bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P3-10-22: BluePrint molecular subtyping versus HER2 assessment by immunohistochemistry and FISH in the real-world diagnostic setting
Authors: Treece T , Audeh W , Navarro F , Wei J
Published: May 2019

Background: Both the Aphinity and ExteNET trials for anti-HER2 targeted agents were challenged in showing significant benefit of added HER2-targeted treatment. The findings suggested biological heterogeneity in HER2+ cancers, not entirely identified by IHC/FISH, requiring more nuanced biomarkers to clearly identify patient subsets who may derive benefit from new HER2-targeted agents. We have previously shown that BluePrint (80-gene) molecular subtyping reclassifies nearly half of HER2+/ER+ patients to Luminal-type with differential neoadjuvant treatment response (Whitworth, Ann Surg Oncol 2014; Whitworth, ASCO 2018). Here, we evaluated the reclassification rate in the real-world diagnostic setting. Methods: Physicians regularly provide pathology reports to Agendia, Inc for samples which are processed for MammaPrint (70-gene signature) and BluePrint molecular assays as part of routine diagnostic care. For this analysis, 4986 sequentially available pathology reports (submitted between October 2016 to October 2017) were reviewed; HER2 and ER IHC results were captured. The molecular subtype was compared to IHC/FISH status. Results: HER2 IHC/FISH results were available for 1568 samples. Of those, 85% (1330/1568) were HER2-nonamplified, 10% (153/1568) were HER2-equivocal, and 5% (85/1568) were HER2-amplified by IHC/FISH. Of the HER2-nonamplified tumors, BluePrint reclassified 0.1% (2/1330) as HER2-type. Of the HER2-equivocal tumors, none were HER2-type by BluePrint; 91% (139/153) were Luminal-type and 9% (14/153) were Basal-type. Of the HER2-amplified tumors, 15% (13/85) were dominant HER2-type, 79% (67/85) were dominant Luminal-type, and 6% (5/85) were Basal-type. Conclusions: In this set of tumors identified as HER2-amplified by IHC/FISH, BluePrint reclassified 85% of tumors to non-HER2 molecular subtypes, mostly Luminal-type for ER-positive tumors and Basal-type for ER-negative tumors. Moreover, BluePrint gave clarity where IHC/FISH could not, classifying all HER2-equivocal tumors to non-HER2 subtypes. Additional therapeutic options should be explored for HER2+/ER+ BluePrint Luminal-type patients who have observed much lower pCR rates versus BluePrint HER2-type patients (12% vs. 51%, respectively; Lee, AACR 2018).

HER2 IHC/FISH vs BluePrint SubtypeHER2 IHC/FISH StatusHER2-type Luminal-type Basal-type Total IHC ER-IHC ER+IHC ER-IHC ER+IHC ER-IHC ER+ Nonamplified (ER-Unknown, n=28)118120353361330Equivocal (ER-Unknown, n=6)  2131104153Amplified (ER-Unknown, n=2)112 664 85Total21310140067401568

Citation Format: Treece T, Audeh W, Navarro F, Wei J. BluePrint molecular subtyping versus HER2 assessment by immunohistochemistry and FISH in the real-world diagnostic setting [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-22.
Abstract P3-10-24: Not presented
Authors: Gasparini E , Bisagni A , Di Cicilia R , Kuhn E , Falco G , Ferrari G , Foroni M , Tamagnini I , Bassano C , Ragazzi M , Gardini G , Cecchi F , Hembrough T , Bisagni G
Published: May 2019

This abstract was not presented at the conference.

Citation Format: Gasparini E, Bisagni A, Di Cicilia R, Kuhn E, Falco G, Ferrari G, Foroni M, Tamagnini I, Bassano C, Ragazzi M, Gardini G, Cecchi F, Hembrough T, Bisagni G. Not presented [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-24.
Abstract P3-10-27: Profiling of metastatic carcinomas to the breast for the biomarkers of immuno-oncology therapy
Authors: Gatalica Z , Senarathne W , Vranic S , Pockaj BA
Published: May 2019

Introduction: Breast metastases from non-mammary carcinomas are very rare and the molecular characteristics of these metastases have not been studied. Due to the uniqueness of this type of metastases, we evaluated whether there was any molecular or immunologic similarities among the patients with metastases to the breast.

Patients and Methods: Fifty-one patients with metastatic carcinomas to the breast (49 female and 2 male) were identified in the files of Caris Life Sciences (Phoenix, AZ). Average age at presentation was 57 years (range, 20-90 years). Immunohistochemistry for PD-L1 was performed using SP142 (Ventana) or 22c3 (DAKO) antibodies; 592 genes sequencing (n=47), DNA microsatellite instability (MSI) (n=6) and total mutational burden (TMB) (n=14) were obtained from a next generation sequencing platform (Illumina).

Results: Three most common primary sites were lung (19, 37%), ovary (15, 29%) and fallopian tubes/peritoneum (7, 14%). Ten metastases (20%) demonstrated neuroendocrine differentiation. When 1% cutoff was applied, tumor cells PD-L1 expression was detected in 10 cases (20%), and immune cells expression of PD-L1 in 19 patients (37%). Three cases (6%) had PD-L1 staining in both cancer and infiltrating immune cells. 50% of the cases exhibited mutations in TP53 gene, while other mutated genes followed the pathways typically seen in their primary sites (e.g. VHL in renal carcinomas, BRCA1 in fallopian tube carcinoma). Total mutational burden was high (≥10 mutations/Mb) in five out of 14 successfully analyzed cases (36%) and no case (0%) exhibited high microsatellite instability. Interestingly, estrogen receptor status was positive in 13/49 patients (26.5%) including 12 adenocarcinomas originating from ovary, Fallopian tube or peritoneum and one duodenal neuroendocrine. No tumor was HER2 positive.

Conclusions: Metastases to the breast proved to be heterogeneous but there were molecular, endocrine and immunologic findings that yielded information that could be used to make therapeutic decisions which including the use of new immunotherapeutic strategies.

Citation Format: Gatalica Z, Senarathne W, Vranic S, Pockaj BA. Profiling of metastatic carcinomas to the breast for the biomarkers of immuno-oncology therapy [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-27.
Abstract P3-10-28: The impact of HER2-directed targeted therapy on HER2-positive DCIS of the breast
Authors: Lewis GD , Haque W , Farach A , Hatch SS , Butler EB , Schwartz MR , Bonefas E , Teh BS
Published: May 2019

Purpose/Objectives: In invasive breast cancer, HER2 is a well-established negative prognostic factor. However, its significance on the prognosis of ductal carcinoma in situ (DCIS) of the breast is unclear. As a result, the impact of adding HER2-directed therapy to HER2-positive DCIS is unknown and is currently the subject of ongoing clinical trials. In this study, we aim to determine the impact of HER2 status on DCIS patient outcomes as well as the possible impact of HER2-directed targeted therapy on survival outcomes for HER2-positive DCIS patients.

Materials/Methods: The National Cancer Data Base (NCDB) was used to retrieve patients with biopsy-proven DCIS diagnosed from 2004-2015. Only patients with known estrogen receptor (ER) status, progesterone receptor (PR) status, and HER2 status were included in the analysis. Patients were divided into two groups based on the adjuvant therapy they received: systemic therapy (assumed to be HER2-directed targeted therapy) or no systemic therapy. Statistics included multivariable logistic regression to determine factors predictive of receiving systemic therapy, Kaplan-Meier analysis to evaluate overall survival (OS), and Cox proportional hazards modeling to determine variables associated with OS.

Results: Altogether, 1927 patients met inclusion criteria; 430 (22.3%) received HER2-directed targeted therapy, while 1497 (77.7%) did not. Patients who received HER2-directed targeted therapy were likely more likely to be ER-negative. Patients who received HER2-directed targeted therapy had a higher 5-year OS compared to patients that did not (97.7% vs. 95.8%, p = 0.043). This survival benefit remained on multivariate analysis. Factors associated with worse OS on multivariate analysis included Charlson-Deyo Comorbidity Score ≥ 2 and no receipt of hormonal therapy.

Conclusions: In the largest study to date evaluating HER2-positive DCIS patients, the receipt of HER2-directed targeted therapy was associated with an improvement in OS. The results of currently ongoing clinical trials are needed to confirm this finding.

Citation Format: Lewis GD, Haque W, Farach A, Hatch SS, Butler EB, Schwartz MR, Bonefas E, Teh BS. The impact of HER2-directed targeted therapy on HER2-positive DCIS of the breast [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-28.
Abstract P3-11-03: Validation of CIN4 in the DBCG 89D clinical cohort
Authors: Spears M , Jensen M-B , Lyttle N , Liao L , Laenkholm A-V , Ejitlertsen B , Bartlett JM
Published: May 2019

Background: Chromosome instability (CIN) in solid tumours is associated with poor prognosis and results in numerical and structural chromosomal aberrations. Our group previously have developed the CIN signatures and have demonstrated the CIN signatures as prognostic biomarkers in breast cancer cohorts. Furthermore, our work in the BR9601 and MA.5 clinical cohorts CIN4 provided level IIB evidence that CIN4 was predictive of anthracycline sensitivity. An analysis of the DBCG 89D clinical trial was now performed to validate the role of CIN gene expression signatures as a marker of anthracycline sensitivity.

Methods: RNA was extracted from patients in DBCG 89D clinical trial analysed through NanoString technology. The prognostic and predictive values of the signatures on distant relapse-free survival (DRFS) were explored using Cox proportional hazard models. Multivariate models included menopausal status, tumour size, nodal status, ER and Her2 status, histological type and grade, and treatment regimen.

Results: All of the 594 samples available from the DBCG 89D we successfully analysed. CIN25 and CIN70 gene expression signatures did not associate with any of the clinicopathological characteristics tested. In addition, CIN25 and CIN70 were not prognostic or predictive of distant relapse free or breast cancer specific survival in this clinical cohort. Low CIN4 score was associated with ER negativity (p=0.02), HER2 normal expression (p<0.05).

Conclusion: In this study we demonstrated that CIN4 was associated with aggressive disease. We were however in DBCG 89D unable to validate the predictive value of CIN4 concerning anthracycline sensitivity.

Citation Format: Spears M, Jensen M-B, Lyttle N, Liao L, Laenkholm A-V, Ejitlertsen B, Bartlett JM. Validation of CIN4 in the DBCG 89D clinical cohort [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-03.
Abstract P3-11-06: HRD (homologous recombination deficiency) score and its clinicopathological features in neoadjuvant chemotherapy treated sporadic breast cancers
Authors: Imanishi S , Naoi Y , Shimazu K , Shimoda M , Kagara N , Tanei T , Miyake T , Kim SJ , Noguchi S
Published: May 2019

[Purpose]Homologous Recombination Deficiency (HRD) score has been developed to evaluate the DNA double-strand break repair function. BRCA1/2 germline mutation-associated breast cancers are known to show a high HRD score and a high sensitivity to platinum-based chemotherapy as well as PARP inhibitor. HRD can be theoretically induced by not only the dysfunction of BRA1/2 but also the dysfunction of the other molecules involved in homologous recombination, and, actually, high HRD score is reported to be seen in a significant proportion of sporadic breast tumors without BRCA1/2 mutation. The aim of the present study was to elucidate the clinicopathological characteristics of sporadic breast tumors with high HRD score as well as their sensitivity to sequential paclitaxel and FEC chemotherapy (P-FEC).

[Methods]Tumor tissues obtained by Mammotome from 132 sporadic breast cancer patients (stage II-III) before neoadjuvant chemotherapy with P-FEC were subjected to assay for HRD score using Oncoscan CNV kit®. HRD score was a simple sum of NtAI, LOH, and LST (cutoff, 42), and were also subjected to the gene expression analysis using the Affymetrix microarray (U133 plus 2.0). BRCA1 promoter methylation was assayed by methylation specific real-time PCR.

[Result]Of the 132 breast tumors, 32% showed a high HRD score which were significantly associated with high histological grade (P=0.001), negative progesterone receptor (P=0.023) and high Ki67 index (P=0.004). Triple negative breast cancer (TNBC)(n=19) showed a significantly (P<0.001) higher HRD score than the other subtypes (HR+/HER2-(n=73), HR+/HER2+(n=12), HR-/HER2+(n=18)). BRCA1 promoter methylation was significantly associated with a high HRD score. There was no significant correlation between HRD score and pCR to P-FEC when all tumors were considered but tumors with a high HRD score showed a significantly (P=0.020) lower pCR rate when only TNBC were considered. In TNBC, majority of tumors showed a high HRD score in five subtypes (BL1, BL2, IM, M, MSL) but no tumor showed a high HRD score in LAR (luminal androgen receptor) subtype.

[Conclusion]Approximately one third of sporadic breast tumors show a high HRD score, indicating the presence of homologous recombination dysfunction, and they are most often seen in TNBC with BRCA1 promoter methylation and never in LAR subtype. Interestingly, breast tumors with a high HRD score were less sensitive to P-FEC.

Citation Format: Imanishi S, Naoi Y, Shimazu K, Shimoda M, Kagara N, Tanei T, Miyake T, Kim SJ, Noguchi S. HRD (homologous recombination deficiency) score and its clinicopathological features in neoadjuvant chemotherapy treated sporadic breast cancers [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-06.
Abstract P3-11-11: Prediction of treatment response to neoadjuvant chemotherapy in breast cancer by subtype using tumor-infiltrating lymphocytes: Study of a large Mexican cohort
Authors: Campos Gomez S , Cabañas Chávez N , Hernández Alvarez J , Campos Gómez K
Published: May 2019

Background/aim:

Tumor-infiltrating lymphocytes (TILs) are emerging as biomarkers mediating tumor response to treatments. Studies have provided evidence that the level of TILs has prognostic value in breast cancer. The level of TILs has been associated with treatment outcome and pathologic response in patients undergoing neoadjuvant chemotherapy. In this study, we analyzed TILs before neoadjuvant therapy and if they correlate with pathological response to treatment

Patients and methods

We retrospectively analyzed the specimen slides of patients with Breast Cancer from the Centro Oncologico Estatal ISSEMyM treated with Neoadjuvant Chemotherapy (NAC) during 2007–2014. We identified 410 patients fulfilling the inclusion criteria of this study. The histological sections had already been evaluated by hematoxylin and eosin slides. They were reassessed by our pathologist for the percentage of intratumoral and stromal TILs. The correlation with pathological response of the tumor after neoadjuvant therapy was also studied in these patients. The proportions of TILs were categorized into high and low groups using a cut-off value of 10%

Resultados

The mean age was 51.4 years. The most frequent type of breast cancer histology was invasive ductal breast carcinoma in 369 (90%) patients, 34 (8.3%) lobular and 1.7% other. The BC subtypes among the 410 patients who received NAC were TNBC in 82 (20%), HER2BC in 70 (17.1%), and HRBC in 258 (62.9%) patients. Treatment response was pCR in 164 (40%) and non-pCR in 245 (60%) patients. Based on subtype, pCR was achieved in 43 (52 %) patients with TNBC, 35 (50.0%) with HER2BC, and 87 (33%) with HRBC. There were 191 (46.5%) patients in the high-TIL group and 219 (53.5%) patients in the low-TIL group. Patients were divided into high-TIL and low-TIL groups, and the clinicopathological characteristics of each group were examined. High TILs that had a pathologically complete response were found in 56 % of patients, and low TILs were found in 44 %. There was a significant correlation between TILs and pathological response in patients with neoadjuvant chemotherapy (p = 0.04).

Conclusion

TILs may be a biomarker for predicting treatment response to NAC in patients with Breast Cancer. Studies assessing outcomes and therapeutic efficacies should consider.

Citation Format: Campos Gomez S, Cabañas Chávez N, Hernández Alvarez J, Campos Gómez K. Prediction of treatment response to neoadjuvant chemotherapy in breast cancer by subtype using tumor-infiltrating lymphocytes: Study of a large Mexican cohort [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-11.
Abstract P3-11-13: On-treatment biomarkers can improve prediction of response to neoadjuvant chemotherapy in breast cancer
Authors: Bownes RJ , Turnbull AK , Cameron D , Sims AH , Oikonomidou O
Published: May 2019

Purpose: Neo-Adjuvant chemotherapy treatment is increasingly being used in breast cancer to preoperatively shrink tumour volumes and facilitate surgical plans. These datasets however, are still scarce, making it difficult to assess the relative value of multiple time point biopsies compared to diagnostic only sampling. This study aims to identify sequential samplings intrinsic value.

Method: A total of 97 samples from a cohort of 50 neoadjuvant chemotherapy treated primary breast cancer patients (aged 29-76 at diagnosis, Allred status 47:53% +/-, Her2 status 80:20% +/-, mixed grade and menopausal status) taken pre- treatment, at 2 weeks on-treatment, mid chemotherapy and at resection were sequenced with Ion Ampliseq transcriptome yielding expression values for 12,635 genes. Differential expression analysis was performed across response groups (16 Responders, 34 Non-Responders) as defined by Pathological Complete Response and over treatment time to identify significantly differentially expressed genes, pathways and markers indicative of response status.

Results: An on-treatment marker for response was identified (AAGAB), which resulted in a testing accuracy of 100% and a validation accuracy of 78% in the I-SPY 1 Trial. AAGAB was predictive of long term survival (p = 0.048 testing, p = 0.031 validation) in both chemotherapy cohorts at the same expression level as defined for treatment response. The single gene on-treatment biomarker, AAGAB proves more performant than established prognostic tests, Mammaprint (Edinburgh NEO trial, pre-treatment 61%, on-treatment 63%. I-SPY 1 trial, pre-treatment 60%, on-treatment 66%) and Pam50 RORS (neo trial pre-treatment 50%, on treatment 58%, Magbanua trial pre-treatment 56%, on-treatment 64%)

Conclusion: Changes in gene expression of on-treatment chemotherapy breast cancer resulted in the identification of a novel gene marker that was as effective in predicting prognostic status as established prognostic tests. These results support the use of on-treatment testing in breast cancer to improve the accuracy of tumour response prediction.

Citation Format: Bownes RJ, Turnbull AK, Cameron D, Sims AH, Oikonomidou O. On-treatment biomarkers can improve prediction of response to neoadjuvant chemotherapy in breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-13.
Abstract P4-01-03: Withdrawn
Authors: Di Cosimo S , Appierto V , Pizzamiglio S , Baselga J , Piccart M , Huober J , Izquierdo M , de la Pena L , Hilbers F , de Azambuja E , Untch M , Pusztai L , Pritchard KI , Nuciforo P , Salomon AV , Symmans FW , Apolone G , de Braud F , Verderio P , Daidone MG
Published: May 2019

This abstract was withdrawn by the authors.

Citation Format: Di Cosimo S, Appierto V, Pizzamiglio S, Baselga J, Piccart M, Huober J, Izquierdo M, de la Pena L, Hilbers F, de Azambuja E, Untch M, Pusztai L, Pritchard KI, Nuciforo P, Salomon AV, Symmans FW, Apolone G, de Braud F, Verderio P, Daidone MG. Withdrawn [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-01-03.
Abstract P4-01-07: A comprehensive liquid biopsy in patients undergoing neoadjuvant therapy
Authors: Sandoval Leon AC , Medina Saenz K , Miller P , Benson A , Calfa C , Mahtani R , Slingerland J , Perez A , Vogel C , Valdes-Albini F , El-Ashry D , Lippman M
Published: May 2019

Background: Precision medicine is revolutionizing breast cancer (BC) care. Comprehensive liquid biopsies are a tool for personalized care in patients with locally advanced breast cancer (LABC). Identifying robust biomarkers as part of a comprehensive liquid biopsy to predict response to treatment is of immense clinical interest.

Methods: After obtaining IRB approval, serial blood samples were collected from patients with LABC undergoing neoadjuvant therapy. Paired biopsies were collected prior to treatment and were sent to Foundation Medicine for next-generation sequencing (NGS). We used a sized-base microfilter technology to capture circulating tumor cells (CTCs) and circulating cancer associated fibroblasts (cCAFs). Patients with one or more CTCs or cCAFs were deemed positive for these tests. Additionally, in collaboration with Foundation Medicine, we extracted circulating tumor DNA (ctDNA) and we analyzed it using the FoundationACT platform. Patients with a detectable genomic alteration in their plasma were considered as having a positive ctDNA test. Our primary objective is to determine if a comprehensive liquid biopsy can serve as a prognostic marker of pathologic complete response (pCR).

Results: For this analysis we describe our findings in the initial blood draw of the first 18 patients enrolled. The mean age is 54 years (38-70). All patients who had their tumors sequenced had a detectable mutation. Consistent with the findings of others, we found TP53 mutations to be the most prevalent at 83.3%. We found that 44% of patients had ctDNA, 68.4% had cCAFs and 78.9% had CTCs. Many patients also had clusters of cells, consisting of one cell type, or co-clusters, consisting of both. 38.9% had CTC clusters, 16.7% had cCAF clusters and 16.7% had co-clusters (CTCs and cCAFs together). Some patients with CTCs did not have cCAFs and vice versa. The number of CTCs and cCAFS did not correlate with stage of disease or receptor status.

Conclusions: We describe a comprehensive liquid biopsy combining a sized-based microfilter technology for CTC and cCAFs identification and the FoundationACT platform for ctDNA analysis is feasible and these biomarkers can be detected in patients with LABC prior to the initiation of neoadjuvant therapy. Our study is accruing rapidly, and we will update our results with the longitudinal collection and the prognostic value of a comprehensive liquid biopsy at the time of the meeting.

Citation Format: Sandoval Leon AC, Medina Saenz K, Miller P, Benson A, Calfa C, Mahtani R, Slingerland J, Perez A, Vogel C, Valdes-Albini F, El-Ashry D, Lippman M. A comprehensive liquid biopsy in patients undergoing neoadjuvant therapy [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-01-07.
Abstract P4-01-12: Low peripheral blood CD4/CD8 ratio at the time of surgery is a negative long-term prognostic factor in women with early stage breast cancer
Authors: Magbanua MJM , Yau C , Scott JH , van't Veer L , Park JW , Esserman L , Campbell M
Published: May 2019

It is hypothesized that cancer prognosis may be related to the functional status of the immune system. We examined the correlation between peripheral blood CD4/CD8 ratio measured at the time of surgery and clinical outcome in patients diagnosed with early stage breast cancer.

Patient and Methods

Peripheral blood from 57 treatment-naïve early breast cancer patients, not eligible for neoadjuvant chemotherapy, was collected on the day of definitive surgery. CD4+ and CD8+ T cells were enumerated using flow cytometry and the ratio between the two immune cell populations was calculated. Cox regression analyses were performed to determine the relationship between CD4/CD8 ratio vs. distant disease-free survival (DRFS), breast cancer-specific survival (BCSS) and overall survival (OS). The median follow-up times were 10.1 years (range: 0.4-17.5) and 15.0 (range: 1.0-18.5) for DRFS and BCSS/OS, respectively.

Results

The patients' mean age at diagnosis was 54 years old (range: 31-78). 82% were hormone receptor-positive, 21% HER2-positive, and 61% node-negative. The median CD4/CD8 ratio was 2; and a ratio ≤ 2 was considered low. CD4/CD8 ratio was not associated with any of the clinicopathologic variable examined. Multivariate analysis using a survival model that adjusted for potential confounding factors (age, tumor size, grade, stage, hormone receptor, HER2, lymph-node status) revealed that patients with low CD4/CD8 ratio have statistically significant increased risk of distant recurrence (DRFS HR 5.3, Wald p=0.0381) and death (OS HR 3.8 Wald p=0.0271).

Conclusions

Immune dysfunction at the time surgery is correlated with long-term increased risk for metastatic recurrence and death. Larger clinical studies are warranted to confirm the results of this study.

Citation Format: Magbanua MJM, Yau C, Scott JH, van't Veer L, Park JW, Esserman L, Campbell M. Low peripheral blood CD4/CD8 ratio at the time of surgery is a negative long-term prognostic factor in women with early stage breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-01-12.
Abstract P4-02-04: Tumor-infiltrating lymphocytes in invasive lobular breast cancer identify a poor prognostic sub-group
Authors: Tille J-C , Saint Martin C , Fuhrmann L , De Koning L , Reyal F , Piccart M , Kirova Y , Cottu P , Carton M , Vincent Salomon A
Published: May 2019

Background:

Tumor infiltrating lymphocytes (TILs) are associated with an improved outcome in triple negative and HER2 breast cancers. Conversely, a recent pioneer study showed that TILs in infiltrating lobular carcinoma (ILC) were associated with a worse prognosis. We aimed at assessing the prognostic impact of TILs in a large cohort of primary surgically treated ILC with a long follow-up in a single institution.

Methods:

We retrospectively reviewed 459 ILC treated at the Curie Institute between 2005 and 2008. Clinico-pathological patients' characteristics were collected and all archived tumor slides were centrally reviewed for TILs quantification. We analyzed stromal TILs scored as a continuous variable. Cox analyses were performed for relapse free survival (RFS) and for overall survival (OS), and chi-square and Fisher test were performed to evaluate the correlation between TILs and other clinico-pathological variables. For statistical analyses, presence or absence of TILs was considered.

Results:

Patients had a mean age of 60.3 years and a median follow-up of 8.8 years, during which 74 local and/or distant relapse events occurred, among which 37 deaths due to the disease. The presence of TILs were significantly associated with greater tumor size (pT), positive nodal status (pN), molecular class (HER2 amplified), and specific morphological nuclear features such as high nuclear grade, multinucleation, macronucleoli and high Nottingham Prognostic Index class. Presence of intra-tumoral lymphocytes is significantly associated with a worse RFS (HR: 2.07 p=0.003) and OS (HR: 4.58 p<0.001) in a multivariate analysis, independently of chemotherapy.

Relapse Free Survival   TILs% (IC 95%)% (IC 95%)No95.3 (92.5-98.2)89.0 (84.3-94.0)Yes89.3 (85.1-93.6)74.0 (67.5-81-1)

Overall Survival OS at 5 yearsOS at 10 yearsTILs% (IC 95%)% (IC 95%)No99.5 (97.8-100)95.8 (92.3-99.4)Yes95.3 (92.3-98.1)83.5 (77.8-89.7)

Conclusion:

TILs are an independent prognostic factor associated with a poor outcome in ILC that are easy to assess on diagnostic biopsies. In order to elucidate the role of TILs in ILC, we will further characterize the immune infiltrate and integrate these results with DNA, RNA and protein analyses of the same carcinomas, in order to gain insight into their molecular features and to facilitate the identification of new therapeutic strategies for these ILC with TILs.

Citation Format: Tille J-C, Saint Martin C, Fuhrmann L, De Koning L, Reyal F, Piccart M, Kirova Y, Cottu P, Carton M, Vincent Salomon A. Tumor-infiltrating lymphocytes in invasive lobular breast cancer identify a poor prognostic sub-group [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-02-04.
Abstract P4-02-14: Digitalizing immunohistochemistry analysis (IHC) with quantitative dot blot analysis (QDB) by absolute quantification of biomarkers in FFPE specimens
Authors: Zhang J , Zhang W , Yu G , Tang F , Lv J
Published: May 2019

Purpose: The feasibility of Quantitative Dot Blot (QDB) method for absolute quantification of diagnostic biomarkers was investigated here using diagnostic antibodies for biomarkers in breast cancer tissues, including 4B5 and EP3 for Her2, MIB1 for Ki67. SP2 for Estrogen Receptor and SP1 for progesterone receptor (PR) in more than 300 FFPE specimens. The specificity and accuracy of this method, and the inter-relationship among biomarkers were explored by the absolute expression levels of these biomarkers.

Methods: FFPE specimens of 332 breast cancer patients were provided by local hospital, tissue slices of 2X15 um was de-paraffined and total protein was extracted for QDB analysis, with each sample measured in triplicate in three indepedent experiments. Purified protein standards were included to convert the measurement into absolute levels. IHC and FISH results from local hospital were used to validate the sensitivity and specificity of QDB method.

Results: The results show high consistency with intra-cv less than 7% and inter-cv less than 14%. With Her2, 99.5% concordance was achieved with IHC method, and 92 to 94% concordance with FISH analysis. Both Her2 level and Ki67 levels were significantly increased in patients at grade III based on Nottingham histologic scores.

Conclusion: The QDB method was used here to achieve digitalization of biomarkersl in breast cancer tissue with existing diagnostic antibodies for IHC analysis. The method is simple, objective, consistent, comprehensive and suitable for high throughput analysis. We expect this method to be extended to other diagnostic IHCs with existing diagnostic antibodies, and the easy adoption of this method in clinical practice should have instant impact on pathological practice.

Citation Format: Zhang J, Zhang W, Yu G, Tang F, Lv J. Digitalizing immunohistochemistry analysis (IHC) with quantitative dot blot analysis (QDB) by absolute quantification of biomarkers in FFPE specimens [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-02-14.
Abstract P4-04-12: TZAP mutation leads to poor prognosis of patients with breast cancer
Authors: Lee J-H
Published: May 2019

The zinc finger protein ZBTB48 is a telomere-associated factor that has been renamed as telomeric zinc finger-associated protein (TZAP). It binds preferentially to long telomeres, competing with telomeric repeat factors 1 and 2. However, its expression in cancers has not been investigated. In the present study, we analyzed the TZAP mutation in 128 breast carcinomas (BCs). In addition, its association with telomere length was investigated. The TZAP mutation (c.1272G>A, L424L) was found in 7.8% (10/128) of the BCs and was associated with the N0 stage (p = 0.035). BCs with the TZAP mutation had longer telomeres than those without this mutation (2.62 ± 1.71 vs. 0.98 ± 0.37, p = 0.009). Telomere elongation was shown in 29.7% (38/128) of BCs and was significantly higher in luminal A and triple negative BCs (p < 0.001). Other characteristics were not associated with TZAP mutations and telomere length. Survival analysis showed that the TZAP mutation resulted in a poorer overall survival (52.8 vs. 87.3 months, χ2 = 4.37, P = 0.037). However, telomeres did not have any prognostic value for BCs (P = 0.908). These results suggest that the TZAP mutation is a possible prognostic marker in BC.

Citation Format: Lee J-H. TZAP mutation leads to poor prognosis of patients with breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-04-12.
Abstract P4-06-02: Chemoimmunotherapy with cyclophosphamide plus a toll-like receptor 9 (TLR9) agonist eradicates triple negative breast cancer in a murine model
Authors: Campbell MJ , Tandon V , Senman B , Li J , Esserman L
Published: May 2019

Background. Toll-like receptors (TLR) recognize conserved molecular patterns expressed by microbes and together with other molecular sensors serve as a first line of defense, inducing soluble and cellular mediators of innate immunity and initiating key steps of an adaptive immune response. The use of TLR agonists for therapeutic purposes relies on the ability of these compounds to induce, at least partially, some of the immune events that occur during natural infections. For example, synthetic CpG-containing oligodeoxynucleotides (ODN) act as TLR9 agonists, mimicking stimulation of the immune system by bacterial or viral DNA. CpG ODNs are being developed for cancer immunotherapy based on their capacity to stimulate innate and adaptive anti-tumor responses. In this study, we examined the efficacy of intratumorally administered CpG-ODN 1826 alone or in combination with immunomodulatory antibodies (anti-PD-1, anti-OX40, anti-CTLA4) or chemotherapy (cyclophosphamide, paclitaxel) in the syngeneic mouse 4T1 breast tumor model.

Methods. Tumors were implanted orthotopically in the mammary fat pads on the left and right flanks while only one tumor was injected with CpG-ODN 1826. Therapy began 12-14 days post tumor injection when tumors were 8-10 mm in diameter. CpG-ODN 1826 (100 ug) was administered intratumorally, while immunomodulatory antibodies (anti-PD-1 [200 ug], anti-CTLA4 [100 ug], anti-OX40 [400 ug]) and chemotherapies (cyclophosphamide [150 mg/kg], paclitaxel [10 mg/kg]) were administered intraperitoneally. Tumor volume was monitored on both flanks to assess direct and abscopal/systemic anti-tumor activity. Tumor tissue obtained during the treatment regimen was used to evaluate therapy-induced changes in the immune microenvironment.

Results. CpG-ODN 1826, administered intratumorally over 5 consecutive days induced complete regressions in ˜50% of the treated tumors, but only delayed growth in the distant lesions. The immunomodulatory antibodies had little effect on their own and did not add to the therapeutic efficacy of CpG-ODN 1826. The best therapeutic efficacy was obtained with a combination of weekly cyclophosphamide + CpG-ODN 1826 resulting in complete regression of both the CpG injected tumors and the contralateral tumors in 100% of the mice.

Conclusions. These data support the clinical investigation of the combination of a TLR9 agonist (CpG-ODN) with cyclophosphamide in women with breast cancer.

Citation Format: Campbell MJ, Tandon V, Senman B, Li J, Esserman L. Chemoimmunotherapy with cyclophosphamide plus a toll-like receptor 9 (TLR9) agonist eradicates triple negative breast cancer in a murine model [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-06-02.
Abstract P4-06-04: Targeting neo-epitopes from PIK3CA and p53 mutations for immunotherapy of breast cancer
Authors: Chen M , Yuvaraj P , Sharma AA , Narang P , Wilson Sayres MA , Anderson KS
Published: May 2019

Introduction: Preclinical data suggests that priming T cell immunity to mutated or overexpressed proteins can induce tumor rejection, and may potentiate the effects of checkpoint blockade. PIK3CA and p53 are the most commonly mutated genes in breast cancer, and are enriched in high-grade and metastatic tumors. Here, we evaluated the immunogenicity of neo-epitopes derived from common PIK3CA and p53 mutations.

Methods: We applied a custom informatics pipeline, EpitopeHunter, to predict neo-epitopes (8-11mers) from twelve PIK3CA and ten p53 mutations restricted to thirty-nine MHC class I alleles. We selected high affinity neo-epitopes (IEDB score < 500) with predicted poor binding of the matched wild type epitope. We used the predicted peptides to stimulate PBMC from healthy donors in vitro, and measured T cell immunity by interferon-γ ELISPOT. We compared the T cell specificity of mutant vs wild type epitopes. A positive response to an epitope was defined as a response that met both of the following criteria: 1) net number of spots (after subtracting background) was >5 spots/50,000 cells and 2) net number of spots exceeded the background spots plus two SDs.

Results: In total, 1,824 PIK3CA and 1,520 p53 derived peptide sequences were generated. Of these, 42 PIK3CA (2%) and 45 p53 (3%) peptides were predicted to bind to the HLA class I molecules included in our study. Over 90% of the neo-epitopes were 9- and 10-mers, and the predicted neo-epitopes varied across mutations (range 2-10 per mutation). PIK3CAH1047L and p53R248W had the highest number of potential binding neo-epitopes (n=10 each). We successfully generated T cell lines specific for A*0201 PIK3CAH1047L, A*1101 PIK3CAH1047L, A*1101 PIK3CAE542K and A*0201 p53R248W.

Conclusions: Common mutations of PIK3CA and p53 can lead to the generation of potential HLA class I restricted neo-epitopes. We identified four immunogenic neo-epitopes, which may serve as candidates for targeted immunotherapy in breast cancer.

Citation Format: Chen M, Yuvaraj P, Sharma AA, Narang P, Wilson Sayres MA, Anderson KS. Targeting neo-epitopes from PIK3CA and p53 mutations for immunotherapy of breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-06-04.
Abstract P4-06-22: Transcription factor T-bet and PD-L1 expression in tumor microenvironment of triple-negative breast cancer
Authors: Mori H , Kubo M , Kai M , Kurata K , Kawaji H , Kaneshiro K , Motoyama Y , Kuroki R , Yamada M , Nishimura R , Okido M , Oda Y , Nakamura M
Published: May 2019

Background: Many analyzes regarding immunotherapies using checkpoint blockade has made it clear that tumor infiltrating lymphocytes (TILs) plays an important role in treating cancers with high levels of somatic mutations such as triple-negative breast cancer (TNBC). We reported the relationship between TILs and PD-L1 expression, and revealed that high-TILs/positive-PD-L1 expression population in TNBC was associated with better prognosis (Oncotarget 2017). However, its molecular mechanism is still unclear. Meanwhile, T-box transcription factor 21 (T-bet) which regulates effecter T-cells activation is derived by stimulation of T-cell receptor and IL-12. Activated T-cells work as antitumor lymphocytes by enhancing the production of cytokines such as INFγ. We focused on T-bet and examined the function of activated T-cells.

Patients and Methods: This study included 242 patients with primary TNBC who underwent resection without neoadjuvant chemotherapy at our three hospitals between January 2004 and December 2014. The immunohistochemistry scoring for CD8 and T-bet expression on TILs was defined as ≥30 per 0.00625mm2. PD-L1 positivity was defined as ≥1% of tumor cells staining positive for PD-L1.

Results: Of the 242 TNBC, CD8 on TILs was expressed as positive in 127 (52.5%) tumors, T-bet on TILs was expressed as positive in 67 (27.7%) tumors, and PD-L1 expression on tumor cells was expressed as positive in 99 (40.9%) tumors. T-bet expression was significantly correlated with CD8 expression (P<0.0001) and PD-L1 expression (P=0.0004). There was no significant difference in recurrence free survival (RFS) and overall survival (OS) regardless of CD8 or PD-L1expression level. Meanwhile, the patients with T-bet-positive tumors had a longer OS, compared to those with T-bet-negative tumors (P = 0.13 in RFS and P = 0.047 in OS). The multivariate analysis revealed that T-bet expression on TILswas an independent and positive prognostic factor for OS(HR = 0.5, 95%CI 0.1-0.9, P = 0.035).

Conclusion: OS was significantly longer among patients with high T-bet expressing TNBC. These results may validate the significance of T-bet as a biomarker for various immunotherapies in TNBC.

Citation Format: Mori H, Kubo M, Kai M, Kurata K, Kawaji H, Kaneshiro K, Motoyama Y, Kuroki R, Yamada M, Nishimura R, Okido M, Oda Y, Nakamura M. Transcription factor T-bet and PD-L1 expression in tumor microenvironment of triple-negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-06-22.
Abstract P4-06-25: B7-H3 and B7-H4 expression in ductal carcinoma in situ of the breast: Associations with clinicopathologic features and T-cell infiltration
Authors: Lee JS , Kim G-E , Park MH , Yoon JH
Published: May 2019

Background: With the clinical success of immune check point blockades in treating malignant tumors, there are intense investigations in identifying new pathways to activate immune system by targeting immunoregulatory molecules. B7-H3 and B7-H4 play an inhibitory role in T cell function by limiting proliferation and cytokine production. Although several studies have investigated the expression of B7-H3 and B7-H4 in invasive breast cancers, the information of the B7-H3 and B7-H4 molecules in ductal carcinoma in situ (DCIS) remains uncertain.

The present work was undertaken to evaluate the expression of B7-H3 and B7-H4 in DCIS and its association with clinicopathological features in patients with DCIS. In addition, the association of B7-H3 and B7-H4 expression with the T-cell infiltration was also assessed to investigate its roles in the regulation of tumor immune surveillance.

Materials and methods: B7-H3 and B7-H4 expression was examined in 8 pairs of DCIS tissues and matched normal adjacent tissues at mRNA and protein levels by RNAscope in situ hybridization (ISH) and immunohistochemistry. Immunohistochemical staining of B7-H3 and B7-H4 was done in 79 DCIS samples with known hormone receptor (HR) and human epidermal growth factor 2 (HER2) expression using tissue microarray. In addition, immunohistochemical staining was also performed for the T cell lineage markers CD3 and CD8 in DCIS.

Results: RNAscope ISH and immunohistochemistry of B7-H3 and B7-H4 confirmed their increases in DCIS tissues compared with their corresponding normal tissues. B7-H3 and B7-H4 mRNA and protein expression appeared to be concentrated mainly in the DCIS carcinoma cells. High B7-H3 and B7-H4 expression was observed in 58 (73.4%) and 62 (78.5%) cases with DCIS, respectively. High B7-H3 expression was significantly associated with high nuclear grade and presence of comedo-type necrosis (P < 0.05 and P < 0.05, respectively). B7-H3 expression was higher in the HR–/HER2+ subtypes than in the HR+/HER2– subtype (P < 0.05). B7-H3 and B7-H4 expression tended to be associated with low stromal tumor infiltrating lymphocytes density and was negatively related to the CD3+ and CD8+ T cell infiltrates density.

Conclusions: These results suggest that B7-H3 and B7-H4 may play an important role in immune surveillance mechanisms of DCIS and may be useful targets for immune-based therapy to alter or prevent DCIS progression.

Citation Format: Lee JS, Kim G-E, Park MH, Yoon JH. B7-H3 and B7-H4 expression in ductal carcinoma in situ of the breast: Associations with clinicopathologic features and T-cell infiltration [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-06-25.
Abstract P4-06-27: LncRNA NKILA promotes tumour immune evasion by sensitizing tumour-specific t cells to activation-induced cell death
Authors: Song E , Su S , Huang D
Published: May 2019

Activation induced cell death (AICD) of T lymphocytes is critical to maintain T-cell homeostasis, which is adopted by malignant tumors to convey immune evasion by eliminating tumor-reactive cytotoxic T cells. In this study, we demonstrated excessive apoptosis of tumor antigen-specific CTLs in breast and lung cancers. However, the mechanism involved in AICD of tumor-specific T cells remains obscure. Here, we demonstrated that NF-κB activity in tumor-specific T cells is high at the early phase of CTL activation induced by breast tumor antigens, but is suppressed at the later phase. This results in massive apoptosis of tumor-specific CTLs challenged by tumor cells. Interestingly, NKILA, an NFκB interacting lncRNA, sensitizes CTLs to AICD by inhibiting NF-κB activities after their activation, leading to tumor immune evasion. In vivo, administering CTLs with NKILA silencing into immunocompromised mice with breast cancer patient derived xenografts (PDXs) effectively inhibits PDX growth by increasing CTL infiltration. Clinically, NKILA was overexpressed in the tumor specific CTLs of breast and lung cancers, which was associated with less CTL infiltration in the tumors and shorter patient survival. Our findings present the first evidence that AICD in tumor-specific CTLs is crucial to cancer immune evasion, and targeting NKILA in CTLs emerges as a novel anti-tumor immunotherapy.

Citation Format: Song E, Su S, Huang D. LncRNA NKILA promotes tumour immune evasion by sensitizing tumour-specific t cells to activation-induced cell death [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P4-06-27.