Author ORCID Identifier
Date of Graduation
Clinical and Translational Sciences
Masters of Science (MS)
Eugenie S. Kleinerman
Robert Bast Jr.
Survival for patients with osteosarcoma has not improved for > 30 years. Despite aggressive multi-agent chemotherapy combined with surgical resection, a significant fraction of patients with localized disease relapse after optimal treatment. We evaluated the occurrence of cytoplasmic LC3B (light chain 3B)-positive puncta (a marker of autophagy) and presence of HSP27 (heat shock protein 27) in cancer cells within pre-treatment biopsy, post-treatment surgical resection, and metastatic osteosarcoma specimens by immunohistochemistry in 260 patients. LCB3+ puncta expression was seen in 34% of pre-treatment. 50% of resection, and 67% of metastasis samples. Sixty-six percent of all specimens were scored positive for HSP27 (85% of pre-treatment. 52% of resection, and 50% of metastasis samples). Among 215 patients with localized disease, pre-treatment HSP27 expression was associated with inferior overall survival (adjusted HR 26.7, p=0.0263) as well as at resection following chemotherapy (adjusted HR 1.85, p=0.039). Lack of LC3B-puncta expression was an independent poor prognostic marker at resection (adjusted HR 1.75, p=0.045). Patients with LC3B+/HSP27- tumors at resection had the best prognosis whereas patients with LC3B-/HSP27+ osteosarcoma had the worst long-term survival. Neither HSP27 nor LC3B expression correlated with tumor necrosis. These findings indicate that HSP27 expression is a negative prognostic biomarker in osteosarcoma. Conversely, presence of autophagy following neoadjuvant chemotherapy, as measured by LC3B-puncta, predicts longer overall survival in osteosarcoma patients with localized disease.
We additionally evaluated the significance of chemotherapy-induced autophagy in 2 human osteosarcoma cell lines: LM7 and CCH-OS-D. Both doxorubicin (DOX) and cisplatin (CDDP) were found to induce autophagy. In LM7 cells, autophagy inhibition with hydroxychloroquine (HCQ) prior to chemotherapy resulted in a trend towards decreased viability consistent with a cytoprotective role of autophagy. In CCH-OS-D cells, autophagy inhibition prior to DOX significantly decreased chemosensitivity suggesting a cytotoxic role of autophagy in this setting. The post-treatment expression of phosphorylated HSP27 was increased in LM7 and decreased in CCH-OS-D following DOX or CDDP. These findings support a dual role of chemotherapy-induced autophagy and potential application of pHSP27 as a predictive biomarker of autophagy inhibitors in osteosarcoma.
Osteosarcoma, autophagy, heat shock protein, biomarkers, LC3B, HSP27