Goetz, Matthew P.

1057 Background: Endocrine therapy (ET), particularly aromatase inhibitors (AIs), for estrogen receptor (ER)-positive breast cancer can lead to acquired ESR1 mutations ( mESR1) driving endocrine resistance and tumor progression. Treatments for mBC with mESR1 are limited, especially after progression on ET/cyclin-dependent kinase 4 and 6 inhibitor (CDK4/6i) combinations. In ELAINE 2, LAS, a next-generation ET (breast ER antagonist) plus Abema had a median progression-free survival (PFS) of ~13 mos, objective response rate (ORR) of 33%, and 24-wk clinical benefit rate (CBR) of 62% with a favorable safety profile in patients with resistant mESR1 mBC (Damodaran S, et al. J Clin Oncol. 2022;40:16 suppl 1022). Here, we report ELAINE 2 data with longer patient follow up (Jan 31, 2023). Methods: In this open-label, phase 2, single-arm study, 29 women with mESR1, ER+/HER2- mBC that progressed on prior ET took oral LAS 5 mg/day and Abema 150 mg BID until disease progression/toxicity. The primary endpoint was safety/tolerability; secondary included PFS, CBR, and ORR. Data were summarized descriptively with no formal hypothesis testing. Results: Patients (median age 60 yrs) had received a median of 2 prior therapy lines for mBC; all but one had prior CDK4/6i exposure and 14 (48%) had prior chemotherapy for mBC. LAS/Abema was well tolerated with primarily grade 1/2 treatment-emergent adverse events (most commonly diarrhea, nausea, fatigue, and vomiting). One treatment withdrawal occurred due to grade 2 diarrhea. No deaths occurred. LAS dose was not reduced; Abema dose was reduced to 100 mg BID in 6 (21%) patients. Median PFS was 56.0 wks (~13 mos) and CBR 65.5% (95% CI, 47.3‒80.1). Median overall survival was not estimable. In 18 patients with measurable lesions, ORR was 55.6% (95% CI, 33.7‒75.4), with median time to response of 5.7 mos and median duration of response of 6.4 mos. Seventy-six percent, 56%, and 39% of patients were progression free at 6, 12, and 18 mos, respectively. Three (10%) patients had scan-identified venous thromboembolic events, with one symptomatic pulmonary embolism and one symptomatic deep vein thrombosis. In 26 patients with evaluable baseline and wk 4 ctDNA, ESR1 mutant allele fraction decreased from baseline to wk 4 in 81% of patients and was not detected in 54%. Conclusions: With longer ELAINE 2 follow up, LAS/Abema continues to be well tolerated with clinically meaningful efficacy in women with mESR1, ER+/HER2- mBC that had progressed on ET and CDK4/6is. Decreases in mESR1 ctDNA suggest effective target engagement of LAS. The PFS (median ~13 mos) and ORR (56%) with LAS/Abema are promising and a confirmatory phase 3 study (ELAINE 3) will begin in 2023. If ELAINE 2 results are confirmed, the combination of LAS and Abema would address a critical unmet need, providing a practice changing option for treating mESR1 breast cancer. Clinical trial information: NCT04432454 .

TPS633 Background: Ovarian function suppression (OFS) when combined with tamoxifen or an aromatase inhibitor is the standard of care for premenopausal estrogen receptor positive (ER+) human epidermal growth factor receptor 2 negative (HER2-) breast cancer (BC) based on SOFT (Francis 2022) and TEXT (Pagani 2022) clinical trials and confirmed in a metanalysis (EBCTCG 2022). However, estrogen deprivation in premenopausal women is associated with significant morbidity and up to 40% of premenopausal women with ER+/HER2- BC are intolerant of OFS. For these women, tamoxifen monotherapy is the only FDA approved option. Therefore, there is substantial room to improve endocrine therapy for premenopausal women with ER+/HER2- BC. (Z)-endoxifen is one of the most active anti-estrogen tamoxifen metabolites, with preclinical data demonstrating superiority to tamoxifen and letrozole in aromatase expressing xenograft models and evidence for antitumor activity in early phase clinical trials in patients with prior progression on endocrine therapy including tamoxifen. In addition to its ability to potently block ERa, (Z)-endoxifen at clinically achievable concentrations inhibits protein kinase C beta 1 (PKCβ1), resulting in downstream AKT inhibition and apoptosis. It is hypothesized that (Z)-endoxifen, when delivered as monotherapy at doses that dually target both ERα and PKCβ1, will potently inhibit BC proliferation while obviating the need for OFS in premenopausal women. If successful, this innovative approach could spare women with endocrine sensitive BC from the side effects of OFS. Methods: To further study (Z)-endoxifen in premenopausal women, a multicenter study in the United States, with possible expansion to other countries was designed (EVANGELINE; NCT05607004). This is a Phase 2, open-label, randomized, neoadjuvant study in premenopausal women with Stage IIA or IIB ER+/HER2- BC. Subjects are randomized 1:1 to either exemestane plus goserelin or (Z)-endoxifen. The primary objective is to determine whether the endocrine sensitive disease (ESD) rate (defined as Ki-67 ≤ 10% after 4 weeks of neoadjuvant therapy) with (Z)-endoxifen is non-inferior to exemestane plus goserelin. Subjects will be enrolled with the intent of surgical treatment in the involved breast(s) after completing 6 months of neoadjuvant treatment. Safety, tolerability, response rates and surgical outcomes will be assessed. Disease progression will be monitored throughout study participation. Paired tumor biopsies and blood sampling will allow for a robust biomarker evaluation. Prior to initiating the Treatment Cohort (162 subjects), up to 12 subjects will be enrolled in a pharmacokinetic (PK) Run-in Cohort to identify the optimal dose for the Treatment Cohort. Enrolment in the PK Run-in cohort began in February 2023. Clinical trial information: NCT05607004 .

581 Background: Several studies have established the crucial role of preexisting immune response measured by tumor-infiltrating lymphocytes (TILs) in triple-negative breast cancer (TNBC). Emerging studies showed that not only the number of TILs but also the location of TILs is as critical. There are 3 distinct immune landscapes described based on the locations of TILs, namely immune enriched (IN), immune excluded (IE), and immune desert (ID), which are associated with outcomes in TNBC treated with immune-checkpoint inhibitors. Here we evaluated characteristics associated with each immune landscape. Methods: NanoString IO360, Digital Spatial Profiling (DSP), and CosMx, a spatial multi-omics single-cell imaging platform, were used. DSP was used to quantify 39 immune-related proteins in stromal and tumor-enriched segments from 44 TNBC samples from the FinXX trial (NCT00114816) and 276 samples from the Mayo Clinic (MC) TNBC cohort (Leon-Ferre BCRT 2018). CosMx was performed in 75 samples from the MC TNBC cohort. First, tumors with TIL quantified by H&E ≤ 30% were classified as ID. The rest of the tumors were categorized according to the intratumoral CD8 protein expression by DSP, with IE having intratumoral CD8 in the lower median and IN having intratumoral CD8 in the upper median. Differential expression listed as log fold change (FC) was estimated from the linear mixed model with significance defined as two-sided p < 0.05. Results: Using DSP in the FinXX trial, intratumoral and stromal higher HLA-DR (FC 1.68, p = 0.001), B2M (FC 0.8, p = 0.005), CD4 (FC 0.74, p = 0.01), and CD40 (FC 1.56, p = 0.001) were associated with IN compared to ID. When comparing IE and IN, higher intratumoral CD11c (FC 0.97, p = 0.01) and stromal CD4 (FC 0.89, p = 0.047), CD20 (FC 0.85, p = 0.016), CD40 (FC 0.95, p = 0.045), and CD27 (FC 0.84, p = 0.024) were associated with IN. Similar findings were observed in the MC cohort. Moreover, intratumoral NY-ESO-1 expression (FC 0.55, p = 0.03) was associated with IN. Using GSEA with IO360 in the FinXX trial, PI3K-Akt signaling was associated with ID compared to IN (p = 0.01). We further evaluated the differential gene expression in a spatially resolved manner using CosMx with single-cell sequencing in the MC cohort. Expressions of MHC class I and class II in tumor cells, including HLA-A, HLA-B, HLA-C, HLA-DRA, HLA-DRB1, HLA-DPA1, and HLA-E, were associated with IN compared to ID and IE. Conclusions: Using an in-depth analysis with spatially defined context, we identified characteristics associated with distinct immune landscapes in TNBC. Our study highlights the potential future implications of intratumoral MHC expression, CD40, and PI3K-AKT as biomarkers and therapeutic targets. Clinical trial information: NCT00114816 .

ABSTRACT Triple negative breast cancer (TNBC) continues to be the subtype of breast cancer with the highest rates of recurrence and mortality. The lack of expression of targetable proteins such as the estrogen receptor and absence of HER2 amplification have made relying on cytotoxic chemotherapy necessary for decades. In the operable setting, efforts to improve outcomes have focused on escalation of systemic therapy and a shift toward preoperative delivery followed by a response adapted approach to postoperative systemic therapy. An improved understanding of tumor biology has resulted in the identification of subsets of patients with specific molecular features, leading to testing and approval of multiple new targeted therapies for this disease. Furthermore, advances in drug development have led to the approval of antibody-drug conjugates that are redefining classification schemes for breast cancer. This review focuses on the modern management of TNBC, with particular focus on recent updates in the treatment of operable disease, and an overview of the most recent promising advances in the therapeutic landscape of metastatic disease. It discusses the practical challenges and unanswered questions resulting from the approval of neoadjuvant immunotherapy and shares an approach in the clinic on topics for which evidence is lacking. In addition, it provides a glimpse into the future, highlighting challenges and opportunities for biomarker based right-sizing of preoperative therapy, refining evaluation of response to preoperative therapy after surgery, early diagnosis and detection of relapse, and areas of needed research for metastatic TNBC.

Abstract Purpose: Clinical trials reported 25% to 30% pathologic complete response (pCR) rates in HER2+ patients with breast cancer treated with anti-HER2 therapies without chemotherapy. We hypothesize that a multiparameter classifier can identify patients with HER2-“addicted” tumors who may benefit from a chemotherapy-sparing strategy. Experimental Design: Baseline HER2+ breast cancer specimens from the TBCRC023 and PAMELA trials, which included neoadjuvant treatment with lapatinib and trastuzumab, were used. In the case of estrogen receptor–positive (ER+) tumors, endocrine therapy was also administered. HER2 protein and gene amplification (ratio), HER2-enriched (HER2-E), and PIK3CA mutation status were assessed by dual gene protein assay (GPA), research-based PAM50, and targeted DNA-sequencing. GPA cutoffs and classifier of response were constructed in TBCRC023 using a decision tree algorithm, then validated in PAMELA. Results: In TBCRC023, 72 breast cancer specimens had GPA, PAM50, and sequencing data, of which 15 had pCR. Recursive partitioning identified cutoffs of HER2 ratio ≥ 4.6 and %3+ IHC staining ≥ 97.5%. With PAM50 and sequencing data, the model added HER2-E and PIK3CA wild-type (WT). For clinical implementation, the classifier was locked as HER2 ratio ≥ 4.5, %3+ IHC staining ≥ 90%, and PIK3CA-WT and HER2-E, yielding 55% and 94% positive (PPV) and negative (NPV) predictive values, respectively. Independent validation using 44 PAMELA cases with all three biomarkers yielded 47% PPV and 82% NPV. Importantly, our classifier's high NPV signifies its strength in accurately identifying patients who may not be good candidates for treatment deescalation. Conclusions: Our multiparameter classifier differentially identifies patients who may benefit from HER2-targeted therapy alone from those who need chemotherapy and predicts pCR to anti-HER2 therapy alone comparable with chemotherapy plus dual anti-HER2 therapy in unselected patients.

Abstract Purpose: Strategies to implement estrogen therapy for advanced estrogen receptor–positive (ER+) breast cancer are underdeveloped. Preclinical data suggest that cycling treatment with 17β-estradiol followed by estrogen deprivation can control tumor growth long-term. Patients and Methods: Postmenopausal women with advanced ER+/HER2− breast cancer with recurrence or progression on ≥ 1 antiestrogen or aromatase inhibitor (AI)-based therapy were eligible. Patients received 17β-estradiol (2 mg orally, three times a day) for 8 weeks followed by AI (physician's choice) for 16 weeks, alternating treatments on an 8-week/16-week schedule until disease progression. Patients then optionally received continuous single-agent treatment until a second instance of disease progression. Endpoints included 24-week clinical benefit and objective response per RECIST, and tumor genetic alterations. Results: Of 19 evaluable patients, clinical benefit rate was 42.1% [95% confidence interval (CI), 23.1%–63.9%] and objective response rate (ORR) was 15.8% (95% CI, 5.7%–37.9%). One patient experienced a grade 3 adverse event related to 17β-estradiol. Among patients who received continuous single-agent treatment until a second instance of disease progression, clinical benefit was observed in 5 of 12 (41.7%) cases. Tumor ER (ESR1) mutations were found by whole-exome profiling in 4 of 7 (57.1%) versus 2 of 9 (22.2%) patients who did versus did not experience clinical benefit from alternating 17β-estradiol/AI therapy. The only two patients to experience objective responses to initial 17β-estradiol had tumor ESR1 mutations. Conclusions: Alternating 17β-estradiol/AI therapy may be a promising treatment for endocrine-refractory ER+ breast cancer, including following progression on CDK4/6 inhibitors or everolimus. Further study is warranted to determine whether the antitumor activity of 17β-estradiol differs according to ESR1 mutation status.

Abstract Introduction: Immune checkpoint blockade (ICB) has clinical activity in triple negative breast cancer (TNBC) but is less effective in the ER+HER2- signature, where there is a cold immune microenvironment (IM) and regulatory T cells (Tregs) may suppress effector T cells. Agents that activate the IM by turning cold tumors hot may support ICB. The biguanides hexyl-benzyl-biguanide (HBB) and its bioisostere hexyl-(cuban-1-yl-methyl)-biguanide (HCB) are candidate agents to activate the IM because they potently inhibit biosynthesis of immunosuppressive epoxyeicosatrienoic acids (EETs) and EET-driven oxidative phosphorylation (OXPHOS), while blocking N-glycosylation of immune checkpoint (IC) proteins. We hypothesized that reversal of hypoxia by biguanides in the ovarian dependent ER+HER2- STAT1 KO SSM2ucd mouse mammary carcinoma (MC) model would suppress Tregs and promote effector T cells in the tumor IM. While the SSM2ucd model did not express immune checkpoint protein PD-L1 (B7-H1), it did express related IC proteins B7-H3 and B7-H4. We hypothesized that by inhibiting OXPHOS and reducing N-glycosylation of immune checkpoint proteins, HBB and HCB may promote efficacy of ICB. We chose the SSM2ucd model to test impact of HCB on the ER+ MC IM. Results: SSM2ucd cells exhibited longer tumor latency (60 days) than the basal 4T1 (10 days) and 67NR (20 days) mouse MC models. SSM2ucd tumor reimplantation shortened latency by more than half, to 20 days. Immunohistochemistry showed that B7-H3 and B7-H4 protein levels were 1.2 (P=0.001) and 1.3-fold (P=0.04) higher in reimplanted tumors vs. control. In SSM2ucd cells, HCB inhibited N-glycosylation of B7-H3 (P=0.01) by 35% and B7-H4 (P=0.02) by 45% and suppressed TGFβ induction of B7-H3 by 21% (P=0.02) and B7-H4 by 79% (P=0.001) at 24 hours, while 14,15-EET promoted N-glycosylation of B7-H3 (1.2-fold; P=0.03) and B7-H4 (1.3-fold; P=0.04) at 4 hours. Effects of HBB and HCB on anti-CD3 and anti-CD28 stimulated mouse splenocytes were assayed. The proliferative effects of HBB on CD4+ and CD8+ cells peaked at 12 uM (p&lt;0.001) and for HCB at 6.25 uM (p&lt;0.001). Tregs decreased with HCB at a threshold of 12 uM (p &lt;0.001) and with HBB at 25 uM (P &lt; 0.001) while 14,15-EET increased Tregs (1.2 fold; P=0.02) and suppressed the CD8+:Treg ratio (0.79; P=0.02). In the SSM2ucd mammary allograft model, although HCB 12mg/kg daily did not inhibit tumor growth, it reduced intratumoral hypoxia by 20% (P=0.01), increased CD8+ TIL by 4.4-fold (P=0.04), decreased Treg:CD4+ TIL ratio by 76% (P=0.02), and decreased Treg:CD8+ TIL ratio by 88% (P=0.01) relative to control. Conclusion: B7-H3 and B7-H4 expression inversely correlated with latency of ER+ MC and may represent targets for immune checkpoint antibodies and their drug conjugates. HCB, an inhibitor of OXPHOS and EET biosynthesis, reduced intratumoral hypoxia, increased CD8+ TIL and reduced the Treg:CD8+ ratio, potentially supporting ICB therapy of ER+ MC by turning cold tumors hot. Supported by CDMRP BCRP Grant BC180596, Award Number W81XWH-19-1-0099 Citation Format: Zhijun Guo, Jianxun Lei, Hrishi Venkatesh, David Owen, Adam Bass, Christine Cannon, Joshua McCarra, Brenda Koniar, Craig Flory, Beverly Norris, Robert J. Schumacher, Swaathi Jayaraman, John Hawse, Emmanuel S. Antonarakis, Emanuel F. Petricoin, Julia Wulfkuhle, Robert D. Cardiff, Elizabeth A. Ambrose, Gunda I. Georg, Kaylee L. Schwertfeger, Michael A. Farrar, Brad St. Croix, Matthew P. Goetz, David A. Potter. Hexyl-(cuban-1-yl-methyl)-biguanide (HCB) suppresses N-glycosylation of immune checkpoint proteins B7-H3 and B7-H4, reverses tumor hypoxia, decreases intratumoral regulatory T cells, and increases intratumoral CD8+ T cells in the ovarian dependent ER+HER2- SSM2ucd mammary cancer allograft model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB102.

Abstract Background: Breast cancer is the most frequently diagnosed cancer and the leading cause of cancer-related death among women worldwide. Triple negative breast cancer (TNBC) represents 15-20% of all breast cancers and is an aggressive subtype. Chemotherapy-based treatments remain the standard of care for TNBC. Unfortunately, chemotherapy resistance is common, and for these patients, outcomes are poor and alternative treatment strategies remain an unmet need. circRNAs are a newly identified class of noncoding RNA molecules with covalently closed circular structures. An increasing number of recent studies including ours have indicated that circRNAs play crucial roles in regulating tumor development and chemoresistance. However, the role of circRNAs in the process of chemotherapy resistance and TNBC progression is not clear. Materials and Methods: As a first step towards identifying circRNAs that participate in the development of chemoresistance in TNBC cells, and to determine if targeting such circRNAs is a novel and efficacious therapeutic strategy, doxorubicin-resistant (Doxo-R), paclitaxel-resistant (PTX-R), and double-resistant (DP-R) cell lines were generated from MDA-MB-231. Human circRNA microarrays were utilized to profile the expression of approximately 14,000 known circRNAs in normal breast tissue, matched patient-derived xenografts (PDX) generated prior to and following neoadjuvant chemotherapy (NAC), and TNBC chemosensitive and chemoresistant cell lines. Top hits were validated using RT-PCR. Results: circRNA microarray profiling identified 429 and 310 transcripts differentially expressed in doxorubicin and paclitaxel resistant cells, respectively, compared to parental chemosensitive cell lines (|FC| ≥ 1.5; p value &lt; 0.05). In comparison to pre-NAC derived xenografts, 1,396 circRNAs were dysregulated among post-NAC PDX models. Further, three circRNAs (hsa_circ_001388, hsa_circ_104652, and hsa_circ_061260) were upregulated in Doxo-R, PTX-R, and post-NAC PDX samples compared to their respective controls. Among these three circRNAs, hsa_circ_001388 (also known as circNSD2) was the only transcript also predicted to be translated into a novel and uncharacterized protein given the presence of a high confidence translation initiation site and IRES sequence. Ongoing studies are aimed at determining the role of circNSD2 protein in breast cancer carcinogenesis, progression, and response to standard of care chemotherapeutics and the mechanistic process by which this protein functions. Additionally, the efficacy of specifically targeting this circRNA as a novel therapeutic approach is being explored. Conclusions: Increasing knowledge of the important functions of circRNAs underlying drug resistance will provide new opportunities for developing efficacious therapeutic strategies and prognostic/predictive biomarkers for TNBC. Citation Format: Xiyin Wang, Michael J. Emch, Xiaojia Tang, Jia Yu, Krishna R. Kalari, Liewei Wang, Matthew P. Goetz, John R. Hawse. The role of circular RNAs in triple negative breast cancer and chemotherapy resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1689.

Abstract Purpose: Triple-negative invasive lobular carcinoma (TN-ILC) of breast cancer is a rare disease and the clinical outcomes and prognostic factors are not well-defined. Methods: Women with stage I-III TN-ILC or triple-negative invasive ductal carcinoma (TN-IDC) of the breast undergoing mastectomy or breast-conserving surgery between 2010 and 2018 in the National Cancer Database were included. Kaplan-Meier curves and multivariate Cox proportional hazard regression were used to compare overall survival (OS) and evaluate prognostic factors. Multivariate logistic regression was performed to analyze the factors associated with pathological response to neoadjuvant chemotherapy. Results: The median age at diagnosis for women with TN-ILC was 67 years compared to 58 years in TN-IDC (p<0.001). There was no significant difference in the OS between TN-ILC and TN-IDC in multivariate analysis (HR 0.96, p=0.44). Black race and higher TNM stage were associated with worse OS, whereas receipt of chemotherapy or radiation was associated with better OS in TN-ILC. Among women with TN-ILC receiving neoadjuvant chemotherapy, the 5-year OS was 77.3% in women with a complete pathological response (pCR) compared to 39.8% in women without any response. The odds of achieving pCR following neoadjuvant chemotherapy were significantly lower in women with TN-ILC compared to TN-IDC (OR 0.53, p<0.001). Conclusion: Women with TN-ILC are older at diagnosis but have similar OS compared to TN-IDC after adjusting for tumor and demographic characteristics. Administration of chemotherapy was associated with improved OS in TN-ILC, but women with TN-ILC were less likely to achieve complete response to neoadjuvant therapy compared to TN-IDC.

ImportanceAurora A kinase (AURKA) activation, related in part to AURKA amplification and variants, is associated with downregulation of estrogen receptor (ER) α expression, endocrine resistance, and implicated in cyclin-dependent kinase 4/6 inhibitor (CDK 4/6i) resistance. Alisertib, a selective AURKA inhibitor, upregulates ERα and restores endocrine sensitivity in preclinical metastatic breast cancer (MBC) models. The safety and preliminary efficacy of alisertib was demonstrated in early-phase trials; however, its activity in CDK 4/6i–resistant MBC is unknown.ObjectiveTo assess the effect of adding fulvestrant to alisertib on objective tumor response rates (ORRs) in endocrine-resistant MBC.Design, Setting, and ParticipantsThis phase 2 randomized clinical trial was conducted through the Translational Breast Cancer Research Consortium, which enrolled participants from July 2017 to November 2019. Postmenopausal women with endocrine-resistant, ERBB2 (formerly HER2)–negative MBC who were previously treated with fulvestrant were eligible. Stratification factors included prior treatment with CDK 4/6i, baseline metastatic tumor ERα level measurement (&amp;lt;10%, ≥10%), and primary or secondary endocrine resistance. Among 114 preregistered patients, 96 (84.2%) registered and 91 (79.8%) were evaluable for the primary end point. Data analysis began after January 10, 2022.InterventionsAlisertib, 50 mg, oral, daily on days 1 to 3, 8 to 10, and 15 to 17 of a 28-day cycle (arm 1) or alisertib same dose/schedule with standard-dose fulvestrant (arm 2).Main Outcomes and MeasuresImprovement in ORR in arm 2 of at least 20% greater than arm 1 when the expected ORR for arm 1 was 20%.ResultsAll 91 evaluable patients (mean [SD] age, 58.5 [11.3] years; 1 American Indian/Alaskan Native [1.1%], 2 Asian [2.2%], 6 Black/African American [6.6%], 5 Hispanic [5.5%], and 79 [86.8%] White individuals; arm 1, 46 [50.5%]; arm 2, 45 [49.5%]) had received prior treatment with CDK 4/6i. The ORR was 19.6%; (90% CI, 10.6%-31.7%) for arm 1 and 20.0% (90% CI, 10.9%-32.3%) for arm 2. In arm 1, the 24-week clinical benefit rate and median progression-free survival time were 41.3% (90% CI, 29.0%-54.5%) and 5.6 months (95% CI, 3.9-10.0), respectively, and in arm 2 they were 28.9% (90% CI, 18.0%-42.0%) and 5.4 months (95% CI, 3.9-7.8), respectively. The most common grade 3 or higher adverse events attributed to alisertib were neutropenia (41.8%) and anemia (13.2%). Reasons for discontinuing treatment were disease progression (arm 1, 38 [82.6%]; arm 2, 31 [68.9%]) and toxic effects or refusal (arm 1, 5 [10.9%]; arm 2, 12 [26.7%]).Conclusions and RelevanceThis randomized clinical trial found that adding fulvestrant to treatment with alisertib did not increase ORR or PFS; however, promising clinical activity was observed with alisertib monotherapy among patients with endocrine-resistant and CDK 4/6i–resistant MBC. The overall safety profile was tolerable.Trial RegistrationClinicalTrials.gov Identifier: NCT02860000