Li, Zhuo

Background: The Clinical Treatment Score post-5 years (CTS5) is a risk stratification tool used to determine the risk of late recurrence in hormone receptor–positive (HR+), HER2-negative breast cancer (BC). Limited data exist on its use in HR+, HER2-positive (HER2+) BC. Patients and Methods: CTS5 was evaluated in HR+, HER2+ BC in the North Central Cancer Treatment Group (NCCTG) N9831 (Alliance) and NSABP B-31 (NRG) trials. Results: A total of 1,862 patients with HR+, HER2+ BC without recurrence 5 years after enrollment were included. Overall, the CTS5 score was significantly associated with recurrence-free survival (RFS), with a hazard ratio (HR) of 1.35 (95% CI, 1.12–1.63; P=.002), but did not reach statistical significance in patients who received trastuzumab (n=829; HR, 1.29; 95% CI, 0.98–1.71; P=.07). CTS5 risk category was not significantly associated with RFS. In patients who received trastuzumab, other variables used in CTS5, including patient age and tumor size, were not significantly associated with RFS. N3 was significantly associated with worse outcomes (HR, 1.86; 95% CI, 1.09–3.17; P=.02) compared with N0–N1. Paradoxically, higher tumor grade was associated with better outcomes after 5 years in the multivariate analysis (HR, 0.71; 95% CI, 0.50–1.00; P=.05). The incidence of recurrences or deaths between years 5 to 10 was 10.6% in the CTS5 low-risk category, 5.6% in the intermediate-risk category, and 9.8% in the high-risk category. Conclusions: The CTS5 model does not accurately predict the risk of late recurrence in HR+, HER2+ BC treated with adjuvant trastuzumab in the N9831 and B-31 trials. This study underlines the need to develop a new prognostic model to better delineate the risk of late recurrence in patients with HR+, HER2+ BC receiving adjuvant trastuzumab. ClinicalTrials.gov identifiers: NCT00005970 (NCCTG N9831) and NCT00004067 (NRG/NSABP B-31).

Abstract Background: Tertiary lymphoid structure (TLS) is an organized form of ectopic lymphoid aggregates (LA) with secondary lymphoid organ structure. Several studies showed that the presence of TLS associates with improved outcomes in multiple cancers when treated with immune checkpoint inhibitors. However, data is currently limited regarding TLS and outcomes in HER2+ breast cancer patients (pts) treated with adjuvant trastuzumab. Furthermore, distinguishing TLS and simple LA is challenging in standard hematoxylin and eosin (H&E) staining, particularly when the germinal center is absent. Emerging studies also showed the prognostic value of MHC expression in breast cancer but mainly in triple-negative breast cancer. In this study, we evaluated integrated pathological quantification and genomic data to assess functional TLS in association with MHC expression and outcomes in the N9831 trial. Methods: Pathological evaluation of LA in H&E slides from pts treated in Arm A (chemotherapy alone) and Arm C (chemotherapy with concurrent trastuzumab) in N9831 was performed. NanoString was used to quantify mRNA of MHC class I and II expression as well as TLS-related immune genes, including IFNG, ICOSLG, CXCL13, CXCR3, BCL6, IL21R, ICOS, PDCD1, CXCR5, CXCL9, TBX21, CD38, CXCL10, CLXCL11, IL21, CD200, CD19, MS4S1. Wilcoxon rank sum test, Chi-squared test, Kaplan-Meier method, and Cox regression model were used to evaluate the association between LA and baseline characteristics, MHC expression, and outcomes. Results: LA was evaluated in 526 pts in Arm A and 485 in Arm C. Greater number of LA was significantly associated with stromal tumor-infiltrating lymphocytes (sTILs), hormone receptor negative, and higher tumor grade, but not stage and age. Using multivariable Cox regression analysis, increasing numbers of LA were associated with improved recurrence-free survival in both arms combined (RFS, p 0.028). However, when evaluating each arm separately, LA was associated with improved RFS only in Arm A (HR 0.6, 95%CI 0.43-0.84, p 0.003) but not in Arm C (HR 0.72, 95%CI 0.47-1.1, p 0.134). We further evaluated TLS-related immune genes in 252 pts in Arm C with LA ≥ 1. As a continuous variable, higher expression of BCL6 (HR 0.61, 95%CI 0.41-0.92, p 0.019) and IL21R (HR 0.78, 95%CI 0.62-0.98, p 0.03) were associated with improved RFS in Arm C pts with LA ≥ 1. However, these genes were not significantly associated with outcomes in Arm C pts without LA with BCL6 (HR1.07, 95%CI 0.67-1.71, p 0.776) and IL21R (HR 0.96, 95%CI 0.71-1.29, p 0.775). Moreover, we evaluated differential gene expression between tumors with LA ≥ 1 with high BCL6 vs. LA 0. All MHC class I and II (HLA A, B, C, E, DQ, DM, DO, DP, and DQ) were significantly higher in tumors with LA ≥ 1 with high BCL6 (p < 0.001). Conclusion: A greater number of LA was associated with improved outcomes in pts with early-stage HER2+ breast cancer, particularly when treated with chemotherapy alone. Using histologic and genomic integration with the combination of pathological LA and TLS-related immune genes, we identified that pts with functional TLS with LA ≥ 1 and higher expression of BCL6 or IL21R had significantly improved outcomes when treated with trastuzumab. High MHC class I and II expressions are associated with the presence of functional TLS, underscoring the crucial role of antigen presentation in the generation of an effective adaptive immune response. Support: U10 CA180821, U10 CA180882, U24 CA196171, https://acknowledgments.alliancefound.org; Genentech; Clinicaltrials.gov Id: NCT00005970 Citation Format: Saranya Chumsri, Tracy Shachner, Zhuo Li, Nadine Norton, Alvaro Moreno-Aspitia, Gerardo Colon-Otero, Edith Perez, Aziza Nassar, E. Thompson, Keith Knutson. MHC Class I and II Expression, Functional Tertiary Lymphoid Structure (TLS), and Outcomes in Early-Stage HER2-Positive (HER2+) Breast Cancer in NCCTG N9831 (Alliance) [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 PO4-25-04.

Abstract Background The current definition, prognosis, and optimal treatment (tx) for oligometastatic breast cancer (OMBC) are not fully known. With advances in multimodality breast cancer (BC) tx and overall improvement in patient (pt) outcomes, it is important to identify baseline pt factors that confer better prognosis in OMBC and assess the impact of local/regional tx and metastasis (mets)-directed tx on survival outcomes. Methods We reviewed 105 Mayo Clinic pts with OMBC (up to 5 mets) from 2003 to 2021. Pts were excluded from analysis if they were misidentified as having OMBC (n=6), did not have a primary breast mass (n=1), developed de novo BC during tx (n=1), or were lost to follow up (n=2). Categorical variables were summarized as counts. Continuous variables were reported as medians. Kaplan-Meier method was used to estimate survival and the time from diagnosis to next tx at 1, 3, and 5 years. Log-rank test was used to compare survival rates between baseline factors. Univariate Cox proportional hazards models were performed on both baseline and time-dependent factors. All tests were two-sided with p-value < 0.05 considered statistically significant. Results Total pts included were 95. Median age was 49 (range, 26-86), most (93.6%) pts were White, and 46.8% were postmenopausal. Invasive ductal carcinoma (89.5%) was the most common BC type. Median survival was 10.8 years; 1-, 3-, and 5-year survival rates were 98.9%, 87.7%, and 81%, respectively. 58 pts (61.1%) required a change in tx due to disease progression. No significant survival difference was observed in pre- vs. postmenopausal pts (p=0.71) or in groups based on hormone receptor and/or human epidermal growth factor receptor 2 status. 47 pts (49.5%) received local/regional tx; no significant difference in survival (HR 0.57, 95% CI 0.23-1.40; p=0.217) or time to next tx (HR 0.67, 95% CI 0.39-1.15; p=0.144) was seen in this subgroup. Of these pts, 36 (76.6%) received neoadjuvant chemotherapy and/or immunotherapy, 5 of whom (10.6%) also started endocrine tx preoperatively. Overall, 11/47 pts (23.4%) received neoadjuvant endocrine tx. 20 pts received systemic tx without eventual surgery, and 5 pts got palliative radiation (RT). Biopsy-confirmed mets were noted in 76 pts (80%), with bone-only mets in 50%, 30 pts (31.6%) with viscera-only mets and 7 pts (7.4%) with both bone and visceral mets. 18 pts had suspected mets on imaging but did not undergo biopsy. The hazard of death was 6.34 times higher in pts with both bone and visceral mets than those with bone-only mets (p=0.008). Pts with viscera-only mets had higher survival at 2 and 3 years than pts with bone-only or both bone and visceral mets (p=0.093). Pts with 3 mets (7/76, 7.4%) had decreased survival at 1, 2, and 3 years compared to pts with 1-2 (65/76, 85.5%) mets (p=0.6). 67 pts (70.5%) received mets-directed tx; RT alone was the most common modality (52/67, 77.6%), followed by surgery (6/67, 9%), RT plus surgery (5/67, 7.5%), and ablation alone (3/67, 4.5%). There was no significant difference in survival (HR 1.27, 95% CI (0.53, 3.07), p=0.589) or time to next tx (HR 0.95, 95% CI (0.56, 1.63), p=0.856) in pts who received mets-directed tx. Multivariate analysis was not performed because most findings were not statistically significant in univariate analysis. Conclusions We did not find any significant differences in survival based on characteristics like menopausal status or site or number of mets in pts with OMBC. There was a trend toward improved survival in pts with viscera-only mets, but this finding requires validation. Local/regional and mets-directed tx did not improve survival; however, survival at 1, 3, and 5 years was excellent in this OMBC pt population. Our study was limited by low pt numbers and heterogeneity in the pt population. Findings need validation in larger studies. Citation Format: Tanmayi Pai, Raza Zarrar, Zhongwei Peng, Zhuo Li, Lauren Cornell, Kostandinos Sideras, Rohit Rao, Alvaro Moreno-Aspitia, Saranya Chumsri, Sarah McLaughlin, James Jakub, Emmanuel gabriel, Sanjay Bagaria, Laura Vallow, Santo Maimone, Pooja Advani. Clinical Features and Survival Outcomes of Oligometastatic Breast Cancer Patients: The Mayo Clinic Experience [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 PO3-06-04.

Abstract Background: Previous studies demonstrated that obesity is associated with poor outcomes and significantly worse pathological complete response (pCR) in triple-negative breast cancer (TNBC) treated with conventional chemotherapy. Emerging studies described a phenomenon termed the obesity paradox, which referred to improved outcomes observed in obese patients treated with immune checkpoint inhibitors (ICIs). Our study aimed to determine distinct immune features in TNBC across body mass index (BMI). Methods: Characteristics and outcomes of TNBC patients treated with neoadjuvant pembrolizumab-based chemotherapy at Mayo Clinic were collected. Normal BMI was defined as 18-25 kg/m2, overweight 25-30, and obese BMI ≥ 30. NanoString CosMx, a spatial multi-omics single-cell imaging (SMI), was performed in 75 samples from the Mayo Clinic TNBC cohort (Leon-Ferre BCRT 2018). Differential expression as log 2-fold change (FC) was estimated from the linear mixed model with significance defined as two-sided p< 0.05. Results: Between 2021-2023, there were 108 TNBC patients treated with neoadjuvant pembrolizumab. 25.9% were obese, 32.4% were overweight, and 41.7% had normal BMI. In contrast to previous studies with conventional chemotherapy, obese patients numerically had a higher pCR rate of 71.4% with pembrolizumab-based chemotherapy compared to 55.6% in normal and 62.9% in overweight patients. However, this difference did not reach statistical significance (obese vs. normal weight, OR 2.26, 95%CI 0.89-6.06, p 0.094). We further evaluated spatially-resolved immune features associated with obesity using CosMx SMI in 75 samples from the MC TNBC cohort. Obese patients had significantly higher intratumoral B cells (log2FC 1.88, p 0.017), stromal B cells (log2FC 2.79, p 0.001), stromal plasmablasts (log2FC 1.94, p 0.047), and stromal macrophages (log2FC 1.80, p 0.017) compared to patients with normal BMI. Obese patients also had significantly lower intratumoral myeloid dendritic cells (mDC) (log2FC -1.27, p 0.037). Similar trends were observed after standardizing cell counts per 100 tumor or stromal cells, including higher intratumoral B cells (log2FC 3.00, p 0.001), intratumoral plasmablasts (log2FC 2.00, p 0.006), intratumoral natural killer (NK) cells (log2FC 1.66, p 0.044), and stromal B cells (log2FC 1.50, p 0.008). Differential gene analyses for intratumoral B cells revealed that genes (CD74, HLA-C, HLA-DRB1, HLA-DRA, HLA-DRB) associated with the positive regulation of type 2 immune or chemokine response were upregulated. Conclusions: Using in-depth analysis with spatially defined context, our study showed that obese TNBC patients had significantly higher B cell infiltration, which is associated with improved outcomes with ICIs. Future research on how obesity alters the immune microenvironment is needed to identify potential pathways to improve outcomes with ICIs in TNBC. Citation Format: Songhan Pang, Yi Liu, Yaohua Ma, Zhuo Li, Roberto A. Leon-Ferre, David Zahrieh, David W. Hillman, Judy C. Boughey, James N. Ingle, Krishna R. Kalari, Fergus J. Couch, Matthew P. Goetz, Keith L. Knutson, Saranya Chumsri, E. Aubrey Thompson. Tumor-immune microenvironment associated with obesity in triple-negative breast cancer in the Mayo Clinic cohort [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1162.

Abstract Background: Previous studies showed conflicting results regarding association between stromal tumor infiltrating lymphocytes (sTIL) and outcome in HER2+ breast cancer. While sTIL were not associated with outcome in patients (pts) treated with trastuzumab in N9831, immune functions genes are linked to outcome in these pts. TLS is an organized form of ectopic lymphoid aggregates (LA) with secondary lymphoid organ structure. Several studies showed that TLS associates with improved outcome in multiple cancers. However, distinguishing TLS and simple LA is challenging, particularly when germinal center is absent. In this study, we evaluated integrated pathological quantification and genomic data to assess functional TLS. Methods: Pathological evaluation of LA in H&E slides from pts treated in Arm A (chemotherapy alone) and Arm C (chemotherapy with concurrent trastuzumab) in N9831 was performed. NanoString was used to quantify mRNA. Wilcoxon rank sum test, Chi squared test, Kaplan-Meier method and Cox regression model were used to evaluate association between LA and baseline characteristics as well as outcomes. Results: LA was quantified in 1011 pts (526 Arm A, 485 Arm C). Greater number of LA was significantly associated with higher tumor grade, ER/PR negativity and increasing sTIL, but not age, tumor size or lymph node status. Increasing numbers of LA were associated with improved RFS in both arms combined (p 0.028). However, using multivariable Cox regression analysis in each treatment arm, LA (≥ 1 vs 0) was associated with improved RFS only in Arm A (HR 0.6, 95%CI 0.43-0.84, p 0.003) but not Arm C (HR 0.72, 95%CI 0.47-1.1, p 0.134). Further evaluation of expression of TLS-related immune genes including IFNG, ICOSLG, CXCL13, CXCR3, BCL6, IL21R, ICOS, PDCD1, CXCR5, CXCL9, TBX21, CD38, CXCL10, CLXCL11, IL21, CD200 was carried out among 252 pts in Arm C with LA ≥ 1. Among these 16 TLS-related immune genes, as a continuous variable, higher expression of BCL6 (HR 0.61, 95%CI 0.41-0.92, p 0.019) and IL21R (HR 0.78, 95%CI 0.62-0.98, p 0.03) were associated with improved RFS in Arm C pts with LA ≥ 1. However, these genes were not significantly associated with outcome in Arm C pts without LA with BCL6 HR1.07 (95%CI 0.67-1.71, p 0.776) and IL21R HR 0.96 (95%CI 0.71-1.29, p 0.775). Conclusion: Similar to sTIL, greater number of LA was associated with improved outcome in HER2+ pts treated with chemotherapy alone but not chemotherapy in combination with trastuzumab. Using histogenomic integration with the combination of pathological LA and TLS-related immune genes, we identified that pts with functional TLS with LA ≥ 1 and higher expression of BCL6 or IL21R had significantly improved outcome when treated with trastuzumab. Future studies are needed to further confirm these findings. Support: U10 CA180821, U24 CA196171, https://acknowledgments.alliancefound.org; Genentech; Clinicaltrials.gov Id: NCT00005970 Citation Format: Saranya Chumsri, Tracy R. Shachner, Zhuo Li, Nadine Norton, Alvaro Moreno-Aspitia, Gerardo Colon-Otero, Edith A. Perez, E. A. Thompson, Aziza Nassar, Keith L. Knutson. Functional tertiary lymphoid structure (TLS) and outcome in HER2-positive (HER2+) breast cancer in NCCTG N9831 (Alliance) [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 5627.

288 Background: Obesity is a risk factor for developing cholangiocarcinoma (CCA). However, the effect of obesity on survival of CCA is unclear. The primary aim of this study was to analyze the impact of obesity upon overall survival of CCA patients. Secondary aims were to analyze impact of obesity upon other disease characteristics such as tumor site, stage, age, sex, BMI and Ca 19-9. Methods: A total of 411 unique pts diagnosed with CCA at Mayo Clinic Florida between 2000 and 2018 were retrieved from our collective SDMS database. Variables evaluated included:demographics, Body Mass Index (BMI), AJCC stage, tumor location and Ca 19-9.A total of 185 pts had all data available pertaining to these variables. We further restricted the analysis to pts with intrahepatic CCA classified BMI as per CDC criteria normal (18.5-25kg/m2), overweight (25-29.9kg/m2) and obese (≥30 kg/m2), thus leaving a total of 152 pts. Continuous and categorical variables were compared across BMI groups using Chi-squared or Fisher’s exact test. Overall survival rates after diagnosis at 1, 2 and 3 years were estimated using Kaplan-Meier method. Results: Among 152 pts included in the study, 28% were normal weight, 40% were overweight and 32% were obese. The overall survival rate at 1, 2 and 3 years for normal weight pts with all stages combined was 54.1%, 35%, and 30.7%, respectively. The overall survival rate at 1, 2 and 3 years for overweight pts with all stages combined was 59.7 %, 32.6%, and 25.4%, respectively. The overall survival rate at 1, 2 and 3 years for obese pts with all stages combined was 63.9%, 37.6%, and 26.7%, respectively(p = 0.8766). Multivariate analysis demonstrated is no significant difference in overall survival for obese pts compared to normal or overweight pts.(Table to be shown) However it showed, gender and Ca19-9 were statistically significant predictors of overall survival, with males and pts with Ca19-9≥100 doing worse (HR1.65 (CI = 1.05, 2.61, p = 0.031) and HR 2.31 (CI = 1.49, 3.59, p = < 0.01), respectively). Conclusions: BMI did not make a significant impact on the overall survival, though there may be a trend toward worse OS for ptswith higher BMI. A larger, stage focused evaluation is warranted for further exploration of this trend.