Preclinical Study Of PD-L1 Blockade Combined With Chemotherapy In The Treatment Of Osteosarcoma

Osteosarcoma is the most frequent malignant neoplasm of bone in children and adolescents.Following the implementation of chemotherapy in the 1970 s,the 5-year overall survival rate of osteosarcoma has increased dramatically to 60%-70%.At present,surgery combined with neoadjuvant chemotherapy is still the conventional treatment strategy for osteosarcoma.However,the chemotherapy of osteosarcoma has reached a plateau for nearly 40 years.In up to 40% of osteosarcoma patients,chemotherapy has not prevented recurrence or distant metastasis,and there is no other effective therapy.Thus,novel therapeutic strategies for osteosarcoma are necessarily to be explored.The aim of this study was to evaluate the potential value of blocking PD-L1 using PD-L1 antibody or sorafenib in combination with chemotherapy for treating osteosarcoma.Part One Checkpoint Blockage Augments Tumor Cell Apoptosis in Combination with DNA Damaging Chemotherapy in OsteosarcomaCombination of different therapeutic modes has achieved better outcomes in malignant tumor treatment than single pattern.However,there is no guideline of how to combine different therapeutic modes.The aim of this study is to provide a theoretic basis that the combination of conventional chemotherapy and immunotherapy would be an effective treatment in osteosarcoma.Here,the expression of programmed death-ligand 1 (PD-L1)in 26 clinical osteosarcoma tissues before and after chemotherapy was analyzed.Effects of doxorubicin,a conventional chemotherapeutic agent,treated osteosarcoma cells on proliferation and apoptosis of CD8+ T lymphocytes were investigated in vitro.Then effectiveness of doxorubicin combined with PD-L1 antibody on osteosarcoma was tested in 24 mouse subcutaneous tumor formation models.The results showed that expression of PD-L1 was up-regulated by chemotherapy in both clinical osteosarcoma tissues and osteosarcoma cell lines.Proliferation of CD8+ T lymphocytes was inhibited and apoptosis of CD8+ T lymphocytes was enhanced by doxorubicin pretreated osteosarcoma cells,while this effect was reversed by PD-L1 antibody.A more effective result was observed when combining doxorubicin with PD-L1 antibody in vivo.In short,combination of conventional chemotherapy and PD-L1 antibody might be an effective option in osteosarcoma treatment since PD-L1 antibody can reverse immunosuppression induced by chemotherapy.Part Two Pre-clinical Study of Sorafenib Combined with Chemotherapy in Osteosarcoma TreatmentSince previous studies revealed that chemotherapeutic agents up-regulate the expression of PD-L1 through ERK pathway in malignant tumors,the aim of this study was to confirm that an effective ERK pathway inhibitor,sorafenib,can significantly enhance the efficiency of doxorubicin in osteosarcoma.It was observed that sorafenib could effectively inhibit the up-regulation of doxorubicin-inducing up-regulation of PD-L1 expressed in osteosarcoma cells.In addition,ERK pathway inhibitor also had the same PD-L1 inhibitory effect as sorafenib.Finally,we confirmed that the expression of PD-L1 in osteosarcoma treated with doxorubicin combined with sorafenib was significantly decreased compared with that treated with doxorubicin alone.These findings indicated that sorafenib in combination with chemotherapy could be a more effective treatment strategy for osteosarcoma.