Cimino‐Mathews, Ashley

PURPOSE Predictive biomarkers to better tailor therapy for patients with early-stage human epidermal growth factor 2 (HER2)-positive breast cancer are a priority. We hypothesized that HER2 and immune-based biomarkers would be predictive of pathologic complete response (pCR) to preoperative trastuzumab/pertuzumab (HP). MATERIALS AND METHODS Patients with stage II/III, estrogen receptor (ER)-negative, HER2-positive breast cancer received neoadjuvant HP in the TBCRC026 clinical trial. The pCR after receiving HP alone was 22% (18/83). Tumor biopsies were performed at baseline. Secondary correlative objectives were to determine the relationship between HER2-based biomarkers and immune processes with pCR. NanoString code sets BC360 and IO360 were used to compare differential gene expression in baseline tumors that underwent pCR versus no pCR. NanoString GeoMx digital spatial profiling was used to assess immune protein abundance in intraepithelial and stromal segments. Stromal tumor-infiltrating lymphocytes and Ki67 were evaluated by hematoxylin and eosin and immunohistochemistry, respectively. RESULTS Intraepithelial HER2 protein abundance was significantly associated with pCR ( P = .001). Low HER2 abundance tumors were primarily basal-like, and essentially all (19/20) failed to achieve pCR. High HER2 abundance tumors that achieved pCR (14/51) exhibited a high degree of immune cell activity, whereas high HER2 abundance tumors that failed to achieve pCR tumors (37/51) were enriched for M-phase processes and epidermal growth factor receptor signaling. Baseline Ki67 was significantly higher in nonresponders ( P = .04). CONCLUSION ER-negative, HER2-positive breast cancer has unique molecular and immunologic features that may predict pCR after neoadjuvant HP. Validation of these potential biomarkers and composite biomarker analyses may guide design of future clinical trials.

530 Background: Robust biomarkers are needed to personalize treatment in early stage HER2-positive breast cancer. The TBCRC026 trial (n=83) demonstrated that early metabolic changes on FDG-PET/CT [>40% decline in SULmax at day 15 post treatment initiation (C1D15); C1D15 SULmax] were associated with response to neoadjuvant trastuzumab/pertuzumab (HP), without chemotherapy, in an ER-negative, HER2-positive cohort (NCT01937117). We investigated the association of RNA sequencing (RNAseq)-based gene expression signatures (GES) at baseline and on-treatment (C1D15), with early metabolic change on PET/CT and clinical outcomes. Methods: RNAseq (664 GES) was performed on baseline (n=56) and C1D15 (n=31) tumor biopsies (25 paired samples available). Association with pCR following HP (secondary), PET/CT parameters and survival (exploratory) was studied. Gene expression changes between baseline and C1D15 samples were tested by Wilcoxon signed-rank test. Association between GES and PET/CT parameters and GES with pCR were evaluated using logistic regression models. Cox regressions were performed to study the association between PET/CT SULmax at C1D15 and a B cell signature and relapse-free survival (RFS). The likelihood ratio test was used to compare the prognostic ability of two nested Cox models. Results: Both RNAseq and clinical outcomes were available in 53/83 patients. HER2-enriched (HER2-E) intrinsic subtype comprised 71% (40/56) of cases at baseline and 61% (19/31) at D15; basal subtype for 27% (15/56) and 39% (12/31), respectively. pCR rate was 19% (10/53) with all but one exhibiting HER2-E subtype. One basal-like sample underwent pCR, with downregulation of HER2 amplicon and IgG signatures slightly more prominent. Gene expression changes from baseline to C1D15 were analyzed; ERBB2was downregulated after HP (p = 0.007), B cell signatures, CD4, CD8, natural killer cells and Treg cells were significantly upregulated at C1D15. GES associated with 40% decline in SULmax at C1D15 included ERBB2 amplicon genes ( ERBB2, GRB7) and correlation to the HER2-E subtype, whereas basal-like signatures were significantly associated with lack of response on PET at C1D15. Univariate logistic regression analysis suggested that FOS-JUN, fibroblast and B cell GES at baseline were significantly associated with pCR (p=<0.05). Finally, the addition of a B cell signature improved the prognostic value of C1D15 SULmax on PET/CT for RFS (p < 0.05). Conclusions: In this unique cohort with ER-negative, HER2-positive breast cancer treated with neoadjuvant HP without chemotherapy, HER2-E tumors and those with increased immune signaling were more likely to respond to HER2-directed therapy. Combining C1D15 SULmax on PET/CT and a B cell signature provided improved prognostic information and should be further explored as candidate biomarkers in this setting. Clinical trial information: NCT01937117 .

PURPOSE Predictive biomarkers to identify patients with human epidermal growth factor receptor 2 (HER2)–positive breast cancer who may benefit from targeted therapy alone are required. We hypothesized that early measurements of tumor maximum standardized uptake value corrected for lean body mass (SULmax) on 18 F-labeled fluorodeoxyglucose positron emission tomography-computed tomography (PET-CT) would predict pathologic complete response (pCR) to pertuzumab and trastuzumab (PT). PATIENTS AND METHODS Patients with stage II or III, estrogen receptor–negative, HER2-positive breast cancer received four cycles of neoadjuvant PT. 18 F-labeled fluorodeoxyglucose positron emission tomography-computed tomography was performed at baseline and 15 days after PT initiation (C1D15). Eighty evaluable patients were required to test the null hypothesis that the area under the curve of percent change in SULmax by C1D15 predicting pCR is ≤ 0.65, with a one-sided type I error rate of 10%. RESULTS Eighty-eight women were enrolled (83 evaluable), and 85% (75 of 88) completed all four cycles of PT. pCR after PT alone was 22%. Receiver operator characteristic analysis of percent change in SULmax by C1D15 yielded an area under the curve of 0.72 (80% CI, 0.64 to 0.80; one-sided P = .12), which did not reject the null hypothesis. However, between patients who obtained pCR and who did not, a significant difference in median percent reduction in SULmax by C1D15 was observed (63.8% v 41.8%; P = .004) and SULmax reduction ≥ 40% was more prevalent (83% v 52%; P = .03; positive predictive value, 31%). Participants not obtaining a 40% reduction in SULmax by C1D15 were unlikely to obtain pCR (negative predictive value, 91%). CONCLUSION Although the primary objective was not met, early changes in SULmax predict response to PT in estrogen receptor–negative and HER2-positive breast cancer. Once optimized, this quantitative imaging strategy may facilitate tailoring of therapy in this setting.

Human epidermal growth factor receptor 2 (HER-2) targeted therapy shows promising results in HER-2-positive uterine serous carcinoma (USC). HER-2 scoring criteria for USC and its associated noninvasive lesion, serous endometrial intraepithelial carcinoma (SEIC), are not well-established. Here, we compare the breast and gastric (GI) HER-2 immunohistochemistry (IHC) scoring criteria for HER-2 with HER-2/neu fluorescence in situ hybridization (FISH) in 68 tumors (17 USC with SEIC, 30 USC, 18 SEIC, 3 metastatic USC). The majority (97%) of lesions displayed intratumoral HER-2 IHC heterogeneity. Breast or GI IHC scoring criteria were performed equivalently. The breast and GI IHC criteria classified 51% and 47% USC as HER-2 negative (IHC 0/1+), 40% and 45% as equivocal (IHC 2+), and 9% each as HER-2 positive (IHC 3+). A quarter of USC classified as HER-2 negative or positive with the breast (25%, n=7/28) or GI IHC criteria (23%, n=6/26) was discordant by FISH. Specifically, 13% to 14% of IHC 0/1+ USC were FISH amplified; 50% of IHC 3+ USC were FISH negative. The majority (77% to 83%) of SEIC were HER-2 IHC 0/1+, and no SEIC was HER-2 IHC 3+. A minority (4% to 7%) of IHC 0/1+ SEIC were FISH positive. Discordant HER-2 status was observed between half (47%,bn=7/15) of synchronous SEIC and USC. In conclusion, USC displays HER-2 intratumoral heterogeneity, a high IHC/FISH discordance rate, and variation in HER-2 status between the SEIC and invasive components. Caution is required when evaluating HER-2 in small biopsies, which should be repeated on excisions. Both IHC and FISH should be performed on USC until clinical trials correlate HER-2 status with clinical response to HER-2-targeted therapy.

AbstractImmune checkpoint inhibitor therapies targeting PD‐1/PD‐L1 are now the standard of care in oncology across several hematologic and solid tumor types, including triple negative breast cancer (TNBC). Patients with metastatic or locally advanced TNBC with PD‐L1 expression on immune cells occupying ≥1% of tumor area demonstrated survival benefit with the addition of atezolizumab to nab‐paclitaxel. However, concerns regarding variability between immunohistochemical PD‐L1 assay performance and inter‐reader reproducibility have been raised. High tumor‐infiltrating lymphocytes (TILs) have also been associated with response to PD‐1/PD‐L1 inhibitors in patients with breast cancer (BC). TILs can be easily assessed on hematoxylin and eosin–stained slides and have shown reliable inter‐reader reproducibility. As an established prognostic factor in early stage TNBC, TILs are soon anticipated to be reported in daily practice in many pathology laboratories worldwide. Because TILs and PD‐L1 are parts of an immunological spectrum in BC, we propose the systematic implementation of combined PD‐L1 and TIL analyses as a more comprehensive immuno‐oncological biomarker for patient selection for PD‐1/PD‐L1 inhibition‐based therapy in patients with BC. Although practical and regulatory considerations differ by jurisdiction, the pathology community has the responsibility to patients to implement assays that lead to optimal patient selection. We propose herewith a risk‐management framework that may help mitigate the risks of suboptimal patient selection for immuno‐therapeutic approaches in clinical trials and daily practice based on combined TILs/PD‐L1 assessment in BC. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

PURPOSE Predictive biomarkers to identify patients with human epidermal growth factor receptor 2 (HER2)–positive breast cancer who may benefit from targeted therapy alone are required. We hypothesized that early measurements of tumor maximum standardized uptake values corrected for lean body mass (SULmax) on [ 18 F]fluorodeoxyglucose positron emission tomography/computed tomography would predict pathologic complete response (pCR) to neoadjuvant pertuzumab and trastuzumab (PT). PATIENTS AND METHODS Patients with stage II/III, estrogen receptor–negative, HER2-positive breast cancer received four cycles of neoadjuvant PT. [ 18 F]Fluorodeoxyglucose positron emission tomography/computed tomography was performed at baseline and 15 days after PT initiation (C1D15). Eighty evaluable patients were required to test the null hypothesis that the area under the curve of percentage of change in SULmax by C1D15 predicting pCR is less than or equal to 0.65, with a one-sided type I error rate of 10%. RESULTS Eighty-eight women were enrolled (83 evaluable), and 85% (75 of 88) completed all four cycles of PT. pCR after PT alone was 34%. Receiver operating characteristic analysis yielded an area under the curve of 0.76 (90% CI, 0.67 to 0.85), which rejected the null hypothesis. Between patients who obtained pCR versus not, a significant difference in median percent reduction in SULmax by C1D15 was observed (63.8% v 33.5%; P < .001), an SULmax reduction greater than or equal to 40% was more prevalent (86% v 46%; P < .001; negative predictive value, 88%; positive predictive value, 49%), and a significant difference in median C1D15 SULmax (1.6 v 3.9; P < .001) and higher proportion of C1D15 SULmax less than or equal to 3 (93% v 38%; P < .001; negative predictive value, 94%; positive predictive value, 55%) were observed. CONCLUSION Early changes in SULmax predict response to four cycles of PT in estrogen receptor–negative, HER2-positive breast cancer. Once optimized, this quantitative imaging strategy may facilitate a more tailored approach to therapy in this setting.