Progesterone Receptor Expression Research Articles

An ovine model for investigation of the microenvironment of the male mammary gland.

The specific biology of the male breast remains relatively unexplored in spite of the increasing global prevalence of male breast cancer. Delineation of the microenvironment of the male breast is restricted by the low availability of human samples and a lack of characterisation of appropriate animal models. Unlike the mouse, the male ovine gland persists postnatally. We suggest that the male ovine mammary gland constitutes a promising adjunctive model for the male breast. In this study, we evaluate the male ovine mammary gland microenvironment, comparing intact and neutered males. Assessment of the glandular histo-anatomy highlights the resemblance of the male gland to that of neonatal female sheep and confirms the presence of rudimentary terminal duct lobular units. Irrespective of neutered status, cell proliferation in epithelial and stromal compartments is similarly low in males, and cell proliferation in epithelial cells and in the intralobular stroma is significantly lower than in pubertal female sheep. Between 42% and 72% of the luminal mammary epithelial cells in the male gland express the androgen receptor and expression is significantly reduced by neutering. Luminal epithelial cells within the intact and neutered male gland also express oestrogen receptor alpha, but minimal progesterone receptor expression is observed. The distribution of leukocytes within the ducts and stroma is similar to the mammary gland of female sheep and females of other species. Both macrophages and T lymphocytes are intercalated in the epithelial bilayer and are more abundant in the intralobular stroma than the interlobular stroma, suggesting that they may have a protective immunological function within the vestigial glandular tissue of the male sheep. Mast cells are also observed within the stroma. These cells cluster near the glandular tissue and are frequently located adjacent to blood vessels. The abundance of mast cells is significantly higher in intact males compared to neutered males, suggesting that hormone signalling may impact mast cell recruitment. In this study, we demonstrate the utility of the male ovine mammary gland as a model for furthering our knowledge of postnatal male mammary biology.

Metaplastic breast cancer with a unique presentation and complete response to chemotherapy: a case report

BackgroundMetaplastic breast carcinomas are a rare variant group of breast carcinomas. They are usually high-grade and triple-negative tumors. They often present with large primary tumor sizes. However, the involvement of axillary lymph nodes is infrequent at the time of diagnosis. Metaplastic breast carcinomas are associated with a worse prognosis and a poorer response to chemotherapy in comparison with other non-metaplastic triple-negative breast cancers. Up until this point, there are no specific treatment recommendations for metaplastic breast carcinomas beyond those intended for invasive breast cancer in general.Case presentationA 40-year-old woman complained of a palpable mass in her left axilla. On ultrasonography, the mass was solid, spindle-shaped, hypoechoic with regular borders, and exhibited decreased vascularity. At first, the mass appeared to be of a muscular origin. There was not any clinical nor ultrasonic evidence of a primary breast tumor. On magnetic resonance imaging, the axillary mass was a well-defined with regular borders, measuring 24 × 35 mm. Needle biopsy showed a spindle cell tumor with mild to moderate atypia. The subsequent surgical resection revealed a spindle cell neoplasm within a lymph node, favoring a metastatic origin of the tumor. The tumor cells lacked expression of estrogen, progesterone, and HER2 receptors. PET-CT scan indicated pathological uptake in the left breast. Accordingly, the patient was diagnosed with metaplastic breast cancer that had metastasized to the axillary lymph node. She commenced a combined chemotherapy regimen of doxorubicin and cyclophosphamide. After six treatment cycles, she underwent left modified radical mastectomy with axillary lymph node dissection. Pathological examination of the specimens revealed a total burn-out tumor in the breast due to excellent treatment response. There were no residual tumor cells. All dissected lymph nodes were free of tumor. At the one-year follow-up, the patient showed no signs of tumor recurrence.ConclusionThis report sheds light on a distinctive presentation of metaplastic breast carcinoma, emphasizing the need for vigilance in diagnosing this rare and aggressive breast cancer variant. In addition, the patient’s remarkable response to chemotherapy highlights potential treatment avenues for metaplastic breast cancer.

MerTK Drives Proliferation and Metastatic Potential in Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is characterized by the absence of the estrogen receptor, progesterone receptor, and receptor tyrosine kinase HER2 expression. Due to the limited number of FDA-approved targeted therapies for TNBC, there is an ongoing need to understand the molecular underpinnings of TNBC for the development of novel combinatorial treatment strategies. This study evaluated the role of the MerTK receptor tyrosine kinase on proliferation and invasion/metastatic potential in TNBC. Immunohistochemical analysis demonstrated MerTK expression in 58% of patient-derived TNBC xenografts. The stable overexpression of MerTK in human TNBC cell lines induced an increase in proliferation rates, robust in vivo tumor growth, heightened migration/invasion potential, and enhanced lung metastases. NanoString nCounter analysis of MerTK-overexpressing SUM102 cells (SUM102-MerTK) revealed upregulation of several signaling pathways, which ultimately drive cell cycle progression, reduce apoptosis, and enhance cell survival. Proteomic profiling indicated increased endoglin (ENG) production in SUM102-MerTK clones, suggesting that MerTK creates a conducive environment for increased proliferative and metastatic activity via elevated ENG expression. To determine ENG’s role in increasing proliferation and/or metastatic potential, we knocked out ENG in a SUM102-MerTK clone with CRISPR technology. Although this ENG knockout clone exhibited similar in vivo growth to the parental SUM102-MerTK clone, lung metastasis numbers were significantly decreased ~4-fold, indicating that MerTK enhances invasion and metastasis through ENG. Our data suggest that MerTK regulates a unique proliferative signature in TNBC, promoting robust tumor growth and increased metastatic potential through ENG upregulation. Targeting MerTK and ENG simultaneously may provide a novel therapeutic approach for TNBC patients.

Abstract PO4-16-02: Can high progesterone receptor (PgR) expression identify tumours with low-risk tumour gene expression scores?

Abstract Background Strong PgR expression predicts favorable outcomes for ER+ve HER2-ve breast cancer and has been proposed as a surrogate marker to distinguish between IHC-defined luminal A and luminal B subtypes. It is therefore possible that strong PgR expression may be able to predict tumor gene expression test results and independently identify tumors that are unlikely to be chemotherapy sensitive. PgR expression is traditionally determined by immunohistochemistry (IHC). Several validated RNA-based PGR expression tests have been developed that may outperform IHC. Methods We compared Oncotype DX RS with Oncotype DX reported PGR in 4 independent datasets which included 407 cases from the OPTIMA prelim trial. We further analyzed 251 OPTIMA prelim cases which had additional tumor gene expression data. All gene expression assays were performed by the test vendor, including PGR gene expression determined using the Mammatyper assay. PgR IHC was determined in a single laboratory on triplicate tissue micro-arrays using quantitative image analysis including a 10% manual quality control check. We analyzed PGR expression using cutoffs that correspond to approximately 20% staining; the standard Oncotype DX PGR assay is reported as positive if the score is >5.4, corresponding to approximately 1% staining by IHC. We used Spearman’s rank correlation coefficient to compare PGR data. Results The four Oncotype DX datasets consistently demonstrated that high Oncotype PGR expression was associated with a low RS (table). Combining the 3 validation data sets consisting of 633 cases, 70.9% had high PGR expression of which 92.7% had a an Oncotype RS ≤25. Approximately 50% of cases with low Oncotype PGR expression had an Oncotype RS >25. Mammatyper and Oncotype PGR were highly correlated (Rs = 0.9258, P< 0.001) in the OPTIMA prelim dataset (n=251), with only 8.4% of tumors having discordant high/low Mammatyper and Oncotype PGR scores. 93.2% of 176 Mammatyper high PGR expression cases had an RS ≤25. The Mammatyper PGR and PgR IHC correlation was weaker (Rs=0.763, P< 0.001); 87.2% of 211 cases with >20% staining had RS ≤25. PgR IHC staining had a bimodal distribution and there was little effect on the prediction of low RS score up to a 67% cut-off. Mammatyper and Oncotype PGR scores both appear to have a normal distribution. We took advantage of this to perform an exploratory analysis using a higher Mammatyper PGR cutoff. We were able to show superior prediction of a low RS (96.8%) but with a reduced proportion (50.2%) of high PGR score tumors. High PGR gene expression was weakly associated with low (≤60) Prosigna ROR_PT score and MammaPrint low risk (72.2% and 65.9% respectively) and with Prosigna and MammaPrint luminal A subtype (both 64.8%). Conclusion High progesterone receptor gene expression measured using locally performed RNA-based assays may allow the reliable prediction of Oncotype DX low-risk tumours. This analysis provides additional information for the clinical utility of PGR measurement. Additional data will be presented on the optimal PGR cutoff. OPTIMA prelim is registered as ISRCTN42400492 and funded by the UK NIHR Health Technology Assessment Programme, award number 10/34/01. Views expressed are those of the authors and not those of the HTA Programme, NIHR, NHS or the Department of Health. Table. Oncotype DX RS and PGR in 4 datasets Distribution of RS according to %cases with high or low PGR Citation Format: Robert Stein, Ralph Wirtz, Andrea Marshall, Jane Bayani, Sebastian Eidt, Claudia Schumacher, Hans-Peter Sinn, Andreas Schneeweiss, Andreas Makris, Iain Macpherson, Luke Hughes-Davies, Tammy Piper, Monika Sobol, Georgina Dotchin, Helen Higgins, Sarah Pinder, Abeer Shaaban, Janet Dunn, John MS Bartlett. Can high progesterone receptor (PgR) expression identify tumours with low-risk tumour gene expression scores? [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-16-02.

Abstract RF01-01: SERENA-3: A randomized pre-surgical window of opportunity study assessing dose and duration of camizestrant treatment in post-menopausal women with ER-positive, HER2-negative primary breast cancer

Abstract Background Camizestrant, a next-generation oral selective estrogen receptor (ER) degrader (SERD) and pure ER antagonist, has demonstrated statistically significant and clinically meaningful progression-free survival (PFS) benefit over fulvestrant at both 75 and 150 mg once-daily doses in the Phase 2 SERENA-2 (NCT04214288) study in post-menopausal women with ER+ HER2- advanced breast cancer. SERENA-3 (NCT04588298) was established to explore the biological effects of three camizestrant dose levels in post-menopausal women with ER+ HER2- primary breast cancer. Methods Post-menopausal women with ER+ HER2- primary breast cancer scheduled to undergo curative intent surgery were randomly assigned in Stages 1 and 2 to pre-surgical treatment with camizestrant at 75, 150 or 300 mg once daily for 5 to 7 days or, in a sequential Stage 3, randomly assigned to camizestrant at 75 or 150 mg once daily for 12 to 15 days. Image-guided tumor biopsies were accessed as pre-treatment samples, while on-treatment ultrasound-guided tumor biopsies were obtained on the last day of camizestrant treatment. The primary objective was to explore the effect of camizestrant on ER expression by comparison of pre-treatment and on-treatment biopsies as assessed by immunohistochemistry (IHC) H-score. Secondary objectives included assessment of Ki67 and progesterone receptor (PgR) expression, and also of safety and tolerability, with exploratory pharmacodynamic (PD) assessment using other transcriptomic and proteomic technologies. The study was undertaken in 17 centers across 3 countries (Georgia, Mexico and the United Kingdom). Results 135 patients were randomized and received treatment. 76 patients received camizestrant for 5 to 7 days at doses of 75 mg (n=30), 150 mg (n=33), and 300 mg (n=13); 59 received camizestrant for 12 to 15 days at doses of 75 mg (n=30) and 150 mg (n=29). Baseline disease characteristics, and ER, Ki67 and PgR levels were well balanced across arms and stages. Pharmacokinetic (PK) observations in all stages were commensurate with steady-state predictions derived from previous studies. PD data for ER and Ki67 are shown in Table 1. Camizestrant reduced ER H-score by approximately 65% irrespective of dose or duration. While the 75 mg dose reduced Ki67 score to a lesser degree than 150 mg and 300mg after 5–7 days exposure, after 12–15 days exposure 75 and 150 mg reduced Ki76 score to a similar substantial degree (82% reduction). PD results assessed by additional transcriptomics and proteomics assays were concordant with the IHC analyses. No serious adverse events were reported. All treatment-emergent adverse events were grade 1, apart from one patient with a grade 2 visual impairment, and one patient with grade 3 diarrhea. Conclusions SERENA-3 is the first study investigating the effect of multiple dose levels of camizestrant with different treatment durations on ER, Ki67 and PgR expression in post-menopausal women with primary ER+ HER2˗ breast cancer. SERENA-3 demonstrated that the 75 mg dose of camizestrant achieves maximal levels of ER degradation and Ki67 suppression. Together with SERENA-2, this provides strong evidence supporting 75 mg as the optimal dose of camizestrant, including for the early disease setting, and highlights the additive value of a specifically designed multi-dose pre-surgical PD study for optimal dose selection. Table 1 Citation Format: John Robertson, Teimuraz Gogitidze, Zaza Katashvili, Juan Enrique Bargallo Rocha, Ekaterine Arkania, Iain Moppett, Kwok-Leung Cheung, Gia Nemsadze, Maxine Ajimi, Itziar Irurzun-Arana, Justin Lindemann, Teresa Klinowska, Alastair Mathewson, Christopher Morrow, Myria Nikolaou, Giorgi Dzagnidze. SERENA-3: A randomized pre-surgical window of opportunity study assessing dose and duration of camizestrant treatment in post-menopausal women with ER-positive, HER2-negative primary breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr RF01-01.

Abstract PO1-27-11: Survival Benefit of Adjuvant Endocrine Therapy in Patients with Estrogen Receptor Low-Positive Breast Cancer

Abstract Background: Estrogen receptor (ER) low-positive (1–10%) breast cancer is a very rare subtype that accounts for only 2–5% of all breast cancer cases and was introduced as a new category in the 2020 American Society of Clinical Oncology/College of American Pathologist Guidelines. There are limited data on the overall benefit of endocrine therapy for patients with ER low-positive cancers. The objective of this study was to analyze the clinicopathological characteristics and treatment patterns in patients with ER low-positive breast cancer and to compare disease-free survival (DFS) between patients who received and did not receive endocrine therapy. Methods: A retrospective search of the prospectively maintained database of our institution identified consecutive women with ER low-positive (1–10%) primary invasive breast cancer who underwent curative surgery at Seoul National University Hospital (Seoul, Korea) between January 2010 and April 2021. Standard immunohistochemical staining techniques were used to assess the level of ER, progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression, and semiquantitative scores (0–100%) were reported for ER and PR. DFS was defined as the time from surgery to the development of any recurrence and was compared between women who received and did not receive adjuvant endocrine therapy using univariable and multivariable Cox proportional hazards regression. Results: A total of 721 women (median age, 51 years; range, 24–83 years) with ER low-positive cancer were included in the analysis. Overall, 28% (203/721) had AJCC stage I, 42% (304/721) had stage II, and 30% (214/721) had stage III. PR expression was negative (< 1%) in 79% (571/721), low-positive (1–10%) in 12% (90/721), and high-positive ( >10%) in 8% (60/721). HER2 expression was negative in 54% (390/721) and positive in 46% (331/721). Most women (88% [634/721]) received adjuvant endocrine therapy, including all women with PR expression >10%. Of the 661 women with PR expression ≤10%, 87 (13%) did not receive adjuvant endocrine therapy. HER2-targeted therapy was performed in 81% (268/331) of women with HER2-positive cancer. During a median follow-up of 5.7 years, 98 (14%) had recurrence (25 locoregional/contralateral and 73 distant metastases). AJCC stage III (HR, 3.6; 95% CI: 2.4, 5.4; P < .001) and receiving total mastectomy (HR, 1.8; 95% CI: 1.2, 2.8; P = .004) were associated with worse DFS. Receiving adjuvant endocrine therapy was independently associated with better DFS in the entire cohort (HR, 0.46; 95% CI: 0.28, 0.76; P = .002) and in the HER2-negative subgroup (HR, 0.40; 95% CI: 0.22, 0.75; P = .004). In the HER2-positive subgroup, receiving adjuvant endocrine therapy was not significantly associated with better DFS (HR, 0.55; 95% CI: 0.23, 1.32; P = .18). Conclusion: In this large cohort of women with ER low-positive breast cancers, adjuvant endocrine therapy was administered in the majority of cases, including those with PR >10% expression. Receiving adjuvant endocrine therapy was associated with better DFS, especially when HER2 expression was negative. Citation Format: Su Hyun Lee, Han Suk Ryu, Myoung-jin Jang, Kyung-Hun Lee, Han-Byoel Lee. Survival Benefit of Adjuvant Endocrine Therapy in Patients with Estrogen Receptor Low-Positive Breast Cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-27-11.

Abstract PO3-01-12: Clinicopathologic Factors affecting response in patients with Estrogen receptor-positive and Human Epidermal Growth Factor Receptor-negative breast cancer receiving neoadjuvant chemotherapy

Abstract Background: The evolution of Neoadjuvant chemotherapy(NAC) regimen has enabled the down-staging of locally advanced breast cancer, allowing patients to undergo breast conserving surgery. NAC is known to be effective in triple-negative breast cancer (TNBC) and HER2-positive breast cancer. However, the role of NAC in hormone receptor-positive and HER2-negative (HR+HER2-) breast cancer patients remains unclear. In this study, we aimed to explore the utility of NAC for estrogen receptor-positive and HER2-negative (ER+HER2-) breast cancer patients. Methods: Clinical data were collected from 120 patients who were diagnosed with estrogen receptor (ER)-positive and HER2-negative breast cancer and received neoadjuvant chemotherapy between July, 2017 and April, 2023 at Seoul St Mary’s Hospital (Seoul, Korea). We analyzed the factors that affect clinical response subgroups, including pathologic complete response (pCR), partial response (PR), stable disease(SD), and progressive disease (PD), following neoadjuvant chemotherapy. The median follow-up period was 17 months. < Results: Among 120 patients after NAC, 5.8% achieved pCR, while 67.5% had a partial response. The median age at the time of surgery was 55 years at pCR group, 49 years at PR group, 47 years at SD group, and 49 years at PD group. Although the age of patients achieving pathological complete response was higher, this differences was not statistically significant. Patients achieving pCR had a higher incidence of progesterone receptor (PR) negativity (pCR 71.4% vs PR 30.9%, p=0.011) and higher levels of Ki-67 expression than those who did not achieve pCR (71.7% vs 35.6%, p=0.003). Also, there was no significant difference in Androgen receptor (AR) and HER-2 expression between the groups. Tumor size was also significantly associated with clinical response (p=0.003). However, no differences in clinical response was observed between patients with lymph node metastasis (p=0.221). Our analysis of 107 patients who were clinically N positive before NAC showed that the Ki-67 index level was the only factor found to affect nodal pCR. In addition, in the non-pCR group, patients with a Ki-67 index of less than 20% after neoadjuvant had a significantly better disease-free survival (5yr DFS, 79.8% vs 65.9%, p=0.039) Conclusion: Our analysis revealed that patients with higher levels of Ki-67 and lower level of PR expression were more likely to achieve pCR or PR. Also, decrease in Ki-67 index was confirmed to be an important prognostic factor of ER+ HER2- breast cancer. A combination of clinical and molecular factors, such as tumor size, progesterone receptor expression, and Ki-67 expression, can be used to optimize treatment plans and improve outcomes for the patient population. Citation Format: Jin Ah Lee, Young Joo Lee, Dooreh Kim, Chang Ik Yoon, Woo-Chan Park, Soo Youn Bae. Clinicopathologic Factors affecting response in patients with Estrogen receptor-positive and Human Epidermal Growth Factor Receptor-negative breast cancer receiving neoadjuvant chemotherapy [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-01-12.

Abstract PO4-15-09: Unraveling the influence of progesterone receptor status on endocrine therapy sensitivity: insight from epigenetic and fragmentomics ctDNA profiling in estrogen receptor positive, HER2 negative metastatic breast cancer (MBC)

Abstract Background: In patients (pts) with estrogen receptor (ER) positive (pos), HER2 negative (neg) MBC, the absence of progesterone receptor (PR) expression is generally associated with a poorer endocrine therapy (ET) response and an unfavorable prognosis. However, real-time molecular changes during ET according to PR status are still not fully understood. The aim of this study was to examine the prognostic impact of PR and its association with the methylation levels of ESR1 promoters A (promA) and B (promB) at different timepoints in pts with ERpos HER2neg MBC who received first-line ET. Methods: Pts were enrolled in the prospective multicenter MAGNETIC.1 trial (CRO-2018-56) between January 2018 and January 2023. As first-line ET, pts received either fulvestrant or AIs with or without CDK4/6 inhibitors (CDK4/6i) as first-line ET. ctDNA samples were collected at baseline (T0) and at 3-month intervals (T3 and T6) and analyzed through methylation-specific (MS) droplet digital PCR (ddPCR), ddPCR fragmentomics and next generation sequencing (NGS). Clinico-pathological differences across the PR expression spectrum were analyzed using Fisher’s exact test. Matched pairs variations across timepoints (T0, T3, T6) of ACTBfragments distribution and ESR1 promA and promB were tested through Wilcoxon signed rank test. Results: Out of the 111 enrolled pts, 28 had PRneg MBC, of which 20 had an invasive ductal carcinoma (IDC) histotype, and 10 were diagnosed as de novo disease. Bone was the most common metastatic site both in the PRpos and PRneg subgroups (75% vs 73%), most pts received ET+CDK4/6i (96% vs 95%). ESR1 and PIK3CA mutations were respectively detected in 8% and 6%, and in 23% and 25% pts respectively for PRpos and PRneg. The prognostic impact of PR status was evident for both PFS (median 38 mos and 14 mos respectively for PRpos and PRneg, P = 0.0048) and OS (median not reached and median 33 mos respectively for PRpos and PRneg, P= 0.001). A significant increase in promB was observed at T3 vs T0 and at T6 vs T3 in the PRpos subgroup (respectively P=0.0158 and P< 0.001) but not in the PRneg. When comparing T6 to T0, no significant differences were observed regardless of PR status. At T3, significant decreases were observed in both the PRpos and PRneg populations for ACTBshort (P < 0.001 and P=0.03, respectively), ACTBmedium (P=0.0049 and P= 0.035, respectively) and ACTBlong(P< 0.001 and P< 0.001, respectively), compared to T0. In the comparison of T6 to T3, a significant variation in ACTBshort levels was observed in both subgroups (P< 0.001 for PRpos and P=0.012 for PRneg). while a significant reduction in ACTBmedium levels was observed only in the PRpos population when comparing T6 to T0 (P< 0.001). Conclusions: PRneg status was confirmed as an unfavorable prognostic factor with respect to PRpos both for PFS and OS, potentially due to different ET sensitivity. Epigenetic ctDNA profiling revealed different dynamics of ESR1 promB between PR-positive and PR-negative MBC, suggesting a distinct role of ESR1 in the onset of ET resistance within the two subgroups. Citation Format: Lorenzo Foffano, Elisabetta Molteni, Arianna Dri, Alessandra Franzoni, Linda Cucciniello, Lucia Da Ros, Silvia Buriolla, Silvia Bolzonello, Claudia Noto, Stefania Russo, Simon Spazzapan, Elena Nascimbeni, Brenno Pastò, Giada Targato, Serena Della Rossa, Marta Bonotto, Alessandro Marco Minisini, Giuseppe Damante, Barbara Belletti, Lorenzo Gerratana, Fabio Puglisi. Unraveling the influence of progesterone receptor status on endocrine therapy sensitivity: insight from epigenetic and fragmentomics ctDNA profiling in estrogen receptor positive, HER2 negative metastatic breast cancer (MBC) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-15-09.

Abstract PO3-05-02: Impact of hormone receptor expression level in HR+/HER2- metastatic breast cancer in the large French ESME cohort

Abstract Background: In patients (pts) with early breast cancer (BC), the level of expression of estrogen receptor (ER) evaluated by immunohistochemistry is prognostic and predictive while progesterone receptor (PR) expression has an additional prognostic value. Allred scoring combines the percentage of positive cells and the intensity of staining. It is widely used for treatment choice for early BC. In metastatic setting, its role is less defined. Methods: The ESME database is a French National cohort of all consecutive pts who initiated a first-line treatment for metastatic BC (MBC) between January 2008 and December 2020 in one of the 18 French Comprehensive Cancer Centers (NCT03275311). For this study, we selected pts with HER2- MBC, known ER expression and intensity percentages, known PR status and who received endocrine-based therapy as first-line metastatic treatment. Primary objective was to evaluate the overall survival (OS) and first line progression-free survival (PFS1) under endocrine therapy according to ER and PR expression level. ER expression was defined using Allred score on the last available tumor sample. PR status was defined using a positive (≥10%)vs negative ( < 10%) threshold. Secondary objective was to evaluate clinical PFS1 whether de novo or recurrent MBC and according to type of first line therapy (mono- vs combined endocrine therapy). Results: In the ER+ ± PR+/HER2- population, 8269 pts had information on ER percentage and staining intensity. Median follow-up was 53.4 months (95%CI [51.9; 54.7]). Allred score distribution, OS and PFS1 are described in table 1. Median OS was 43.1 months [41.1; 45.0] for the PR- group compared with 52.3 months [50.5; 54.4] for the PR+ group. After adjustments, patients with a PR- tumour had still a worse prognosis (HR=1.20, 95%CI [1.12-1.28], p<0.001). Median PFS1 was 10.2 months[9.6; 10.8]and 14.8 months[14.2; 15.4] for pts with PR- vs PR+ tumour (HR=1.36 (1.29-1.43), p<0.001) and PR status remained significant prognostic factor independently of de novo or recurrent status and association or not with a targeted therapy. Conclusion: ER expression evaluated using the Allred scoring, as well as PR positivity, retain prognostic value in first-line MBC patients. Table 1 : OS and PFS according to Allred status, de novo status and type of endocrine therapy. * Selected variables on multivariate model include PR status, age, SBR grade, histological subtype, number of metastatic site, type of metastasis (non visceral, visceral, brain visceral), HER 2 (0 vs 1-2), metastatic-free interval, mono- vs combined endocrine therapy. Citation Format: Marie Alexandre, Florence Castan, Thibault De La Motte Rouge, Audrey Mailliez, Etienne Brain, Thomas Bachelot, Monica Arnedos, Florence Dalenc, Jean-Marc Ferrero, Vincent Massard, Isabelle Desmoulins, Marie-Ange Mouret-Reynier, Christelle Levy, Anthony Gonçalves, Anca Berghian, Aude-marie Savoye, Jean-Sébastien frenel, Lise Bosquet, Suzette Delaloge, William Jacot. Impact of hormone receptor expression level in HR+/HER2- metastatic breast cancer in the large French ESME cohort [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-05-02.

Abstract PO1-01-13: Clinical characterization, prognostic and predictive values of HER2-low in early breast cancer in the PALLAS trial

Abstract Background: Low levels of HER2 expression (HER2-low) have recently emerged as a therapeutic target in patients (pts) with breast cancer (BC). The aim of this study was to evaluate clinical and demographic variables associated with HER2 expression, as well as the value of HER2-low as a prognostic and predictive biomarker for palbociclib benefit in pts enrolled in the PALLAS trial. Methods: The phase III PALLAS trial investigated the addition of palbociclib to adjuvant endocrine therapy (ET) in pts with stage II-III hormone receptor-positive BC. The aims of this analysis are (1) to assess the association between the presence of low levels of HER2 expression with demographic and clinicopathological parameters, (2) to test the prognostic value of HER2-low status on invasive disease-free survival (IDFS), distant relapse-free survival (DRFS), and overall survival (OS) and (3) to assess the value of HER2 expression as a predictive biomarker of response to palbociclib. HER2-low was defined as HER2 immunohistochemistry (IHC) 1+ or IHC 2+ with negative in situ hybridization; all pathologic evaluation was performed locally. Pts with HER2-0 and Her2-low BC were included in this analysis; those with HER2-positive BC or missing HER2 status were excluded. Association of HER2 with other baseline covariates were tested with Chi-squared tests. Prognostic and predictive power of HER2 were assessed with Cox Models. Multivariable models included age, T-stage, N-stage, grade and progesterone receptor (PR) expression. Hazard ratios (HR) and 95% Confidence intervals (CI) are reported. Results: From the original PALLAS intention-to-treat population (N = 5,753), 5304 pts (92.2%) were included in this analysis. Among these, 2,254 pts (42.5%) were classified as HER2 IHC 0 (HER2-0) and 3,050 (57.5%) as HER2-low (1,838 with IHC 1+ and 1,212 with IHC 2+). For this analysis, median follow-up was 59.8 months. Compared to HER2-0, pts with HER2-low tumors had statistically higher estrogen receptor expression levels (mean expression 89.2% vs 88.2%, p = 0.019), lower PR expression levels (65.9% vs 68.2%, p = 0.007). Importantly, HER2-low status varied significantly across 21 participating countries (range 16.7% to 75.6%; p< 0.001) and was more frequent in pts enrolled in North America (63.1%) than in Europe (53.4%) and other regions (53.4%) (p < 0.001). No differences in HER2 expression were observed according to anatomic stage, T-stage, N-stage, histological grade, age, menopausal status, primary surgery type, prior radiation, prior chemotherapy, or baseline performance status. There were no statistically significant differences in IDFS, DRFS or OS according to HER2 status in univariate or multivariable Cox models (prognostic value, HR provided in the Table). There was no significant interaction between the HER2 status and the benefit to palbociclib in IDFS, DRFS or OS (Table). Similar results were obtained when HER2-0 was compared to HER2 1+ and HER2 2+ separately. Conclusions: In this large, prospective and international cohort, no differences were observed in clinical parameters, prognosis, or differential benefit from palbociclib (predictive value) between HER2-0 and HER2-low tumors. Significant geographic variability was observed in the prevalence of HER2-low status, suggesting a high degree of variation in pathologic assessment of HER2 expression without impact on outcomes. Support: AFT, Abbvie Pfizer; Clinicaltrials.gov: NCT02513394 https://acknowledgments.alliancefound.org Citation Format: Guilherme Nader-Marta, Christian F. Singer, Dominik Hlauschek, Angela DeMichele, Paolo Tarantino, Georg Pfeiler, Miguel Martín, Justin Balko, Zbigniew Nowecki, Marija Balic, Adam Brufsky, Arlene Chan, Patrick G. Morris, Tufia Haddad, Sibylle Loibl, Yuan Liu, Lidija Sölkner, Christian Fesl, Erica Mayer, Michael Gnant. Clinical characterization, prognostic and predictive values of HER2-low in early breast cancer in the PALLAS trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-01-13.

Abstract PO4-25-10: Towards personalized medicine for DCIS - the role of hormone receptors, HER2, and Ki67 status in high-grade DCIS

Abstract Objective: This study aimed to investigate the molecular details in breast ductal carcinoma in situ (DCIS). Expression status of estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), and Ki67 were investigated among luminal A (LumA), luminal B HER2-negative (LumB HER2-), luminal B HER2-positive (LumB HER2+), HER2-enriched, and triple-negative (TPN), subtypes of DCIS. Materials & Methods: The study comprised formalin-fixed paraffin-embedded (FFPE) specimens of 357 DCIS grade 3 cases diagnosed between 1996 – 2018. Routine diagnostic immunohistochemical (IHC) staining were performed. DCIS cases were classified as LumA, LumB HER2-, LumB HER2+, HER2-enriched or TPN, according to the 2013 St. Gallen guidelines, which is used for molecular subtyping of invasive breast carcinoma (IBC). Each subtype was sorted into three subcategories: “pure” meaning those without an invasive component; “w/invasive” meaning those with an invasive component; and “all” meaning the entire group of the given subtype. Furthermore, ER and PR-receptor expression was registered for LumA, LumB HER2- and LumB HER2+ cases as intervals. The distribution of Ki67 was analyzed within each subtype. For some analyses, we combined the LumA and LumB HER2- subtypes into one entire group. The inter-observer variability of Ki67 was calculated by setting a cut-off value of 20% (2013 St. Gallen). A cohort (n = 47) of DCIS cases with a median Ki67 value of 18% was selected. Ki67 was calculated by analyzing 200 DCIS cells in two separate hotspots. Cohen's Kappa coefficient was calculated based on these data. HER2 (IHC) was scored based on ASCO/CAP guidelines established for routine diagnostic work-up for IBC. 16 equivocal cases (2+) were further investigated using dual SISH. Results: 98% of “all” cases of the LumA subtype showed an ER ≥ 50%. PR expression ≥ 50% was found in 91% of cases in this subtype. The incidence of ER-receptor at a cut-off ≥ 50% in the LumA subtype was significantly higher than that in the LumB HER2- and LumB HER2+ subtypes (p < 0.0001, Chi-square). In contrast, there was a statistically significant reduction in ER-receptor expression at a cut-off < 10% in LumA compared to the latter subtypes. The proportion of cases with PR-receptors with a cut-off of < 20% showed significant differences between LumA, LumB HER2- and LumB HER2+ subtypes (1.6%; 47% and 37%, respectively). There was also a significantly higher proportion of PR-receptor ≥ 50% cases among the LumA subtype. We found a significant association between PR < 20% and HER2 (3+) in luminal subtypes (p < 0.0004, Fisher's exact). There was no significant difference in ER/PR expression in “pure” cases of luminal subtypes of DCIS versus those with an invasive component (p = 0.1831, Chi-square). The Ki67 in the entire population (n = 357) varied from < 1% to > 80%. The mean and median were around 20% in those subtypes whose classifications were not depended on Ki67. There was a significant difference in the distribution of Ki67 when cases of LumA “all” and those of LumB HER2- “all” were combined as one entire group and were compared to LumB HER2+ “all” and HER2-enriched “all” (p-adjusted < 0.0001, Kruskal Wallis). The assessment of Ki67 among different observers showed a Cohen's kappa coefficient of 0.29 – 0.31 (fair agreement). We compared the HER2 (IHC) 0, 1+ and 2+ score among LumA and LumB HER2- subtypes and did not find a statically significant difference, when the “pure” and “w/invasive” were compared (p = 0.603, Chi-square). Conclusions: Ki67 was highly variable in DCIS grade 3. Inter-observer agreement was (as expected) suboptimal, and the cut-off at 20% defined by the 2013 St Gallen guidelines for IBC is not reliable for the distinction of LumA and LumB HER2 subtypes of DCIS. The LumB subtype of DCIS is heterogeneous with considerable variability among the four IHC markers used in the present study. A low PR is strongly associated with HER2 (3+), in luminal subtypes (p < 0.0004, Fisher's exact). Citation Format: Hossein Schandiz, Lorant Farkas, Daehoon Park, Solveig Norheim Andersen, Jürgen Geisler, Torill Sauer. Towards personalized medicine for DCIS - the role of hormone receptors, HER2, and Ki67 status in high-grade DCIS [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-25-10.

Abstract PO4-26-02: Evaluation of Tumor Infiltrating Lymphocytes as a predictive biomarker of recurrence in patients with Ductal Carcinoma In Situ of the breast

Abstract Introduction: Ductal carcinoma in situ (DCIS) represents around 20% of early breast cancer diagnoses globally. Some studies evaluated risk factors associated with DCIS recurrence. Among these, age, surgical margins, comedonecrosis (CN), high nuclear grade, negative hormone receptors and HER2 expression stand out. Recent approaches have been evaluating the role of the immune microenvironment and its association with recurrence and progression of DCIS. Objective: To evaluate the association between Tumor Infiltrating Lymphocytes (TILs) in DCIS samples and disease recurrence. To characterize the immunological microenvironment of DCIS, analyzing the presence of TILs, “touching TILs” and desmoplastic reaction and their association with recurrence. Epidemiological, clinical, histological and immunohistochemical (IHC) characteristics were also evaluated. Methods: This is a retrospective cohort study with patients diagnosed with DCIS and treated at Sao Paulo State Cancer Institute and Hospital das Clinicas of University of Sao Paulo. We included women over 18 years old with a diagnosis of DCIS who underwent treatment from Jan/2007 to Dec/2020. Male patients, patients with a diagnosis of invasive or microinvasive disease in the anatomopathological examination of the surgical specimen or patients with history of any neoplasm were excluded. The main outcome was survival analysis according to the quantification of TILs, adjusted for potential confounders. For that, we collected data on age, presence of palpable mass, imaging findings, histological and IHC characteristics, such as expression of estrogen receptor (ER), progesterone receptor (PR) and hyperexpression of HER2, type of surgical approach, type of radiation therapy, use of endocrine therapy, length of follow-up, recurrence, and its type. Two pathologists evaluated TILS in the sample with the highest tumor representation and numerically quantified it as percentage. They also evaluated the presence of “touching tils” (lymphocytes in contact with the basement membrane) and desmoplastic reaction in the tumor stroma. Kaplan-Meier curves, log-rank tests and Cox regression models were used to evaluate survival. Chi-square tests were used to evaluate the association between categorical variables. Results: 283 patients met the eligibility criteria. Mean age of patients was 55 years. 15% had a palpable nodule at physical examination. Clustered amorphous microcalcifications were the most prevalent mammographic presentation, found in 41% of the patients. The most frequent histological and IHC features were cribriform presentation (73%) and ER positivity (86%), respectively. Desmoplastic reaction was absent in 10.5%, discreet in 51.9%, moderate in 24% and intense in 13.6% of the patients. Breast conserving surgery was performed in 189 patients, and 100 of them received adjuvant radiation therapy. Mean follow-up was 77.2 months, with a recurrence rate of 9.2%. We observed that tumors with focal necrosis (HR 6.4 [1.39-34.71] p 0.018) or CN (HR 4.53 [1.34 – 15.28] p 0.015) had higher risks of recurrence. Patients with a percentage value of TILs greater than or equal to 17% also had a higher risks of recurrence (HR 2.97 (1.17-7.51) p 0.02). These patients were mostly under 65 years of age (OR 0.45 [0.21 - 0.97] p 0.049). In a multivariate model, CN and TILs>17% remained significantly associated with recurrence (p=0.034 and p=0.035 respectively). There was a trend for invasive recurrence in 76.9% of the patients that relapsed when TILS were greater than or equal to 17% (p 0.062). “Touching TILs” were present in 12.9% of the patients but were not associated with recurrence (p=0.575). Conclusion: In our cohort, high value of TILs (>= 17%) and presence of CN were independently associated with DCIS recurrence. Our findings suggest that TILS are a prognostic immunological biomarker in DCIS and warrant further investigation in prospective trials to determine the most adequate cut-point. Citation Format: Camila Pasetto, Fernando Aguiar, Marcella Peixoto, Maira Doria, Bruna Mota, Jonathan Maesaka, Jose Roberto Filassi, Edmund Baracat, Rodrigo Gonçalves. Evaluation of Tumor Infiltrating Lymphocytes as a predictive biomarker of recurrence in patients with Ductal Carcinoma In Situ of the breast [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-26-02.

Abstract PO1-24-08: Targeting of HER3 potentiates the antitumor activity of paclitaxel against triple negative breast cancer

Abstract Background: Triple negative breast cancer (TNBC) is characterized by the absence of expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) overexpression. This aggressive form of breast cancer carries a higher risk of recurrence compared to other subtypes, presenting a significant clinical challenge. Although chemotherapy is commonly used as the primary treatment for a substantial number of TNBC patients, it is often accompanied by drug resistance and frequent tumor recurrence. Urgent efforts are needed to identify novel molecular targets specific to TNBC and develop effective therapies to combat this aggressive disease. The ultimate objective is to discover treatments capable of overcoming drug resistance and improving the overall survival of patients with TNBC. Methods: Immunohistochemistry was performed to examine the expression of HER3 in TNBC samples. Western blots were used to assess protein expression and activation. Cell proliferation and viability were determined by cell growth (MTS) assays. TCGA databases were analyzed to correlate HER3 mRNA expression with the clinical outcomes of TNBC patients. Specific shRNA was used to knockdown HER3 expression. LIVE/DEAD Cell Imaging was to detect live and dead cells. Orthotopic tumor models were established in nude mice to determine the capability of TNBC cells forming tumors and to test if our newly developed anti-HER3 monoclonal antibody (mAb) 4A7 could potentiate the antitumor activity of paclitaxel in vivo. Results: The chemo-resistant subline HCC1806-TR was obtained by subjecting TNBC HCC1806 cells to prolonged treatment with paclitaxel. In comparison to the parental cell line HCC1806-P, HCC1806-TR exhibited enhanced HER3 expression and activation. It is noteworthy that approximately half of the tested TNBC specimens and cell lines displayed elevated expression of HER3. Analyzing TCGA databases revealed that TNBC patients with high levels of HER3 mRNA expression in their tumors had significantly poorer overall survival (OS) and relapse-free survival (RFS) compared to those with low HER3 expression. Specifically knockdown of HER3 markedly inhibited TNBC cell proliferation and mammosphere formation in vitro and tumor growth in vivo. Blockade of HER3 signaling with our mAb (4A7) in combination with paclitaxel exerted significant antitumor activity against TNBC in vitro and in vivo. Conclusions: Our data demonstrate that increased HER3 is an effective therapeutic target for TNBC and our anti-HER3 mAb (4A7) may enhance the efficacy of paclitaxel in TNBC. Keywords: HER3, Anbibody, Chemotherapy, Triple Negative Breast Cancer Citation Format: Hui Lyu, Sanbao Ruan, Congcong Tan, Ann Thor, Bolin Liu. Targeting of HER3 potentiates the antitumor activity of paclitaxel against triple negative breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-24-08.

Abstract PO4-06-12: Distinction of basal-like and triple-negative basal-like breast cancers utilizing a novel comprehensive single-cell liquid biopsy-based test

Abstract Introduction: Breast cancer is a highly heterogeneous disease and is a leading cause of death for women worldwide. Triple-negative breast cancer (TNBC), the most aggressive form of the disease, is clinically defined by the lack of expression of estrogen receptor (ER) and progesterone receptor (PR), as well as lack of over-expression of human epidermal growth factor receptor-2 (HER2). Approximately 70% of all TNBC cases exhibit basal-like cells, characterized by high basal expression of keratins as well as extensive genomic instability with copy number gains and losses across most chromosomes. In this study we show that novel single cell genomic evaluation coupled with immunofluorescence (IF) protein analysis of CTCs can detect TNBC with basal-like genomic features as well as basal-like tumors with expression of ER and HER2. Materials and Methods: Epic Sciences’ DefineMBC™ liquid biopsy test characterizes CTC proteomics and genomics as well as ctDNA for the treatment of metastatic breast cancer. Blood samples from 813 metastatic breast cancer patients, with either newly diagnosed or progressing disease, with historical tissue subtype [HR(+)HER2(-), HR(+/-)HER2(+), and TNBC] supplied by the referring physician were collected and tested. Up to 5 CTCs were selected and isolated from each sample based on morphology and IF signal. DNA was extracted from individual cells, and whole genome amplified and sequenced at low coverage. Copy number analysis was performed to determine copy number gains and losses across chromosomes, as well as large scale-state transitions (LSTs) and ERBB2 amplification. CTCs characterized with high chromosomal instability (e.g., exhibiting >30 LSTs; 1Q firestorm and 8Q stair step) and high levels of cytokeratin expression (mean fluorescence intensity signal >10K) were categorized as basal-like. Results: By previous metastatic tissue biopsy, 77% were HR(+), 8% were HER2(+), and 12% were TNBC. CTCs categorized in DefineMBC as basal-like subtype (9.8 % of all samples tested) showed to be highly present in patients called as TNBC (79% basal-like). Additionally, 54% of HR(+) and 23% of HER2(+) had basal-like CTCS. Patients characterized with TNBC by metastatic tissue biopsy and by DefineMBC exhibited basal-like cells in 88% of the cases. Conclusion: The results show a novel clinical assay which utilizes CTC protein detection as well as single-cell genomic analysis to identify tumors driven by basal-like cells. Generally, TNBC is classified with negative biomarker expression. We show that CTCs in TNBC display genomically basal-like cells. CTCs from HR(+) and HER2(+) tumors may express basal-like features with preserved ER and HER2 expression. The comprehensive nature of DefineMBC™ utilizing genomic analysis of detected CTCs has provided a new tool to diagnose both basal-like TNBC and basal-like hormone receptor positive and HER2 over-expressing tumors. Citation Format: Alessandra Cunsolo, David Bourdon, Ernest Lam, Giuseppe Di Caro, Nilesh Dharajiya, Timothy Pluard, Lee Schwartzberg. Distinction of basal-like and triple-negative basal-like breast cancers utilizing a novel comprehensive single-cell liquid biopsy-based test [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-06-12.

Abstract PO2-25-11: Incidence of Breast Cancer in Louisiana Compared to the United States

Abstract Aside from skin cancer, breast cancer is the most common cancer in American women. More specifically, one in eight women will develop breast cancer during their lifetime. Breast cancer is classified into subtypes defined by immunohistochemistry (IHC) expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). ER+/PR+/HER2- tumors are the most common form of breast cancer and are typically associated with risk factors including older age, nulliparity, obesity following menopause, alcohol and tobacco use, sedentary lifestyle, and hormone therapy. ER-/PR-/HER2+ tumors comprise approximately 15-20% of breast cancers and are caused by overexpression of the HER2 gene. These tumors have similar risk factors to ER+/PR+/HER2- tumors, except for age. Women with HER2+ breast cancer are likely to be younger compared to ER+PR+/HER2-. Tumors lacking hormone receptors are classified as ER-/PR-/HER2-, otherwise known as triple-negative breast cancer (TNBC). TNBC represents 10%-20% of breast cancers and carries a poorer prognosis, shorter survival, and unresponsiveness to hormone therapy compared to other forms of breast cancer. TNBC has been associated with African American race, socioeconomically deprived population, younger age at diagnosis (most frequently in women ages 40-50), more advanced disease stage, higher grade, high mitotic indices, family history of breast cancer and BRCA1 mutations. The primary aim of this study is to compare incidence of ER+/PR+/HER2-, ER-/PR-/HER2+, and ER-/PR-/HER2- (TNBC) in Louisiana to the national population. The Louisiana Tumor Registry (LTR) was utilized to obtain state and national level data. The LTR is a participant of the National Cancer Institute’s Surveillance, Epidemiology and End Results Program, and the Centers for Disease Control and Prevention’s National Program of Cancer Registries. Our study evaluated women in Louisiana diagnosed with ER+/PR+/HER2-, ER-/PR-/HER2+, and ER-/PR-/HER2- from 2010-2019. The populations were subdivided by age at time of diagnosis. Data was subdivided into ages 0-49, and >50. Rates were calculated per 100,00 and age-adjusted to the 2000 United States Standard Population (19 age groups - Census P25-1130). A p-value of < 0.05 indicates a significantly different rate in Louisiana comparative to the United States (SEER). Our results are as follows. Unexpectedly, Louisiana has a significantly lower rate of ER+/PR+/HER2- cancer in women ages 0-49 and ages >50 compared to the United States. Based on epidemiological risk factors, Louisiana should have a much higher rate of ER+/PR+/HER2- comparative to the United States population. Louisiana has a significantly higher rate of ER-/PR-/HER2+ cancer in women ages >50 and ER-/PR-/HER2- (TNBC) in women ages 0-49 and ages >50 compared to the United States. These findings are discordant with traditional epidemiologic studies regarding breast cancer and require further investigation. Breast Cancer Incidence Rates1 by Age from 2010-2019 in Louisiana and the United States Table 1: Breast Cancer Incidence Rates in women ages 0-49 and ages >50 from 2010-2019 in Louisiana Compared to the United States. Citation Format: Avery Daily, Julie Cupp, Krystle Trosclair. Incidence of Breast Cancer in Louisiana Compared to the United States [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-25-11.

Abstract PO1-14-03: Discovery of proliferation essential gene signature and ACTL6A as potential biomarker for predicting prognosis and neoadjuvant therapy response in triple-positive breast cancer

Abstract Background: Based on the expression of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2), a special subtype of breast cancer which expresses all three receptors can be distinguished, namely triple-positive breast cancer (TPBC). TPBC patients has a higher risk of recurrence and lower survival rate compared to other luminal breast cancers. However, there are few studies on predicting molecules of prognosis and treatment response for TPBC. Methods: Proliferation essential genes (PEGs) were identified using CRISPR-Cas9 dataset of DepMap. The expression profiles and clinical data of TPBC were obtained from the TCGA database, GEO datasets and our center cohort. To develop a TPBC-PEG prognostic signature, cox regression and Lasso (Least absolute shrinkage and selection operator) regression analysis were applied. Functional analysis was performed using Gene Set Enrichment analysis. Finally, the relationship between candidate genes and neoadjuvant therapy (NAT) sensitivity was explored using real-time qPCR (RT-qPCR) and immunohistochemistry (IHC) based on clinical samples. Results: Among 900 TPBC-PEGs, 437 genes showed significant differential expression between TPBC and normal tissues. Subsequently, we identified 3 prognostic PEGs (ACTL6A, CCT2, TARS) and used them to construct a PEG signature. Patients with high PEG signature scores exhibited worse overall survival and lower sensitivity to NAT compared to those with low PEG signature scores. RT-qPCR indicated that in patients who lacked sensitivity to NAT, ACTL6A and CCT2 were significantly upregulated. The IHC results showed that ACTL6A protein was highly expressed in the NAT-insensitive group and non-pathological complete response patients. Conclusion: In this study, we developed an efficient PEG prognostic model that can predict the outcome for TPBC. Furthermore, we found that the expression of ACTL6A is associated with neoadjuvant therapy sensitivity in TPBC patients and can serve as an important factor in predicting patient prognosis and drug sensitivity. Figure 3. Construction and validation of a PEG signature for TPBC patients. Figure 6. The predictive value of ACTL6A on NAT efficacy and its correlation with clinical characteristics. Citation Format: Xiaofen Li, Wenfen Fu, Shiping Luo, Jie Zhang, Qingshui Wang, Chuangui Song. Discovery of proliferation essential gene signature and ACTL6A as potential biomarker for predicting prognosis and neoadjuvant therapy response in triple-positive breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-14-03.

Abstract PO2-18-08: Identification of novel genetic/therapeutic vulnerabilities in breast cancer

Abstract Background: Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer death among women worldwide. Triple negative breast cancer (TNBC) is an aggressive subtype that is defined by the lack of expression of estrogen and progesterone receptors in the absence of HER2 amplification or overexpression. TNBC represents 15-20% of all breast cancers and occurs more frequently in young premenopausal women. TNBC often ends with a poor clinical outcome due to high histological grade, rapid recurrence in many patients combined with few treatment options. These realities highlight the urgent need to identify novel therapeutics vulnerabilities that can be leveraged to improve TNBC patient outcomes. Methods: We utilized the publicly available DepMap and Kaplan Meier datasets to identify essential genes that were unique to TNBC cells and whose expression levels were associated with worse patient outcomes. siRNA based approaches were used to confirm the relevance of top hits in mediating cancer cell phenotypes including viability, cell-cycle profiles and induction of apoptosis among others. A novel drug currently being developed was identified for the top hit in these studies and was subsequently employed in multiple in vitro approaches to characterize the mechanisms of action in TNBC as well as initial efficacy studies using clinically relevant ex vivo patient derived models. Results: Using the DepMap database, we interrogated genome-wide sgRNA knockout screens conducted in over 40 breast cancer cell lines to identify specific genes whose expression was required for cell viability in TNBC cells, but not other breast cancer cell lines. This endeavor identified 6 genes (CTPS1, HUS1, PRKRA, RAD1, RAD9A and RHOA) that were preferentially essential in TNBC cells. Further profiling of these genes revealed that CTPS1 expression is higher in cancer relative to normal tissue and that its expression is further upregulated in chemotherapy- and PARPi-resistant models. Knockdown of CTPS1 in TNBC cells was shown to decrease cell proliferation and viability with no effects observed in estrogen receptor+ cell lines. Using a first-in-class, highly selective and orally bioavailable CTPS1 inhibitor (STP938) developed by Step-Pharma, we identified IC50s in the low nM range for many TNBC cell lines, including cell models that are resistant to existing standard-of-care therapies. Inhibition of CTPS1 was shown to arrest cells in S phase. Conclusions: Our studies have identified CTPS1 as an essential gene in TNBC. CTPS1 encodes an enzyme responsible for the catalytic conversion of UTP (uridine triphosphate) to CTP (cytidine triphosphate), an essential step in the biosynthesis of nucleic acids. These findings are timely in light of the recent and ongoing development of the first-in-class CTPS1 inhibitor, STP938, for which first-in-human studies are underway for refractory B- and T-cell lymphomas. Based on our initial studies using this drug, we provide pre-clinical evidence of its efficacy in multiple models of primary and advanced TNBC. Efforts are currently underway to move this work toward a clinical trial for TNBC patients. Citation Format: Xiyin Wang, Michael Emch, Lauren Voll, Rebecca Russell, Esther Rodman, Philip Beer, John Hawse. Identification of novel genetic/therapeutic vulnerabilities in breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-18-08.

Abstract PO5-06-05: DISCORDANCE BETWEEN HORMONE RECEPTORS AND HER2 STATUS IN BREAST CANCER PATIENTS RELAPSE

Abstract Introduction: Breast cancer (BC) is a heterogeneous disease, and during its progression, the tumor phenotype can undergo changes that are often associated with a poor prognosis. In this study, we aim to compare the discordance in estrogen receptor (ER), progesterone receptor (PgR), human epidermal growth factor receptor 2 (HER-2), and Ki-67 expression between primary and recurrent/metastatic lesions. Methods: This study is a retrospective analysis of 132 non-advanced breast cancer patients who experienced confirmed recurrence/metastasis at the Instituto Nacional de Enfermedades Neoplasicas between 2011 and 2022. Descriptive analysis was conducted to assess the expressions of ER, PgR, HER-2, and Ki-67 in primary and metastatic breast cancer, as well as clinical variables. Normality tests, including Kolmogorov-Smirnov and Shapiro-Wilk, were performed. The paired samples Wilcoxon test was used to determine significant differences between primary and recurrent/metastatic tumors. Additionally, Spearman's rank correlation coefficient was employed to evaluate the relationships of expression discordance. Results: Fifty-nine percent of patients presented locoregional recurrence, while 41% presented visceral metastasis. In the primary tumor, ER and PR expression were found in 63.6% and 55.3% of cases, respectively. HER2 status was determined as 0 (36.3%), +1 (22%), +2 (14.4%), +3 (26.5%), and not determined (0.8%) by IHC. For the recurrent/metastatic tumor, ER and PR expression were found in 59% and 34% of cases, respectively. HER2 status was determined as 0 (27.3%), +1 (19.7%), +2 (25%), +3 (25%), and not determined (3%). The median Ki-67 levels were 37% in the primary tumor and 40% in the recurrent/metastatic tumor. Significant differences were found for ER (p=0.014), PR (p< 0.001), and HER2 (p=0.019), but not for Ki-67 percentage (p=0.139). The discordance rates of ER, PR, HER2, and Ki-67 expression were 27.9%, 47.7%, and 26.3%, respectively. Conclussions: Discordance in hormone receptor expression between primary and recurrent/metastatic tumors is a common phenomenon, with ER showing a discordance rate of 27.9% and PR showing a higher rate of 47.7%. HER2 status also displayed significant discordance, with a 26.3% rate of discordant expression between primary and recurrent/metastatic tumors. The findings from this study emphasize the dynamic nature of breast cancer and the importance of comprehensive evaluation of biomarker expression in both primary and recurrent/metastatic tumors, to choose the best treatment options. Citation Format: Katia Roque, Iris Otoya, Natalia Valdivieso, Zaida Morante, Hugo Fuentes, Silvia Neciosup, Henry Gómez, Jorge Cotito, Gonzalo Ziegler, Estefania Peña, Tatiana Vidaurre, Ramon Andrade de Mello, Carlos Castañeda. DISCORDANCE BETWEEN HORMONE RECEPTORS AND HER2 STATUS IN BREAST CANCER PATIENTS RELAPSE [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-06-05.

Abstract PO1-01-11: Development and Validation of a Breast Cancer Recurrence Model Demonstrates Accurate Identification of Patients with Favorable Long-Term Outcomes

Abstract Background: The Oncotype DX (ODX) test is a 21-gene expression assay widely used for the prediction of risk recurrence in early-stage breast cancer, but it may be possible to identify patients who can forgo testing using only clinicopathologic variables. In 2018, the National Cancer Database (NCDB) began reporting quantitative histologic parameters for estrogen receptor (ER), progesterone receptor (PR), and Ki-67 expression in breast cancer patients. Inclusion of these variables may improve the development of nationally applicable models to predict ODX results using clinicopathologic variables alone. Methods: Using a cohort of patients from the NCDB diagnosed from 2018–2020 with hormone receptor (HR)-positive, HER2-negative, Stage I-III breast cancer, we trained machine learning models to predict high-risk (26-100) ODX score. A subset comprising 80% of patients was used for model training, while the remaining data were set aside for internal validation. An external validation cohort was selected from the University of Chicago Medical Center (UCMC), including patients diagnosed from 2009–2021. Feature selection, model architecture selection, and hyperparameter tuning were performed using 10-fold cross-validation within the NCDB training set. We compared a model with quantitative ER, PR, and Ki-67; a model with only quantitative ER and PR, and a model without quantitative immunohistochemistry – to best reflect the likely data available in a variety of practice patterns. The primary endpoint was the area under the receiver operating characteristic curve (AUROC) for prediction of high-risk ODX results in the UCMC validation cohort. Models were also evaluated as rule-out tests to identify low-risk patients who did not require further ODX testing, using a high (90%) sensitivity threshold, fit in the NCDB training dataset. Results: We identified 53,346 patients from the NCDB cohort meeting the inclusion criteria; 7% had a high risk ODX score, with a median follow-up time of 28 months. The UCMC validation cohort included 896 patients, and was more diverse, with 30% non-Hispanic Black patients (versus 8% in NCDB), more high-risk patients (18% with high ODX), and a longer median follow-up time of 55 months. In the NCDB validation cohort, models incorporating quantitative ER/PR (AUROC 0.78, 95% CI 0.77–0.80) and quantitative ER/PR/Ki-67 (AUROC 0.81, 95% CI 0.80–0.83) both performed better than the non-quantitative model (AUROC 0.70, 95% CI 0.68–0.72). These results were preserved in the external UCMC cohort, where the ER/PR model (AUROC 0.86, 95% CI 0.80–0.92, p = 0.032) and the ER/PR/Ki-67 model (AUROC 0.87, 95% CI 0.81–0.93) outperformed the non-quantitative model (AUROC 0.80, 95% CI 0.73–0.87, p = 0.009). The high sensitivity rule-out threshold of the ER/PR model predicted that 30% of patients in the UCMC cohort would be low ODX, and the ER/PR/Ki-67 model predicted 44% as low risk – negative predictive value was over 96% for prediction of high ODX. Of the patients predicted to be low risk by the quantitative models, none had a documented high ODX score, and recurrence was < 3% at 5 years. The hazard ratio for recurrence free interval, adjusted for age and comorbidity score, of patients predicted to be high risk by this threshold was 2.96 (95% CI 1.02–8.58) for the ER/PR model and 3.84 (95% CI 1.48–9.97) for the ER/PR/Ki-67 model. Conclusions: We present externally validated and nationally applicable models that identify approximately half of HR-positive/HER2-negative breast cancer patients who are unlikely to have high ODX results using widely available quantitative clinicopathologic variables. Patients identified as low risk by these models have excellent long-term outcomes and may be able to forgo adjuvant chemotherapy without further genomic testing. Citation Format: Asim Dhungana, Augustin Vannier, Fangyuan Zhao, Jincong Freeman, Poornima Saha, Megan Sullivan, Katharine Yao, Elbio Flores, Olufunmilayo Olopade, Dezheng Huo, Alexander Pearson, Frederick Howard. Development and Validation of a Breast Cancer Recurrence Model Demonstrates Accurate Identification of Patients with Favorable Long-Term Outcomes [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-01-11.

Abstract PO2-05-14: Prognosis and clinicopathological characteristics of HER2-negative metastatic breast cancers with low expression of hormone receptors: real world evidence from the French ESME cohort

Abstract Background: HER2-negative breast cancers expressing low (1-9 %) immunohistochemistry levels of hormone receptors (HR, estrogen and progesterone receptors), remain an uncertain category, being considered either as triple negative breast cancers (TNBC) or HR-positive (HR+) tumors across guidelines or approvals. The issues regarding this clinical subgroup are both prognostic, with the need of a better definition of their clinical outcome, as well as predictive. Methods: The ESME database is a French National cohort of all consecutive patients who initiated a first-line treatment for metastatic breast cancer (MBC) from 2008 on, in one of the 18 French Comprehensive Cancer Centers. Patients with HER2-negative (0 to 2+, ISH negative) MBC and known estrogen receptor (ER) and progesterone receptors (PR) expression levels were selected. Primary objective was to evaluate crude overall survival (OS) in HER2-/HR-low (ER and PR 0-9 and not TNBC) MBC pts compared with those with TNBC and HR+ disease defined by ER and PR < 1%, and ER ± PR ≥ 10%, respectively. In these 3 populations, predefined secondary objectives were to evaluate PFS under first line chemotherapy (CT), to describe clinical characteristics and the distribution of genetic abnormalities identified at the time of MBC diagnosis. Results: Out of 30,459 patients in the ESME database initiating their MBC treatment between 01/2008 and 01/2021, 19109 patients (2113 TNBC; 228 HR-low; 16768 HR+) were eligible for this analysis. Median follow-up was 58.0 months (95%CI [56.6; 59.0]). Median OS were 15.7 months [15.0-16.8], 19.1 months [15.5-22.4] and 44.6 months [43.8-45.5] in the TNBC, HR-low and HR+ subgroups, respectively. The multivariable analysis adjusted on age at MBC diagnosis, pathological grade and subtype, metastatic-free interval, number and sites of metastases and HER2 status (0 vs. 1-2+), identified no significant OS difference between HR-low and TNBC subgroups (HR=0.95, 95%CI 0.79-1.14), while HR+ patients had a better OS than TNBC patients (reference) (HR=0.50, 95%CI 0.47-0.53). Of these 19109 pts, 8910 received first-line CT regimen (1960 TNBC; 195 HR-low; 6755 HR+): taxanes-based: 66.7%, anthracyclines-based: 32.9%, capecitabine: 18%. Median PFS under first line CT were 5.3 (5.1-5.6), 5.1 (4.1-6.2) and 10.2 months (9.9-10.6) for TNBC, HR-low and HR+ subgroups, respectively. In the multivariable analysis adjusted on the same variables, the HR-low subgroup had a statistically poorer PFS compared to TNBC (HR=1.24, 95% CI 1.04-1.49). On the other hand, the HR+ subgroup had a better PFS than the TNBC (HR=0.74, 95%CI [0.70; 0.78]). Baseline germline BRCA1 mutations (at MBC diagnosis) were present in 4.5% of the TNBC cases, 2.6% of the HR-low cases, and 0.4% of the HR+ cases. The distribution of germline BRCA2 mutations was more homogeneous between the three groups (0.9% of TNBC cases, 1.3% of HR-low cases and 1.1 % of HR+ cases). Conclusions: In this large cohort, patients with HER2-negative, HR-low MBC seem to share similar general prognosis, oncogenetic features and PFS under standard chemotherapy treatment with TNBC patients. Citation Format: Marie Alexandre, Ludovic Gauthier, Audrey Mailliez, Thibault DE LA MOTTE ROUGE, Thomas Bachelot, Monica Arnedos, Florence Dalenc, Etienne Brain, Jean-Marc Ferrero, Vincent Massard, Isabelle Desmoulins, Marie-Ange Mouret-Reynier, Christelle Levy, Anthony Gonçalves, Marianne Leheurteur, Thierry Petit, Jean-Sébastien Frenel, Lise Bosquet, Suzette Delaloge, William Jacot. Prognosis and clinicopathological characteristics of HER2-negative metastatic breast cancers with low expression of hormone receptors: real world evidence from the French ESME cohort [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-05-14.