Ki-67 Response Research Articles

A Multicenter Physician Survey Evaluating the Use of Ki-67 in Breast Cancer Management in Canada.

Ki-67's response to pre-operative endocrine therapy (ET) in early breast cancer is an evidence-based tool to guide adjuvant treatment decisions. Physicians across Canada were surveyed to explore current practice patterns and perceived barriers to the use of Ki-67 in practice. Physicians were invited to participate in an anonymous survey and were eligible if they prescribed systemic therapy for breast cancer in Canada. Respondents were asked to describe their usage of Ki-67, perceptions of the evidence surrounding Ki-67 ET response, and interest in future trials using this approach. The survey received 48/163 responses (29.4%). The majority of respondents (97.6%) reported access to Ki-67 testing upon request. Treatment decisions for adjuvant Abemaciclib was the most common reason (97.6%), followed by adjuvant chemotherapy decisions (16.7%). Only 19.0% had used Ki-67's response to pre-operative ET in practice. Common barriers to this approach that were identified included a lack of awareness from other providers (54.8%), an increased resource requirement (54.8%), and a lack of timely medical oncology consultation (52.4%). The majority of physicians (85.3%) reported that they would participate in future trials using the Ki-67 endocrine response, and that rate of treatment decision change (95.2%) and cost analysis (42.3%) were important endpoints. Despite the widespread availability of Ki-67 testing, few physicians in Canada currently use it to assess endocrine response, predominantly due to logistical and resource constraints. There is a high level of interest in participating in future trials using this strategy, which should focus on disease related outcomes, feasibility, and the financial impact on the public healthcare system.

Clinical and immune responses to neoadjuvant fulvestrant with or without enzalutamide in ER+/Her2- breast cancer.

Most ER+ breast cancers (BC) express androgen receptors (AR). This randomized phase II trial of 4 months of neoadjuvant fulvestrant (Fulv) alone or with enzalutamide (Combo) assessed whether adding AR blockade to Fulv would limit residual tumor at the time of surgery, as measured by modified preoperative endocrine predictive index (PEPI) score. Eligible patients were women with ER+/HER2- primary BC cT2 or greater. Stratification factors were clinical node and T-stage. Fresh tumor biopsies were required at study entry, after 4 weeks on therapy (W5), and at surgery. Laboratory analyses on tumors included immunochemistry (IHC) for ER/PR/AR/GR and Ki67 protein, evaluation of gene expression, multiplex for myeloid lineage immune cells, reverse-phase protein array, and plasma metabolomic analyses. Of 69 consented patients, 59 were evaluable. Toxicity was as expected with endocrine therapy. Combo achieved PEPI = 0 more frequently (24%: 8/33) than Fulv (8%: 2/26). Ki67 was ≤10% across arms by W5 in 76% of tumors. Activation of mTOR pathway proteins was elevated in tumors with poor Ki67 response. Tumors in both arms showed decreased estrogen-regulated and cell division gene sets, while Combo arm tumors uniquely exhibited enrichment of immune activation gene sets, including interferon gamma, complement, inflammation, antigen processing, and B and T cell activation. Multiplex IHC showed significantly reduced tumor-associated macrophages and CD14+/HLADR-/CD68- MDSCs in Combo tumors at W5. In summary, Combo tumors showed a higher PEPI = 0 response, Ki67 response, and more activated tumor immune microenvironment than Fulv. The odds of response were 4.6-fold higher for patients with ILC versus IDC. (Trial registration: This trial is registered at Clinicaltrials.gov ( https://www.clinicaltrials.gov/study/NCT02955394?id=16-1042&rank=1 ). The trial registration number is NCT02955394. The full trial protocol is available under Study Details at the Clinicaltrials.gov link provided).

Abstract PS08-01: Antibody-Drug Conjugates (ADCs) in Breast Cancer: Real World Analysis of Outcomes

Abstract Methods: We estimated the distribution of progression-free survival (PFS), overall survival (OS), and time to treatment failure (TTF) using the Kaplan-Meier method. Differences in survival curves between groups categorized by initial ADC treatment and subsequent ADC treatments were assessed using the log-rank test. Cox proportional hazards regression models were employed to evaluate the association between each survival outcome and various measures of interest, such as age, gender, race, number of prior lines of treatment, hormone receptor status, and HER2 status. Results: The analysis included a total of 469 breast cancer patients, with the distribution of clinical characteristics summarized in Table 1. The median age at the start of ADC treatment was 50 years (range: 20-85). Among the patients, 263 patients (56%) received SC as their initial ADC treatment, while 44% received T-DXd. Additionally, 29 patients received both ADCs during metastatic treatment. The median follow-up time for all patients was 7.9 months (range: 0.1- 39.1). Out of the total patient population, 29% died, while 71% were still alive at the last follow-up. The median OS for all patients was 21.6 months. Median OS of patients receiving SC or T-DXd as their initial ADC treatment, was 14 and 37.1 months, respectively. Patients who received both SC and T-DXd had a median OS of 26.1 months. The median PFS for all patients was 6.0 months. Patients initially treated with SC had a median PFS of 4.7 months, whereas those treated with T-DXd had a median PFS of 9.0 months. In patients who received both, median of PFS was 4.9 months for T-DXd after SC, and 5.0 months for SC after T-DXd. The median TTF for all patients was 6.1 months. Patients initially treated with SC had a median TTF of 4.7 months, while those treated with T-DXd had a median TTF of 9.0 months. In patients who received T-DXd after SC, TTF was 4.9 months and SC after T-DXd was 5.0 months. Clinician-judged responses were seen in 180 (74%)of patients with SC only, 80 (41%) with T-DXd only, 9 (100%) with SC as 2nd ADC and 20 (100%) with T-DXd as 2nd ADC. Furthermore, the analysis explored the association of OS, PFS, and TTF with various measures of interest, including age, gender, race, number of prior lines of treatment, type of treatment, hormone receptor status, HER2 status, Ki67, and best response to ADC. Some notable associations were observed, such as ER-positive status being associated with marginally longer OS compared to ER-negative status (median 24.7 vs. 16.4 months; p=0.06), and HER2-positive status being associated with longer PFS compared to HER2-negative status (median 14.2 vs. 5.4 months; p< 0.001). However, these associations should be interpreted cautiously as they were obtained from univariate analysis. Conclusions: Overall, these results highlight the significant differences in survival outcomes between different ADC treatments in breast cancer patients, likely due to differing patient characteristics. Distribution of clinical characteristics across ADCs Citation Format: Akshara Singareeka Raghavendra, Zhongya Wang, Roland Bassett, Debu Tripathy. Antibody-Drug Conjugates (ADCs) in Breast Cancer: Real World Analysis of 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 PS08-01.

Can Ki-67 predict radiotherapy response in neuroendocrine tumors? Retrospective analysis of a monocentric series of patients.

The impact of radiotherapy (RT) in neuroendocrine neoplasms is still unknown, and outcomes could be improved by a better insight in RT response predictors. This retrospective analysis investigates the potential correlation between Ki-67 and RT response to evaluate its role as biological marker of radiosensitivity. Data from patients treated at an Italian NET-referral center between 2015 and 2020 were retrieved. Inclusion criteria included: histologically-proven diagnosis of NEN, Ki-67 status, indication (symptomatic and/or ablative) and at least one post-RT radiological assessment. Forty-two patients and 63 different treatment lines were included. Primary tumors presented Ki-67 values < 3% in 21% of cases, between 3 and 20% in 45% and >20% in the remaining 33%. Almost all patients were metastatic at the time of RT, which was performed with symptomatic purpose in 43% of cases. At a median time of three months, a complete response on the target lesion was observed in nine cases (14%), a partial response in 17 (27%), stability in 23 (37%) and local progression in 14 (22%). With median FU of 22.8 months, OS does not show statistically significant differences among three Ki-67 groups. Considering all lines of therapy, the relationship between ORR and Ki-67, did not show statistically significant differences, even following adjustments for drug types and delivered RT doses. No association between Ki67 and local tumor response to RT could be observed in the present cohort, regardless of whether the evaluation was performed on a categorical or continuous scale.

Abstract PD13-02: PD13-02 Exploratory gene expression analysis of coopERA Breast Cancer (BC): a study evaluating neoadjuvant giredestrant versus anastrozole alone and in combination with palbociclib in ER-positive, HER2-negative untreated early BC

Abstract Background: Endocrine therapy remains the mainstay treatment for ER+ BC. CDK4/6 inhibitors induce cell cycle arrest and decrease tumor cell proliferation, as measured by the biomarker Ki67, when used in combination with aromatase inhibitors (AI) such as anastrozole (A). Giredestrant is an oral, well-tolerated, and highly potent selective ER degrader (SERD) that achieves robust ER suppression and has demonstrated antitumor activity in the metastatic setting either as monotherapy or in combination with the CDK4/6 inhibitor palbociclib. The randomized, phase 2 coopERA Breast Cancer study (NCT04436744) evaluated giredestrant in postmenopausal women with untreated ER+/HER2- early BC and met its primary endpoint, demonstrating superior Ki67 suppression with giredestrant vs A after two weeks of single agent treatment. This suppression was maintained at surgery where giredestrant vs A was evaluated in combination with palbociclib. Here, we present gene expression analysis and associations with Ki67 response. Methods: 221 eligible patients with measurable ER+/HER2– untreated early BC and baseline Ki67 ≥ 5% were randomized 1:1 to receive 30 mg oral daily (PO QD) giredestrant or 1 mg PO QD A on Days 1–14 of a neoadjuvant window-of-opportunity phase, followed by four 28-day cycles of PO QD giredestrant or A with 125 mg PO QD palbociclib on Days 1–21 before surgery. FFPE specimens were collected at baseline, week 2 and surgery; and RNA-sequencing (seq) was performed. Gene expression analysis included ER pathway activity, PAM50 intrinsic subtypes, and other pathway analyses, assessed by Ki67 response. Results: 112 and 92 patients had paired tumor samples at baseline/week 2 and baseline/surgery, respectively, that were evaluable for RNA-seq and Ki67. The trend for greater Ki67 protein suppression by giredestrant vs A from baseline to week 2 was maintained in the RNA-seq evaluable subset. Interestingly, the same subset revealed similar suppression of both proliferation gene signatures and ER pathway activity between A and giredestrant. PAM50 subtyping showed that 69% of tumors were luminal (Lum) A and 29% were LumB at baseline. Less than 1% were classified as basal or HER2. Interestingly, giredestrant (G) showed greater suppression of both Ki67 and ER pathway activity vs A in LumB tumors (Ki67: -82% [G] vs -62% [A]; ER activity: -0.83 [G] vs -0.66 [A]) compared to LumA at week 2 (Ki67: -74% [G] vs -71% [A]; ER activity: -0.60 [G] vs -0.70 [A]). Moreover, at week 2, 83% (13/18) of LumB tumors at baseline transitioned into a LumA subtype after giredestrant treatment compared to 46% (5/11) of A-treated tumors. Giredestrant-treated tumors also achieved lower mean ER pathway activity compared to those treated with A at surgery (p=0.023). Gene set enrichment analysis showed downregulation of cell-cycle and ER-related pathways at week 2 and surgery in both treatment arms. A subset of cytokine signaling and immune response pathways were increased at week 2 compared to baseline after treatment with A but not with giredestrant. These pathways were also associated with Ki67 resistance (Ki67 ≥ 7.4%) in A-treated tumors. This was consistent with differential expression analysis of samples collected at week 2, in which cytokine signaling pathways were enriched in A compared to giredestrant. Notably, IL12 signaling was enriched in tumors resistant to A but not giredestrant. Conclusions: Giredestrant has a greater effect on Ki67 protein suppression in ER+/HER2- early BC compared to A, which is more pronounced in LumB tumors. This benefit may involve differential regulation of cytokine and immune responses. These exploratory findings reveal novel mechanisms that may differentiate the activity of SERDs vs AIs, which warrant further validation. Citation Format: Alejandro M. Chibly, Tharu M. Fernando, Ciara Metcalfe, Marc Hafner, Gilbert Owusu-Manu, Sara Hurvitz, Aditya Bardia, Peter A. Fasching, Yeon H. Park, Vanesa Quiroga, Jutta Steinseifer, Pablo Perez-Moreno, Heather M. Moore. PD13-02 Exploratory gene expression analysis of coopERA Breast Cancer (BC): a study evaluating neoadjuvant giredestrant versus anastrozole alone and in combination with palbociclib in ER-positive, HER2-negative untreated early BC [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 PD13-02.

Abstract TC1-3-1: Ki 67 Ki67 – a clinically relevant biomarker or just nice to have information?

Abstract Ki67 is an immunohistochemical marker indicating the growth fraction of cycling cells from G1 to S-phase in tumor specimens. In breast cancer, Ki67 is a prognostic marker with high values indicating poor outcome. Over the last decades, obstacles for its wide-spread use in clinical practice included lack of methodological standardization and lack of specific cut-off values for clinical decision making, i.e. discrimination of luminal A vs. B early breast cancer (EBC), chemotherapy indication in HR+/HER2- EBC. However, recently, the International Ki67 working group has put forward consensus statements for a standardized methodology. In general, reproducibility of Ki67 values at extreme sides of the spectrum such as 10% or ≥ 35% is much higher than in the intermediate range and quality assurance programs substantially improve laboratory performance. Moreover, clinical utility has also become apparent in two specific settings of HR±/HER2- EBC. First, the monarchE trial evaluating adjuvant abemaciclib in node-positive high-risk disease used a Ki67 cut-off of 20% as a criterion for aggressive disease and as a sole inclusion criterion for patients with 1-3 lymph nodes (cohort2). While data for this cohort have not yet been reported, Ki67 was a strong prognostic factor in the overall trial population with patients with high Ki67 having worse outcome than those with Ki67 &amp;lt; 20%. Benefit from abemaciclib was independent of Ki67 index. Nevertheless, some health authorities included Ki67 in their abemaciclib label as the absolute benefit was greater in tumors with Ki67 ≥ 20%. Second, Ki67 response after a short (2-4 week) preoperative endocrine therapy allows endocrine response assessment by determining Ki67 in the surgical specimen. This information is widely used in drug development. Ki67post 10% has been defined as endocrine response. In the POETIC trial, endocrine response was associated with improved outcome compared to tumors with Ki67 ≥ 10% after 2-weeks of preoperative AI in postmenopausal patients. In the WSG ADAPT trial, patients with 0-3 lymph nodes, Recurrence Score 25 and endocrine response had excellent outcome with adjuvant endocrine therapy alone independent of menopausal status (5-year dDFS ≥ 95%). In postmenopausal women, probability of endocrine response with an AI is around 80% whereas in premenopausal patients with tamoxifen it is only around 40%. Yet, in premenopausal women addition of GnRH substantially improves endocrine response probability reaching about 80% with GnRH and AI as recently shown by interim analysis of the WSG ADAPTcycle trial (about 2500 patients). Endocrine response probabilities seen in the ADAPT trial (≥ 5000 patients) were validated in ADAPTcycle demonstrating clinical validity of this biomarker. In conclusion, Ki67 is a clinically relevant biomarker that can be used for clinical decision making, particularly in HR+/HER2- EBC. As there is no single generally accepted Ki67 cut-off value to discriminate prognostically favorable from aggressive EBC, Ki67 needs to be used in the context of tumor burden and tumor biology. Currently, next to its use in allocating patients to adjuvant abemaciclib in countries where Ki67 index is included in the label, it provides clinically important information after short-term preoperative endocrine therapy regarding endocrine response in HR+/HER2 EBC. Given its easy and inexpensive determination method, its analytical validity within quality assurance programs, and its clinical usefulness, Ki67 needs to be integrated in our biomarker portfolio in EBC. Citation Format: Nadia Harbeck, Torsten Nielson. Ki 67 Ki67 – a clinically relevant biomarker or just nice to have information? [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 TC1-3-1.

Abstract PD10-07: PD10-07 Low plasma estradiol, low expression of estrogen responsive genes and TP53 mutations are associated with poor anti-proliferative response to aromatase inhibitors

Abstract Background: Aromatase inhibitors (AIs) are highly effective at reducing recurrences and mortality in postmenopausal patients with estrogen receptor positive breast cancer (ER+ BC). Poor anti-proliferative (Ki67) response or ER+ BCs to AIs after 2 weeks is associated with worse long-term outcomes. Factors that relate to the degree of the response may identify markers and/or mechansims of resistance. Methods: The PeriOperative Endocrine Therapy for Individualizing Care (POETIC) trial randomized 4,480 with ER+ BC to 2 weeks’ AI before surgery or no presurgical treatment. All patients within the bottom 15% of Ki67 responders to AI (poor responders [PRs]; n=177 with RNA extracted) were selected from and matched to good responders (GRs) within the 50% showing the best response (n=190). Matching was based on baseline Ki67 levels as measured by immunohistochemistry (IHC). Response to AI was measured by the percentage change in Ki67 after 2 weeks’ treatment. PRs were further divided into groups expressing high ESR1 (PRs ESR1HIGH; n=119) and low ESR1 (PRs ESR1LOW; n=58) levels since there were very few GRs with low ESR1. RNAseq, targeted exome DNA sequencing of 87 BC/resistance related genes and measurement of plasma estradiol levels by mass spectrometry were performed to understand mechanisms of de novo resistance. Intrinsic subtypes were estimated from RNAseq data. Results: More than 90% of PRs ESR1LOW were non-luminal subtypes with low expression of estrogen-responsive genes. In contrast, 11% of PRs ESR1HIGH were non-luminal compared to 4% of GRs but only HER2-enriched subtypes were significantly higher in PR ESR1HIGH (p=0.05, Fisher exact). While AI treatment had limited impact on Ki67 IHC values in PRs ESR1HIGH, PGR expression was more than 2-fold lower after 2 weeks of AI. Gene-set enrichment analysis showed significantly lower expression of estrogen-response genes in PRs ESR1HIGH compared to GRs (FDR&amp;lt; 10-9) at baseline despite similar percentage of Luminal subtypes in PRs ESR1HIGH and GRs. Plasma estradiol levels were correlated with expression of estrogen-response genes (FDR=0.01) and levels were significantly lower in PRs ESR1HIGH compared to GRs (p=0.003, Mann Whitney). PRs ESR1HIGH had significantly more mutations in RB1, TP53, ARID1B and DNAH11 genes (p&amp;lt; 0.05, Fisher exact). TP53 mutations were significantly enriched in Luminal-A PRs ESR1HIGH compared to GRs (22% and 3% respectively; p=0.003, Fisher exact), but not in Luminal-B tumors (23% and 15% mutated respectively). Discussion and conclusions: In approximately 33% of PRs, de novo AI resistance was associated with and most likely due to low expression of ER/ESR1 and estrogen-responsive genes in non-luminal tumors. In the remaining tumors, AI treatment still impacted some estrogen responsive genes but had limited downstream impact on suppressing proliferation. This might be due to mutations including in TP53 that limit suppression of proliferation downstream of estrogen signaling. The proportion of Luminal tumors in GRs and PRs ESR1HIGH was similar, suggesting better outcome of Luminal-A tumors on AI is likely due to their better intrinsic prognosis rather than better response to endocrine therapy. Citation Format: Eugene F. Schuster, Elena López-Knowles, Anastasia Alataki, Lila Zabaglo, Elizabeth Folkerd, David Evans, Kally Sidhu, Holly Tovey, Perry Maxwell, Nicholas Turner, Stephen Johnston, Manuel Salto-Tellez, Maggie Chon U Cheang, John Robertson, Ian Smith, Judith Bliss, Mitch Dowsett. PD10-07 Low plasma estradiol, low expression of estrogen responsive genes and TP53 mutations are associated with poor anti-proliferative response to aromatase inhibitors [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 PD10-07.

Abstract OT2-20-01: Neoadjuvant survivin immunotherapy maveropepimut-S (MVP-S) to increase Th1 immune response in Ki67-high hormone receptor positive (HR+) early-stage breast cancer

Abstract Background: HR+ early stage breast cancer (ESBC) is associated with suboptimal pathologic complete response rate (pCR, ~10%) following neoadjuvant cytotoxic chemotherapy. Neoadjuvant endocrine therapy with aromatase inhibitors (AI) may serve as an effective alternative as gauged using the surrogate Ki67 cell proliferation histologic marker. Patients with poor Ki67 response (defined as Ki67&amp;gt;10%) following neoadjuvant AI exhibit poor prognosis and therapeutic resistance to both endocrine therapy and chemotherapy. In a genomic analysis of Ki67-high HR+ tumors, we identified 8-fold upregulation of BIRC5 (survivin), a gene that regulates apoptosis and the cell cycle and is associated with poor clinical outcome. Maveropepimut-S (MVP-S) leverages the non-aqueous, lipid-based DPX delivery platform to educate a specific and persistent T cell-based immune response to 5 HLA-restricted peptides from Survivin. Treatment with MVP-S and intermittent, low-dose cyclophosphamide (CPA) has shown to increase tumor infiltration of survivin-specific T cells. Previous clinical trials have shown that MVP-S is well-tolerated, immunogenic, and could lead to clinical response in several cancer indications. Further exploration of the regimen in breast cancer could extend the application of this immunotherapy for the unmet medical need of improving clinical response in high ki67 HR+ ESBC prior to surgery. Trial Design: NCT04895761 is phase I trial evaluating the safety and immunologic effects of neoadjuvant MVP-S plus letrozole (arm A, n=6), with/without tumor-directed MR-guided radiotherapy (arm B, n=6), or intermittent low-dose cyclophosphamide (CPA, arm C, n=6). Postmenopausal patients with T1c+ HR+HER2- breast cancer with Ki67&amp;gt;10% will receive two doses of MVP-S and 7 weeks of neoadjuvant letrozole prior to surgery (all arms), arm B will be treated additionally with concurrent 10Gy x 2 tumor boost radiation to facilitate immunogenic cell death, and arm C (n=6) will be treated additionally with intermittent low-dose CPA (50mg BID) to facilitate regulatory T cell depletion. Specific Aims: The primary objective is safety. Biomarker objectives are to evaluate for each treatment arm: 1) systemic type I survivin-specific immune response, as measured by IFN-γ ELISPOT; 2) changes in immune environment by GeoMx digital spatial genomic profiling; 3) and changes in tumor infiltrating lymphocytes (TILs) and Ki67. These data will be used to identify the most immunogenic MVP-S combination therapy for study in phase II trial powered to assess clinical outcome (pCR). Accrual: 3 of 6 patients in the MVP-S+ letrozole arm have been enrolled. Arm B and C will enroll after completion of arm A. Citation Format: Sasha E. Stanton, Lisa D. MacDonald, Stephan Fiset, Staci Mellinger, Nicole Moxon, Heather Hirsch, Tracy L. Kelly, Kristina H. Young, David B. Page. Neoadjuvant survivin immunotherapy maveropepimut-S (MVP-S) to increase Th1 immune response in Ki67-high hormone receptor positive (HR+) early-stage 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 OT2-20-01.

HER2-enriched subtype and novel molecular subgroups drive aromatase inhibitor resistance and an increased risk of relapse in early ER+/HER2+ breast cancer.

SummaryBackgroundOestrogen receptor positive/ human epidermal growth factor receptor positive (ER+/HER2+) breast cancers (BCs) are less responsive to endocrine therapy than ER+/HER2- tumours. Mechanisms underpinning the differential behaviour of ER+HER2+ tumours are poorly characterised. Our aim was to identify biomarkers of response to 2 weeks’ presurgical AI treatment in ER+/HER2+ BCs.MethodsAll available ER+/HER2+ BC baseline tumours (n=342) in the POETIC trial were gene expression profiled using BC360™ (NanoString) covering intrinsic subtypes and 46 key biological signatures. Early response to AI was assessed by changes in Ki67 expression and residual Ki67 at 2 weeks (Ki672wk). Time-To-Recurrence (TTR) was estimated using Kaplan-Meier methods and Cox models adjusted for standard clinicopathological variables. New molecular subgroups (MS) were identified using consensus clustering.FindingsHER2-enriched (HER2-E) subtype BCs (44.7% of the total) showed poorer Ki67 response and higher Ki672wk (p<0.0001) than non-HER2-E BCs. High expression of ERBB2 expression, homologous recombination deficiency (HRD) and TP53 mutational score were associated with poor response and immune-related signatures with High Ki672wk. Five new MS that were associated with differential response to AI were identified. HER2-E had significantly poorer TTR compared to Luminal BCs (HR 2.55, 95% CI 1.14–5.69; p=0.0222). The new MS were independent predictors of TTR, adding significant value beyond intrinsic subtypes.InterpretationOur results show HER2-E as a standardised biomarker associated with poor response to AI and worse outcome in ER+/HER2+. HRD, TP53 mutational score and immune-tumour tolerance are predictive biomarkers for poor response to AI. Lastly, novel MS identify additional non-HER2-E tumours not responding to AI with an increased risk of relapse.FundingCancer Research UK (CRUK/07/015).

Abstract 6151: Tumor microenvironment heterogeneity identifies potential biomarkers of response in ER-positive breast cancers treated with palbociclib

Abstract Despite the breast cancer microenvironment being recognized as a critical participant in tumor progression and therapeutic response, current technologies for tumor cell-to-cell interaction analyses are limited to qualitative and descriptive histological methods. Here, we utilized multiplex immunofluorescence (MxIF), RNA sequencing (RNA-seq), and microarray gene expression to quantitatively assess, using the BostonGene automated pipeline, the immune microenvironment composition of clinical stage 2 or 3 primary estrogen receptor-positive (ER+) HER2-negative (HER-) breast tumors (n = 29) from patients treated with neoadjuvant palbociclib in combination with anastrozole. Microenvironment characteristics of pretreatment tumor biopsies were associated with molecular subtype, cytokine signatures, and Ki67 response. MxIF analysis (21 markers, 2-13 ROIs per slide) of the tumor immune composition revealed that while the cellular composition was similar for each tumor region of the same patient, significant cellular heterogeneity was observed in tumor biopsies from different patients. The spatial heterogeneity was found in the distribution of both immune infiltrate cells and stromal components. T cells, CD68+ and CD163+ macrophages, and B cells were the predominant cell populations in the stromal component. Further MxIF analysis showed an increase in Ki67+ tumor cells in nonresponders (NR) versus responders (R). The number of immune cells was similar between Luminal A and B breast cancer molecular subtypes. A lower number of Ki67+ tumor cells was found in Luminal A compared to Luminal B tumors, as expected. MxIF analysis of cell-to-cell interactions revealed that distinct groups of cellular neighborhoods were significantly increased in NR compared with R: the immune inflammation cluster with a high density of CD4 T cells, Tregs, and CD8 T cells in the stromal component (p = 0.03); CD163+ macrophage-enriched cluster (p &amp;lt; 0.001); and tertiary lymphoid structures cluster (p &amp;lt; 0.001). Transcriptomic analysis showed that while CXCL1, CCL17, CCL13, CXCL9, and CCL5 expression correlated with the high density of CD4 and CD8 T cells in the stromal component cluster, VEGFA, IL1B, CXCL16, CCL3, and CCL7 were increased in the CD163+ and CD68+ macrophage-enriched clusters, indicating that differential cytokine expression is associated with the tumor architecture.Comprehensively characterizing, via MxIF, the immune microenvironment of ER+ HER2- breast cancer provided resolution of cellular architecture and elucidated spatial relationships among the immune and stromal cells of the tumor. Quantitatively assessing breast tumor cell-to-cell interactions has the potential to identify imaging markers of response, ultimately leading to the development of effective therapy options. Citation Format: Maria Bruttan, Souzan Sanati, Anna Belozerova, Ilia Galkin, Akshaya Ramachandran, Pavel Ovcharov, Grigory Perelman, Viktor Svekolkin, Ekaterina Postovalova, Alexander Bagaev, Krystle Nomie, Shana Thomas, Jeremy Hoog, Ravshan Ataullakhanov, James Hsieh, Cynthia Ma. Tumor microenvironment heterogeneity identifies potential biomarkers of response in ER-positive breast cancers treated with palbociclib [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6151.

Neoadjuvant survivin-targeted immunotherapy maveropepimut-S (MVP-S) to increase Th1 immune response in Ki67-high hormone receptor-positive (HR+) early-stage breast cancer (ESBC).

TPS1119 Background: HR+ ESBC is associated with suboptimal pathologic complete response rate (pCR, ̃10%) following neoadjuvant cytotoxic chemotherapy. Neoadjuvant anti-endocrine therapy with aromatase inhibitors (AI) may serve as an effective alternative. Efficacy can be gauged using the surrogate Ki67 cell proliferation histologic marker. Patients with poor Ki67 response (defined as Ki67 &gt; 10%) following neoadjuvant AI exhibit poor prognosis and therapeutic resistance to both anti-endocrine therapy and chemotherapy. In a genomic analysis among Ki67-high HR+ tumors, we identified 8-fold upregulation of BIRC5 (survivin), a gene that regulates apoptosis and the cell cycle and that is associated with poor clinical outcome. Maveropepimut-S (MVP-S, previously named DPX-Survivac) leverages the non-aqueous, lipid-based DPX delivery platform to educate a specific and persistent T cell-based immune response to 5 HLA-restricted peptides from Survivin, a cancer-associated protein commonly upregulated in several cancers. Treatment with MVP-S and intermittent, low-dose cyclophosphamide (CPA) has shown tumor infiltration of survivin-specific T cells. Previous clinical trials have shown that MVP-S is well-tolerated, immunogenic, and could lead to clinical response in several cancer indications. Further exploration of the regimen in breast cancer could extend the application of this immunotherapy for this unmet medical need. Methods: NCT04895761 is phase I trial evaluating the safety and immunologic effects of neoadjuvant MVP-S plus letrozole (arm A, n = 6), with/without tumor-directed MR-guided radiotherapy (arm B, n = 6), or intermittent low-dose cyclophosphamide or CPA (arm C, n = 6). Postmenopausal patients with T1c+ HR+HER2- breast cancer with Ki67 &gt; 10% will receive two doses of MVP-S and 7 weeks of neoadjuvant letrozole prior to surgery (all arms), whereas arm B will be treated additionally with concurrent 10Gy x 2 tumor boost radiation to facilitate immunogenic cell death, and arm C (n = 6) will be treated additionally with intermittent low-dose CPA (50mg BID) to facilitate regulatory T cell depletion. The primary objective is safety. Biomarker objectives are to evaluate for each treatment arm: 1) systemic type I survivin-specific immune response, as measured by IFN-γ ELISPOT; 2) changes in immune environment by GeoMx digital spatial genomic profiling; 3) and changes in tumor infiltrating lymphocytes (TILs) and Ki67. These data will be used to identify the most immunogenic MVP-S combination therapy for study in phase II trial powered to assess clinical outcome (pCR). Clinical trial information: NCT04895761.

LBA2 Assessment of Ki67 and FOXC1-based response predictor tracking proliferation and plasticity as a complementary diagnostic for neoadjuvant olaparib+paclitaxel+durvalumab in primary triple-negative breast cancer: Retrospective analysis of the I-SPY2 trial

Immune checkpoint inhibitors (ICIs) have shown clinical efficacy when administered in combination with neoadjuvant chemotherapy (NACT) for the treatment of triple negative breast cancer (TNBC). However, suitable complementary diagnostics to help guide and tailor treatment recommendations are still lacking. Ki67 is a well accepted and routinely used marker that tracks proliferation and has been shown to predict efficacy of neoadjuvant chemotherapy. Forkhead Box C1 (FOXC1), a transcriptional driver of cell plasticity/partial EMT/metastasis/immune evasion has proven prognostic value, but remains of uncertain predictive value.

Abstract PD15-02: HER2-enriched subtype and novel molecular subgroups drive aromatase inhibitor resistance and an increased risk of relapse in early ER+/HER2+ breast cancer

Abstract Background: Approximately 10% of estrogen receptor positive (ER+) early breast cancers (EBC) are human epidermal growth factor positive/overexpressed (HER2+). Resistance to aromatase inhibitors (AIs) is poorly understood among ER+HER2+ BC. We aimed to identify biomarkers of response and resistance using the HER2+ tumours in the POETIC trial. Methods: POETIC was a phase III trial of post-menopausal women with ER+ EBC randomized 2:1 to 2-weeks of peri-surgical AI vs control, followed by standard-of-care. Baseline formalin-fixed paraffin-embedded samples from 342 ER+HER2+ BC (237 treated/105 controls) were gene expression profiled using Breast cancer 360™ panel (NanoString) covering intrinsic subtypes and 40 key biological signatures. New molecular subgroups were identified using consensus clustering (CC).Proliferation rate was estimated as percentage of cancer-cells staining for Ki67. Ki67 response was defined by percentage-changes from baseline to surgery: poor (PR; reduction &amp;lt;50%), intermediate (IR; 50-75%) and good (GR; &amp;gt;75%). Residual Ki67 at 2 weeks timepoint (Ki672w) was defined as High (H) (≥ 10%) vs Low (L) (&amp;lt;10%). Association of molecular features with Ki67 response was assessed using logistic regression models and with time to recurrence (TTR) using multivariable Cox regression models adjusted for post-surgery clinicopathological variables: grade, tumour size, nodal status and age. Results: Ki672w was measured for all treated samples (n = 227, 48% H, 52% L) and a fraction of controls (n = 50, 96% H, 4% L). At baseline, 45% of tumours were HER2-enriched (HER2-E), 53% Luminal A/B and 2% Basal-Like. HER2-E associated with high Ki672W (84% H, 16% L) compared to non-HER2-E (29% H, 71% L, p&amp;lt;0.001). Amongst the 40 BC360 signatures, 9 were significantly associated with Ki67 response (Table). High expression of endocrine-related and apoptosis signatures associated with GR while HRD and TP53 with PR. These features were also related to Ki672W, with additional signatures involved in immune-checkpoint component and immune-enrichment (IM) (IDO1, PD-L1, IFN-gamma and T-reg) associated with high Ki672W.. Using CC, 4 molecular subgroups predicting differential response to AI were identified. Cluster (C) 1 was characterized by overexpression of IM features and low ER signaling (42%); C2 low IM but highest ERBB2 expression (15%); C3 ESR1 high and PgR low (4%) and C4 high endocrine signaling and lowest ERBB2 expression (39%). Distribution of Intrinsic subtypes varied across these molecular subgroups with the majority of C1 being HER2-E (62%) and Luminal B (27%); C2 HER2-E (74%); C3 Luminal B (64%) and C4 Luminal B (55%) and Luminal A (30%). HER2-E had significantly poorer TTR (HR 2.14; 95%CI1.11-4.17; p=0.0224) compared to Luminal tumours, remaining significant (HR 2.55, 95%CI1.14-5.69; p=0.0222) in the multivariable analysis. C2 (HR 4.67, 95%CI 1.90-11.51; p&amp;lt;0.001) and C4 (HR 2.42, 95%CI1.05-4.5.58; p=0.0385) were independent predictors of shorter TTR compared to C1, adding significant value beyond intrinsic subtypes (Likelihood ratio test, p=0.00429). Conclusions: Our results confirm that HER2-E, TP53 mutation, HRD and IM features are the main components driving poor early response to AI in ER+/HER2+ EBC. Novel molecular subgroups identify additional non-HER-E tumours not responding to AI with an increased risk of relapse. The appropriate additional treatment warrants further investigation. List of significant gene expression signatures associated with response as defined by reduction of Ki67. Odds ratio (OR) were determined by ordinal logistic regression analyses. OR: &amp;lt;1 indicating associated with good response and &amp;gt; 1.0 associated with worse response.Odds Ratiop-valuep-adjusted (False Discovery Rate)ESR10.65&amp;lt; 0.001&amp;lt; 0.001Estrogen Receptor Signaling0.44&amp;lt; 0.001&amp;lt; 0.001ERBB21.52&amp;lt; 0.001&amp;lt; 0.001TP53 mutant signature2.65&amp;lt; 0.001&amp;lt; 0.001FOXA10.42&amp;lt; 0.001&amp;lt; 0.001Apoptosis0.26&amp;lt; 0.0010.00228Homologous Recombination Deficiency (HRD)2.33&amp;lt; 0.0010.00349PgR0.820.001480.00737Hypoxia2.180.001660.00737 Citation Format: Milana A Bergamino Sirvén, Elena López-Knowles, Gabriele Morani, Holly Tovey, Lucy Kilburn, Chris Holcombe, Anthony Skene, Ian Smith, John Robertson, Gene Schuster, Judith M Bliss, Mitch Dowsett, Maggie CU Cheang, POETIC investigators. HER2-enriched subtype and novel molecular subgroups drive aromatase inhibitor resistance and an increased risk of relapse in early ER+/HER2+ breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD15-02.

Abstract PL1: Genomic Profiling in Early Stage ER Positive Breast Cancers/Precision Medicine

Abstract Genomic profiling has become mainstream for the management of early ER+ breast cancer and has become a favoured approach for research into pathways that govern prognosis and response and resistance to medical therapies.Management: Several commercial tests are available for the assessment of residual risk of recurrence of patients receiving hormonal therapies. We have compared several of these in postmenopausal patients treated with anastrozole or tamoxifen in the ATAC trial, including OncotypeDX (ODX), Prosigna (PAM50) and EndoPredict (EP). While PAM50 and EP provide estimates of risk that by design integrate tumour size and nodal status, estimates from ODX are also improved substantially by integration with clinicopathologic factors. The 3 tests include genes related to proliferation and to dependence on estrogen signalling with the ODX being dominated by the latter and PAM50 by the former. This may explain the better performance of the PAM50 than ODX at cessation of endocrine treatment after 5 years when the outcome for patients with high ER appears to worsen. While the tests have largely been validated in postmenopausal cases they are used in premenopausal cases as well. We found that while individual gene modules varied in expression across the menstrual cycle there was little systemic variability in the overall estimates of risk. Our recent work has indicated that mutations in TP53 add significant prognostic information to that from the commercial profiling tests.Research: We have conducted genomic analyses during presurgical therapy of ER+ breast cancer to determine the key pathways of endocrine resistance using change in Ki67 as the end-point of response. Very few tumours with low levels of ER by IHC (&amp;lt;10% positive cells) or rtPCR showed a Ki67 response. A group of the most widely studied genes in breast cancer including FOS and JUN show very marked increases in expression during surgical procedures and can lead to erroneous interpretation in the absence of controls. Considering intrinsic subtypes, tumours categorised as basal-like or HER2-enriched have little chance of Ki67 response; luminal B tumours showed a slightly lower chance of response. GSEA revealed that several hallmarks reflecting immune signalling were those most strongly associated poor Ki67 response with the interferon gamma and alpha and IL6-JAK-STAT3 signalling hallmarks being particularly involved. TME deconvolution indicated a broad involvement of many immune cell types. Other hallmarks of resistance included E2F target genes and hypoxia. Mutation of TP53 was strongly associated with Ki67 poor response. RNAseq profiling of patients treated with palbociclb as well as letrozole revealed high CCNE1 expression as associated with endocrine resistance. Citation Format: M Dowsett. Genomic Profiling in Early Stage ER Positive Breast Cancers/Precision Medicine [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PL1.

Abstract P2-13-08: Combined peri-operative lapatinib and trastuzumab in early HER2-positive breast cancer - Long term results of the randomized UK EPHOS-B trial

Abstract Background: EPHOS-B is a multi-centre randomized trial designed to investigate whether anti-HER2 therapy given 11 days pre-surgery inhibited proliferation and/or increased apoptosis in HER2-positive early breast cancer. Significant differences in Ki67 response (≥30% change between baseline and surgery) were observed between treatment groups and control, although no significant increases in apoptosis were seen [1]. We report here 5-year outcomes and their association with peri-operative response, including post-hoc exploratory analyses on stromal tumour infiltrating lymphocytes (TILs). Methods: Patients were randomized (1:2:2) to no treatment (control), trastuzumab or lapatinib (Part-1), and (1:1:2) to control, trastuzumab, or lapatinib and trastuzumab combination (Part-2). Relapse free survival (RFS) is defined as the time from randomisation to local, regional, or distant tumour recurrence or death from any cause, with second primary cancers censored. Five-year RFS rates were estimated by Kaplan-Meier across treatment and peri-operative response groups, and compared by Log-Rank tests. Central scoring of stromal tumour infiltrating lymphocytes (TILs) on scanned H&amp;E baseline and surgery slides was conducted post-hoc according to the International TILs Working Group. Baseline TILs (bTILS) were explored across treatment groups, and associated with trial outcomes. Changes in TILS in patients without pathological regression were correlated with Ki67 changes and RFS. Results: Overall, 257 patients were randomized (Part-1: control 22, trastuzumab 57, lapatinib 51; Part-2: control 29, trastuzumab 32, trastuzumab+lapatinib 66). Central pathology review identified six patients achieving complete pathological response (pCR), and 13 RCB1 (as per the Residual Cancer Burden score), all but two in the combination group. After 6 years median follow-up, 28 women (11%) had recurrence, and 19 (7%) died; no differences between groups were found (Table). No recurrences were observed amongst pCR patients; only 1 local recurrence observed amongst RCB1. Patients with Ki67 falls ≥50% had better RFS than those with modest falls or no falls (Table). In patients with no evidence of disease regression at surgery, those with higher TILs (≥20%) at surgery had better RFS than those with lower TILs. Conclusions: Early response (pCR/RCB1 or Ki67 reductions &amp;gt;50%) after 11 days pre-operative anti-HER2 dual therapy identifies cancers dependent on the HER2 pathway and provides a strategy for individualising treatment, including de-escalation of therapy. [1] Bundred N, Cameron D, Amstrong A et al (2016) Effects of perioperative lapatinib and trastuzumab, alone and in combination, in early HER2+ breast cancer − the UK EPHOS-B trial (CRUK/08/002) European Journal of Cancer;57:S1-S8. Table - Relapse-free survival by treatment and by peri-operative responseN randomisedNRFS event5-year RFS95%CILog-rank p-valueRandomised treatmentTrastuzumab (P1)577 (12.3%)88%76-94%T vs L p=0.75Lapatinib (P1)515 (9.8%)90%77-96%L vs C p=0.41Control (P1)221 (4.6%)95%92-99%Trastuzumab (P2)327 (21.9%)87%69-95%T vs T+L p=0.048Combination (P2)665 (7.6%)92%83-97%T+L vs C p=0.64Control (P2)293 (10.3%)90%71-97%%Ki67 change:&amp;gt;50% decrease722 (2.8%)99%90-100%0.002&amp;gt;10% decrease7717 (22.1%)80%69-88%&amp;lt;10% or no decrease827 (8.5%)91%83-96%Disease responseRCB060100%-0.63RCB1131 (7.7%)92%57-99RCB2/3/no evidence of regression23626 (11%)90%85-93Baseline TILS&amp;lt;=20%18023 (13%)89%83-930.10&amp;gt;20%502 (4%)96%85-99Surgery TILS (only RCB2/3)&amp;lt;=20%12220 (16.4%)86%78-910.021&amp;gt;20%653 (4.6%)95%86-98Change in TILS (only RCB2/3)No significant increase15221 (13.8%)88%81-920.16Increase &amp;gt;20%352 (5.7%)94%79-99 Citation Format: Nigel Bundred, Nuria Porta, Adrian Murray Brunt, Angela Cramer, Andrew Hanby, Abeer Shaaban, Emad Rakha, Anne Armstrong, Ramsey Cutress, David Dodwell, Marie Emson, Abigail Evans, Sue Hartup, Kieran Horgan, Stuart McIntosh, Jay Naik, Samkaran Narayanan, Jane Oii, Anthony Skene, David Cameron, Judith Bliss. Combined peri-operative lapatinib and trastuzumab in early HER2-positive breast cancer - Long term results of the randomized UK EPHOS-B trial [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-13-08.

Abstract PD15-06: Estrogen receptor expression thresholds by IHC and mRNA for Ki67 response to aromatase inhibition: A POETIC study

Abstract Background. Estrogen receptor (ER) expression is the key determinant for endocrine treatment of breast cancer (BC). In clinical practise ER analysis is currently almost entirely by immunohistochemistry (IHC). ASCO/CAP recommends cut-offs of &amp;lt;1% (negative) and 1-10% (low) to minimise the risk of false negatives. There is uncertainty in the level of responsiveness of those with low ER. mRNA assessment could be acceptable or preferable if a similar or better threshold was shown to identify response to endocrine therapies. Change of Ki67 after 2 weeks’ presurgical treatment with aromatase inhibitors (AIs) relates to reduction in risk of recurrence. We aimed to define IHC and mRNA cut-points for predicting change in Ki67 in response to AI. Method. POETIC is a phase III trial in postmenopausal patients with ER+ (by local assessment) BC randomized to receive 2-weeks’ pre-surgical AI vs no presurgical treatment. Cases were selected from the AI treatment group. All HER2+ cases were analysed. To enrich the HER2- study set for lower ER values, we selected all of the 15% poorest Ki67 suppression (ie PR, &amp;lt;40% suppression) and 30% of the remainder categorised as intermediate (IR, 40-79%) and good-responders (GR, &amp;gt;79%). 770 baseline FFPE samples were analysed (582 HER2-, 188 HER2+). IHC used the NCL-L-ER6F11 (Novocastra, Leica) antibody with the DAKO Flex kit and were scored centrally. mRNA was assessed by rtPCR. Values were expressed according to an arbitrary scale (“A units”). Results. ER IHC was available from 517 HER2- and 188 HER2+ tumors and ER mRNA from 376 HER2- and 173 HER2+ tumors. The correlation between ER levels by IHC and rtPCR was moderately high (Rho=0.616, p&amp;lt;0.00001). In IHC &amp;lt;1%, 1-10% and &amp;gt;10% tumors median (range) mRNA levels were 1.290 (0.188-11.346), 2.368 (0.575-13.863) and 111.150 (0.826-1001.423), respectively. The tables show the %age of PR, IR and GR for each category of ER by IHC or mRNA. The number of cases was adjusted to cater for the analysis of only 30% of the HER2- IR and GR , by multiplying the HER2- IR and GR number of cases by 3.3, resulting in this not being an integer for some categories. IHC: 87% of cases had &amp;gt;60% of cells +ve and only 11-12% of these were PRs. For cases with &amp;lt;10% cells +ve (4.9% of the population) over 90% of cases were PRs and there were no GRs. Cases between 1 and 10% +ve were not less responsive than those &amp;lt;1% +ve. For cases with &amp;gt;10% &amp;lt;20% cells +ve there was a substantial number of GRs and IRs (14.6% and 19.0%, respectively). mRNA: 82% of cases had ≥ 40 A units and only 10-14% of these showed PR. For cases with 5 A units (4.2% of the population) there were 9.5% % IR and no GR. In the next category, ≥5 &amp;lt;10 A units, 34.4% of cases were IR or GR. The levels of ER that distinguished PRs from IR/GRs was no different between HER2+ and HER2- cases. Conclusions. Ki67 responsiveness increases with increasing levels of ER by both IHC and mRNA. There was little responsiveness at IHC&amp;lt;10% and no distinction in responsiveness between &amp;lt;1% and 1-10% cells positive. Above 10% positive cells substantial numbers of patients showed IR or GR. Similar separation of PRs from IR/GRs was achieved by IHC and mRNA. IHC (%)0 &amp;lt; 1≥1 &amp;lt; 10≥10 &amp;lt; 20≥20 &amp;lt; 40≥40 &amp;lt; 60≥60 &amp;lt; 80≥80No. of cases42.32922.625.671.7239.81010.6PR87.596.566.462.541.811.710.8IR12.53.419.020.741.640.933.1GR0.00.014.616.816.647.456.1 mRNA (A units)&amp;lt;5≥5 &amp;lt; 10≥10 &amp;lt; 20≥20 &amp;lt; 40≥40 &amp;lt; 80≥80 &amp;lt; 120≥120No. of cases45.318.336.589.2168.1177.6550.4PR90.565.638.426.913.710.710.7IR9.416.458.947.946.835.226.8GR0.018.02.725.239.554.162.5 Citation Format: Elena Lopez Knowles, Simone Detre, Margaret Hills, Gene F Schuster, Maggie CU Cheang, Holly Tovey, Lucy Kilburn, Judith Bliss, John Robertson, Ian Smith, Mitch Dowsett, POETIC investigators. Estrogen receptor expression thresholds by IHC and mRNA for Ki67 response to aromatase inhibition: A POETIC study [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD15-06.

Relationship between ER expression by IHC or mRNA with Ki67 response to aromatase inhibition: a POETIC study

BackgroundIn clinical practice, oestrogen receptor (ER) analysis is almost entirely by immunohistochemistry (IHC). ASCO/CAP recommends cut-offs of < 1% (negative) and 1–10% (low) cells positive. There is uncertainty whether patients with ER low tumours benefit from endocrine therapy. We aimed to assess IHC and mRNA cut-points for ER versus biological response of primary breast cancer to 2 weeks’ aromatase inhibitor treatment as measured by change in Ki67.MethodsCases were selected from the aromatase inhibitor treatment group of POETIC. We selected the 15% with the poorest Ki67 response (PR, < 40% Ki67 suppression, n = 230) and a random 30% of the remainder categorised as intermediate (IR, 40–79% Ki67 suppression, n = 150) and good-responders (GR, ≥ 80% Ki67 suppression, n = 230) from HER2 − group. All HER2 + cases available were selected irrespective of their response category (n = 317). ER expression was measured by IHC and qPCR.ResultsER IHC was available from 515 HER2 − and 186 HER2 + tumours and ER qPCR from 367 HER2 − and 171 HER2 + tumours. Ninety-one percentage of patients with ER IHC < 10% were PRs with similar rates in HER2 − and HER2 + cases. At or above ER IHC 10% substantial numbers of patients showed IR or GR. Similar proportions of patients were defined by cut-points of ER IHC < 10% and ER mRNA < 5 units. In addition, loss of PgR expression altered ER anti-proliferation response with 92% of PgR − cases with ER IHC < 40% being PRs.ConclusionsThere was little responsiveness at IHC < 10% and no distinction between < 1% and 1–10% cells positive. Similar separation of PRs from IR/GRs was achieved by IHC and mRNA.

Biomarkers of Response and Resistance to Palbociclib Plus Letrozole in Patients With ER+/HER2- Breast Cancer.

To determine (i) the relationship between candidate biomarkers of the antiproliferative (Ki67) response to letrozole and palbociclib alone and combined in ER+/HER2- breast cancer; and (ii) the pharmacodynamic effect of the agents on the biomarkers. 307 postmenopausal women with ER+/HER2- primary breast cancer were randomly assigned to neoadjuvant treatment with letrozole for 14 weeks; letrozole for 2 weeks, then letrozole+palbociclib to 14 weeks; palbociclib for 2 weeks, then letrozole+palbociclib to 14 weeks; or letrozole+palbociclib for 14 weeks. Biopsies were taken at baseline, 2 and 14 weeks and surgery at varying times after stopping palbociclib. Immunohistochemical analyses were conducted for Ki67, c-PARP, ER, PgR, RB1, CCNE1, and CCND1. Higher baselines ER and PgR were significantly associated with a greater chance of complete cell-cycle arrest (CCCA: Ki67 <2.7%) at 14 weeks and higher baseline Ki67, c-PARP, and CCNE1 with a lower chance. The interaction with treatment was significant only for c-PARP. CCND1 levels were decreased c.20% by letrozole at 2 and 14 weeks but showed a tendency to increase with palbociclib. CCNE1 levels fell 82% (median) in tumors showing CCCA but were unchanged in those with no CCCA. Only 2/9 tumors showed CCCA 3-9 days after stopping palbociclib. ESR1 mutations were found in 2/4 tumors for which surgery took place ≥6 months after starting treatment. High CCNE1 levels were confirmed as a biomarker of resistance to letrozole+palbociclib. Ki67 recovery within 3-9 days of discontinuing palbociclib indicates incomplete suppression of proliferation during the "off" week of its schedule.

Top 3 abstracts concerning hormone-receptor-positive early breast cancer

SummaryThe three top abstracts at the 2020 virtual San Antonio Breast Cancer Symposium regarding hormone-receptor-positive early breast cancer, from our point of view, were the long-awaited results from PenelopeBand RxPONDER as well as the data from the ADAPT trial of the West German Study Group. PenelopeBfailed to show any benefit by adjuvant palbociclib when added to standard endocrine therapy in patients without pathologic complete response after neoadjuvant chemotherapy. RxPONDER demonstrated that postmenopausal patients with early hormone receptor positive (HR+)/human epidermal growth factor receptor 2 negative (HER2−) breast cancer, 1–3 positive lymph nodes and an Oncotype DX Recurrence Score of less than 26 can safely be treated with endocrine therapy alone. In contrast, in premenopausal women with positive nodes, adjuvant chemotherapy plays still a role even in case of low genomic risk. Whether the benefit by chemotherapy is mainly an indirect endocrine effect and if ovarian function suppression would be similarly effective, is still a matter of debate. The HR+/HER2− part of the ADAPT umbrella trial investigated the role of a Ki-67 response to a short endocrine therapy before surgery in addition to Oncotype DX—performed on the pretreatment biopsy—to identify low-risk patients who can safely forgo adjuvant chemotherapy irrespective of menopausal status.

Ki-67 and Its Relation With Complete Pathological Response in Patients With Breast Cancer.

IntroductionKi-67 is a nuclear antigen present in the synthesis phase of the cell cycle. Studies have shown that a high value of Ki-67 results in greater response to chemotherapy with higher incidence of complete pathological response, which ultimately results in improved overall survival.Methods and materialsThe objective of the study was to determine the frequency of high Ki-67 levels in breast cancer patients and to find the correlation of complete pathological response in breast cancer with Ki-67 levels. It is a descriptive case series with a correlational study design done at Fauji Foundation Hospital Rawalpindi. Eighty patients with locally advanced breast cancer who underwent neoadjuvant chemotherapy followed by surgery were recruited. Their Ki-67 levels were determined on trucut biopsy. Pathological response in the post-op sample was correlated with Ki-67 levels.ResultsThe results showed 27 (33%) patients out of the 80 had high Ki-67 values. Among them 17 (63%) had complete pathological response, seven (26%) showed partial pathological response whereas three (11%) had disease progression. In contrast, out of the 53 patients having low Ki-67 values, only nine (17%) had complete pathological response, 31 (58%) showed partial pathological response and 13 (25%) had progressive disease. A Chi-square test was applied which showed significant correlation between Ki-67 and complete pathological response, with a p value of 0.00018.ConclusionTherefore high Ki-67 values in patients with breast cancer correlated well with attainment of complete pathological response. We can incorporate Ki-67 in the initial clinical assessment of breast cancer patients to help predict effectiveness as well as response to chemotherapy.