissue samples were also collected and analysed by Foundation but this analysis will be reported in other communications. Results: From October 2020 to May 2023, a total of 273 blood samples were obtained from 253 patients diagnosed with ABC, with the majority being HR+/HER2- (81%), followed by HER2-positive (13%) and triple-negative (6%). Mean turnaround time was 10.9 days from blood collection to final result. Despite nearly 40% of patients being SD or PR, in 79.9% of the LBx (n=218), one or more genomic alterations were detected, and among these, 74.0% (n=202) were classified as pathogenic. Among the 74 genes analyzed by the Guardant360 panel, 8 genes were classified as tiers I-III according to the ESCAT levels of clinical relevance. In 150 LBx samples (54.9%), one or more pathogenic alterations were associated with ESCAT levels I-III, while only the remaining 52 LBx samples (19.1%) with pathogenic alterations were not associated with any ESCAT levels I-III. Among the analyzed LBx samples, a total of 928 distinct gene alterations were identified. Of these, 650 alterations (70.1%) were classified as pathogenic, while 278 (29.9%) were categorized as variants of unknown significance or lacked a reported impact. Notably, 25% (n=323) of the pathogenic gene alterations were further classified as ESCAT I-III. Out of the 273 LBx samples, 140 were obtained at PD, 30 at PT, and 103 at SD/PR. Circulating tumor DNA (ctDNA) was detected in 91.4% of PD samples, 86.7% of PT samples, and 62.1% of SD/PR samples (p< 0.001). Alterations classified as ESCAT levels I-III were observed in 70% of PD samples, 60% of PT samples, and 33% of SD/PR samples (p< 0.001). In 20 patients, LBx was repeated, with 16 of them having the first LBx obtained during SD/PR and the second LBx collected during PD or PT. The detection rate of ctDNA increased significantly from 18.7% (n=3) to 87.5% (n=14) (p< 0.001). The number of LBx samples with ESCAT levels I-III alterations also showed a significant increase from 12.5% (n=2) to 68.7% (n=11) (p=0.0032). Conclusions: Our findings highlight the importance of obtaining LBx samples during disease progression -compared to SD/PR- to gain greater genomic informativeness and better guide personalized treatment approaches for patients with advanced breast cancer. Citation Format: Tomás Pascual, Elia Seguí, Ruben Olivera-Salguero, Juan Miguel Cejalvo, Mafalda Oliveira, Pablo Tolosa, Maria Vidal, Marcos Malumbres, Joaquín Gavilá, Cristina Saura, Sonia Pernas, Rafael López, Mireia Margelí, Judith Balmaña, Montserrat Muñoz, Isabel Blancas, Valentina Boni, Eva Ciruelos, Elena Galve, Antonia Perelló, Rodrigo Sánchez-Bayona, Susana De La Cruz, Miguel De La Hoya, Aura Blanch-Torras, Rosana Sposito, Jordi Canes, Helena Masanas, Rosa Olmos, Margarita Forns, Maria José Prieto, Ana Casas, Aleix Prat. Enhancing Informative Outcomes with Liquid Biopsy in a Real-World Population of Patients with Advanced Breast Cancer: Analysis of the SOLTI-1903 HOPE Study [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 PO4-14-02.

Abstract PO4-02-02: Genomic Landscape of ER positive HER2-low early breast cancers in the FLEX Study: MammaPrint, BluePrint and whole transcriptome analysis

Abstract Background: Antibody-drug conjugates (ADCs) continue to emerge for the treatment of a new subset of patients with HER2-low breast cancer. There is limited evidence to demonstrate HER2-low tumors as a distinct biological subtype and why/if these tumors benefit from ADCs. To improve our understanding of this newly defined HER2 category of breast cancers, we evaluated clinical characteristics, MammaPrint (MP), BluePrint (BP), and the whole transcriptomic profile of HER2-low breast cancers in FLEX study. Methods: FLEX (NCT03053193) is a prospective, observational trial that includes stage I-III breast cancer patients who undergo MammaPrint testing (with or without BluePrint) as standard of care, and consent to full transcriptome and clinical data collection. In this study, clinically ER+/HER2- tumors were analyzed. The HER2-low cohort group (n=1698) was defined as HER2 IHC 1+ (ISH positive excluded) and IHC 2+, ISH Negative, and the HER2-0 group (n=1181) was defined as HER2 IHC 0. MP classified tumors as Low Risk or High Risk (further stratified as High 1 and High 2). MP together with BP categorized tumors as Luminal A (MP Low Risk), Luminal B (MP High Risk), HER2 or Basal. Two-tailed proportional z-test was used to compare clinical features and genomic subtypes of HER2-low vs. HER2-0 and the limma R package for differential gene expression analysis (DGEA). P-values were adjusted for multiple testing by Benjamini-Hochberg; significant differentially expressed genes (DEGs) had a p-value < 0.05 and a fold change >2. Results: In this FLEX study, clinically ER+/HER2- tumors showed that the clinical characteristics between HER2-low and HER2-0 did not differ significantly except higher percentage of premenopausal within HER2-low (23% vs 17%, p < 0.01). Of all HER2-low patients, 46% were MP High Risk as within HER2-0 patients (44%, p = 0.27). In both groups with MP High Risk tumors, there was a higher frequency of MP High 1 (83% in HER2-low, 80% in HER2-0) compared to MP High 2 (17% in HER2-low, 20% in HER2-0). BP subtyping showed similar distribution for Luminal subtype between HER2-low and HER2-0, but the frequency of ER+, Basal subtype was lower in HER2-low (2.5%), compared to HER2-0 (4.5%) (p=0.005). Principal component analysis (PCA) of the 500 most variable genes did not reveal a separation of HER2-low and -0 tumors, but clustering was apparent when tumors were classified by BP. DGEA within Basal tumors revealed no DEGs. Within Luminal A tumors, more than 1800 DEGs were identified, and within Luminal B tumors, nearly 300 DEGs were identified. However, all DEGs were < 1.5-fold change: mean, max (1.09, 1.38) for Luminal A and (1.12, 1.44) for Luminal B. In addition, a significant difference (p< 0.01) towards increased ERBB2 (HER2) expression was detected from HER2-0 to HER2-low, but there was a large overlap of expression between the 2 groups. Conclusion: Our study showed that HER2-low and HER2-0 tumors are clinically and biologically similar. The differences in ERBB2 and low-level genome wide expression detected between HER2-low and HER2-0 should be further investigated to determine how these changes affect tumor biology and treatment benefit. The biological heterogeneity among IHC-defined HER2-negative tumors was better captured by MammaPrint and BluePrint than IHC. MammaPrint identified 53% of HER2-low tumors as Low Risk, a subgroup of patients known to have good outcomes without chemotherapy. Our data suggest a subset of patients with HER2-low, MP Low Risk tumors could be spared from the potential toxicities of ADCs. Future studies will investigate the utility of MammaPrint and BluePrint in predicting chemosensitivity and benefit from ADCs, such as T-DXd, in patients with HER2-low tumors. Citation Format: Abirami Sivapiragasam, Adam Brufsky, Hannah Linden, Natasha Hunter, Cynthia Osborne, Sami Diab, Robert Maganini, Joyce O'Shaughnessy, Manojkumar Bupathi, Sung Ho Lee, Theodore Y Kim, William Audeh, Josien Haan, Andrea Menicucci, Lavanya Samraj, FLEX Investigators' Group. Genomic Landscape of ER positive HER2-low early breast cancers in the FLEX Study: MammaPrint, BluePrint and whole transcriptome analysis [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 PO4-02-02.

Abstract PO5-14-05: Circulating tumor DNA fraction correlates with residual cancer burden post-neoadjuvant chemotherapy in triple negative breast cancer patients

Abstract Background: Pathologic response after preoperative/neoadjuvant chemotherapy is a continuum that can range from no residual cancer to extensive disease. Viable cancers can shed tumor DNA into the blood and we hypothesized that plasma ctDNA levels can reflect pathologic response as continuum. Patients and methods: Post-neoadjuvant plasma samples were obtained after completion of chemotherapy but before surgery from patients with stage I to III TNBC who received treatment with durvalumab concurrent with weekly nab-paclitaxel followed by dose dense doxorubicin/cyclophosphamide in a single arm neoadjuvant clinical trial (NCT02489448). Up to 3 mL of blood was collected into EDTA- and heparin- containing vacutainer tubes, and plasma cfDNA was extracted and twice purified in the lab at Predicine Inc. Pathologic response was quantified on a continuous scale using the Residual Cancer Burden (RCB) score, and also categorized as RCB-0 (complete pathologic response, n=21), -I (minimal residual cancer, n=7), -II (moderate residual cancer, n=13), and -III (extensive residual cancer, n=3). Median follow-up was 35 months (range 25-70 months). Tumor whole exome sequencing (WES) data was previously generated using HiSeq 4000 at the Yale Center for Genome Analysis (dbGaP: PRJNA558949). A personalized MRD panel of up to 60 personalized variants per patient was designed based on the tumor WES data, and ultra-deep next-generation sequencing (100,000X) of plasma cfDNA was performed using the PredicineBEACON assay for MRD detection by Predicine. CtDNA positivity (MRD) was defined as a weighted score equivalent to 2 or more tumor variants detected in the plasma. cfDNA tumor fraction was measured based on mutant allele fraction of observed variants. Adjusted tumor fraction could be confidently measured for 37 out of 44 patients that had enough confidently called personalized ctDNA variants. We compared tumor fraction versus RCB score as continuous variables, and also compared tumor fraction between RCB categories as ordinal variables. Invasive disease-free survival was examined by Kaplan-Meier analysis. Bioinformatic data analysis was performed using Graphpad Prism and R. Results: 44 of 48 plasma samples yielded successful cfDNA generation and sequencing. Six patients (13.6%) were MRD positive. Among these 6 cases, 3 had RCB-0 response (14.3% of RCB 0 cases), 1 had RCB II response (7.7% of RCB II cases), and 2 had RCB III response (66.7% of RCB III cases). The median cfDNA tumor fraction of patients with pathological complete response to treatment (pCR) was significantly lower than patients with residual disease (RD) (0.06% vs. 0.3%, p=0.02). There was a positive correlation between RCB scores and tumor fraction (r= 0.45, Spearman). Tumor fraction was more strongly correlated with later recurrence than a binary ctDNA positivity call. Tumor fraction was significantly higher in patients that later experienced local or metastatic recurrence (n=8) compared to those with no evidence of disease (n=29) (Wilcoxon test, p=0.02). Both ctDNA positive RCB III cases had metastatic recurrence while the remaining 4 ctDNA positive cases currently remain recurrence-free. Mutational genomic landscape of both tumor WES and plasma ctDNA samples identified hotspot mutations in 19 cancer-associated genes. Novel hot-spot mutations were observed in the plasma of two patients following treatment. TP53 mutations were highly prevalent in tissue and plasma, however no significant association of mutation of any gene with treatment response was detected. Conclusions: PredicineBeacon MRD assay was successfully performed on 3 mL plasma samples collected using from EDTA and heparin-containing tubes. Tumor fraction correlated with residual disease, RCB score and disease recurrence. Citation Format: Naing Lin Shan, Billie Gould, Kim Blenman, Julia Foldi, Pan Du, Frank Zhang, Myles Walsh, Lajos Pusztai. Circulating tumor DNA fraction correlates with residual cancer burden post-neoadjuvant chemotherapy in triple negative breast cancer patients [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 PO5-14-05.

Abstract PO5-15-02: Integration of synonymous mutations and variants of unknown significance for basal and longitudinal characterization of metastatic breast cancer by circulating tumor DNA

Abstract Background: Despite how commonly synonymous mutations (SYN) and unknown significance (VUS) are detected in tissue and circulating tumor DNA (ctDNA) next-generation sequencing (NGS), the potential for these somatic changes to impact personalized approaches in metastatic breast cancer (MBC) is unclear. Emerging evidence suggests that SYN mutations may play a role in cancer biology through changes in mRNA stability, splicing, and gene expression. The aim of this study was to characterize SYN and VUS in a large multicenter consortium and to analyze their interplay with pathogenic variants (PATH). Methods: The study retrospectively analyzed a multi-institutional cohort comprising 1189 patients with MBC characterized for ctDNA using NGS (Guardant360™, Guardant Health) before treatment start (BL) and at disease progression (PD). Pathway classification was defined based on prior research (Sanchez-Vega F et al, Cell. 2018). Single Nucleotide Variations (SNVs) were annotated for oncogenicity (OncoKB) and protein domain (UniProtKB). Associations between clinical characteristics and pathway classifications for SYN and VUS were explored by multinomial logistic regression; survival was tested through Cox regression in terms of overall survival (OS). Results: Hormone-receptor positive (HR+)/HER2 negative (luminal-like) was the most represented subtype (68.7%) followed by HER2-positive (14.6%), and triple-negative (TNBC) (16.7%). PATH were mainly detected in the TP53 DNA binding domain, ESR1 ligand binding domain, helical and catalytic domains of PIK3CA, SYN in the ligand binding domain of ESR1, MET Sema, KIT protein kinase domain, and FGFR1 Ig-like C2-type 1. In contrast, VUS were mainly found in the ESR1 ligand binding domain, MET Sema, ATM PI3K/PI4K catalytic domain, DDR2 Protein kinase and PIK3CA C2 PI3K-type. Analyzing pathways, after multivariable multinomial logistic regression, RAS SYN, PI3K and ESR PATH were significantly associated with HER2+ MBC (respectively RRR=4.14, P=0.046; RRR = 0.474, P < 0.001 and RRR = 0.306, P < 0.001). TNBC was associated with PATH PI3K (RRR = 0.366, P < 0.001), PATH P53 (RRR = 4.71, P < 0.001) and PATH RAS (RRR = 0.362, P = 0.026, 95% CI 0.147 - 0.888). In luminal-like MBC, a significant impact on OS was observed for SYN P53 (HR = 2.53, P = 0.008) and PATH ER (HR = 1.54, P < 0.001), PATH P53 (HR = 1.56, P < 0.001), PATH cell-cycle (HR = 1.64, P = 0.0299), and PATH RAS (HR = 1.62, P = 0.010). An impact on OS was observed in TNBC for SYN cell-cycle (HR = 5.41, P = 0.006), VUS ESR (HR = 2.87, P = 0.018), PATH P53 (HR = 1.84, P = 0.012) and PATH cell-cycle (HR = 5.07, P < 0.001). VUS MYC (HR = 38.5, P = 0.001) and PATH PI3K3 (HR = 2.32, P = 0.022) were significant among HER2 positive. Differences in SYN, VUS and PATH alterations were then compared between baseline and progressive disease. While a significant increase in mutant allele frequency (MAF) was observed for PATH (P=0.0010) and VUS (P=0.0152) at PD, no differences were highlighted for SYN (P=0.8362). Conclusions: In our cohort we characterized the MBC for PATH, VUS and SYN utilizing the ctDNA. In this study, considering the complex genomic landscape of MBC, we demonstrated that both SYN and VUS mutations had an impact on the survival outcome. Therefore, further analysis to validate the prognostic and predictive role of SYN and VUS, with or without PATH mutations, are needed to better characterize and monitoring of MBC through ctDNA. Moreover, functional studies are needed to better understand their role in the nuanced biology of MBC. Citation Format: Elisabetta Molteni, Carolina Reduzzi, Andrew Davis, Lorenzo Foffano, Arielle Medford, Katherine Clifton, Whitney L. Hensing, Marko Velimirovic, Ami N. Shah, Laura Munoz Arcos, Charles S. Dai, Jennifer C. Keenan, Elyssa Denault, William Gradishar, Giuseppe Damante, Amir Behdad, Lorenzo Gerratana, Fabio Puglisi, Cynthia Ma, Aditya Bardia, Massimo Cristofanilli. Integration of synonymous mutations and variants of unknown significance for basal and longitudinal characterization of metastatic breast cancer by circulating tumor DNA [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 PO5-15-02.

Abstract PS12-02: Exploration of ctDNA Dynamics in the PACE Trial: A Randomized Phase II Study of Fulvestrant, Palbociclib, and Avelumab for HR+/HER2- Metastatic Breast Cancer

Abstract Background The PACE trial for HR+/HER2- metastatic breast cancer (MBC) prospectively evaluated whether continuation of the CKD4/6 inhibitor (CDK4/6i) palbociclib (P) beyond progression on prior CDK4/6i and aromatase inhibitor (AI), with a change in endocrine therapy (ET) to fulvestrant (F), improved outcomes beyond change to F alone, as well as explored the activity of a palbociclib, fulvestrant, and avelumab (P+F+A) triplet. The primary analysis did not show improvement in progression-free survival (PFS) with F+P versus F alone. In this analysis, an evaluation of serial translational PACE samples was performed to gain insights into the mutational landscape of HR+/HER2- MBC after prolonged exposure to CDK4/6i, evaluate early mutational dynamics with ongoing therapy, and identify potential genetic markers predictive of treatment response. Additionally, the identification of acquired mutations at the time of tumor progression could provide valuable information about mechanisms of treatment resistance. Methods PACE is a multicenter randomized open-label investigator-initiated phase II trial. Eligible patients (pts) had HR+/HER2- evaluable MBC with prior progression on AI and any CDK4/6i after > 6 months (mo) of therapy in the MBC setting, or during/within 12 mo in the adjuvant setting, with no more than 1 prior line of chemotherapy for MBC. Pts were randomized 1:2:1 to F alone, F+P, or F+P+A, with tumor assessments every 8 weeks. Correlative samples including circulating tumor DNA (ctDNA) were collected at baseline (BL), times of tumor assessments (C3D1), and end of study/progression (EOT). Targeted NGS (Guardant360) to evaluate mutations (somatic single nucleotide variants and indels) was performed. Kaplan-Meier survival curves were used for assessing PFS and the log-rank test for comparisons. McNemar’s test with continuity correction was used to test the changes in the frequency of mutations. Results Of the 220 randomized pts, 211 contributed ctDNA samples (200 at BL, 129 at C3D1, 146 at EOT). Characteristics of the correlative science population were similar to the intention to treat population. At BL, the most common mutations observed were ESR1 (57%), TP53 (38%), PIK3CA (36%), GATA3 (20%) ATM (12%) and Rb1 (12%). The presence of a PIK3CA mutation vs WT-PIK3CA at BL was associated with shorter PFS in the F alone arm (p=0.001) but not in the F+P or F+P+A arms. ESR1 mutations present at baseline were D538G (34%), Y537S (21%), Y537N (15%), E380Q (10%) and L536H (4%). At C3D1, the most common mutations observed were ESR1 (47%), TP53 (33%), PIK3CA (26.1), GATA3 (18.1), ATM (13%) and PTEN (8.6%). Including only BL and C3D1 matched samples (N=124), the prevalence of ESR1 mutations decreased after 2 cycles of treatment (57% at BL vs 44% at C3D1), predominantly observed in the F+P+A arm. For those in any arm with at least 6 months of disease stability on study (N=53), the presence of an ESR1 D538G mutation decreased during therapy, with 38% having a mutation at BL, 24% at C3D1, and no pts gaining ESR1 D538G at C3D1. The prevalence of PIK3CA mutations also decreased on treatment (32% at BL to 24% at C3D1), with the greatest decrease in the F+P arm. A predictive model correlating early genomic dynamics and associations with outcome will be presented. Conclusions The PACE trial offers one of the largest clinical trial cohorts describing the genomic landscape of HR+/HER2- MBC post-CDK4/6 exposure. Results from this correlative study provide a comprehensive mutational landscape post-CDK4/6i, demonstrate associations between mutations and clinical outcomes, and suggest the potential value of early ctDNA dynamics in this setting. Citation Format: Rinath Jeselsohn, Jingxin Fu, Yue Ren, Reshma Mahtani, Cynthia Ma, Angela DeMichele, Massimo Cristofanilli, Jane Meisel, Kathy Miller, Yara Abdou, Elizabeth Riley, Rubina Qamar, Priyanka Sharma, Sonya Reid, Naomi Ko, Yuan Liu, Harold Burstein, Michelle DeMeo, Sara Tolaney, Erica Mayer. Exploration of ctDNA Dynamics in the PACE Trial: A Randomized Phase II Study of Fulvestrant, Palbociclib, and Avelumab for HR+/HER2- Metastatic Breast Cancer [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 PS12-02.

Abstract PO2-04-06: Investigation of the genomic evolution of HER2-positive breast cancer following progression on dual HER2-targetted therapy with Trastuzumab and Tucatinib

Abstract Background: Advanced HER2-positive breast cancer remains an incurable and highly morbid condition. One of the key challenges has been treatment of central nervous system (CNS) metastases, commonly seen in those with HER2-positive disease. Tucatinib is a recently approved treatment HER2-targetting tyrosine kinase inhibitor with proven CNS activity. There has been little analysis of potential genomic mechanisms of drug resistance. Method: We retrospectively identified patients that were treated with tucatinib, capecitabine and trastuzumab. We collated baseline clinical data and treatment history and all tumour and plasma samples available were obtained for testing. DNA was extracted from plasma samples using the QIAamp DNA Mini Kit (Qiagen) on 4ml plasma samples. Targeted next generation sequencing (NGS) of serial tumour and plasma samples was performed using a custom hybrid capture assay that covered 180 genes known to be recurrently mutated in breast cancer focusing on the known genomic landscape of metastatic disease as well as the inclusion of specific actionable targets. Survival analyses was performed using Cox regression models. Results: We identified 11 patients at our institution. Plasma samples were available for all 11 patients with a total of 107 serial samples across multiple time points, with a median of 8 (range 2-19) samples per patient. All 11 patients had baseline and End of Treatment (EOT) samples. In addition, 6/11 (54%) patients had tumour sequenced on the same NGS platform prior to commencement of therapy, with 12 individual tumour samples tested. The median PFS of our cohort was 9.6 months (4.0 - NR) with a median OS of 36.2 months (10.4-NR). 7/11 (64%) patients had CNS disease at baseline with their median PFS being 13.5 months (3.9-24.4) and median OS of 36.2months (10.4-NR). Overall, we observed an average of 18.4 (4-35) mutations per patient in tumour, 5.1 (1-15) in baseline plasma and 5.8 (2-14) in EOT plasma. The most frequent mutations seen in baseline tumour samples were ERBB2 (92%), KMT2C (67%) and NCOR1 (58%) compared with KMT2C (64%), NOTCH4 (36%) and PIK3CA (27%) in baseline plasma. Patients with a lower number of ctDNA mutations at baseline showed a trend towards improved survival (PFS 14 months vs 7.5 months). In this small cohort of patients we identified a higher than expected rates of KMT2C mutations. A KMT2C mutation was identified in 67% of samples which is higher than previously described rates of 7-15% in TCGA and METABRIC cohorts respectively. The presence of a concordant ctDNA KMT2C mutation in baseline and progression samples was a poor prognostic indicator (mPFS 7.5 months) when compared with no KMT2C mutation (mPFS 15.5 months) p =0.14 suggesting a possible mechanism of resistance to tucatinib. Conclusions: In this retrospective analysis of patients treated with Tucatinib, Trastuzumab and Capecitabine we established the genomic landscape of heavily pre-treated, advanced HER2-positive disease is complex. We observed there may be a correlation with KMT2C mutations and poorer clinical outcomes, however given the very small number of patients this data remains hypothesis generating, requiring validation with larger datasets. Citation Format: Elizabeth Blackley, Courtney van Geelen, Yi-An Ko, Stephen Wong, Miriam Yeung, Stephen Luen, Sarah-Jane Dawson, Sherene Loi. Investigation of the genomic evolution of HER2-positive breast cancer following progression on dual HER2-targetted therapy with Trastuzumab and Tucatinib [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 PO2-04-06.

Abstract PO4-15-05: Genomic characterization and molecular predictive biomarkers for chemotherapy in a real-world population with metastatic triple-negative breast cancer

Abstract Background: The genomic landscape of metastatic triple-negative breast cancer (mTNBC) in a real-world population is poorly defined and despite recent advances in precision oncology, molecular biomarkers for chemotherapy are largely lacking. The presence of specific genomic alterations and alterations in oncogenic pathways have the potential to provide prognostic as well as predictive information for chemotherapy, which remains the backbone treatment strategy for mTNBC. Methods: We conducted next-generation sequencing (NGS) with the FoundationOne CDx panel on tissue from the primary tumor and/or metastasis of 112 consecutive patients with mTNBC that were treated between 1998 and 2019 at 4 different Swedish hospitals. Every genomic alteration was, if applicable, subdivided into 1 of 10 canonical oncogenic pathways and noted for its involvement in the homologous recombination pathway (HRP). Frequently altered genes and pathways were then correlated with overall survival (OS) and evaluated regarding their association with progression-free survival (PFS) and response rate (RR) in patients treated with different chemotherapy agents. Tumor samples of patients with rapid progression or exceptional response to chemotherapy underwent exploratory comparison with regards to frequently altered genes and pathways and known clinical prognostic factors were compared between the two patient groups. Results: After excluding 15 patients due to insufficient quantity/quality of tumor tissue or absence of clinical data, 97 patients were analyzed. The median age at diagnosis of metastatic disease was 61 (range: 28-90), 80 % had visceral disease and 26 % received platinum-based chemotherapy in the first line setting. The most frequently altered genes were: TP53 (82 %), RAD21 (25 %), PIK3CA (23 %), MYC (22 %) and BRCA 1 or 2 (16 %). The most frequently altered pathways were TP53 (86 %), PI3K (60 %), RTK/RAS (47 %) and cell cycle (36 %). In total, 26 % of patients had an alteration in the HRP. None of the most frequently occurring genomic alterations were associated with OS. Variants of clinical significance in the HRP and BRCA1/BRCA2 genes were associated with a longer PFS in patients treated with platinum-based chemotherapy in the first line setting (HR 0.12-0.84 and 0.12-0.92). Exceptional responders to chemotherapy exhibited a more favorable clinical profile than rapid progressors (median age 51.5 vs 66.5, median Charlson comorbidity index 0.5 vs 3) whereas some numerical difference in the genomic profiling in terms of genomic alterations in MYC and RAS/RTK pathways (more often in exceptional responders) could be observed. Conclusions: We found no genomic alteration with prognostic significance in a cohort of mTNBC patients treated with chemotherapy in a real-world setting. Somatic variants of clinical significance in BRCA 1 and 2 and other HRP-related genes seem to define subgroups of patients with mTNBC that respond favorably to platinum-based chemotherapy. Further research into the genomic landscape of tumors from patients with rapid progression or exceptional response to specific treatment strategies can provide insights into mechanisms of resistance and identify new predictive biomarkers. Citation Format: Erik Olsson, Henrik Lindman, Evangelos Digkas, Viktoria Thurfjell, Haidar Mir Ali, Ute Krüger, Anna-Karin Wennstig, Marie Sundqvist, Antonis Valachis. Genomic characterization and molecular predictive biomarkers for chemotherapy in a real-world population with metastatic triple-negative breast cancer [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 PO4-15-05.

Abstract PO1-15-11: Molecular and clinical disparities in breast cancer among East Asian populations

Abstract Background Breast Cancer (BC) is a complex disease with profound genomic aberrations. However, the underlying molecular disparity influenced by age and ethnicity remains elusive. Method We aimed to investigate the molecular properties of 843 primary and metastatic BC patients enrolled in the K-MASTER program. We explored the unique molecular profiles of breast cancer patients by categorizing them into two distinct age subgroups. The first subgroup comprised young breast cancer (YBC) patients, defined as women under the age of 40, while the second subgroup consisted of older breast cancer (OBC) patients. Our study aimed to characterize the molecular differences between these age-defined groups, providing insights into the underlying mechanisms of breast cancer in different age cohorts. Additionally, we leveraged large-scale genomic data from the TCGA and MSK-IMPACT studies to examine the ethnic-driven molecular and clinical disparities. Results The K-MASTER patients were mainly comprised of triple-negative breast cancer (TNBC) and HER2-positive tumors, while the TCGA and MSK-IMPACT cohorts exhibited a predominance of hormone receptor-positive (HR+) subtype. We observed a high prevalence of PI3KCA mutations in K-MASTER HER2+ tumors, particularly in older patients. Moreover, we identified increased mutation rates in DNA damage response molecules, including ARID1A, MSH6, and MLH1. Importantly, GATA3 mutations were less frequently observed in East Asian patients, which correlated with poor clinical outcomes. Our analysis also revealed specific associations between genomic aberrations, organotropisms, and metastatic breast cancer. In addition to characterizing the molecular disparities, we developed a gradient-boosting multivariable model to identify a new molecular signature that could predict the therapeutic response to platinum-based chemotherapy. Conclusion Our findings collectively provide unprecedented insights into the significance of age and ethnicity on the molecular and clinical characteristics of BC patients. Figure 1. Genomic landscape of KM BRCA. (A) Overall characteristic of KM BRCA sample and clinical data. Clinical features of KM BRCA sample. (B) Genomic landscape of somatic mutations and copy number alterations of BRCA by age group. Figure 2. Genomic landscape of KM BRCA: Significantly mutated genes by age group in KM BRCA. Figure 3. Survival Curve Utilizing a curated Gene Set Developed by a Gradient-Boosting Multivariable Model (A) OS of platinum-based therapy treated KM with curated Gene set. (B) OS of platinum-based therapy treated MSK with curated Gene set. Citation Format: Kim Ju Won, LEE JIWON, Kyong Hwa Park, Seung Pil Jung, Jason Sa, SHIN SANG WON, An Jung Seok. Molecular and clinical disparities in breast cancer among East Asian populations [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 PO1-15-11.

Abstract PO1-15-08: Genomic characterization of triple-negative breast cancer metastases reveals PKD1 as a novel biomarker for immunotherapy

Abstract Background: While primary triple-negative breast cancer (TNBC) garners significant research attention, the genomic alterations that occur in metastasis remain insufficiently understood, especially within Asian populations. Furthermore, the genomic information obtained from the primary tumor inadequately guides metastatic cancer treatment, highlighting the critical need for in-depth investigations into metastatic TNBC. Methods: We constructed the largest cohort of TNBC metastases (n = 296) among advanced TNBC patients treated at Fudan University Shanghai Cancer Center (FUSCC) between October 2018 and December 2020. Comprehensive DNA sequencing was conducted on the collected metastatic samples to analyze genomic alterations associated with treatment response. The underlying mechanisms of specific biomarkers were also explored. Results: We presented the genomic landscape of 296 TNBC metastases, encompassing mutant genes, mutation sites and copy number variations. Through multidimensional analysis, significant disparities in TNBC were observed between Western and Asian populations, primary and metastatic tumors, as well as different metastatic sites. Notably, our findings underscore the importance of sequencing TNBC metastases to guide precision therapy, which was associated with longer progression-free survival compared to physician-chosen treatments, shedding light on the pivotal clinical value of genomic studies in metastatic settings. Furthermore, efficacy analysis suggested that PKD1 mutations enriched in metastases mediated resistance to immunotherapy. These findings were further validated through three clinical trials (NCT03805399, NCT04129996, and NCT04395989). Mechanistic studies unveiled the involvement of PKD1 in TNBC immune evasion by upregulating CCL2, thereby facilitating the recruitment of M2-type tumor-associated macrophages. Conclusion: Our study emphasizes the critical significance and necessity of genomic profiling of metastases in guiding precision therapy for TNBC. Moreover, our findings reveal PKD1 as a novel and promising biomarker for immunotherapy. Citation Format: Xiu-Zhi Zhu, Yi-Fan Zhou, Yun-Yi Wang, Xiao-Hong Ding, Xi Jin, Zhi-Ming Shao, Yi-Zhou Jiang, Zhong-Hua Wang. Genomic characterization of triple-negative breast cancer metastases reveals PKD1 as a novel biomarker for immunotherapy [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 PO1-15-08.

Abstract PO4-24-08: Genomic Landscape of Malignant Phyllodes Tumors Identifies Subsets for Targeted Therapy

Abstract Introduction: Phyllodes tumors (PT) are rare fibroepithelial tumors of the breast, comprising ~1% of all breast cancers, a subset of which have considerable malignant potential. Malignant phyllodes tumors (MPTs) have aggressive biological behavior and high local and distant recurrence rates. Distant metastases of MPTs have a poor prognosis given the limited treatment options available. Surgery remains the primary treatment modality for these patients; however, initial investigations suggest a potential for targeted therapies in managing this disease. The identification of targetable opportunities and a comprehensive understanding of the mutational profiles of MPTs could greatly enhance treatment options. Therefore, we aimed to assess the molecular landscape of these tumors. Methods: Phyllodes tumor samples underwent molecular profiling at Caris Life Sciences through DNA (592-gene panel or whole exome) and RNA sequencing (whole transcriptome). The MPTs were classified into primary and metastatic based on the specimen biopsy site. PD-L1+ expression was tested by IHC (SP142; ≥2+, ≥5%). Tumor mutational burden (TMB)-High was defined as ≥10 mutation/Mb. Immune cell fractions in the tumor microenvironment (TME) were estimated using quanTIseq (Finotello, 2019). Mann-Whitney U, Chi-square, and Fisher-Exact tests were applied where appropriate. Results: We identified 57 analyzed MPTs, all from female patients, with a median age of 53 years (range: 19-88). Approximately half of samples (53%, n=30) were labeled as from primary breast sites, and the remaining sites were metastatic (47%, n= 27). Of metastatic sites, lung was the most common location (74.1%, n=20/27), followed by bone, sacrum, pancreas and small bowel. Alterations in MPT include TERT promoter (56.8%, n=21/37), MED12 (51.6%, n=16/31), TP53 (40.0%, n=22/55), and NF1 (31.8%, n=14/44) mutations, with less frequent mutations of EGFR (10.5%, n=6/57), PIK3CA (7.0%, n=4/57), and BRAF (3.5%, n=2/57). As compared to primary MPT, metastatic MPT had roughly 2-fold higher prevalence of NF1 (45.5%: n=10/22 vs 18.2% n=4/22, p=0.05), KMT2D (25.9%: n=7/27 vs 10.7%: n=3/28, p=0.18), and RB1 (25.0%: n=6/24 vs 9.5%: n=2/21, p=0.25) though not significant. Multiple gene mutations were observed exclusively in lung metastases, as compared to non-lung sites including: TERT promoter (64.3%: n=9/14 vs 0.0%, p=0.08), MED12 (60.0%: n=6/10 vs 0.0%, p =0.044), and RB1 (35.3%: n=6/17 vs 0.0%, p=0.01). Conversely, NF1 (43.8%: n=7/16 vs 50.0%: n=3/6, p=1.0) and KMT2D (20.0%: n=4/20 vs 42.9%: n=3/7, p=0.32) mutation prevalence was higher in other metastatic sites. PD-L1+ expression was observed in 13.3% (n=7) of MPT overall, with similar prevalence among local and metastatic sites. No dMMR/MSI-H was observed, while low LOH (those with genomic LOH in < 16% of segment analyzed) was seen across MPT. Compared to a large cohort of breast adenocarcinoma samples (n=9,926) representing both HER2+ and HER2- tumors, MPT had low ERBB2 expression comparable to HER2- samples. B cells, M2 macrophages and neutrophils had the highest median cell fractions in the TME of MPT. Additionally, one MPT sample harbored a pathogenic NTRK1 fusion (TPM4:NTRK1), and treatment with larotrectinib for over 16 months suggests a clinical response to therapy. Conclusions: Our study demonstrates the importance of employing next generation sequencing (NGS) in MPTs to detect actionable genomic alterations. To effectively analyze fusions, an NGS panel that include RNA sequencing is recommended, considering the occurrence of NTRK1 fusion reported herein. Although HER2 transcriptional expression was low, further investigations examining HER2 IHC in PTs are still necessary. These finding therefore highlight the importance of NGS in phyllodes tumors research, as it may uncover potential targeted treatment options for patients. Citation Format: Rani Bansal, Tolulope Adeyelu, Andrew Elliott, Antoinette Tan, Jane Meisel, Matthew Oberley, Stephanie Graff, Jorge Reis-Filho, George Sledge Jr, Sarah Sammons, Laura Rosenberger. Genomic Landscape of Malignant Phyllodes Tumors Identifies Subsets for Targeted Therapy [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 PO4-24-08.

Abstract PO1-08-12: Genomic Characterization of the Carolina Breast Cancer Study

Abstract Background: Breast cancer is a heterogeneous disease defined by distinct subtypes, mutational profiles, and genomic characteristics. Previous analyses in The Cancer Genome Atlas (TCGA) have been instrumental in understanding the genomic landscape of breast cancer, including identification of key driver mutations, mutational signatures, and distinguishing features of breast cancer subtypes. However, TCGA may not be representative of other population-based breast cancer cohorts, particularly those of earlier stage and including a more diverse set of patients. Our aim was to characterize the genomic landscape of breast cancer in the Carolina Breast Cancer Study (CBCS), a cohort which oversamples Black and young women. Methods: We used targeted sequencing to profile somatic alterations in 1,175 genes from 275 formalin fixed paraffin embedded primary breast tumors in CBCS, 52% of which came from Black women, 41% from women under age 50, and 29% of which were Basal-like based on bulk RNA profiling. We also evaluated TP53 mutation as a source of breast cancer heterogeneity, comparing the type and location of TP53 mutations in each cohort. We assessed chromosome arm-level gains and losses using GISTIC and defined genomic instability as the total number of aneuploid chromosome arms. Intratumoral heterogeneity was assessed with PyClone-VI and defined as the total number and Shannon’s Diversity Index of tumor-specific subclones. For genes mutated in more than 5% of samples and chromosomal aberrations significantly enriched in GISTIC, we estimated the prevalence of alterations and compared to previous distributions in 981 TCGA tumors. Results: Seven genes [TP53 (N=101, 37%), PIK3CA (N=76, 28%), GATA3 (N=32, 12%), CDH1 (N=31, 11%), MAP3K1 (N=22, 8%), KMT2C (N=20, 7%), and CBFB (N =19, 7%)] were mutated in more than 5% of CBCS tumors; all but CBFB which were also highly prevalent in TCGA. While most TP53 hotspot mutations observed in CBCS were previously reported in TCGA, CBCS had twice as many Y220C mutations, one-third fewer R175H mutations, and a non-significantly higher proportion of nonsense mutations than TCGA (20% vs 12%, p = 0.05). Hotspot mutation prevalence in other Luminal-associated driver genes (e.g. PIK3CA, GATA3, CDH1) did not differ by dataset. CBCS tumors showed a higher degree of genomic instability (8 arms vs. 5 arms, p < 0.001) and subclonal diversity than TCGA (48% vs 12% comprised of two or more subclones, p < 0.05). Both datasets showed significant amplifications of 1q and 8q, and deletions of 13q and 17p (where BRCA2 and TP53, respectively, are located), with CBCS having non-significantly higher prevalence of each of these changes. Conclusions: While comparisons should be interpreted in light of technical and population differences between TCGA and CBCS, the overall results show that the suite of commonly altered genes and chromosome arms were highly consistent between TCGA and the diverse CBCS cohort. CBCS tumors tended to display higher genomic instability, intratumoral heterogeneity, and number and diversity of TP53 mutations. This may reflect distinctions in tumor evolutionary state between CBCS and TCGA tumors, and/or could reflect differences in the study populations. Results underscore the importance of considering population characteristics – particularly stage and race – in large-scale genomic contexts, and highlight the importance of diverse cohorts in genomic research. Citation Format: Sarah Van Alsten, Ebonee Butler, Benjamin Calhoun, Michael Love, Charles Perou, Katherine A Hoadley, Melissa Troester. Genomic Characterization of the Carolina Breast Cancer Study [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 PO1-08-12.

Abstract PO5-15-05: Clinical and genomic landscape of ERBB2 and ERBB3 mutated breast cancer

Abstract Background: Our understanding of tumorigenesis in ERBB2 (HER2) non-amplified breast cancer has recently evolved through genome sequencing efforts identifying somatic mutations in ERBB2 and its ERBB3 coreceptor (ERBB2/3). These alterations result in upregulation of HER2 activity and have been implicated in resistance to certain targeted therapies. To develop efficacious therapeutic strategies in this context, further delineation of the clinical and genomic landscape of ERBB2/3-mutated (ERBB2/3-mut) breast cancers is paramount. Methods: Patients (pts) with breast cancer who underwent tumor and normal tissue sequencing using the MSK-IMPACT assay between April 2015 to August 2019 were surveyed in this analysis. We performed gene enrichment analyses to assess the frequency, type and pattern of ERBB2/3-mut, and compared demographics and clinicopathologic characteristics of this cohort with those of pts with ERBB2/3-wild-type (wt) tumors. The landscape of co-occurring and mutually exclusive genomic alterations in oncogenic (as per OncoKB) ERBB2/3-mut vs. ERBB2/3-wt was studied in the context of receptor status. Finally, we investigated the association between ERBB2/3-mut status and clinical outcomes in HR+/HER2- pts. Progression free survival (PFS) on first line CDK4/6 inhibitors and endocrine therapy (CDK4/6i-ET) was assessed utilizing Cox proportional hazard models adjusted for endocrine therapy partner. Results: Of 4,458 evaluable pts with breast cancer, 260 were ERBB2/3-mut (ERBB2 n=182, 4.1%; ERBB3 n=90, 2.0%, with both ERBB2/3 n=12, 0.27%). A total of 368 ERBB2/3 alterations were identified in 321 samples, of which 324 were SNVs (88.0%) and 44 indels (12.0%). Of the 256 ERBB2-mut, 185 (72.3%) were pathogenic variants (PVs), with the most frequent alterations being L755S (28.6%), V777L (13.5%), G778_P780dup (10.3%) and S310F (12.4%). Of the 112 ERBB3-mut, 53 (47.3%) were PVs; E928G (47.3%) and G284R (11.3%) were the most common. ERBB2/3-mut tumors were enriched in lobular (n=72, 13.3%) over ductal histology (n=164, 4.6%; Odds ratio [OR]: 3.15, 95% confidence interval [CI]: 2.31–4.25; p=8.2 e-13). When duplicates from each category were excluded, ERBB2/3 variants were more prevalent in metastatic tumors (n=165, 7.3%) compared to primaries (n=119, 4.7%; OR: 1.61, 95% CI: 1.25–2.06, p=0.0001). A similar trend was observed in the HR+/HER2- subgroup (metastatic: 6.7% vs. primary: 4.3%; OR: 1.61, 95% CI: 1.12–2.2, p=0.0024). In the HR+/HER2- subgroup (n=3,039), ERBB2-mut (4.0%, PVs: 3.1%) were significantly co-altered (q< 0.1) with CDH1, TBX3, ERBB3, RUNX1, CBFB, MAP3K1, FOXA1 and NF1 and mutually exclusive with GATA3. In the HER2+ subgroup (n=640), ERBB2-mut (7.2%, PVs: 4.7%) were significantly co-altered with CDH1, TBX3, and CDKN2A/B. In the TNBC cohort (n=779), ERBB2-mut (2.3%, PVs: 1.7%) were mutually exclusive with TP53, and co-altered with PIK3CA, ARID1A, MAP3K1, CBFB, and CDH1. Overall, the ERBB3-mut were co-altered with ERBB2, CDH1, and CBFB. In HR+/HER2- pts with tumor sequencing performed prior to receiving first-line CDK4/6i-ET (n=468), PFS for ERBB2/3-mut (n=23) was not significantly different than ERBB2/3-wt (hazard ratio: 1.19, 95% CI: 0.75–1.90, p=0.46). Conclusion: Here we describe the clinical and genomic characteristics of a large cohort of ERBB2/3-mut breast cancers. We identify a notable enrichment of ERBB2/3-mut in lobular histology and metastatic tumors and tendency for co-alteration with CDH1 and multiple transcription factors reflecting the unique biology of ERBB2/3-mut breast cancers. Further analyses on an expanded cohort (n >6000 pts), including outcomes on HER2-directed antibody-drug conjugates (T-DXd) and targeted therapies such as PI3K inhibitors, will be presented at the 2023 SABCS Annual Meeting. Citation Format: Nadeem Bilani, Jacqueline Tao, Anton Safonov, Dana Casey, Joshua Drago, Mehnaj Ahmed, Barbara Acevedo, Komal Jhaveri, Jorge Reis-Filho, Mark Robson, Eneda Toska, Ariella Hanker, Carlos Arteaga, Sarat Chandarlapaty, Hanna Y Wen, Pedram Razavi. Clinical and genomic landscape of ERBB2 and ERBB3 mutated breast cancer [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 PO5-15-05.