Purpose: Even though prognostic and predictive significance of tumor-infiltrating lymphocytes (TILs) in triple-negative breast cancer (TNBC) have been demonstrated widely, the cause of TIL influx is not clear. High mobility group (HMG) N1, one of the three subtypes of HMGs which are nucleosome-binding proteins is recently known as a potent alarmin that binds to TLR4 and induces Th1 immune responses. The mechanisms which involve transportation of HMGN1 are not identified. We, in this study, suggest that extracellular secretion of HMGN1 might be associated with influx of TILs and subsequent formation of TLSs, as well as dendritic cell maturation and activation in human TNBC.

Experimental Design: We reviewed the hematoxylin and eosin (H&E)-stained slides of 767 patients with TNBC and evaluated the degree of TILs. We also measured the degree of expression of HMGs (HMGB1 and HMGN1) and several endoplasmic reticulum stress (ERS)-associated molecules (PERK, p-eIF2a, and XBP1) by immunohistochemical (IHC) staining in tissue microarrays of the patients. The hypothesis that HMGN1 migrates from nucleus to cytoplasm and extracellular space as the cell gets variable ER stresses was confirmed by Western blot analyses using human TNBC cell lines and pharmacological ER stress inducers. Results: On IHC staining, high cytoplasmic expression of HMGB1, HMGN1, PERK, p-eIF2a, and XBP1 were identified in 43.1%, 25.7%, 45.9%, 46.9%, and 41.8% of TNBCs, respectively. Cytoplasmic expression of HMGB1 was significantly correlated with cytoplasmic expression of HMGN1, amount of TILs, expression of PERK and XBP1 (p<0.001∼0.039). Cytoplasmic expression of HMGN1 was also significantly correlated with amount of TILs, expression of p-eIF2a and XBP1 (p<0.001∼0.004). Higher nuclear expression of both HMGB1 (p<0.001) and HMGN1 (p<0.001) showed significant association with ER stress. On Western blot analyses using TBNC cell lines, treatment of a ERS inducer resulted in the secretion of HMG proteins to the extracellular space. Conclusions: To the best of our knowledge, it is the first study that elucidated the associations among ERS, extracellular secretion of HMGs, and degree of TILs in TNBCs. Understanding the mechanisms of TIL influx will help to develop effective immunotherapeutic agents for treatment of TNBC.

Citation Format: Sun-Hee Heo, In Ah Park, Hee Jin Lee, Young-Ae Kim, Gyungyub Gong. Endoplasmic reticulum stress induces secretion of high mobility group (HMG) proteins and is associated with tumor-infiltrating lymphocytes in triple-negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-082.