Abstract P4-09-25: Exploring immunomodulatory effects of zoledronic acid in breast cancer from clinical trial result of neoadjuvant chemotherapy with zoledronic acid: JONIE-1 study
Authors: Sangai T , Sato E , Ishikawa T , Kaise H , Hasegawa Y , Miura D , Takao S , Suzuki M , Tanino H , Horiguchi J , Akazawa K , Yamada A , Kohno N 
Published: May 2016

Background: We previously reported that the addition of zoledronic acid (ZOL) to neoadjuvant chemotherapy (CT) is potentially beneficial in postmenopausal patients with triple-negative breast cancer (BC) in JONIE-1 Study (0% pCR rate in CT versus 50% in CT combing ZOL [CTZ], p=0.029). In order to find biomarkers of response to ZOL, a pilot study was performed using 30 core needle biopsy (CNB) samples at diagnosis from JONIE-1 study. We found that Src activation was highly associated with pCR compared to non-pCR especially in CTZ group. CD8+ TIL was also highly associated with pCR compared to non-pCR. None of CD8+ TIL low patients resulted in pCR in CT group but four out of ten CD8+ TIL low patients experienced pCR in CTZ group. From the result of pilot study, we consider Src activation as a promising biomarker for response to chemotherapy especially in combination with ZOL; however, Src inhibition by ZOL in vivo and the mechanism to explain tumor inhibition is unknown. Because ZOL was effective even in CD8+ TIL low group, the possibility of immune response alteration by ZOL still remains. Not all tumors with Src activation and CD8+ TIL had benefit from ZOL, emphasizing the need for additional markers of response. In the present study, we focused on Src and multiple molecules relating to cancer immune response and analyzed associations with pCR.

Patients and Methods: We investigated the relationship between clinicopathological features and tumor shrinkage in Stage IIA-IIIB HER-2-negative BC patients from the JONIE-1 adjuvant phase III trial comparing CT (FEC100 q3w × 4 cycles followed by weekly paclitaxel for 12 cycles) versus CT combining ZOL (4mg q3-4w). To access Src activation, HLA class I expression, PD-1 and PD-L1 expression, we performed immunohistochemistry (IHC) and stained slides were evaluated by pathologists. Infiltration of immature dendritic cells, macrophages, cytotoxic T cells, and regulatory T cells in the tumor were assessed by IHC as well. Formalin fixed, paraffin embedded CNB sample at diagnosis and surgical specimen after neoadjuvant CT were processed for HE staining and IHC using primary antibodies as follows; anti-Src Ab (36D10), anti-pan HLA-class I Ab (EMR8-5), anti-CD8 Ab (Clone C8/144B), anti-FOXP3 Ab (Clone 236A/E7), anti-CD68 Ab (Clone PGM1), anti-CD1a Ab (Clone O10).

Results: All IHC were successfully performed. Stained slides are now under evaluation by pathologist. Associations between clinicopathological features and the effect of CT with ZOL will be under investigation.

Discussion: Src activation was observed in more than 70% of triple negative BC. Multiple cellular functions of Src are mediated by Ras which is the main target of ZOL in osteoclasts. It has been reported that ZOL reverted tumor-infiltrating macrophages (TAM) phenotype from M2 to M1. Moreover, recent study showed that the activation Src and MAPK in melanoma cells promotes PD-L1 expression. Therefore the addition of ZOL might help cytotoxic T cells infiltrate the tumor for inhibiting tumor growth. We will present the results and discuss antitumor effects of ZOL through immunomodulation at the meeting.

Citation Format: Sangai T, Sato E, Ishikawa T, Kaise H, Hasegawa Y, Miura D, Takao S, Suzuki M, Tanino H, Horiguchi J, Akazawa K, Yamada A, Kohno N. Exploring immunomodulatory effects of zoledronic acid in breast cancer from clinical trial result of neoadjuvant chemotherapy with zoledronic acid: JONIE-1 study. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P4-09-25.