Citations

569 Background: Several studies established the prognostic role of both the amount and locations of tumor-infiltrating lymphocytes (TILs) in TNBC. Three distinct immunotypes were described based on the amount and locations of TILs: immune enriched (IN), immune excluded, and immune desert. Using single-cell spatial transcriptomic analysis in the Mayo Clinic TNBC cohort, our previous studies showed the central role of interferon (IFN) signaling in IN phenotype. Herein, we evaluated the association between IFN and outcomes in TNBC in 3 independent datasets. Methods: NanoString IO360 was performed in 114 samples from FinXX (NCT00114816) to generate 22-gene IFNα and 33-gene IFNγ signatures. RNA sequencing was performed in 388 samples from CALGB 40603 (NCT00861705). 3038 TNBC samples were tested by WTS (NovaSeq; Caris Life Sciences, Phoenix, AZ). Median values were used as cut-offs for high vs low IFNγ RNA expression and 18-gene IFNγ signature scores. Caris Life Science CODEai was used to evaluate real-world overall survival (OS) obtained from insurance claims and calculated from tissue collection to last contact using Kaplan-Meier estimates. Chi-square, Mann-Whitney U, ANOVA, and Cox regression were used. Results: A high 22-gene IFNα signature score was associated with significantly improved recurrence-free survival (RFS) in FinXX (hazard ratio [HR] 0.32, 95% confidence interval [CI] 0.14-0.74, p 0.007) and OS (HR 0.28, 95%CI 0.12-0.66, p 0.003). Similar findings were observed with 33-gene IFNγ signature with significant improvement in RFS (HR 0.21, 95%CI 0.09-0.51, p < 0.001) and OS (HR 0.18, 95%CI 0.08-0.44, p < 0.001). Furthermore, in CALGB 40603, both IFNα and IFNγ scores were positively associated with pathologic complete response (pCR: IFNα p 0.019 and IFNγ p 0.007) and residual cancer burden (RCB: IFNα p 0.044 and IFNγ p 0.013). Using the Caris data platform to further validate, we identified 2899 TNBC patients (pts) with genomic and clinical outcome data. High IFNγ expression was associated with significant improvement in OS (25.95 vs 17.43 months; HR 0.65, 95% CI 0.59 – 0.72, p < 0.001). Similarly, pts with high IFNγ signature scores had significant improvement in median OS (25.79 vs 16.22 months; HR 0.66, 95% CI 0.6 – 0.73, p < 0.001). Conclusions: This study underscores the pivotal role of IFN signaling in TNBC. High IFNα and IFNγ signatures were consistently associated with improved RFS, OS, higher pCR rates, and lower RCB across clinical trial cohorts and real-world data. These findings signify IFN signaling as a potential key biomarker and therapeutic target in TNBC. Support: U10CA180821, U10CA180882, U24CA196171; Breast Cancer Research Foundation, Mayo Clinic Breast Cancer SPORE (P50CA116201-17), Bankhead Coley, W81XWH-15-1-0292, P50CA015083, R35CA253187; https://acknowledgments.alliancefound.org . Genentech. Clinical trial information: NCT00114816 and NCT00861705 .

548 Background: TNBC is associated with higher risk of recurrence and poorer survival rates than other breast cancer subtypes. For node-positive TNBC or tumors larger than 0.5 cm, chemotherapy is generally recommended. For stage I TNBC, the benefit from chemotherapy, the optimal regimen, and whether to administer it in the neoadjuvant versus adjuvant setting remain controversial. Here, we report the treatment patterns and clinical outcomes of patients with stage I TNBC treated at Mayo Clinic. Methods: Using the Mayo Clinic Tumor Registry data, we identified patients with stage I TNBC treated between 2010 and 2021. We used the pathologic tumor (T) category for patients treated with upfront surgery and the clinical T category for patients receiving neoadjuvant chemotherapy. We used the Kaplan-Meier method and log-rank test to compare recurrence-free survival (RFS) according to treatment groups, measured from the day of surgery. RFS was reported at a median follow-up of 3.9, 6.2, and 7.3 years for patients who received chemotherapy neoadjuvantly, adjuvantly, and surgery alone with no chemotherapy, respectively. Results: A total of 602 patients with Stage I TNBC were included, with a median age of 62 years. 290 (48%) underwent upfront surgery followed by adjuvant chemotherapy (pT1a: 2%, pT1b: 25%, pT1c: 74%), 127 (21%) received neoadjuvant chemotherapy followed by surgery (cT1a: 2%, cT1b: 11%, cT1c: 87%), and 185 (31%) underwent primary surgery without any chemotherapy (pT1mi/T1a: 33%, pT1b: 32%, pT1c: 35%). Most patients treated with neoadjuvant therapy received anthracycline/cyclophosphamide + taxane (70%), while most patients treated in the adjuvant setting received a non-anthracycline containing regimen (60%). The 5-year RFS was 96% (95% CI: 93-100%) for patients who received chemotherapy neoadjuvantly, 95% (95% CI: 92-98%) for those who received chemotherapy adjuvantly, and 85% (95% CI: 80-91%) for those treated with surgery alone and no chemotherapy ( P < 0.0001). 71 (56%) of the 127 patients who received neoadjuvant therapy achieved a pCR, with only 1 RFS event (at 14 months) in that group, compared to 4 RFS events among those not achieving pCR (at 12, 20, 28 and 75 months, respectively). Among patients not receiving chemotherapy (either adjuvantly or neoadjuvantly), the 5-year RFS rates were 94% for pT1mi/T1a, 92% for pT1b and 72% for pT1c (T1mi/a/b vs T1c, P = 0.009). Conclusions: In this large cohort of stage I TNBC, patients who received chemotherapy (adjuvantly or neoadjuvantly) had better 5-year RFS compared with those treated with locoregional therapy only. Among patients receiving chemotherapy, 5-year RFS was nearly identical regardless of whether chemotherapy was administered before or after surgery. Interestingly, patients undergoing upfront surgery were more likely to receive an anthracycline-sparing chemotherapy regimen.

570 Background: Major histocompatibility complex class I (MHC I) plays a critical role in immune surveillance by binding peptides derived from intracellular proteins and presenting them on the cell surface for recognition by CD8+ T cells. Loss or downregulation of MHC I expression has been identified as a key mechanism of immune evasion in cancers. Here, we evaluated MHC I expression and outcomes in all subtypes of breast cancer (BC). Methods: 9,038 BC samples were analyzed via NGS (592-gene panel, NextSeq; WES/WTS, NovaSeq; Caris Life Sciences, Phoenix, AZ), including triple-negative BC (TNBC) 3,038, HER2-positive (HER2+) 1,082, and hormone receptor-positive (HR+HER2-) 4,918. Immune cell fractions were estimated using WTS deconvolution (Quantiseq). MHC I ( HLA-A / HLA-B / HLA-C )-high (H) and -low (L) were classified by RNA expression above or below the 25th percentile. Real-world overall survival (OS) was derived from insurance claims and calculated from tissue collection to last contact using Kaplan-Meier. NanoString IO360 was performed in 114 samples from the FinXX trial (NCT00114816). Statistical significance was assessed using chi-square, Mann-Whitney U, ANOVA, and Cox regression with multiple comparison adjustments (q<.05). Results: TNBC had higher expression of HLA-A and HLA-B (median TPM: 169 and 191) compared to HER2+ (146.6 and 170, q<0.05) and HR+HER2- (141.7 and 157.5, q<0.05). However, there was no significant difference in HLA-C expression across 3 BC subtypes. In TNBC, MHC I-H tumors had higher frequencies of PD-L1 positivity (66.2% vs. 13.1%) as well as higher infiltration of B cells (4.5% vs. 3.2%), M1 macrophages (5% vs. 1.5%), M2 macrophages (4% vs. 2.1%), Tregs (2.8% vs. 0.8%), CD8 + T cells (1.8% vs. 0%), dendritic cells (3.2% vs. 2.8%), higher T-cell inflamed score (137 vs. -144), and IFN g score (0.02 vs. -0.49) compared to MHC I-L TNBC (all q<.05). MHC I-H TNBC was associated with significant improvement in median OS (30.1 vs. 15.2 months, HR 0.55, 95% CI 0.46-0.65, p<0.0001). However, this survival difference was not observed in patients with MHC I-H vs. MHC I-L in HER2+ (HR 1.04, 95% CI 0.74-1.47, p = 0.81) and HR+HER2- (HR 0.87, 95% CI 0.75-1.02, p= 0.09) BC subtypes. We further validated the MHC I expression in the FinXX trial. Similarly, patients with MHC I-H had significant improvement in recurrence-free survival (HR 0.27, 95%CI 0.11-0.66, p = 0.002) and OS (HR 0.23, 95% CI 0.09-0.57, p = 0.0005) compared to MHC I-L. Conclusions: Our findings demonstrate that higher MHC I expression is associated with higher immune infiltration and improved outcomes in TNBC but not in HER2+ or HR+HER2- BC subtypes. These results suggest that MHC I expression plays a critical role in the tumor microenvironment of TNBC. Future studies are needed to evaluate the prognostic value and potential therapeutic target of MHC I in TNBC. Support: Breast Cancer Research Foundation, Bankhead Coley, W81XWH-18-1-0564. Clinical trial information: NCT00114816 .

604 Background: The addition of OFS to standard endocrine therapy (ET) improves iDFS for preM women with early-stage hormone receptor positive (HR+) breast cancer (BC). Incomplete OFS occurs in a subset of women, the long-term clinical significance of which is unclear. This study evaluates local sE2 levels in preM women receiving OFS with an oral SERD in the neoadjuvant (NA) setting. Methods: The EOP is a I-SPY2 sub-study investigating NA endocrine-based strategies in pts with HR+ HER2- Stage 2/3 BC predicted to have lower benefit from chemotherapy. Pts were randomized to oral amcenestrant 200 mg/d, alone or in combination with either letrozole 2.5 mg/d or abemaciclib 150 mg bid. Additional pts were randomized to a standard of care (SOC) arm consisting of an aromatase inhibitor. All PreM pts received OFS with monthly GnRHa up to 2 wks prior to C1D1 of study therapy. Pts were treated for 6 months prior to surgery. sE2 levels were collected locally prior to each GnRHa injection. Tumor Ki67 was assessed at 3 weeks and surgery, and associated with sE2 level. Statistical significance was determined using a two-sided Student’s t-test and a significance level 0.05. Results: Between 5/2021-8/2022, 74 pts were enrolled to an amcenestrant containing arm, 40 of whom were preM. 38/40 pts had at least one local sE2 level measured in follow up. Of these 38 pts (median age 45 years), 37 suppressed sE2 into the postmenopausal (postM) range at one or more follow-up timepoints. Of the 37 pts that completely suppressed into the postM range, 6 pts had sE2 levels rebound into the preM range (mean 319 pg/mL, range 20-848 pg/mL) with peak sE2 levels occurring at a median of 12 weeks from C1D1. One pt never suppressed to the postM range (peak sE2 1227 pg/mL) and was found to have an ovarian cyst after 3 months on amcenestrant, requiring surgery. One additional pt had multiple ovarian cysts (sE2 84 pg/mL). Median age of the 7 pts who had sE2 rebound was 43 years. There was no significant difference in Ki67 at 3 weeks and surgery between pts whose sE2 levels remained suppressed (median Ki67 2.0%) compared to those whose estradiol rebounded into the preM range or never suppressed (median Ki67 1.5%). In the 5 pts whose sE2 rebounded to > 200 pg/mL, all pts had tumor Ki67 < 10% at 3 weeks and surgery. Between 12/2022 and 8/2023, 20 pts were enrolled to AI +/- OFS and 11 pts were preM. Of the 11 preM pts, all pts suppressed sE2 to the postM range. No SOC patients had sE2 levels rebound into the preM range. Conclusions: In this study of neoadjuvant oral SERD with monthly OFS, 7/38 (18.4%) preM pts had sE2 levels remain or rebound into the preM range. sE2 levels did not appear to impact Ki67 suppression on NA ET. Work up for ovarian cysts should be considered in pts with symptoms, or significantly elevated or persistent sE2 levels. Clinical trial information: NCT01042379 .

612 Background: EOP, an I-SPY2 sub-study, evaluates the tolerability and activity of novel endocrine strategies in stage 2/3 breast cancer (BC) patients (pts) predicted to have lower chemotherapy benefit. Laso, a selective estrogen receptor modulator (SERM), has shown favorable toxicity profile and activity in HR+/HER2- endocrine-resistant metastatic BC. Methods: Pts with Stage 2/3 HR+/HER2-, MammaPrint (MP) low risk BC were enrolled. Pts with MP High1 BC were included if clinically node-negative. Pts received oral laso 5 mg daily for six 28-day cycles. Laso was continued until the day prior to surgery. Premenopausal pts received ovarian function suppression (OFS) starting C2D1. The primary endpoint was feasibility (>75% of patients completing >75% study therapy). Baseline (T0), 3-wk (T1) biopsies, and the surgical specimen (T3) was assessed centrally for Ki-67. Breast MRI functional tumor volume (FTV) was performed at T0, T1, 12 weeks (T2), and pre-operatively (T3). Blood was collected for tumor informed ctDNA at T0, T1, T2, T3. Advance event (AE) was assessed using CTCAE V5. Results: From 3/2023 to 5/2024, 20 pts were enrolled. Median age 50.5 years, 50% premenopausal, and 1 male pt. 60% cN0, 80% MP low-risk signature. 18 (90%) pts completed >75% study therapy. Two pts discontinued treatment due to pt preference. Median Ki67 at T0 was 14.7%. At T1, 87.5% of pts remained or suppressed Ki67 to <10% and 37.5% suppressed to <2.7%. Ki67 at T1 was similar between pre-and postmenopausal pts despite OFS (Table). The median MRI FTV was 8.4 cc at T0, and 3.4 cc at T3. Median % FTV reduction from T0 to T3 was -47.5%. 2/20 pts (10%) achieved a modified PEPI score of 0. No patients achieved completed pathological response. Of the 16 pts with RCB results, 2 (12.5%) RCB-1, 6 (37.5%) RCB-2, 8 (50%) RCB-3 disease. 14 pts had ctDNA available at T0. 4/14 were ctDNA+ at T0, 2 of whom became ctDNA negative, and 2 remained ctDNA+. 10/14 pts were ctDNA negative at T0, 8 of whom remained negative, 2 became positive at T1 then cleared. All AEs were grade(G) 1 except 1 pt with G2 hot flashes. Most common AEs include hot flashes (85%), constipation (50%), fatigue (50%), and nausea (35%). One pt had G3 hypersensitivity and hypertension, both unrelated to therapy. Conclusions: Neoadjuvant laso demonstrates a favorable AE profile and promising anti-tumor activity in suppressing 3-wk Ki67 and MRI FTV change in pts with HR+ HER2-negative early BC. Ki67 suppression in premenopausal pts was seen in the absence of OFS. Clinical trial information: NCT01042379 . Ki67 expression at pre-treatment, and 3-wk time point. All Patients(n=20) Premenopausal (n=10) Postmenopausal (n=9) Median Ki67 expression Baseline 10.0% 12.5% 10.0% 3-wk 5.1% 3.0% 6.0% Number of pts with Ki67 expression <10% at 3-wk 87.5% 1 87.5% 85.7% Number of pts with Ki67 expression <2.7% at 3-wk 37.5% 50% 28.6% 1 Include the one male pt.

3008 Background: Numerous studies have demonstrated the prognostic value of ctDNA analysis after neoadjuvant chemotherapy for early-stage high-risk breast cancer. Few studies have characterized ctDNA in pts receiving NET for early-stage hormone receptor positive (HR+)/HER2- BC that is predicted to benefit less from chemotherapy. Methods: Cell-free DNA (cfDNA) was isolated from 432 plasma samples from 108 pts enrolled in the I-SPY2 EOP trial. Pts had Stage 2/3 HR+/HER2-, MammaPrint low or high risk 1 BC. Pts were randomized to one of 7 neoadjuvant-based treatment arms including arms containing AI, Z-endoxifen, Lasofoxifene, vepdegestrant (ARV-471), and Abemaciclib. Pts were treated for 6 months prior to surgery. Blood was collected at baseline (T0), 3 weeks (T1), 12 weeks (T2), and 6 months (T3). A personalized ctDNA test (Signatera) was designed to detect up to 16 patient-specific mutations (from whole-exome sequencing of pretreatment tumor) in cfDNA by ultra-deep sequencing. The chi-square test was used to assess associations between categorical variables, and the Wilcoxon rank-sum test was used to evaluate differences in medians. Results: ctDNA information was available for 101 patients at baseline (T0) (Table 1). At T0, 36 (35.6%) patients were ctDNA-positive. 23/36 (63.9%) became ctDNA-negative and 13/36 (36.1%) remained ctDNA-positive. At T0, 65/101 patients (64.4%) were ctDNA-negative. Of these, 57 (87.7%) remained negative, while 8 (12.3%) became ctDNA-positive at T1 before reverting to ctDNA-negative. A higher percentage of ctDNA-positive patients at T0 were cN+ compared to ctDNA-negative pts (p = 0.036, 64% vs. 40%). Additionally, ctDNA-positivity at T0 was strongly associated with higher Ki67 (p = 0.03) and larger functional tumor volume by MRI at baseline (p = 0.03). T3/T4 and high-grade tumors at baseline were also associated with having ctDNA positivity at baseline, though this was not statistically significant (p = 0.34 and 0.058, respectively). Conclusions: In this study of pts with Stage 2/3 HR+ HER2- BC with largely MammaPrint low risk signatures, over one-third of pts had detectable ctDNA at baseline. Detectable ctDNA at baseline was associated with cN+ disease, larger FTV, and high baseline Ki67. The majority of pts with positive ctDNA at baseline cleared the ctDNA on NET. Clinical trial information: NCT01042379 . Clinicopathologic characteristics of EOP patients. All Patients(N=101) Age at screening 55 (27-80) years* Clinical N Stage Node+ (cN+) 49 (48.5%) Node- (cN-) 52 (51.5%) SET status High 84 (83.2%) Low 14 (13.9%) Missing 3 (2.97%) MammaPrint (MP) risk** High risk 1 (H1) 15 (14.9%) Low risk 86 (85.1%) *Mean(min-max). **H1: MP score between 0 and -0.57; low: MP score between 0 and 0.355.

e13042 Background: The conventional treatment paradigm for metastatic breast cancer (MBC) is to continue systemic therapy until disease progression. With improved systemic therapy options, some patients (pts) with MBC attain radiographic remission. Tumor-informed circulating tumor (ct)DNA detects molecular disease burden and may guide decision-making. We report on an institutional cohort of pts with MBC in which tumor-informed ctDNA testing was utilized prospectively to guide decisions on systemic therapy or surveillance imaging frequency. Methods: Utilizing an IRB-approved protocol, we retrospectively identified pts monitored with tumor-informed ctDNA using the Signatera™ mPCR-NGS assay (Natera, Inc) as part of routine clinical care in Mayo Clinic, Rochester. We then identified a subset with biopsy-proven MBC who attained radiographic remission of disease following systemic therapy, deemed by the treating medical oncologist as “exceptional responders”. Real-time ctDNA testing was used to inform a decision to either discontinue systemic therapy, treatment de-escalation (remove a component of multi-drug therapy), increase the time interval between restaging scans, or stay off therapy previously stopped due to toxicity . Data on breast cancer characteristics, imaging results, ctDNA test results, and outcomes were collected. Descriptive statistics were used to analyze the data. Results: 10 pts met our study criteria. The median age at MBC diagnosis was 42 years (range 28-61); 5 pts had TNBC, 3 pts had HER2+, and 2 pts with ER+/HER2- MBC. Most pts (60%) were on first or second line therapy (range 1-5). The median duration of the most recent regimen was 27.5 months (range 10-70 months). All pts were without radiographic evidence of active cancer at the time of ctDNA testing with negative results (0 MTM/mL) at the time of clinical decision-making. 4 pts stopped all systemic therapy and proceeded with observation (1 trastuzumab, 1 trastuzumab deruxtecan, 1 pembrolizumab, and 1 talazoparib). 2 pts discontinued a targeted therapy (1 olaparib, 1 abemaciclib) while continuing endocrine therapy. 1 patient stopped targeted therapy (abemaciclib) due to a blood clot, and ctDNA testing guided the decision to not start another CDK4/6 inhibitor while continuing endocrine therapy. In 3 pts the frequency of restaging scans was decreased to either annually or semi-annually. The median follow-up after a ctDNA-informed decision was 11 months (range 1-41 months). At this time, no patient has converted to ctDNA-positivity, experienced resurgence or progression of disease, needed to change systemic therapy or increase imaging frequency. Conclusions: Tumor-informed ctDNA may offer additional confidence to oncologists and pts to deescalate systemic therapy or decrease the interval of imaging frequency in exceptional responders with MBC. Additional data to support this approach is needed.

Abstract Introduction: Heme-binding biguanides inhibit cytochrome P450 monooxygenase-mediated biosynthesis of the epoxy fatty acid (EpFA) (±)14,15-EET, which promotes progression of ER+HER2- breast cancer as well as regulatory T cell (Treg) proliferation. However, there are no known strategies to exploit this vulnerability. Metformin has shown promise but lacks sufficient potency and clinical utility. Through structure-based design, heme binding hexyl cubane derivatives of metformin were identified, which potently inhibited (±)14,15-EET biosynthesis. Fluorination of the hexyl moiety improved pharmacokinetics (PK) and reduced toxicity. Among the fluorinated derivatives, C5F2-HCB potently inhibited (±)14,15-EET biosynthesis (IC50=3.1 uM for C5F2-HCB vs. 5 mM for metformin). C5F2-HCB and metformin are candidate agents for TME activation from “cold” to “hot” due to their inhibition of EETs biosynthesis and EET-driven oxidative phosphorylation (OXPHOS) which promotes tumor hypoxia. We then tested whether C5F2-HCB or metformin could inhibit immune exclusion in the immunologically “cold” ovarian dependent STAT1 KO SSM2ucd mammary carcinoma model, in part, by reducing Tregs and/or increasing CD8 or CD4 tumor infiltrating T cells (TIL). We hypothesized that biguanides, by reducing EETs, would suppress Tregs and promote effector T cells in the TME. We also hypothesized that C5F2-HCB would be more effective than metformin in modulating the TME, based on greater potency for inhibition of (±)14,15-EET biosynthesis. Both biguanides were tested at their maximum tolerated dose (C5F2-HCB; 18 mg/kg daily and metformin; 50 mg/kg every other day). Results: In the SSM2ucd allograft, all combination treatment groups with biguanides reduced tumor growth relative to the IgG control at the endpoint (day 63) : metformin + IgG (51.5±9.8% of control), C5F2-HCB + IgG (53.3±12.8%), metformin + anti-PD-1 (50.4±11.5%), and C5F2-HCB + anti-PD-1 (49.6±12.8%) [(P &lt; 0.05) for all comparisons]. In contrast, anti-PD-1 antibody alone failed to reduce tumor growth at the endpoint. Flow cytometry analysis of the TME revealed that only C5F2-HCB + IgG treatment increased CD8+ tumor-infiltrating lymphocytes (TILs) compared to all other treatment groups (&gt;2-fold; P &lt; 0.05). Additionally, only C5F2-HCB + anti-PD-1 reduced intratumoral Tregs (&gt;3-fold; P &lt; 0.05). C5F2-HCB in combination with IgG or anti-PD-1 reduced intratumoral macrophages by &gt;50% (P &lt; 0.001) and reduced the M2/M1 macrophage ratio by 32-61% (P &lt; 0.05) relative to other groups. No effects on CD4+ T cells were observed. Furthermore, C5F2-HCB and IgG or anti-PD-1 increased intratumoral neutrophils by 75- 120% compared to the anti-PD-1 or IgG (P &lt; 0.05). (Multiple comparisons tested by ANOVA). Conclusion: Relative to IgG alone, C5F2-HCB and metformin inhibited growth of the SSM2ucd allograft in combination with either anti-PD1 or IgG. In contrast to metformin, only C5F2-HCB modulated the TME, including increased CD8+ TILs with IgG, reduced Tregs with anti-PD-1, and decreased M2/M1 macrophage ratio with IgG and anti-PD-1. C5F2-HCB, in contrast tometformin, was effective in modulating an immune excluded TME and may contribute to a strategy for clinical translation. Citation Format: Zhijun Guo, Jianxun Lei, Joshua McCarra, Qing Cao, Michael J. Pryzbilla, Brenda Koniar, Beverly Norris, Robert J. Schumacher, Swaathi Jayaraman, John R. Hawse, Antonino B. D'Assoro, Ilia G. Denisov, Stephen G. Sligar, Kathryn L. Schwertfeger, Michael A. Farrar, Dipak Panigrahy, Hammock D. Bruce, Elizabeth A. Ambrose, Gunda I. Georg, Goetz P. Matthew, David A. Potter. C5F2-hexyl-(cuban-1-yl-methyl)-biguanide (C5F2-HCB) overcomes an immune excluded tumor microenvironment (TME) and suppresses tumor growth in the ovarian dependent ER+HER2- SSM2ucd mammary carcinoma allograft model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB010.

Abstract Background: Endocrine therapy (ET) with aromatase inhibitors (AI) plus ovarian function suppression (OFS) is standard for premenopausal women with ER+/HER2- breast cancer. However, many premenopausal women are intolerant of AI + OFS, leaving tamoxifen (with or without OFS) as their only option. Extensive studies have demonstrated that premenopausal women treated with neoadjuvant ET whose tumors exhibit Ki-67 ≤ 10% following 4 weeks of therapy (defined as endocrine sensitive disease (ESD)) achieve 5-year distant disease-free survival &gt; 96%. Problematically, only 41% of patients reach this ESD threshold with tamoxifen compared to 78% with AI+OFS (Nitz JCO 2022). (Z)-endoxifen (ENDX) is a potent selective estrogen receptor modulator (SERM) that inhibits ERα at plasma concentrations of 3-5 ng/ml and dually targets PKCβ1, a suspected oncogene in breast tumors, at concentrations ≥ 500 ng/ml. We therefore developed the EVANGELINE study (NCT05607004) is an open label, randomized neoadjuvant phase II study that includes a pharmacokinetic (PK) Run-in, aiming to evaluate whether ENDX +/- goserelin is non-inferior to exemestane plus goserelin with respect to ESD rate. Following the completed PK run-in portion of the study, where 40mg/day or 80mg/day ENDX were assessed, the 40 mg/day dose was selected for further evaluation in the second part of the trial. The second part is comprised of a randomized treatment cohort, for patients with a Ki-67% &gt;10% at preregistration randomized to either 40mg (Z)-endoxifen + goserelin or exemestane + goserelin neoadjuvant treatment and a Single Arm Treatment Cohort for patients who have a Ki-67 ≤ 10% at pre-registration assigned to 40mg/day (Z)-endoxifen monotherapy neoadjuvant treatment. Methods: To investigate ENDX further in premenopausal women, a multicenter study in the United States, was designed(EVANGELINE; NCT05607004). This phase 2, open-label, randomized, neoadjuvant trial includes a PK run-in phase designed to assess whether the 4-week ESD rate with (Z)-endoxifen + goserelin is non-inferior to that of exemestane + goserelin in premenopausal women with Stage IIA or IIB ER+/HER2- breast cancer with diagnostic Ki67 &gt; 10%. Additionally, a single arm Phase 2, open-label neoadjuvant Cohort was designed to assess the 4-week ESD rate with ENDX monotherapy in premenopausal women with Stage IIA or IIB ER+/HER2- BC with diagnostic Ki67 ≤10%. Subjects will undergo neoadjuvant treatment for six months before receiving surgical treatment in the involved breast. This study will additionally assess safety, tolerability, response rates, surgical outcomes, and monitor disease progression. Comprehensive biomarker evaluations will be conducted using paired tumor biopsies and blood samples collected at baseline and week four, as well as 6 month surgical specimens. Enrollment in the PK run-in phase (n-22) was completed in Fall 2024 with the second portion of the trial slated to begin early in 2025. Citation Format: Matthew P. Goetz, Vera J. Suman, Claudia Lopez, Hayley Erickson, Lonnissa Nguyen, Sandra S. Hammer, Lida Mina, Pooja Advani, Roberto Leon-Ferre, Karthik Giridhar, Felipe Batalini, Daniel Flora, Jason M. Jones, Nusayba A. Bagegni, Katie N. Hunt, Mara Piltin, Amy Degnim, James N. Ingle, Judy C. Boughey, Sarah A. Buhrow, Joel M. Reid, Matthew Schellenberg, John R. Hawse, Steven C. Quay. A randomized phase 2 non-inferiority trial of (Z)-endoxifen + goserelin and exemestane + goserelin as neoadjuvant treatment for premenopausal women with ER+/HER2- breast cancer (EVANGELINE) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr CT237.