Rapid Fire 4

O. Metzger¹, S. Mandrekar², T. Dockter³, L. Gianni⁴, E. Ciruelos⁵, A. De Michele⁶, J. Gligorov⁷, E. Lim⁸, S. Loibl⁹, X. Gonzàlez Farré¹⁰, F. Lynce¹¹, J. Lanzillotti¹², L. Carey¹³, S. Goel¹⁴, A. Partridge¹, E. Winer¹⁵; ¹Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, ²Biostatistics Shared Resource & Alliance Statistics Data Management Center, Mayo Clinic, Rochester, MN, ³Department of Quantitative Health Sciences / Biostatistics Core, Mayo Clinic, Rochester, MN, ⁴Medical Oncology, Michelangelo Foundation, Milan, ITALY, ⁵Medical Oncology, Hospital Universitario 12 de Octubre, Madrid, SPAIN, ⁶Medical Oncology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, ⁷Medical Oncology, UCBG; Tenon Hospital IUC AP-HP Sorbonne Université, Paris, FRANCE, ⁸Medical Oncology, St. Vincent’s Hospital; Garvan Institute of Medical Research;, Sydney, AUSTRALIA, ⁹Medical Oncology, German Breast Group, Heidelberg, GERMANY, ¹⁰Medical Oncology, Hospital Universitari General de Catalunya, Barcelona, SPAIN, ¹¹Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, ¹²Operations, Alliance Foundation Trials, Boston, MA, ¹³Medical Oncology, University of North Carolina (UNC) Lineberger Comprehensive Cancer Center, Chapel Hill, NC, ¹⁴Medical Oncology, Peter MacCallum Cancer Centre, Melbourne, AUSTRALIA, ¹⁵Medical Oncology, Yale Cancer Center; Yale School of Medicine, New Haven, CT.

Background Central nervous system (CNS) metastases are a major clinical concern in patients with HER2-positive metastatic breast cancer (MBC). The PATINA phase III study showed improved PFS with palbociclib added to maintenance anti-HER2/endocrine therapy (44.3 vs. 29.1 months; hazard ratio 0.75; 95% CI 0.59-0.96; 1-sided P=0.0109). Preclinical and early clinical data suggest palbociclib may have CNS activity. We conducted a pre-specified secondary analysis to evaluate the incidence and timing of CNS progression. Methods CNS progression was defined as the emergence of new intracranial lesions per investigator assessment. Baseline CNS imaging was not mandated but recommended if symptomatic. Competing risks analyses evaluated time from registration to first CNS progression or death (CNS PFS), treating non-CNS progression as a competing event. All randomized patients were included in the primary analysis, regardless of known baseline CNS involvement. Patients with baseline CNS metastases were not censored if progression involved the CNS. A secondary analysis excluded patients with CNS metastases present at baseline. Cumulative incidence functions were estimated and compared using 2-sided Gray’s test. Results Data cutoff for this analysis was on October 15, 2024. 518 participants were enrolled between June 2017 and July 2021, 261 to the palbociclib arm and 257 to the control arm. CNS metastases at baseline were present in 11 patients in the palbociclib arm and 9 patients in the control arm. Among all randomized patients, CNS progression occurred in 35 of 261 (13.4%; 95% CI: 9.5-18.2) in the palbociclib arm and 50 of 257 (19.5%; 95% CI: 14.8-24.8) in the control arm. When excluding patients with CNS metastases at baseline, CNS progression was observed in 32 of 250 (12.8%; 95% CI: 8.9-17.6) in the palbociclib arm and 47 of 248 (19.0%; 95% CI: 14.3-24.4) in the control arm. The median follow-up for patients who were alive and progression-free was 53.5 months. At 36 months, the cumulative incidence of CNS PFS was 13.4% vs. 19.9% for palbociclib versus control (Gray’s test p=0.0386). Importantly, when the analysis was restricted to pts without evidence of CNS disease at baseline (Table 1), —the CNS benefit with palbociclib was maintained (13.0% vs. 19.2% at 36 months, Gray’s test p = 0.0378). Conclusion The addition of palbociclib to anti-HER2 and endocrine therapy was associated with a lower incidence of CNS metastases. Among patients without known baseline CNS involvement, the absolute reduction in CNS progression at 36 months was 6.2% in favor of palbociclib. This benefit was observed at 12 months and sustained over 3 years (Table 1). These findings suggest that palbociclib may help delay or prevent CNS involvement in HR+/HER2+ MBC, warranting confirmation in future studies. Table 1. Cumulative Incidence of CNS Progression or Death Among Patients Without CNS Metastases at Baseline

W. Janni¹, T. Fehm², V. Müller³, J. Blohmer⁴, A. De Gregorio⁵, T. Decker⁶, A. Hartkopf⁷, N. Ditsch⁸, M. Schmidt⁹, P. Wimberger¹⁰, T. Engler⁷, M. Banys-Paluchowski¹¹, P. A. Fasching¹², B. Rack¹, A. Schneeweiss¹³, K. Pantel¹⁴, T. W. Friedl¹, F. Schochter¹, J. Huober¹⁵; ¹Department of Gynecology and Obstetrics, University Hospital Ulm, Ulm, GERMANY, ²Department of Gynecology and Obstetrics, University Hospital Düsseldorf, Düsseldorf, GERMANY, ³Department of Gynecology and Obstetrics, University Hospital Hamburg-Eppendorf, Hamburg, GERMANY, ⁴Department of Gynecology and Breast Center, Charité University Hospital Campus, Berlin, GERMANY, ⁵Klinik für Gynäkologie und Geburtshilfe, SLK-Kliniken Heilbronn, Heilbronn, GERMANY, ⁶Oncological Practice, Oncological Practice, Ravensburg, GERMANY, ⁷Department of Women’s Health, Tübingen University Hospital, Tübingen, GERMANY, ⁸Gynecology, Obstetrics and Senology, Faculty of Medicine, University Hospital Augsburg, Augsburg, GERMANY, ⁹Department of Obstetrics and Gynecology, University Hospital Mainz, Mainz, GERMANY, ¹⁰Department of Gynecology and Obstetrics, University Hospital Dresden, Dresden, GERMANY, ¹¹Department of Gynecology and Obstetrics, University Hospital Schleswig-Holstein Campus Lübeck, Lübeck, GERMANY, ¹²Department of Gynecology and Obstetrics, University Hospital Erlangen, Erlangen, GERMANY, ¹³National Center for Tumor Diseases, University Hospital and German Cancer Research Center, Heidelberg, GERMANY, ¹⁴Institute of Tumor Biology, University Hospital Hamburg-Eppendorf, Hamburg, GERMANY, ¹⁵Breast Center St Gallen, HOCH Cantonal Hospital, St. Gallen, SWITZERLAND.

Background: Current data support that metastatic hormone receptor-positive (HR+) and HER2-positive (HER2+) breast cancer offers distinct treatment options targeting both the hormone receptor and the HER2-pathway. This might lead to increased treatment efficacy with the potential of omitting chemotherapy in these patients. The aim of the randomized multicenter phase III DETECT V trial was to analyze the efficacy of chemotherapy free and CDK4/6 inhibitor containing treatment strategies compared to current standard treatment. Methods: Between 09/2015 and 11/2022, 271 patients with HER2+ and HR+ MBC in the 1st-3rd line setting were randomized 1:1 to receive trastuzumab and pertuzumab combined with either endocrine therapy or chemotherapy followed by maintenance therapy with dual HER2 targeted and endocrine therapy. Chemotherapy and the endocrine agents could be chosen from a variety of available regimens according to physicians’ choice. In January 2019, the protocol was amended with the addition of the CDK4/6 inhibitor ribociclib to both treatment arms. The primary objective of DETECT V was to compare tolerability between the treatment arms, and secondary objectives comprised the comparison of overall survival (OS), progression-free survival (PFS) and safety, as well as the evaluation of the effect of adding ribociclib to both treatment arms. Here we report results of the final efficacy analyses as based on the modified ITT set of 262 patients (9 patients did not receive any study treatment and were excluded). Of these, 120 patients were recruited before the addition of ribociclib (59 and 61 patients in the chemotherapy-free and chemotherapy-containing treatment arm, respectively), and 142 patients were recruited after the addition of ribociclib (73 and 69 patients in the chemotherapy-free and chemotherapy-containing treatment arm, respectively). Results: Median patient age was 60 years, 202 (77.1%) of patients were in the first line setting and 137 (52.3%) patients had a metastasis-free interval (from primary diagnosis) exceeding 12 months. The overall comparison between all patients receiving chemotherapy-free and chemotherapy-containing treatment showed no significant difference with regard to OS and PFS (median OS not reached vs 46.1 months, HR 0.98, 95% CI 0.60 – 1.59, p = 0.93; median PFS 19.5 vs 23.0 months, HR 1.15, 95% CI 0.82 – 1.62, p = 0.42). However, both OS and PFS were significantly improved by the (non-randomized) addition of ribociclib (OS: median OS not reached vs 46.1 months, HR 0.43, 95% CI 0.26 – 0.72, p = 0.001; PFS: median PFS 29.7 vs 15.6 months, HR 0.48, 95% CI 0.34 – 0.67, p < 0.001). A separate analysis for the two treatment arms showed that the addition of ribociclib to chemotherapy-containing treatment significantly improved both OS (median OS not reached vs 38.7 months, HR 0.30, 95% CI 0.14 – 0.64, p = 0.002) and PFS (median PFS 33.1 vs 15.4 months, HR 0.35, 95% CI 0.21 – 0.59, p < 0.001), while the addition of ribociclib to chemotherapy-free treatment significantly improved PFS (median PFS 27.2 vs 15.6 months, HR 0.61, 95% CI 0.38 – 0.98, p = 0.040) but not OS (median OS not reached in both groups, HR 0.60, 95% CI 0.30 – 1.23, p = 0.163). An exploratory cross-cohort comparison revealed significantly improved outcome for patients receiving chemotherapy-free treatment plus ribociclib vs chemotherapy-containing treatment without ribociclib (OS: median OS not reached vs 38.7 months, HR 0.48, 95% CI 0.24 – 0.94, p = 0.033; PFS: median PFS 27.2 vs 15.4 months, HR 0.53, 95% CI 0.33 – 0.85, p = 0.008). Conclusion: Our results suggest that chemotherapy-free treatment for patients with HER2+ and HR+ MBC may be a valuable and effective treatment alternative, and may improve survival with the addition of ribociclib.

M. Balic¹, G. Tang², G. Young³, P. Rastogi⁴, J. Acosta⁵, A. Schneeweiss⁶, C. Hilton⁷, T. Freeman⁸, C. Denkert⁹, M. Reinisch¹⁰, M. R. Palomares¹¹, A. A. Bradley¹², S. Murali¹³, J. Boileau¹⁴, D. Boudreau¹⁵, P. J. Polewski¹⁶, S. Hassan¹⁷, J. Mouta¹⁸, W. C. Darbonne¹⁹, F. L. Baehner¹¹, E. P. Mamounas²⁰, N. Wolmark²¹, S. Loibl²², C. E. Geyer, Jr²³; ¹Oncology, NSABP Foundation, Inc.; UPMC Hillman Cancer Center; University of Pittsburgh School of Medicine, Pittsburgh, PA, ²Department of Biostatistics, NRG Oncology Statistics and Data Management Center, and University of Pittsburgh School of Public Health, Department of Biostatistics and Health Data Science, Pittsburgh, PA, ³Oncology, NSABP Foundation, Inc.; UPMC Hillman Cancer Center; University of Pittsburgh School of Medicine, Redwood City, CA, ⁴Hematology and Medical oncology, NSABP Foundation, Inc.; UPMC Hillman Cancer Center; University of Pittsburgh School of Medicine, Pittsburgh, PA, ⁵Physician, Allegheny Health Network; NSABP Foundation, Inc, Pittsburgh, PA, ⁶Division Gynecologic Oncology, National Center for Tumor Diseases (NCT); Heidelberg University Hospital and German Cancer Research Center, Heidelberg, GERMANY, ⁷Oncology, NSABP Foundation, Inc.; Allegheny Health Network, Pittsburgh, PA, ⁸Department of Pathology, NSABP Foundation, Inc.; University of Pittsburgh School of Medicine, Pittsburgh, PA, ⁹Pathology, Institute of Pathology, Philipps-Universität Marburg and University Hospital Marburg; German Breast Cancer Group (GBG) Forschungs GmbH, Marburg, GERMANY, ¹⁰Breast Unit or Gynecology Medical University Mannheim, Interdisciplinary Breast Unit, University Medical Center Mannheim, University of Heidelberg, Mannheim, Germany, Mannheim, GERMANY, ¹¹Medical, Exact Sciences Corporation, Redwood City, CA, ¹²Medical Oncology, Exact Sciences Corporation, Redwood City, CA, ¹³Oncol, Southern California Permanente Medical Group, San Marcos, CA, ¹⁴Surgical Oncology, Jewish General Hospital Segal Cancer Centre McGill University, Montréal, QC, CANADA, ¹⁵Oncol, Hôpital du Saint-Sacrement, Québec, QC, CANADA, ¹⁶Hem/Onc, Aurora Cancer Care, Advocate Aurora Health, Milwaukee, WI, ¹⁷Surg/Onc., Centre Hospitalier de l’Université de Montréal (CHUM); Centre de Recherche de CHUM; Université de Montréal, Montréal, QC, CANADA, ¹⁸Product Development Medical Affairs, F. Hoffmann – La Roche Ltd, Basel, SWITZERLAND, ¹⁹Department of Oncology Biomarker Development, US Medical Affairs Oncology, Genentech, Inc., South San Francisco, CA, ²⁰Surgical Oncology, AdventHealth Cancer Institute, Orlando, FL, ²¹Div Surgical Oncology, Dpt of Surgery, NSABP Foundation, Inc.; UPMC Hillman Cancer Center; University of Pittsburgh School of Medicine, Pittsburgh, PA, ²²Chair, German Breast Group; Exec Officer, GBG Forschungs GmbH, German Breast Cancer Group (GBG) Forschungs GmbH, and Goethe University Frankfurt, Neu-Isenburg, GERMANY, ²³Malignant Hematology and Medical Oncology, NSABP Foundation, Inc.; UPMC Hillman Cancer Center; University of Pittsburgh School of Medicine, Pittsburgh, PA.

Background: Detection of occult micro-metastatic cancer—molecular residual disease (MRD)—in patients following treatment for early triple-negative breast cancer (TNBC) using circulating tumor DNA (ctDNA) is associated with a high risk of recurrence. Dynamics of ctDNA clearance in response to neoadjuvant therapy (NAT) has also shown potential for predicting pathologic compete response (pCR) and potentially improving the prognostic ability of pCR status. NSABP B-59/GBG-96-GeparDouze was a phase III placebo-controlled trial which evaluated the efficacy and safety of adding atezolizumab or placebo to NAT followed by atezolizumab or placebo as adjuvant therapy to complete 1 year of therapy in 1,550 patients with Stage II/III TNBC. A sub-study collected serial blood samples for ctDNA analysis during the first 2 years following randomization, which were analyzed with a whole-exome sequencing of the primary tumor with variants selection informed by parallel sequence analysis of the patient’s normal germline DNA obtained from peripheral blood mononuclear cells. Patient-specific bespoke assays with 50-200 variants were used to assess each available sample for presence or absence of ctDNA. Methods: This prospective sub-study included patients enrolled within the NSABP B-59/GBG-96-GeparDouze trial (Clin Cancer Res [2025] 31 [12_Suppl]: GS3-05) following an amendment to incorporate serial ctDNA collections at baseline prior to neoadjuvant therapy (NAT), immediately before surgery, 3-6 weeks post-surgery, 12-months after randomization, 24-months after randomization, and at recurrence. The primary objective of this sub-study was to demonstrate the association between a positive ctDNA result at the post-surgery timepoint and distant recurrence-free interval (DRFI), defined as time from surgery to first occurrence of distant metastasis. The log-rank test was used to compare DRFI between patients with and without ctDNA detection post surgery, with the lag between surgery and blood sample test considered as delayed entry. The Cox proportional hazards model was used to assess the strength of relationship between ctDNA detection and risk of distant recurrence. Secondary endpoints included recurrence-free interval (RFI) and pCR. Results: The mean age of the patients in the sub study was 50, 78.9% had high tumor grade, 61.2% were node positive at presentation, and 51% received atezolizumab. 160 patients with baseline blood samples had sufficient DNA yield to perform WES for development of the bespoke assay for ctDNA. We detected ctDNA at presentation in 153 (96%) of these patients. At the end of NAT prior to surgery, 155 blood samples were available and ctDNA was detected in 14 (9.0%) indicating substantial clearance on the NAT. Following surgery, 147 blood samples were available to assess the primary endpoint of DRFI. Median follow-up after surgery was 37 months. Distant recurrences had developed in 14 patients (9.5%). Following NAT and surgery, 138 (93.8%) were ctDNA negative, and of those 131 (95%) were free of distant recurrence. Among the 9 patients who were ctDNA positive (MRD+) after surgery, 7 developed distant recurrences (log-rank p<1E-16, HR=30.3, 95% CI=10.4, 88.7). Similar finding were observed in RFI. Conclusions: Detection of ctDNA (MRD+) using a bespoke MRD assay was highly prognostic for distant recurrence in TNBC patients following neoadjuvant therapy and surgery. The assay also demonstrated a high prevalence of ctDNA at presentation with substantial clearance on the chemotherapy regimen +/- atezolizumab employed in the trial.

D. R. Sudhan¹, C. Salinas-Souza², V. Cutano², I. De Toma³, S. Dunn², J. Bradley², P. Neven⁴, M. Beeram⁵, E. Hamilton⁶, B. Pistilli⁷, A. Raskov Kodahl⁸, P. Lau⁹, V. F. Borges¹⁰, M. Campone¹¹, T. Foukakis¹², E. Lim¹³, I. Ługowska¹⁴, P. Wysocki¹⁵, I. Oshiomogho¹⁶, V. Bhagawati Prasad², C. Gresty², U. McDermott², S. Barry², E. de Bruin², J. Armenia², H. S. Rugo¹⁷; ¹-, AstraZeneca, Gaithersburg, MD, ²-, AstraZeneca, Cambridge, UNITED KINGDOM, ³-, AstraZeneca, Barcelona, SPAIN, ⁴-, Universitaire Ziekenhuizen Leuven, Leuven, BELGIUM, ⁵-, South Texas Accelerated Research Therapeutics, San Antonio, TX, ⁶-, Sarah Cannon Research Institute, Nashville, TN, ⁷-, Gustave Roussy Cancer Campus, Villejuif, FRANCE, ⁸-, Odense University Hospital, Odense, DENMARK, ⁹Linear Clinical Research, Perth, WA, Harry Perkins Institute for Medical Research, Perth, WA, University of Western Australia, Perth, WA, AUSTRALIA, ¹⁰-, University of Colorado Cancer Center, Aurora, CO, ¹¹-, Institut de Cancérologie de l’Ouest – Centre René Gauducheau, Saint-Herblain, FRANCE, ¹²-, Karolinska Institutet and Breast Cancer Centre, Cancer Theme, Karolinska University Hospital, Karolinska Comprehensive Cancer Center, Solna, SWEDEN, ¹³-, St Vincent’s Hospital Sydney and University of New South Wales, Sydney, NSW, AUSTRALIA, ¹⁴-, Maria Skłodowska-Curie National Institute of Oncology, Warsaw, POLAND, ¹⁵Department of Clinical Oncology, Jagiellonian University Medical College Hospital, Cracow, POLAND, ¹⁶-, AstraZeneca, Mississauga, ON, CANADA, ¹⁷-, City of Hope Cancer Centre, Duarte, CA.

Background PI3K/AKT pathway activation is implicated in resistance to endocrine therapy (ET) + CDK4/6i. Simultaneous inhibition of PI3K/AKT and CDK4/6 pathways may delay ET and/or CDK4/6i resistance, and/or re-sensitize tumors to ET plus CDK4/6i. Results from Phase 1b of the CAPItello-292 Phase 1b/3 trial (NCT04862663) have shown that triplet therapy with pan-AKT inhibitor capivasertib + CDK4/6i (palbociclib or ribociclib) + fulvestrant is tolerable in patients with HR+/HER2− ABC, with preliminary evidence of clinical activity. Most Phase 1b patients were heavily pre-treated and had received prior CDK4/6i treatment.This exploratory analysis from Phase 1b of CAPItello-292 examined matched pre- and post-treatment circulating tumor DNA (ctDNA) to identify potential genomic mechanisms associated with resistance to the triplet treatment. Methods Baseline and end-of-treatment (EOT) ctDNA were tested using the Guardant360 Liquid assay on the Guardant Infinity platform. Somatic alterations of known/likely significance with variant allele fraction ≥0.3% (limit of quantification) were evaluated. Copy number changes, single nucleotide variants, fusions, and indels in EOT ctDNA were considered to be acquired only if absent in baseline ctDNA and archival tumor tissue. Separately, genome-wide CRISPR-Cas9 knockout screens were performed in estrogen receptor-positive (ER+) breast cancer cell lines (MCF7, CAMA-1, and T47D) to discover genomic modifiers of response/resistance following 3 weeks of treatment with combination treatments: capivasertib + fulvestrant and fulvestrant + CDK4/6i. Moreover, a stringent genome-wide CRISPR knockout screen (10 days treatment) was conducted in ER+ breast cancer cells to elucidate mechanisms of resistance to capivasertib + fulvestrant + palbociclib. Results Across the CAPItello-292 Phase 1b population, paired ctDNA samples from 48 patients were analyzed; 42/48 patients discontinued treatment due to disease progression. 259 genetic alterations were detected at baseline and 304 alterations at the EOT. 30 patients had ≥1 newly detected alteration at EOT (30/48, 63%). Alterations in targets downstream of AKT were observed in 15% (7/48) at EOT, compared with 8% (4/48) at baseline, particularly loss-of-function mutations in TSC1, a tumor suppressor that controls mTORC1 signaling, and amplifications in RICTOR, which encodes the catalytic subunit of the mTORC2 complex. RAS pathway alterations were present in 25% (12/48) of patients at EOT versus 17% (8/48) at baseline. Cell cycle gene alterations were detected at EOT in 46% (22/48) versus 42% (20/48) at baseline. Newly detected PIK3CA/PTEN alterations were found at EOT in 6% (3/48) of patients. The specific PIK3CA/PTEN alterations identified at EOT included PIK3CA E545K, PIK3CA H1047R, and PTEN rearrangement, known to be sensitive to capivasertib. CRISPR resistance screen identified loss of TSC1, TSC2 and STK11 (AMPK pathway regulator) as key genes limiting response to capivasertib + fulvestrant. Resistance to the fulvestrant + palbociclib combination was associated with loss of cell cycle control in particular loss of RB. Identified mechanisms of resistance to capivasertib + fulvestrant + palbociclib combination further confirmed that loss of TSC1, TSC2 and STK11 attenuates the effectiveness of the triplet therapy. Conclusions Consistent with AKT being a pivotal node in the PI3K/AKT signaling pathway, these data show that treatment with capivasertib + fulvestrant + CDK4/6i resulted in emergence of alterations associated with activation of alternate mechanisms, such as mTORC1, RAS, and AMPK. The CRISPR data provides additional support to clinical observations.

S. Fernando¹, S. Sisoudiya², I. Barlow-Busch³, J. Tao⁴, M. Fine⁴, M. Cohen⁵, S. Sivakumar², E. Sokol², J. Burke³, N. Vasan¹; ¹laura and Isaac Perlmutter Cancer Center, NYU Langone, New York, NY, ²Foundation Medicine Inc, Foundation Medicine Inc, Cambridge, MA, ³Department of Biochemistry and Molecular Biology, University of Victoria, Victoria, BC, CANADA, ⁴Department of Medicine, Columbia University Irving Medical Center, New York, NY, ⁵Herbert Irving Comprehensive Cancer Center, Columbia University Irving Medical Center, New York, NY.

Background: PI3K pathway alterations occur in ~40% of estrogen receptor-positive metastatic breast cancer (ER+ MBC). These form the basis for the approved PI3Kα inhibitors alpelisib and inavolisib in patients with PIK3CA-altered tumors, and the AKT inhibitor capivasertib in patients with PIK3CA, AKT1, or PTEN-altered tumors. On-target toxicities such as hyperglycemia and rash limit PI3Kα and AKT inhibitor use, and next-generation mutant-selective PI3K inhibitors (e.g., STX-478, RLY-2608) are showing promising clinical activity with reduced metabolic toxicity. We previously demonstrated that PIK3CA double mutations confer heightened PI3Kα inhibitor sensitivity (Vasan et al., Science 2019), underscoring the predictive potential of novel genomic alterations. PIK3R1 encodes the p85α regulatory subunit of PI3Kα, which restrains p110α activity, yet its role in breast cancer is poorly defined. Notably, patients with PIK3R1-altered ER+ MBC were excluded from registrational trials of PI3K and AKT inhibitors and are currently ineligible for these therapies. We sought to define the biological and therapeutic relevance of PIK3R1 mutations in breast cancer. Methods and Results: We analyzed targeted sequencing data from the largest cohort of breast cancer patients to undergo comprehensive genomic profiling in routine clinical care (n = 51,574) and identified PIK3R1 alterations in 3.2% of tumors (n=1,662), evenly distributed between ER+ and ER- disease. Nearly half (49%) of these alterations were in-frame multi-residue insertions and deletions (indels), primarily clustered within the inter-SH2 (iSH2) region, which directly interacts with the adaptor binding domain and C2 domain of p110α. Truncating (20%) and missense (7%) mutations were also frequent. In MCF10A breast epithelial cells, PIK3R1 mutations induced two tiers of increased growth relative to wild-type: a moderate effect from single amino acid substitutions and a stronger effect from multi-residue indels. These two tiers of increased growth were paralleled by similarly graded increases in downstream PI3K signaling. Across cancer cell lines (Cancer Cell Line Encyclopedia) and patient tumors (The Cancer Genome Atlas), PIK3R1 alterations were associated with elevated PI3K pathway activity by phosphoproteomic analysis. Biochemically, PIK3R1 iSH2 alterations increased lipid kinase activity and membrane binding. Phosphopeptides did not appreciably enhance lipid kinase activity, mimicking the activation mechanism of p110α adaptor binding domain and C2 domain mutations, which relieve p85α-mediated inhibition. Strikingly, PIK3R1 mutations conferred pan-sensitivity to active-site PI3Ki (alpelisib, inavolisib), AKTi (capivasertib), and mutant-selective PI3Ki (STX-478, RLY-2608) in vitro. Additional biochemical data, structural studies using hydrogen-deuterium exchange mass spectrometry (HDX-MS), and in vivo data will be presented. Conclusion: This is the first comprehensive study of PIK3R1 alterations in breast cancer. Our findings establish PIK3R1 as a functional driver of oncogenic PI3K signaling and show that PIK3R1 alterations confer sensitivity to both established and investigational PI3K and AKT inhibitors. These findings nominate PIK3R1 alterations as an actionable genomic biomarker and support the inclusion of patients with PIK3R1-altered breast cancer in clinical trials testing PI3K and AKT inhibitors.

J. M. Specht¹, F. Duan², H. Jacene³, E. Q. Konnick⁴, F. Brescia⁵, A. R. Pantel⁶, S. J. Galgano⁷, J. Romanoff², L. M. Peterson¹, M. Muzi⁴, N. U. Lin³, A. K. Abou Hussein⁸, W. Razaq⁹, W. R. Gwin¹, A. N. Shah¹⁰, M. A. Cherian¹¹, B. H. Mavromatis¹², C. L. Wright¹³, D. G. Stover¹¹, A. M. Fowler¹⁴, H. M. Linden¹, A. M. DeMichele⁶, J. E. McConathy⁷, C. Comstock¹⁵, A. C. Wolff¹⁶, D. A. Mankoff⁶; ¹University of Washington/Fred Hutch Cancer Center, Seattle, WA, ²Brown University, Providence, RI, ³Dana Farber Cancer Institute, Boston, MA, ⁴University of Washington, Seattle, WA, ⁵Medical University of South Carolina, Charleston, SC, ⁶University of Pennsylvania/Abramson Cancer Center, Philadelphia, PA, ⁷University of Alabama at Birmingham, Birmingham, AL, ⁸Cooper University Medical Center, Camden, NJ, ⁹University of Oklahoma Health Sciences Center, Oklahoma City, OK, ¹⁰Northwestern University, Chicago, IL, ¹¹The Ohio State University, Columbus, OH, ¹²UPMC Western Maryland, Cumberland, MD, ¹³University of Cincinnati Cancer Center, Cincinnati, OH, ¹⁴University of Wisconsin, Carbone Cancer Center, Madison, WI, ¹⁵Weill Cornell Medicine, New York, NY, ¹⁶Johns Hopkins University, Sidney Kimmel Cancer Center, Baltimore, MD

Background: Patients (pts) with bone-only (BO) or bone-dominant (BD) metastatic breast cancer (MBC) represent a large patient population who are often excluded from clinical trials using RECIST 1.1 as the primary response assessment because bone lesions are classified as non-measurable, non-target lesions. FDG-PET/CT provides a direct measure of tumor metabolic activity, and we hypothesized that categories of metabolic response assessed by FDG-PET/CT will be predictive of important clinical endpoints: progression free survival (PFS), time to skeletal related events (SRE), and overall survival (OS). Methods: Pts with hormone receptor positive MBC, known HER2 status, and radiologically confirmed BD or BO disease receiving systemic therapy for MBC were eligible for the study. Pts underwent a baseline FDG-PET/CT scan (T0) then started systemic therapy within 21 days of the T0 scan. Pts underwent a second FDG-PET/CT scan 12 wks after initiating therapy (T1). Response assessment by serial FDG-PET/CT was completed using mPERCIST criteria (where measurable lesions on the T0 scan were defined as having SUL_peak greater than 1.5 × mean liver SUL). Change in SUL_peak was calculated as the percentage change in SUL_peak of the single hottest lesion at the T0 scan and the single hottest lesion at the T1 scan. Pts responses were classified as mPERCIST responders (complete metabolic response, partial metabolic response, or stable metabolic disease) or non-responders (progressive metabolic disease [PMD]). Unequivocal progression was defined as progressive disease (PD) per RECIST 1.1 (with some modifications related to the MD Anderson criteria for bone metastases for non-measurable disease) or death from any cause. To determine PD, pts were followed serially with standard-of-care imaging at 12-wk intervals for five intervals following therapy initiation, and at 24-wk intervals thereafter. Following disease progression, pts continued to be followed every 6 months to assess vital status and SRE. Results: 138 pts were enrolled at 35 NCTN/NCORP sites. After enrollment, 1 pt was deemed ineligible due to pleural effusion seen on the pre-enrollment CT scan. Of the 137 eligible pts, 2 pts did not undergo the T0 FDG-PET/CT scan, 1 pt did not start systemic therapy, 7 pts did not have a T1 FDG-PET/CT scan, and 11 pts did not have an evaluable mPERCIST response due to absence of measurable lesions on T0. Hence, the analysis set consists of 116 pts. The median age was 59.5 (IQR: 51.5-66) years. 72.4% (84/116) of pts identified as White and 9.5% (11/116) identified as Black or African American, and 5.2% (6/116) identified as Hispanic or Latino. Of 116 mPERCIST response evaluable pts [data cutoff 1 Sep 2025; median follow-up 14.4 months (mos); (IQR: 6.2-25.9)], 21 had PMD and 95 did not have PMD. PFS was compared between pts with and without PMD. The median PFS was 3.0 mos (95% CI: 2.8 to 5.6 mos) for the PMD group and 19.4 mos (95% CI: 15.1 to 30.5 mos) for the non-PMD group (log-rank test P<0.001). Univariate Cox regression analysis yielded a hazard ratio (HR) of 0.16 (95% CI: 0.10 to 0.28; P<0.001), indicating a significantly lower risk of progression in pts with non-PMD compared to PMD pts. A multivariable Cox regression model was fit to estimate the HR of the dichotomized mPERCIST response controlling for age, menopausal status, ECOG performance, HER2 status, metastatic presentation, line of therapy, and type of therapy. The HR for the mPERCIST response was 0.17 (95% CI: 0.09 to 0.33; P<0.001), confirming a statistically significant longer PFS for mPERCIST responders versus non-responders. Conclusions: Our analysis demonstrates that the 12-wk mPERCIST response (dichotomized as PMD versus non-PMD) is significantly associated with PFS. These findings support the role of serial FDG PET/CT mPERCIST response assessment in pts with BD and BO MBC.

S. Yazaki¹, E. Ferraro², M. Acikel¹, P. Razavi², S. Shen², X. Pei¹, S. Modi², S. Chandarlapaty², S. Powell¹, N. Riaz¹, A. Khan¹; ¹Radiation Oncology, Memorial Sloan Kettering Cancer Center, New York, NY, ²Medicine, Memorial Sloan Kettering Cancer Center, New York, NY.

Background: Metastatic breast cancers (MBC) can present as either de novo (dn) stage IV disease or metastatic recurrence of prior stage I-III cancers. Historically, 30% of human epidermal growth factor receptor 2-positive (HER2+) MBC was dnMBC; however, due to recent improvements in adjuvant systemic therapy, over 50% of HER2+ MBC now represents dnMBC. We previously reported that 34% of patients with HER2+ dnMBC had local-regional progression only, with sustained systemic control. Although HER2+ dnMBC shows distinct clinical characteristics and a favorable prognosis following anti-HER2 therapy compared to relapsed MBC, genomic features associated with treatment response and patterns of failure remain poorly understood. Methods: Patients with HER2+ dnMBC who received first-line trastuzumab and pertuzumab (HP)-based therapy from May 2011 to February 2023 and had MSK-IMPACT sequencing data were included. Clinicopathologic data were obtained from medical records. Overall survival (OS) was defined from the first dose of therapy to death from any cause. Tumor samples were categorized as breast, axillary lymph node, and distant metastasis pre- and post-HP therapy. Progression patterns were classified as local-only (breast/ipsilateral axillary lymph nodes), systemic (distant ± local), or no progression. Mutational signature was analyzed on samples with ≥5 single-nucleotide variants using SigMA.Results: We identified 162 patients with HER2+ dnMBC treated with first-line HP-based therapy. Median age was 48.5 years; 61.7% were estrogen receptor (ER)-positive. The median follow-up was 51.2 months. Forty-two patients (25.9%) remained on first-line therapy (durable responders), while 114 (70.4%) progressed; 76 had systemic and 45 had local-only progression. In total, 209 tumor samples were analyzed, including 142 pre-treatment samples (61 breast, 6 axillary lymph nodes, 75 distant) and 67 post-treatment samples (29 breast, 38 distant). In pre-treatment samples, ERBB2 amplification, TP53, and PIK3CA mutations were observed in 71.8%, 66.9%, and 21.8%, respectively. There was no difference in TMB and FGA between pre-treatment breast and distant tumors. Alterations in JAK-STAT signaling pathway genes were enriched in pre-treatment breast tumors compared to pre-treatment distant tumors (q < .001), and were also enriched in pre-treatment tumors that showed local-only progression compared to those with systemic progression (q = .003). Pre-treatment TP53 and PIK3CA mutations were less common in durable responders compared to non-durable responders (PIK3CA; 9.5% vs. 32.6%, p = .005, TP53; 50% vs. 75%, p = .006). PIK3CA mutations were independently associated with worse OS in a multivariable analysis adjusted for age and ER status (HR 2.29, 95%CI 1.09-4.79, p = .03). Tumors with dominant non-clock-like signatures tended to be more frequent in non-durable responders (75% vs. 56%, p = .068). Post-treatment tumors had higher TMB (p = 0.001) and a lower frequency of ERBB2 amplification (58.2% vs. 77.9%, p = 0.005). Paired analysis of pre- and post-treatment samples showed a decrease in dominant signature 3 tumors (p < .001), and an increase in dominant APOBEC signatures tumors (p = .01) after treatment. Conclusions: Pre-treatment breast and metastatic tumors in HER2+ dnMBC exhibited similar genomic characteristics. PIK3CA was an independent prognostic factor in this de novo metastatic setting. JAK-STAT alterations in breast tumors may contribute to local progression. The shift in mutational signatures from signature 3 to APOBEC signature in post-treatment indicates potential mechanisms of acquired resistance. These findings may inform future strategies for tailored treatment in HER2+ dnMBC.