Receptor-positive Triple-negative Breast Cancer Research Articles

Abstract P1-13-14: Increased androgen receptor expression as a mechanism of acquired anti-androgen resistance in androgen receptor-positive triple negative breast cancer

Abstract Background: Triple negative breast cancer (TNBC) represents the most aggressive breast cancer subtype, and despite recent treatment advances, clinical outcomes remain poor, particularly in the metastatic setting. TNBC is a biologically heterogeneous entity, and up to 50% of TNBC express the androgen receptor (AR). Gene expression subtyping has identified a subset of AR-TNBC with a luminal AR-driven (LAR) phenotype. Consequently, there has been great interest in translating androgen receptor signaling inhibitors (ARSIs) approved for use in prostate cancer such as enzalutamide and bicalutamide into the management of LAR-TNBC. However, early phase clinical trials of anti-androgens in metastatic AR-TNBC have had modest results. While multiple mechanisms of anti-androgen resistance have been identified in prostate cancer, including AR amplification and over-expression, emergence of AR splice variants (AR-Vs), and transition to AR independent growth, molecular determinants of anti-androgen resistance in TNBC remain poorly understood, and pre-clinical models to study acquired ARSI resistance in LAR-TNBC have been limited. Methods: A novel pre-clincial model of acquired ARSI resistance was derived from the LAR subtype MDA-MB-453 cell line through serial passaging with increasing concentrations of enzalutamide. Celltiter Blue viability assays were used to determine enzalutamide IC50 of parental and enzalutamide-resistant (EnzR) cell lines. RNA was extracted from the parental and resistant cells, reverse transcribed and expression of AR, splice-variants and canonical AR target genes were evaluated using RT-qPCR. Immunofluorescence was used to investigate the amount and nuclear localization of the AR protein within the parental and enzalutamide resistant cells. Results: EnzR MDA-MB-453 cells were derived through culture in increasing concentrations of enzalutamide for >6 months, and found to have a significant increase in enzalutamide IC50 compared to the parental cell line. EnzR MDA-MB-453 cells had a 4.5 fold increase in full-length AR expression at the transcriptional level compared to the parental cell line. AR protein expression and nuclear localization were also increased in EnzR cells compared to the parental cell line. Canonical AR targets including NKX3.1 were downregulated in response to enzalutamide in parental but not EnzR cells. While pathogenic AR splice variants (AR-Vs) implicated in ARSI resistance in prostate cancer were detected at low levels in MDA-MB-453 cells, no increase in AR-V expression was observed in the EnzR cell line. Conclusions: We have developed a novel pre-clinical model of anti-androgen/ARSI resistance in LAR-TNBC, and identified AR overexpression and persistent AR signaling associated with acquired enzalutamide resistance in LAR-TNBC. While pathogenic AR splice variants have been implicated in ARSI resistance in metastatic prostate cancer and are detected in the MDA-MB-453 cell line model, enzalutamide resistance was not associated with increased AR splice variant expression in this model. Future work will investigate mechanisms leading to AR overexpression as well as combination therapeutic strategies to overcome acquired ARSI resistance. Citation Format: Hannah Krause, Marina N. Sharifi, Serena K. Wolfe, Jamie M. Sperger, Kari B. Wisinski, Ruth O’Regan, Joshua Lang. Increased androgen receptor expression as a mechanism of acquired anti-androgen resistance in androgen receptor-positive triple negative breast cancer. [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P1-13-14.

Hsa-mir-3163 and CCNB1 may be potential biomarkers and therapeutic targets for androgen receptor positive triple-negative breast cancer.

Breast cancer (BC) is the most common malignancy in female, but the role of androgen receptor (AR) in triple-negative breast cancer (TNBC) is still unclear. This study aimed to exam the performance of innovative biomarkers for AR positive TNBC in diagnosis and therapies. Four datasets (GSE42568, GSE45827, GSE54002 and GSE76124) were analyzed by bioinformatic methods and the differential expression genes (DEGs) between the AR positive TNBC tissues and normal tissues were firstly identified by limma package and Venn diagrams. Next, Gene Ontologies (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed to explore the relationship between these DEGs. Then, the Protein-protein interaction (PPI) network was constructed. CytoHubba and bioinformatic approaches including Molecular Complex Detection (MCODE), Gene Expression Profiling Interactive Analysis (GEPIA), the Kaplan-Meier (KM) plotter and The Human Pro-tein Atlas (THPA) were used to identify the hub genes. Lastly, a miRNA-hub-gene regulatory axis was constructed by use of Target Scan database and ENCORI database. As a result, a total of 390 common DEGs were identified, including 250 up-regulated and 140 down-regulated. GO and KEGG enrichment analysis showed that the up-regulated DEGs were mostly enriched in the cell division, mitotic nuclear division, nucleosome, midbody, protein heterodimerization activity, cadherin binding involved in cell-cell adhesion, systemic lupus erythematosus and alcoholism, while the down-regulated DEGs were mainly enriched in carbohydrate metabolic process, extracellular space, extracellular region, zinc ion binding and microRNAs in cancer. Then, 13 hub genes (CCNB2, FOXM1, HMMR, MAD2L1, RRM2, TPX2, TYMS, CEP55, AURKA, CCNB1, CDK1, TOP2A, PBK) were selected. The survival analysis revealed that only CCNB1 was associated with significantly poor survival (P <0.05) in TNBC patients. Finally, we found that hsa-miR-3163 took part in the regulation of CCNB1 and constructed a potential hsa-miR-3163-CCNB1 regulatory axis. The results of current study suggest that CCNB1 and hsa-miR-3163 may serve as highly potential prognostic markers and therapeutic targets for AR positive TNBC. Our findings may make contributions to the diagnosis and therapies of AR positive TNBC.

Abstract 590: Multiplex liquid biopsy for AR pathway activity in metastatic androgen receptor-positive triple negative breast cancer

Abstract Introduction: The androgen receptor (AR) is expressed in up to 50% of patients with triple negative breast cancer (TNBC), and a luminal androgen receptor (LAR) subtype has been identified that is dependent on androgen receptor signaling, but AR protein expression by immunohistochemistry (IHC) has been a suboptimal biomarker for response to anti-androgen therapies in AR-TNBC and there remains an unmet need for biomarkers to identify patients likely to benefit from anti-androgens. We have developed a multiplex liquid biopsy encompassing AR protein expression and downstream gene expression analysis in circulating tumor cells (CTCs). We report here the longitudinal evaluation of this liquid biopsy as part of a phase I trial (NCT03090165) of the anti-androgen bicalutamide with selective CDK4/6 inhibitor ribociclib in metastatic AR-TNBC. Methods: Peripheral blood was collected (n=11) prior to treatment, after two cycles of treatment, and at progression and processed using the VERSA (Versatile Exclusion-based Rare Sample Analysis) microfluidic chip that integrates CTC capture and downstream analysis. For protein expression, fixed CTCs were captured immunomagnetically and stained on chip for AR followed by quantitative fluorescent microscopy. For gene expression analysis, live CTCs were captured immunomagnetically followed by RNA isolation on chip, reverse transcription, and RT-qPCR for a panel of 40 luminal AR genes. Results: AR protein and transcript were detected in CTCs from 10/11 patients, and AR target gene expression in 8/11 patients. The pathologic ligand independent AR splice variant AR-V7 was detected in 2/11 patients. Pretreatment CTC number was higher in patients with anti-androgen resistant (range 3-753) than anti-androgen sensitive (range 0-30) disease. Among patients with low AR IHC (&amp;lt;10%) on biopsy, high AR protein and transcript expression in pre-treatment CTCs was associated with anti-androgen sensitivity. Among patients with high AR IHC (&amp;gt;50%) on biopsy, AR-V7 was detected in CTCs of 2/3 of patients with resistant disease and 0/2 of patients with sensitive disease. Conclusions: This study demonstrates the feasibility of a longitudinal multiplex liquid biopsy as a pharmacodynamic biomarker of AR pathway activity in AR-TNBC, and suggests that AR pathway activity in CTCs may provide additional information to solid tumor biopsy AR expression in predicting sensitivity to anti-androgens. This work also demonstrates for the first time the detection of AR-V7 in CTCs in anti-androgen resistant AR-TNBC, suggesting that AR-V7 expression may be a mechanism of anti-androgen resistance in AR-TNBC as it is in advanced prostate cancer. Citation Format: Marina N. Sharifi, Serena K. Wolfe, Jamie M. Sperger, Jennifer L. Schehr, Saswati Bhattacharya, Kari B. Wisinski, Joshua M. Lang, Ruth M. O'Regan. Multiplex liquid biopsy for AR pathway activity in metastatic androgen receptor-positive triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 590.

Abstract P6-03-12: Characterization of anti-androgen resistant androgen receptor-positive triple negative breast cancer cells

Abstract Triple negative breast cancer (TNBC) has a poor prognosis in part due to a lack of targeted therapeutics that have improved outcome for other breast cancer subtypes. Gene expression analysis has identified several subtypes of TNBC, including subtypes that express androgen receptor (AR). However, to date the use of AR inhibitors (ARI) in AR+ TNBC has produced disappointing results, likely due to inherent resistance. Additionally, patients that initially benefit from ARIs ultimately develop acquired resistance with resultant progressive disease. In order to better understand the mechanisms of resistance to ARIs in AR+ TNBC, we developed cell lines with acquired resistance to standard ARI therapy. Methods: Bicalutamide (Bica) and enzalutamide (Enza)-resistant cell lines (Bica-R and Enza-R) were generated via continuous passaging of the wild type MDA-MB- 231 (Wt-231) cells in drug-containing media. Cell viability assays were conducted over a 72-hour period using respective anti androgen drugs at varying concentrations. IC-50 for respective drugs was determined via linear regression on normalized control values. Immuno-histochemical analysis and Western blotting for AR and other proteins were compared between the sensitive and resistant lines. Real time PCR was used to detect AR variants (AR-Vs). Additionally we evaluated the efficacy of a CDK inhibitor abemaciclib (Abe) in the resistant cell lines in comparison to the Wt-231 cells. Results: The ARI-resistant cell lines showed higher IC-50 values compared to the Wt cells. For the Bica-R and Enza-R cells the values were at 729 μM and 496μM with Bica and Enza respectively, whereas the values are at 134 μM and 91μM with Bica and Enza respectively for the Wt-231 cells. The resistant lines also exhibited a different morphology, compared to the sensitive cell lines with a more mesenchymal appearance. Expression of AR-Vs differed between the Wt and resistant AR+ TNBC cells. Confocal microscopy revealed different AR localization in the resistant cells along with changes in cell size: compared to Wt cells, Bica-R cells show more diffuse distribution of AR while still maintaining high nuclear levels; Enza-R cells are 3-4 fold size larger than Wt cells and show predominantly nuclear AR localization. Chromogranin A, a protein known for its protective role in regulating endothelial barrier function against increased production of tumor necrosis factor during cancer progression, was found to be down regulated in the resistant cells compared to the sensitive cells. The CDK inhibitor Abe inhibited the growth of the Bica-R cells (IC50 5.74 μM) and was equally effective in inhibiting growth of Wt cells (IC50 5.4 μM). Enza treatment inhibited growth of the Bica-R cells with IC-50 values of 173 μM compared to 91 μM in Wt cells. Interestingly, synergistic effects on cytotoxicity were observed between Abe and Enza in the Bica-R cells, where combination of Enza and 0.1 μM Abe inhibited the growth of the bicalutamide-resistant cells at even lower Enza concentrations (71 μM) than when used as single agents (173 μM). Conclusion: ARI-resistant AR+ TNBC is distinct from AI-sensitive AR+ TNBC at the protein and genomic level. CDK inhibition appears effective in AR+ TNBC, even in cells with acquired-resistance to ARIs and the combination of CDK inhibition and AR inhibition is currently being evaluated in several ongoing clinical trials for patients with AR+ TNBC. Citation Format: Saswati Bhattacharya, Noah Siegel, Ruth O'Regan. Characterization of anti-androgen resistant androgen receptor-positive triple negative breast cancer cells [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-03-12.

34P - Verification of metabolic regulatory mechanisms in androgen receptor-positive triple negative breast cancer

BackgroundAndrogen receptor-positive breast cancer, characterized by overexpression of androgen receptors (ARs), is attracting attention as a subcategory of triple-negative breast cancer (TNBC). The involvement of androgen signaling and the presence of endocrine activity in these TNBC cells make them potential treatment targets. Recent improvements in metabolomics measurement techniques have also revealed the metabolic characteristics of cancer cells. Different cancer cell subtypes exhibit different metabolic balances, and it is believed that a shift to aerobic glycolysis occurs in TNBC, although oxidative phosphorylation has been heavily implicated in endocrine-dependent breast cancer. In this study, we used metabolomics to investigate metabolic regulatory mechanisms in androgen receptor-positive TNBC. MethodsWe introduced ARs into the MDA-MB-231 strain of TNBC cells by using the pEGFP-C1-AR plasmid vector, establishing a stable AR-forced expression TNBC line (MDA-MB-231-AR). Positively ionized metabolites were purified from the parental cell line and the AR forced expression cell line. Metabolome analysis was conducted using a capillary electrophoresis–mass spectrometer (CE-TOFMS/-QqQMS). Data analysis was carried out by extracting, cross-checking, and ordering peaks using MasterHands. ResultsHierarchical clustering analysis of metabolite peak values showed that the MDA-MB-231-AR strain had improved biological malignancy compared to the parental strain. An evaluation of energy metabolism pathways (the glycolysis system, the pentose phosphate pathway, and tricarboxylic acid cycle) showed that the MDA-MB-231-AR strain had a higher NADH/NAD+ ratio and a lower lactic acid/pyruvic acid ratio than the parental strain, suggesting improved hypoxic metabolism. The glutathione/oxidized glutathione ratio was also lower in the MDA-MB-231-AR cell line than in the parental strain, indicating release from oxidative stress. ConclusionsImproved hypoxic metabolism was observed in the energy metabolism pathways of AR-positive TNBC cells. This suggests a possible shift from aerobic glycolysis to oxidative phosphorylation might in TNBC cells. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

Abstract P2-06-04: Enhancing abiraterone acetate efficacy in androgen receptor-positive triple negative breast cancer: Chk1 as a potential target

Abstract Purpose: Our aim was to identify predictive factors of abiraterone acetate (AA) efficacy and putative new druggable targets in androgen receptor (AR)-positive triple-negative breast cancer (TNBC) treated in the UCBG 2012-1 trial. Material and methods: We defined AA response as either complete or partial response, or stable disease at 6 months. Using Ampliseq, we sequenced 91 general and breast cancerassociated genes from the tumor DNA samples. We analyzed transcriptomes from the extracted RNA samples on a Nanostring platform and performed immunohistochemistry (IHC) on tumor samples using tissue microarrays. We assessed AA and CHK1 inhibitors (GDC-0575 and AZD7762) efficacies, either alone or in combination, on cell lines grown in vitro and in vivo. Results: Classical IHC apocrine markers, including AR, FOXA1, GGT1 and GCDFP15, allowed identifying AA responders and non-responders. All responders have clear apocrine features. Transcriptome analysis revealed that 31 genes were differentially expressed in the two subgroups, 9 of them being linked to proliferation and DNA damage repair. One of the most significant differences was the overexpression in non-responders of CHEK1, a gene encoding Chk1, a protein kinase that can be blocked by specific inhibitors. In vitro, AA and Chk1 inhibitor combination showed additive or slightly synergistic effect on cell viability, cell cycle, apoptosis and accumulation of DNA damages. In vivo, orthotopic xenograft experiments confirmed the efficacy of this combination therapy. Conclusions: This study suggests that apocrine features can be helpful in the identification of AA-responders. We identified Chk1 as a putative drug target in AR-positive TNBCs. Citation Format: Grellety T, Callens C, Richard E, Pulido M, Goncalves A, Gestraud P, MacGrogan G, Bonnefoi H, Cardinaud B. Enhancing abiraterone acetate efficacy in androgen receptor-positive triple negative breast cancer: Chk1 as a potential target [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-06-04.

Abstract OT1-02-01: Phase I trial of bicalutamide and ribociclib in androgen receptor-positive triple negative breast cancer

Abstract Background: To date, the use of anti-androgens in the subset of triple negative breast cancers (TNBC) that express androgen receptor (AR) has shown modest response rates, indicating anti-androgen-resistance in the majority of these tumors. Based on data that Cyclin D kinase (CDK) inhibitors reverse resistance to anti-androgens in prostate cancer cell lines, we hypothesize that the use of CDK inhibition may enhance the activity of anti-androgens in AR-positive TNBC. Methods: Key eligibility include: patients with centrally confirmed AR-positive TNBC, defined as AR expression &amp;gt;0%; 0 to 1 line of prior therapy for metastatic disease; and measurable disease. Patients are treated with bicalutamide 150mg orally once daily plus ribociclib at one of 3 dose levels (see table). Table 1Dose levelBicalutamideRibociclib1150mg orally once daily (D1 to 28)400mg daily (D1 to 21)2150mg orally once daily (D1 to 28)400mg daily (D1 to 28)3150mg orally once daily (D1 to 28)600mg daily (D1 to 21) Results: AR expression was positive by trial criteria in 74% of screened patients. Three patients have been accrued at each dose level. Median age is 56 and 6 and 3 patients were treated in first and second-line settings, respectively. Median AR expression was 50% (range 5 to 75%). Toxicity data is available for 6-patients treated on dose levels 1 and 2. No dose-limiting toxicities were noted. As anticipated with ribociclib, the most common toxicity is neutropenia (1 patient grade 4 and 2 patients grade 3). Two patients experienced grade 3 hypertension and 1 experienced grade 3 lymphopenia. Grade 2 or lower toxicities included fatigue, nausea, hyperglycemia and mucositis. One patient experienced grade 1 QT interval prolongation. Conclusion: The combination of bicalutamide and ribociclib is tolerable without unexpected toxicities. Data on the 3-patients treated at dose level 3 and dose expansion will be included. Phase 2 dosing schedule will be decided based on phase 1 results. Citation Format: Sharifi M, Wisinski KB, Burkard ME, Tevaarwerk AJ, Tamkus D, Chan N, Truica C, Danciu O, Hoskins K, O'Regan RM. Phase I trial of bicalutamide and ribociclib in androgen receptor-positive triple negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr OT1-02-01.

Abstract CT058: A phase 2 open-label study to evaluate the efficacy and safety of VT-464 in patients with androgen receptor positive triple-negative breast cancer patients, and men with ER positive breast cancer

Abstract Background: The androgen receptor (AR) is expressed in 20-30% of patients with triple negative breast cancer (TNBC). In phase 2 trials androgen blockade with bicalutamide demonstrated a clinical benefit rate (CBR = Partial Response + Complete Response + Stable Disease) of 19% at 24 weeks and enzalutamide showed a CBR of 35% at 16 weeks. VT-464 is an oral non-steroidal, small molecule that is a potent anti-androgen through both inhibition of CYP17 lyase and through direct inhibition of AR. Studies in breast cancer cell lines have shown that VT-464 inhibits growth in a soft agar assay of MCF7 (ER positive/low AR expression), tamoxifen-resistant MCF7, and MDA-MB-453 (ER negative/AR positive) cells in a dose dependent manner and with higher potency than enzalutamide. Patients with AR+ TNBC may benefit from treatment with VT-464. In addition, since VT-464 will deplete androgen available for aromatization, men with ER-positive (ER+) breast cancer may benefit from use of VT-464. Methods: This is a Phase 2, open label study evaluating the potential benefit of VT-464 in female patients with TNBC with AR 1 to 9% (cohort 1) and AR ≥ 10% (cohort 2), and men with ER+ breast cancer (cohort 3) (NCT02130700; NIH 14-c-0090). VT-464 will be administered once daily in continuous 28-days cycles. Due to differences in drug metabolism, women with TNBC will receive VT-464 450mg by mouth daily while men with ER+ breast cancer will receive VT-464 600mg by mouth daily. In the two female TNBC cohorts the primary endpoint is CBR at 16 weeks. In the ER+ male breast cancer cohort the primary endpoint is progression free survival at 24 weeks. Secondary objectives include evaluation of response rate and the safety profile of the drug in both populations. Women with TNBC with &amp;gt; 1% AR expression per IHC are eligible. Males with ER+ breast cancer, who have failed at least one prior endocrine therapy and are undergoing gonadal suppression using LHRH agonists or antagonists are eligible for the study. All patients must be at least 18 years old and have a ECOG of 0 to 1. Consenting patients must have biopsiable disease which will be used for genomic and transcriptomic evaluation before and after treatment with VT-464. Correlative studies including serum hormone levels, circulating tumor cells, and immune subset will be analyzed. Patients must also have adequate hematopoietic, hepatic and renal function. Exclusions include symptomatic CNS metastases, radiotherapy within 28 days of study entry and active HIV, hepatitis B or hepatitis C infection. This trial opened in December 2016. While no patients have been enrolled at this time, multiple patients are currently being screened. Citation Format: Alexandra Zimmer, Margaret E. Gatti-Mays, Stan Lipkowitz, Fatima Karzai, James Gulley, William Dahut, Seth Steinberg, William D. Figg, Ravi Madan. A phase 2 open-label study to evaluate the efficacy and safety of VT-464 in patients with androgen receptor positive triple-negative breast cancer patients, and men with ER positive breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr CT058. doi:10.1158/1538-7445.AM2017-CT058

Abstract OT2-01-07: A phase 2 open label, multi-center, multinational study investigating the efficacy and safety of GTx-024 on advanced, androgen receptor-positive triple negative breast cancer (AR+ TNBC)

Abstract Background: The androgen receptor (AR) is the most highly expressed steroid receptor in breast cancer (BC) with expression seen in up to 95% of estrogen receptor positive (ER+) and up to 50% of ER negative disease. Historically, steroidal androgens exhibited virilizing side effects, thus limiting clinical use. In TNBC, the expression of AR and androgen synthesizing enzymes is associated with lower proliferation, lower tumor grade, better overall survival, and more favorable clinical outcomes as compared to those patients with TNBC not expressing AR. Data from two trials targeting AR in TNBC indicates low level but encouraging clinical activity. A non-steroidal, tissue-selective, AR modulator (SARM), such as GTx-024, offers a targeted approach of AR activation in AR+ TNBC without virilization or estrogenic effects. Trial Design: Open label, multicenter, multinational, Phase 2 study for the treatment of advanced or metastatic TNBC. Subjects will receive GTx-024, 18 mg orally (PO) daily, continued until evidence of disease progression or toxicity. Subjects whose tumors demonstrate Clinical Benefit (CB) will be treated on study drug until progression. Eligibility Criteria: Inclusion: Female, ≥18 years old, confirmed AR+ (≥10% staining) TNBC (confirmed by medical history), up to 1-2 prior chemotherapy regimens, available archived tumor tissue, measurable or bone-only disease, ECOG 0 or 1, with prior toxicities from chemotherapy resolved. Exclusion: Life expectancy &amp;lt;4 months, uncontrolled CNS metastases, radiotherapy ≤14 days prior to enrollment, active hepatitis, HIV positive, prior treatment with anti-androgens, testosterone or testosterone-like agents, or estrogens or megesterol acetate, or prior treatment for a different cancer (other than BC or non-melanoma carcinoma of the skin) within the past 2 year Specific Aims: The primary aim is to measure the proportion of AR+ subjects with CB at 16 weeks; defined as subjects with a complete response (CR), partial response (PR), or stable disease (SD); per modified RECIST 1.1. Secondary endpoints include: objective response rate, progression free survival, and time to progression. Statistical Methods: Simon's two-stage (optimal) design will be used to assess primary efficacy, requiring up to 41 evaluable subjects; i.e., subjects with centrally confirmed AR+ who receive at least one dose of study drug. The first stage will be assessed among the first 21 evaluable subjects. If at least 2/21 achieve CB per modified RECIST 1.1 at 16 weeks, then the trial will proceed to the second stage of recruitment of up to a total of 41 subjects in the evaluable subset of the Full Analysis Set. Otherwise, the trial will be discontinued for lack of efficacy. The trial will test for an unacceptably low CBR of ≤5% versus a CBR more consistent with ≥20%. Target Accrual: Up to 55 subjects will be enrolled. Trial Information: www.gtxinc.com Citation Format: Rugo H, Overmoyer B, Schwartzberg L, Palmieri C, Taylor R, Hancock M, Small S, Johnston MA. A phase 2 open label, multi-center, multinational study investigating the efficacy and safety of GTx-024 on advanced, androgen receptor-positive triple negative breast cancer (AR+ TNBC). [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr OT2-01-07.

Abstract P3-04-03: PIK3CA mutations in androgen receptor-positive triple negative breast cancer confer sensitivity to the combination of PI3K and androgen receptor inhibitors

Abstract Triple negative breast cancer (TNBC) is a heterogeneous collection of biologically diverse cancers, which contributes to variable clinical outcomes. Previously, we identified a TNBC subtype that has a luminal phenotype and expresses androgen receptor (AR+). TNBC cells derived from these luminal AR+ tumors have high frequency PIK3CA mutations. The purpose of this study was to determine if targeting PI3K alone or in combination with an AR antagonist is effective in AR+ TNBC. Methods: We determined the frequency of activating PIK3CA mutations in AR+ and AR- TNBC clinical cases. Using AR+ TNBC cell lines and xenograft models we evaluated the effectiveness of PI3K inhibitors, used alone or in combination with an AR antagonist, on tumor cell growth and viability. Results: PIK3CA kinase mutations were highly clonal, more frequent in AR+ vs. AR- TNBC (40% vs. 4%), and often associated with concurrent amplification of the PIK3CA locus. PI3K/mTOR inhibitors had an additive growth inhibitory effect when combined with genetic or pharmacological AR targeting in AR+ TNBC cells. We also analyzed the combination of bicalutamide +/- the pan-PI3K inhibitor GDC-0941 or the dual PI3K/mTOR inhibitor GDC-0980 in xenograft tumor studies and observed additive effects. Conclusions: While approximately one third of TNBC patients respond to neoadjuvant/adjuvant chemotherapy, recent studies have shown that patients with androgen receptor positive (AR+) TNBC are far less likely to benefit from the current standard of care chemotherapy regimens and novel targeted approaches need to be investigated. In this study, we show that activating PIK3CA mutations are enriched in AR+ TNBC; and, the growth and viability of AR+ TNBC cell line models is significantly reduced after treatment with PI3K inhibitors used in combination with an AR antagonist. These results provide rationale for pre-selection of TNBC patients with a biomarker (AR expression) to investigate the use of AR antagonists in combination with PI3K/mTOR inhibitors. Citation Format: Brian D Lehmann, Joshua A Bauer, Johanna M Schafer, Christopher S Pendleton, Luojia Tang, Kimberly C Johnson, Xi Chen, Justin M Balko, Henry Gomez, Carlos L Arteaga, Gordon B Mills, Melinda E Sanders, Jennifer A Pietenpol. PIK3CA mutations in androgen receptor-positive triple negative breast cancer confer sensitivity to the combination of PI3K and androgen receptor inhibitors [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-04-03.

A Case of Androgen Receptor-positive Triple Negative Breast Cancer with Good Response to Anti-androgen Therapy

A Case of Androgen Receptor-positive Triple Negative Breast Cancer with Good Response to Anti-androgen Therapy

PIK3CA mutations in androgen receptor-positive triple negative breast cancer confer sensitivity to the combination of PI3K and androgen receptor inhibitors.

IntroductionTriple negative breast cancer (TNBC) is a heterogeneous collection of biologically diverse cancers, which contributes to variable clinical outcomes. Previously, we identified a TNBC subtype that has a luminal phenotype and expresses the androgen receptor (AR+). TNBC cells derived from these luminal AR + tumors have high frequency phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) mutations. The purpose of this study was to determine if targeting phosphoinositide 3-kinase (PI3K) alone or in combination with an AR antagonist is effective in AR + TNBC.MethodsWe determined the frequency of activating PIK3CA mutations in AR + and AR- TNBC clinical cases. Using AR + TNBC cell line and xenograft models we evaluated the effectiveness of PI3K inhibitors, used alone or in combination with an AR antagonist, on tumor cell growth and viability.ResultsPIK3CA kinase mutations were highly clonal, more frequent in AR + vs. AR- TNBC (40% vs. 4%), and often associated with concurrent amplification of the PIK3CA locus. PI3K/mTOR inhibitors had an additive growth inhibitory effect when combined with genetic or pharmacological AR targeting in AR + TNBC cells. We also analyzed the combination of bicalutamide +/- the pan-PI3K inhibitor GDC-0941 or the dual PI3K/mTOR inhibitor GDC-0980 in xenograft tumor studies and observed additive effects.ConclusionsWhile approximately one third of TNBC patients respond to neoadjuvant/adjuvant chemotherapy, recent studies have shown that patients with AR + TNBC are far less likely to benefit from the current standard of care chemotherapy regimens and novel targeted approaches need to be investigated. In this study, we show that activating PIK3CA mutations are enriched in AR + TNBC; and, we show that the growth and viability of AR + TNBC cell line models is significantly reduced after treatment with PI3K inhibitors used in combination with an AR antagonist. These results provide rationale for pre-selection of TNBC patients with a biomarker (AR expression) to investigate the use of AR antagonists in combination with PI3K/mTOR inhibitors.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-014-0406-x) contains supplementary material, which is available to authorized users.