Analysis of HSP27 and the Autophagy Marker LC3B Puncta Following Preoperative Chemotherapy Identifies High-Risk Osteosarcoma Patients

Chemotherapy-induced autophagy is a proposed mechanism of chemoresistance and potential therapeutic target in osteosarcoma. We evaluated heat shock protein 27 (HSP27) and autophagy-related proteins as predictors of pathologic treatment response and prognostic markers among osteosarcoma patients who received standard chemotherapy. We analyzed 394 tumor specimens (pre-treatment, post-treatment, and metastases) from 260 osteosarcoma patients by immunohistochemistry for cytoplasmic light chain 3B (LC3B)-positive puncta, sequestosome 1 (SQSTM1), high mobility group box 1 (HMGB1), and HSP27 expression. The staining percentage and intensity for each marker were scored and the extent to which marker expression was correlated with pathologic response, relapse-free survival (RFS), and overall survival (OS) was assessed. LCB3 puncta in post-treatment primary tumors (50%) and metastases (67%) was significantly higher than in pre-treatment biopsy specimens (30%; = 0.023 and <0.001). Among 215 patients with localized osteosarcoma, both pre-treatment [multivariate hazard ratio (HR), 26.7; 95% confidence interval (CI), 1.47-484; = 0.026] and post-treatment HSP27 expression (multivariate HR, 1.85; 95% CI, 1.03-3.33; = 0.039) were associated with worse OS. Lack of LC3B puncta at resection was an independent poor prognostic marker in both univariate (HR, 1.78; 95% CI, 1.05-3.03; = 0.034) and multivariate models (HR, 1.75; 95% CI, 1.01-3.04; = 0.045). Patients with LC3B/HSP27 tumors at resection had the best 10-year OS (75%) whereas patients with LC3B/HSP27 tumors had the worst 10-year survival (25%). Neither HSP27 expression nor the presence of LCB3 puncta was correlated with pathologic treatment response. Our findings establish HSP27 expression and LC3B puncta as independent prognostic markers in osteosarcoma patients receiving standard chemotherapy and support further investigation into strategies targeting HSP27 or modulating autophagy in osteosarcoma treatment. .