Citations

Abstract Background: DNA methyltransferase (DNMT) isoforms 1, 3A and 3B are enzymes that regulate DNA methylation, controlling gene expression, chromatin stability and genetic imprinting. We have previously shown that DNMT3A protein expression in early triple negative breast cancer (TNBC) is associated with more aggressive clinicopathologic features (higher tumor grade, tumor proliferation and frequency of nodal metastases) and worse clinical outcomes in the absence of chemotherapy (Leon-Ferre et al, SABCS 2024). Pre-clinically, we have shown that DNMT3A protein expression predicts sensitivity to DNMT inhibitors (Yu J et al., J Clin Invest 2018), and that DNMT inhibitor treatment leads to downregulation of DNMT3A, enhanced expression of immune-related pathways, and halted tumor growth, particularly in combination with immunotherapy and paclitaxel. Methods: This is a phase 1, open-label, multicenter study evaluating the safety and recommended phase 2 dose (RP2D) of the oral DNMT inhibitor ASTX727 + paclitaxel + pembrolizumab in metastatic TNBC. ASTX727 is an oral fixed dose combination (FDC) tablet of 35 mg decitabine (active agent) and 100 mg cedazuridine (a cytidine deaminase inhibitor that prevents decitabine degradation in the bowel and liver, allowing oral delivery). The study consists of two parts: Part 1 is a dose-finding cohort, using a 3+3 design, with 5 dose levels (Table), and its primary objective is to determine the RP2D. Part 2 is an expansion cohort (n=6), with the primary objective to confirm tolerability of the regimen at the RP2D. Secondary objectives are to record the antitumor activity of the regimen, describe the AE profile, explore the association of baseline gene expression profiles with clinical benefit, and to evaluate impact of treatment on tumor and ctDNA methylation. Exploratory objectives will evaluate the association of baseline DNMT3A protein expression and antitumor activity, and the impact of treatment on the tumor and peripheral blood immune phenotype. Eligible patients are adults 18 years or older with metastatic TNBC (ER/PR ≤10%, and HER2-negative per ASCO/CAP guidelines), independent of PD-L1 expression. Patients are allowed to have received 0-3 prior lines of systemic therapy in the metastatic setting, including prior immune checkpoint inhibitors in the early and/or metastatic settings. Patients must be eligible for taxanes and have adequate organ and bone marrow function. Patients with treated brain metastases are eligible if previously treated without evidence of progression after 4 weeks. Leptomeningeal carcinomatosis is not allowed. Patients must have an ECOG performance status of ≤2 and have no gastrointestinal disorders impacting absorption of oral medications. As of June 20, 2025, the study is enrolling patients, with 10 patients dosed in the dose-finding phase. Citation Format: R. Leon-Ferre, D. M. Zahrieh, P. P. Advani, D. Quiroga, K. V. Giridhar, P. J. Dizona, D. Stover, X. Wang, A. J. Tevaarwerk, G. M. Choong, S. Yasir, K. R. Kalari, J. Foldi, R. Parajuli, M. H. Hackney, H. K. Chew, R. Wajeeha, H. Bear, E. Sharon, B. Ko, G. I. Shapiro, S. Gore, L. Pelosof, L. Wang, M. P. Goetz. Trial in progress: Phase 1 Study Targeting DNA Methyltransferases in Metastatic Triple-Negative Breast Cancer (NCI Protocol #10546) [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS5-07-20.

Abstract Breast cancer is the most common female malignancy in the U.S. and the leading cause of cancer-related deaths in women worldwide. Nearly 80% of breast tumors express estrogen receptor alpha (ERα) which drives tumor progression. While CDK4/6 inhibitors in combination with endocrine therapy improve overall survival in the metastatic setting and disease-free survival in the adjuvant setting, many patients eventually relapse/progress with resistant forms of disease. In the post-CDK4/6 inhibitor setting, second line endocrine therapy responses are limited, highlighting the need for alternative treatment approaches. Through a genome-wide CRISPR knockout screen, we identified Lemur Tail Kinase 2 (LMTK2) as the top hit exhibiting synthetic lethality in the presence of the tamoxifen metabolite, endoxifen, in both endocrine-sensitive and -resistant cell line models. We further studied multiple other endocrine resistant models and identified that LMTK2 mRNA and protein expression were both increased in ERα cell lines resistant to estrogen deprivation, tamoxifen, fulvestrant, abemaciclib and palbociclib, as well as in endocrine resistant patient derived models. We then studied LMTK2 in patient samples and identified copy number gain/amplification in 67% of ERα+ human metastatic breast tumors, while only 1% of primary tumors harbored these genomic alterations. From a prognostic standpoint, elevated tumoral LMTK2 expression was associated with worse recurrence-free survival and overall survival in early-stage breast cancer. Through proteomic studies, we identified the MET-PI3K-AKT pathway as overactivated in LMTK2 overexpressing cells. Unlike activating PIK3CA mutations that drive AKT signaling, LMTK2 was found to employ an alternative mechanism to activate AKT signaling by inhibiting PP1α, an AKT-inhibitory phosphatase. Further, LMTK2 overexpression was sufficient to drive endocrine and CDK4/6 inhibitor resistance in vitro and resulted in dramatic increases in AKT and ERα phosphorylation and activity. While knockout of LMTK2 in treatment naïve ERα+ breast cancer cells neither altered AKT/ERα phosphorylation/activity nor endocrine/CDK4/6 inhibitor responsiveness, attempts to knockout LMTK2 in endocrine/CDK4/6 inhibitor-resistant models failed, further confirming its essentiality in this setting. In summary we have uncovered LMTK2 as a novel biomarker of endocrine/CDK4/6 inhibitor resistance that is mechanistically linked to the AKT pathway. LMTK2 may be a novel drug target for the effective treatment of resistant and advanced forms of ERα+ breast cancer. Citation Format: A. M. Detry, S. Marigliano, J. Heinen, A. Bass, C. Jones, R. S. Muthyala, M. P. Goetz, J. R. Hawse. Lmtk2: a novel driver of endocrine resistance in ER+ breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS2-11-01.

Abstract Background: We have identified two potentially predictive signatures of palbociclib resistance: the RBsig, composed of E2F1/E2F2 dependent genes, which is associated with genetic loss of RB1, and the ratio between the gene expression levels of CCNE1 to RB1 (CCNE1/RB1). Both signatures have been previously analyzed in vitro and in neoadjuvant and metastatic studies with palbociclib. The present analysis aims to explore the role of RBsig and CCNE1/RB1 in the phase III adjuvant PALLAS trial, which randomized patients to palbociclib plus endocrine therapy, versus endocrine therapy alone. Materials and methods: Gene expression data from PALLAS were generated using the HTG EdgeSeq Oncology BM Panel. Of the 87 genes composing RBsig, 46 were available within the EdgeSeq dataset and were used for the analyses; CCNE1 and RB1 were both available. RBsig was calculated as the mean of the Z-score scaled gene expression (log) of the 46 genes; CCNE1/RB1 was computed as the log ratio between the mRNA expression of CCNE1 and RB1. The prognostic/predictive effect of the continuous signatures in terms of invasive disease-free survival (IDFS), distant recurrence-free survival (DRFS), and overall survival (OS) was evaluated using Cox proportional hazard models. Results: HTG data were available from 2,587 of 5,748 patients from the entire PALLAS intention-to-treat (ITT) cohort. As RBsig and CCNE1/RB1 levels are affected by pre-surgical treatments, we limited the analyses to 1612 pre-treatment samples (530 core biopsies and 1082 surgical specimens) (analysis population). Compared with the entire PALLAS ITT cohort, the analysis population was significantly enriched for patients with low clinical risk (41.3% vs 59.4%) who did not receive prior chemotherapy (17.5% vs 55.9%). In keeping with the PALLAS ITT cohort, no significant benefit from the addition of palbociclib was observed in the analysis population. Higher baseline RBsig or CCNE/RB1 values were significantly and independently associated with worse IDFS (hazard ratio [HR] 1.63, 95% confidence interval [CI] 1.3-2.0; HR 1.22, 95%CI 1.1-1.3, respectively), DRFS (HR 1.94, 95%CI 1.5-2.5; HR 1.26, 95%CI 1.1-1.4, respectively) and OS (HR 1.79, 95% CI 1.3-2.4; HR 1.26, 95%CI 1.1-1.4, respectively) in the analysis population. No predictive effect of palbociclib benefit was observed for either signature in the analysis population, nor according to clinical risk or chemotherapy receipt. Interestingly, higher RBsig was strongly prognostic in patients who did not receive adjuvant chemotherapy (HR 1.83, 95%CI 1.4-2.3) but not in those who received it (HR 1.01, 95%CI 0.6-1.6) (interaction p=0.02) suggesting a potential predictive role of RBsig for chemotherapy benefit. Conclusions: In the PALLAS trial, higher baseline RBsig or CCNE1/RB1 values are associated with worse prognosis, but are not predictive of benefit from palbociclib in the adjuvant setting. RBsig might be associated with differential benefit from chemotherapy. Further study of these biomarkers may strengthen their role in risk stratification.https://acknowledgments.alliancefound.org Citation Format: L. Malorni, D. Hlauschek, E. Mayer, M. Benelli, L. Biganzoli, I. Migliaccio, A. Kermanidis, G. Pfeiler, E. P. Mamounas, A. M. Brufsky, M. Bellet-Ezquerra, K. Clifton, G. Rubovszky, T. Foukakis, M. Goetz, S. Lee, M. Ruiz-Borrego, W. Symmans, K. V. Ballman, F. Liu, J. Machacek-Link, C. Denkert, A. DeMichele, M. Gnant. Prognostic and predictive role of RBsig and CCNE1/RB1 gene-expression signatures for patients with early breast cancer treated with endocrine therapy with or without palbociclib in the PALLAS trial (ABCSG-42, AFT-05, BIG 14-03) [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS2-07-13.

Abstract Significance: Emergence of resistance to endocrine therapies represents a major challenge in theclinical management of estrogen receptor positive (ER+) breast tumors. CDK4/6 inhibitors(palbociclib, ribociclib, abemaciclib) have improved survival when combined with endocrine therapy. Nonetheless, resistance to CDK4/6 inhibitors eventually occurs. One of the emerging mechanisms ofresistance toward endocrine therapy and CDK4/6 inhibitors is characterized by the enrichment ofbreast cancer stem cells (CSCs) that show stem-like features such as self-renewal capacity, highCD44 expression and ALDH activity. We have demonstrated that Aurora-A kinase (AURKA) has apivotal role in promoting cancer cellular plasticity and clonal expansion of endocrine resistantCD44high/ALDHhigh CSCs. Our preclinical studies on AURKA-driven tumor stemness provided thetherapeutic foundation for a Phase-II clinical trial (TBCRC041) of the AURKA inhibitor, alisertib, whichdemonstrated clinically meaningful activity in patients with endocrine and CDK4/6 inhibitors resistantER+ metastatic breast cancer (MBC). Despite such activity, alisertib resistance finally occurred. Therefore, we hypothesize that up-regulation of IL6R/STAT3 and PR stemness oncogenic pathwaysmay sustain the clonal expansion of CD44high/ALDHhigh CSCs after tumor progression on AURKAinhibitors. Experimental Design: We used variant MCF-7 breast cancer models that show resistance toendocrine therapy (letrozole and fulvestrant) and alisertib (MCF-7 LR/FR, MCF-7 LR/FR-AlisR) todefine in vitro and in vivo the efficacy of stemness-targeted therapies to enhance alisertib activity byimpairing the enrichment of hormone refractory CD44High/ALDHHigh CSCs. Parental MCF-7 cells wereused as control. We employed exclusive PDX-derived 3D-Mammospheres (MPS) that wereestablished from the Phase-II clinical trial TBCRC041 to test the efficacy of stemness-targetedtherapies in enhancing the therapeutic activity of alisertib. In Vitro Studies: Immunofluorescence andimmunoblot assays were used to assess the expression of breast cancer stemness biomarkers. Enrichment of ALDHHigh CSCs was measured using Red-Aldefluor kit. Cell proliferation and Red-Annexin-VHigh apoptotic cells were quantified in real-time using IncuCyte S3. In Vivo Studies: 1x106breast cancer cells were injected into the mammary fat pad of NSG-female mice and treated withfulvestrant, alisertib and tocilizumab as monotherapy or in combination. Control groups were treatedwith saline solution placebo. Drug Treatments: Tocilizumab and Onapristone were used to inhibit theIL6R/STAT3 and PR stemness pathways, respectively. Results: Our study demonstrates for the first time that nuclear STAT3/PR complex mediatesAURKA-induced expression of KLF4 stemness gene and enrichment of endocrine resistantCD44high/ALDHhigh CSCs. Alisertib resistant MCF-7 LR/FR cells showed increased expression ofIL6R, T288-AURKA, S727-STAT3 and S294-PR compared to alisertib naïve cells. MCF-7 LR/FR-AlisR-derived 3D-MPS had increased KLF4 and CD44 expression. Alisertib resistant PDX modelsalso showed high levels of CD44, IL6R/STAT3 and KLF4 expression. Remarkable, pharmacologicalblockade of PR and STAT3 transcriptional regulatory networks enhanced the efficacy of alisertib byreducing the enrichment of CD44High/ALDHHigh CSCs. Conclusions: These findings provide the strong preclinical rationale for the development of novelcombinatorial therapeutic strategies targeting IL6R/STAT3 and PR stemness pathways to enhancethe efficacy of AURKA inhibitors that will result in the selective eradication of CD44High/ALDHHighCSCs with consequent benefits on the overall survival of patients with advanced ER+ breast tumorsrefractory to endocrine therapies and CDK4/6 inhibitors.輀 Citation Format: A. B. D'ASSORO, M. Drago, T. Haddad, M. GOETZ, K. Giridhar, K. Kalari, L. Manzella, P. Vigneri, F. Drago, C. Vancheri, C. Lange, J. Ingle. Stemness-targeted therapies to enhance the efficacy of AURKA inhibitors in endocrine resistant er+ breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS2-11-08.

Abstract Introduction Serial assessments of circulating tumor cells (CTCs) in metastatic breast cancer (MBC) provide independent prognostic significance for progression free survival and overall survival. A low CTC burden (<5 CTCs / 7.5mL blood) at baseline is favorable prognostically, and reduction of CTCs during treatment indicates response to treatment (Janni, CCR 2025). Despite increasing evidence supporting its clinical validity, real-time serial CTC enumeration has not been utilized broadly in clinical practice in the U.S. The ACT-MBC study is a prospective observational study to evaluate the impact of serial CTC testing on treatment decisions and response assessment (NCT05662345). Provider (physician or advanced practitioner) perception of CTC testing is measured with questionnaires at baseline (PQ1), and for each pt during the first restaging visit (PQ2), and at the end of the study (PQ3). We previously reported PQ1 results (Giridhar, SABCS 2024). Here, we report on results of PQ2. Methods ACT-MBC prospectively enrolled patients (pts) at Mayo Clinic Rochester and Mayo Clinic Health Systems sites (Mankato, Albert Lea and Eau Claire). Pts with MBC were enrolled at time of progressive disease during any line of therapy (TNBC) or ≥2nd line metastatic therapy for ER+/HER2- disease. Pts were tested for CTCs using the FDA-cleared CellSearch® assay which included enumeration and biomarker testing on CTCs for ER, HER2, and PD-L1. The treating clinician determined the regimen and timing of restaging scans. CTC testing was performed once per cycle (every 3-4 weeks) with results returned to the treating provider within 7 days. We descriptively summarized PQ2 responses from providers who had at least one on-treatment CTC result available by the first restaging visit. We report the proportion (90% CI) of providers who agreed or strongly agreed that assessing CTCs enhanced their ability to determine response to therapy and that the CTC results helped guide their decision to continue or change therapy (co-primary endpoints) while accounting for within provider correlation. Results As of June 25, 2025, 15 providers enrolled a median of 2 pts each (range: 1 to 15 pts) and completed PQ2 surveys on 40 pts. 7 providers (47%) practiced in the community setting and 4 (27%) were nurse practitioners. Most pts were female (98%), White (98%), and had ER+/HER2- MBC (80%). Baseline (C1) CTC results were elevated (≥5 CTCs) in 24 (60%) pts (CTC range 0-609). At C2 (n=39), CTC values remained undetected in 7 pts, decreased in 21 pts, increased in 8 pts, and remained unchanged in 3 pts. At PQ2 (n=40), based on clinician interpretation of radiographic findings, results showed 2.5% of pts with complete response, 20% with partial response, 25% with stable disease, 17.5% with mixed response, and 35% with progressive disease. Clinicians reported that CTC results were concordant with imaging results in 34 (85%) responses, discordant with imaging in 7.5% of responses, and unclear in the remaining 7.5% of responses. Most clinicians (82.5%) agreed or strongly agreed that assessing CTCs enhanced their ability to determine response to therapy (90% CI: 68.3%, 96.7%). Further, 70% of providers agreed or strongly agreed that the CTC results helped guide their decision to continue or change therapy (90% CI: 50.8%, 89.2%). 32.5% of the time, CTCs results influenced the timing of the first restaging scans. In 11/25 pts without progressing disease, CTC results influenced the timing of future scans. In the 7/8 pts with a mixed response at first restaging scans with decreasing CTCs, 4 remained on the same systemic therapy. Conclusions Oncologists in academic and community practice responded that serial CTC assessments enhance ability to determine response of therapy, guide decisions to continue or change treatments, and influence timing of radiographic assessments. Citation Format: K. V. Giridhar, D. Zahrieh, M. Hanna, A. Singh, E. Al-Hattab, P. Schumacher, J. Carroll, J. Hoppenworth, T. Haddad, A. Tevaarwerk, B. Rudder, G. M. Choong, S. Basu, R. A. Leon-Ferre, T. O'Brien, A. Lewis, F. de Snoo, L. Stork-Sloots, F. K. Kuhr, S. Lazaro, R. Prendergast, D. S. Spinner, M. P. Goetz. Assessing the impact of real-time serial circulating tumor cell results on the management of patients with metastatic breast cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS4-03-10.

Abstract Background Invasive lobular carcinoma (ILC) differs from invasive ductal carcinoma (IDC) in its estrogen receptor (ER) signaling pathways, response to systemic therapy, and imaging features. We evaluated differences in early change in the proliferation marker Ki67 and features on magnetic resonance imaging (MRI) after novel endocrine based neoadjuvant therapy in lobular versus non-lobular cases on the I-SPY2 EOP Trial. Methods EOP is a prospective, randomized NET trial enriched for patients with ILC. Patients with stage II-III ER positive HER2 negative breast cancer with low molecular risk (Mammaprint low or Mammaprint Hi1 plus high Sensitivity to Endocrine therapy Index) were randomized to one of several endocrine therapy-based arms. Treatment arms include novel oral SERDS with or without CDK4/6 inhibitor, aromatase inhibitors, and SERMs with or without ovarian function suppression in pre-menopausal patients. Patients undergo repeat core needle biopsy with central Ki67 assessment after 3 weeks of treatment and serial breast MRI. Here, we compare endocrine therapy responsiveness as measured by change in Ki67 at 3 weeks and functional tumor volume (FTV) on pre-surgical MRI in lobular versus non-lobular cases. Results We evaluated 253 patients of whom 148 (58.5%) had non-lobular cancer (93.9% IDC), and 105 (41.5%) had lobular cancer (17.1% mixed ILC/IDC). Overall, mean age was 53.3 years, 63.8% of tumors were grade 2, and 86.5% were Mammaprint (MP) Low with the remaining being Hi1, with higher rates of MP Low in the lobular versus non-lobular cohorts (92.4% versus 83.4%, p=0.0358). At baseline, tumors in the lobular cohort had significantly lower mean Ki67 than those in the non-lobular cohort (11.2% [SD 12.8] versus 16.4% [SD 14.4%], p=0.0038). Consistent with this, the proportion with baseline tumor Ki67 <10% was significantly higher in the lobular cohort than the ductal cohort (60.4% versus 39.3%, p=0.001). At 3-week biopsy, mean Ki67 remained lower in the lobular versus non-lobular group (3.4% versus 5.4%, respectively, p=0.0415). The proportion of patients with Ki67 <10% at 3 weeks did not differ significantly between lobular and non-lobular cases (88.6% versus 79.7%, respectively, p=0.08). Among the 40 lobular cases and 88 non-lobular cases with baseline Ki67 ≥10%, week 3 Ki67 decreased to <10% in 90% of lobular cases and 72.7% of non-lobular cases (p=0.07). On MRI, mean FTV at baseline was similar among lobular and non-lobular cases (21 cc3 versus 16.3 cc3). On pre-surgical MRI, patients in the lobular cohort had significantly higher mean residual FTV than those in the non-lobular cohort (5.7 cc3 versus 3.8 cc3, p=0.034). Interestingly, when splitting the lobular cohort into mixed lobular and pure lobular, mean residual MRI FTV was 7.5 cc3 in mixed cases and 3.8 cc3 in pure lobular cases, reflecting that FTV decreased the least in mixed lobular cases (decrease of 51.1% in mixed cases, 69.7% in pure lobular cases, and 73.9% in non-lobular cases, p=0.0255). Conclusions These results highlight differences in lobular versus non-lobular breast cancer. Lobular cases had lower baseline Ki67, and similar rates of decrease in Ki67 after 3 weeks of NET than non-lobular cases. Despite this, mean FTV after completion of 6 months of NET was higher in lobular tumors. These findings may reflect differences in early versus late indicators of response to NET, or differential response patterns by histology. Alternatively, standard response measures such as Ki67 and FTV may need lobular-specific modifications. Ongoing work includes central assessment of Ki67 from surgical specimens, analysis of changes in gene expression signatures, and evaluation of surgical outcomes such as breast conservation rates by histologic subtype. Citation Format: R. Mukhtar, P. Norwood, A. Borowsky, S. Alkhafaji, K. Giridhar, C. Vaklavas, A. Elias, M. Wei, M. Goetz, N. Onishi, O. Olopade, L. Van 't Veer, L. Huppert, W. F. Symmans, L. Brown Swigart, C. Wu, C. Yau, D. Yee, M. Magbanua, E. Price, N. Hylton, L. Esserman, J. Chien. Differential early response to neoadjuvant endocrine therapy in lobular versus non-lobular breast cancer on the Endocrine Optimization Pilot (EOP) of the I-SPY2 Trial [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS4-02-01.

Abstract Background: Tumor-infiltrating lymphocytes (TILs) in HER2+ and triple-negative breast cancer (BC) are prognostic and associated with therapy response; however, there are few data regarding distribution and significance of TIL values in ER+ HER2-negative (HER2-) MBC patients (pts) treated with CDK4/6 inhibitors (CDK4/6i). CDK4/6 inhibition may promote antitumor immunity by production of type III interferons and tumor antigen presentation, and by inhibition of regulatory T cells (Goel, Nature 2017). Further, the addition of immune checkpoint therapy to palbociclib numerically prolonged PFS in the 2nd line (2L) setting (Mayer, JCO 2024). We aimed to investigate the prognostic role of TILs derived from fresh metastatic tumor biopsies in pts with ER+HER2- MBC prior to starting CDK4/6i and endocrine therapy (ET) in the first line (1L) and 2L setting. Methods: PROMISE (NCT0281902) is a multicenter, prospective study that enrolled women commencing the CDK4/6i palbociclib (Pb) with letrozole (1L) or fulvestrant (2L) for HR+HER2- MBC. The primary goal was to identify novel genomic variants and pathways associated with PFS. Here, we performed a secondary analysis to quantitate stromal TILs using protocol mandated pretreatment tumor biopsies and determine their association with PFS. Using the Kaplan-Meier method, we reported median PFS (mPFS) and 3-year PFS percentage (%) (95% confidence intervals [CIs]) by TILs [≥20% vs <20%] separately within 1L and 2L. As sample TIL values were generally concentrated at zero (27/58 = 46.6%), coupled with wide spread of sample TIL values ≥20% [range: 20%- 80%], we applied a ≥20% threshold for PFS analysis (high vs low TIL group) as described by Luen, Salgado et al. Lancet Oncol 2017. Statistical significance was based on the Wilcoxon test of the null hypothesis that there was no difference in the PFS curves by TILs. PFS is reported at median follow-up of 21 months. Results: Pretreatment tumor biopsies from 58/68 (85.3 %) pts were evaluable (42 in 1L and 16 in 2L). Median [Q1, Q2] TIL values were higher in 1L (5% [0, 10]) vs 2L (0% [0, 15]). The most common biopsy sites were bone (44.8%), lymph node (17.2%), liver (17.2%) and breast (12%). Overall, TILs frequency (>20%) by the 4 most common biopsy sites were bone (2/20, 10%), lymph node (4/7, 57%), liver (2/6, 33%), and breast (1/7, 14%). In 1L, although PFS was not significantly different (P = 0.237), mPFS (44 vs 34 months) and 3-year PFS 75.0%, [95% CI: 50.3%, 100%] vs 46.4% [95% CI: 31.0%, 69.5%]) were longer in pts with high versus low TILS, respectively. In 2L, the 3-year PFS was numerically similar comparing high and low TILs groups, (25.0%, [95% CI: 4.6%, 100%] vs 22.5% [95% CI: 6.7%, 76.1%]) with no significant difference in PFS (P = 0.761). Conclusions: In pts with ER+HER2- MBC treated with Pb and ET, PFS was numerically longer comparing high vs low TIL values in 1L but not 2L settings. Future studies evaluating TILs in larger datasets are needed to validate these findings. Genomic analysis (DNA/RNA) and its association with TIL levels is underway and will be presented at the meeting. Funding: The Center for Individualized Medicine, Mayo Clinic; Mayo Clinic Breast Specialized Program of Research Excellence (SPORE P50CA116201); Mayo Clinic Cancer Center (CA15083-40A2); TEMPUS (https://www.tempus.com/) and Conquer Cancer Foundation of ASCO (Career Development Award 2018, Dr. O’Sullivan) Citation Format: C. C. O'Sullivan, A. M. Moyer, A. Nassar, D. M. Zahrieh, V. Suman, X. Tang, K. J. Thompson, R. C. Hentz, J. J. Harrington, A. Moreno-Aspitia, D. W. Northfelt, T. C. Haddad, S. Chumsri, P. P. Peethambaram, K. J. Ruddy, K. V. Giridhar, R. A. Leon-Ferre, R. Weinshilboum, L. Wang, K. R. Kalari, M. P. Goetz. Tumor-infiltrating lymphocytes in advanced ER-positive, HER2-negative breast cancer prior to treatment with palbociclib and endocrine therapy: a retrospective analysis of the PROMISE study [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS1-10-22.

Abstract Background: TheTAILORx trial established that adjuvant endocrine therapy (ET) is non-inferiorto chemotherapy plus ET (CET) in T1-2N0 HR+/HER2-negative early breast cancer(EBC) with an Oncotype DX (ODX) 21-gene recurrence score (RS) of 11-25, RS 0-10is associated with low distant recurrence (DR) rates with ET alone, and RS26-100 is associated with higher DR rates despite CET. Although ODX isprognostic for 10-year DR, it has limited value for late DR >5 years. Inorder to develop improved DR risk prognostication, we developed multimodal AImodels integrating clinical, molecular, and histopathology features to provideprognostic information for early (<5years), late (>5 years), and overall DR at 15 years using primary tumorsamples and clinical data from volunteers who participated in TAILORx. Methods: DigitizedH&E slides were generated and nucleic acids extracted and sequenced from∼5000 primary tumor samples using Caris MI Tumor Seek-Hybrid, of which 2,808(56%) were used for model training and 5-fold nested cross validation (60/20/20split). Models included single-modality (clinical [C], image [I], molecular [M:ODX genes]), dual-modality (CM, IC), and multimodal (ICM). An expandedmolecular model (M+) model was also derived from expanded molecular inputsderived from whole transcriptome sequencing (WTS) data, including 5 commercialgene signatures (ODX, MammaPrint, Prosigna, EndoPredict, BCI) and 57 high-variancegenes; only EndoPredict, BCI, and ODX gene signatures were retained for model M+.Model-CM+ combined clinical plus expanded molecular features, while ICM/ICM+integrated image, clinical, and (expanded) molecular inputs. Risk scores were dichotomized into high vs. low risk groups usingthresholds chosen to yield risk group proportions closely matching thosedefined by the ODX RS cutoff of 25 for distinguishing high vs. low genomicrisk. Theprimary endpoint was distant recurrence (DR). Prognostic performance wasassessed using truncated concordance index (C-index, 90th percentile of eventtimes), hazard ratios (HRs), and log-rank tests. DNA-level whole exomesequencing (WES) features were also tested but did not provide additionalprognostic information and thus excluded from final models. Results: Theactual ODX continuous RS achieved a C-index of 0.625 for overall DR and 0.743early DR, but no prognostic value for late DR (0.52). Similar results for theC-index were observed for ODX+C for overall DR (0.619), early DR (0.724), andlate DR (0.514). Of all modelsevaluated, model-ICM+ gave the best overall prognostic accuracy for overall DR (C-index0.713; hazard ratio [HR] 3.56 for comparison of high vs. low risk, p<0.001)and late DR (C-index 0.655; HR 2.36, p<0.001), and ranked second for earlyDR (C-index 0.772; HR 6.59, p<0.001) behind the CM+ model (C-index 0.782). Amongsingle-modality models, Model-M+ improved early DR prognostication comparedwith a molecular model including only ODX genes(C-index 0.767 vs 0.744 for M),whereas Model-I was strongest for late DR prognostication (C-index 0.635). Conclusions: Molecularfeatures primarily drove prognostic accuracy for early DR within 5 years,whereas histopathology features strengthened the prognostic accuracy for lateDR after 5 years. Multimodal models captured these complementary signals, withICM+ giving the best overall accuracy for individualized prognostic assessment.Additional validation of the optimized ICM+ model and other models is beingperformed in an intendent validation sample of 2000 TAILORx volunteers, withresults to be presented at the meeting.Supported by the Breast CancerResearch Foundation and the U.S. NIH/NCI U10CA180820 and 5U24CA196172, and theU.S. Postal Service Breast Cancer Stamp Fund. Citation Format: J. A. Sparano, V. Wang, R. J. Gray, D. F. Makower, K. S. Albain, D. F. Hayes, C. E. Geyer, E. C. Dees, M. P. Goetz, J. A. Olson, S. S. Badve, T. J. Saphner, T. J. Whelan, M. Radovich, V. G. Kaklamani, G. W. Sledge. Multimodal artificial intelligence (AI) models integrating image, clinical, and molecular data for predicting early and late breast cancer recurrence in TAILORx [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr GS1-08.

Importance Molecular analyses of biospecimens collected from study participants are essential for identifying biomarkers that can tailor treatments to specific subsets of patients who are most likely to benefit. Sharing of data and biospecimens from clinical trials enables personalized, patient-centric use of cancer therapies and accelerates the development of new treatments. Objective To describe obstacles to sharing data and biospecimens and to propose strategies to enhance access and collaboration. Evidence Review This is a Special Communication authored by 53 academic investigators and patient representatives from the breast cancer community with extensive experience in conducting clinical and translational research. The article also evaluates the impact of biomarker research on specifying responsive subpopulations in the 29 registrational clinical trials that have led to approval of a new drug for treatment of breast cancer between 2017 and 2024. Findings Clinical trial participants are increasingly asked to provide tissue and/or body fluid biospecimens for biomarker research that is typically controlled by the sponsoring pharmaceutical company, but published biomarker studies are rare. Among 29 breast cancer registrational studies reported in the past 8 years, none resulted in biomarker research that restricted a drug’s approved indication. Herein, strategies to maximize the value of clinical data and biospecimens contributed by participants are proposed, thereby supporting the shared goals of the pharmaceutical industry and academia to improve patient care. These strategies include (1) establishing coleadership structures involving academia and patients in clinical trial design and conduct, (2) ensuring that informed consent forms state that data and biospecimens will be shared with academia for future research, (3) requiring the sharing of clinical data as a condition for regulatory approval, and (4) enabling access to biospecimens and translational research data for independent studies on biomarkers that may indicate drug efficacy and toxicity. Conclusions and Relevance Data and biospecimen sharing from registrational trials has been suboptimal. Improving clinical data, biospecimens, and biospecimens’ related data sharing requires concrete actions and a multidimensional stakeholder approach to accelerate the impact of clinical cancer research on the quality of patient care.