Dendritic Cell Immunotherapy is Effective for Lung Metastasis from Murine Osteosarcoma

Two-thirds of patients with osteosarcoma survive when treated with the current standard protocol of neoadjuvant chemotherapy. However, lung metastasis is associated with a fatal outcome in most patients. We examined the efficacy of dendritic cell immunotherapy for lung metastasis using a previously established murine osteosarcoma cell line, LM8, which exhibits high metastatic potential after subcutaneous inoculation. Vaccination of dendritic cells generated from syngeneic mice increased lymphocyte interferonproduction, particularly when dendritic cells were pulsed by tumor antigens, and induced lymphocyte proliferation and cytotoxicity against LM8 cells. We performed three experimental protocols. Dendritic cell vaccination before tumor inoculation prevented tumor development. Dendritic cell vaccination after the primary tumor developed suppressed growth of subcutaneous tumors and lung metastases. Dendritic cell vaccination after surgical resection of the primary tumor also suppressed of lung metastases. In addition, in vitro Boyden chamber assay revealed dendritic cell-treated lymphocytes attenuated tumor invasiveness. Our data suggest dendritic cell-based immunotherapy may be an effective postoperative therapeutic approach to prevent metastasis in patients with osteosarcoma. Osteosarcoma is the most frequent primary malignant bone tumor in adolescents and young adults. Current multimodal treatment includes the standard protocol of neoadjuvant chemotherapy combined with wide local excision, which improves the 5-year disease-free survival 60% to 70%. However, survival rates have remained poor with lung metastasis associated with fatal outcome for most patients. If lung metastases or unresectable tumors are present at diagnosis, the prognosis is dependent on sensitivity of the malignancy to chemotherapy, which can be extremely poor because of limited treatment strategies. Lack of effective treatment means complete remission cannot be achieved by many patients. Nonetheless, some investigators have reported surgical resection of all operable lesions improved prognosis in patients with osteosarcoma with lung metastasis. Kempf-Bielack et al reported complete removal of the resectable lesion is a prerequisite for a good outcome for the second-line chemotherapy against recurrence of osteosarcoma. Dendritic cells are powerful antigen-presenting cells, exhibiting high potency in induction of T-lymphocyte response. With progress in our understanding of the immune system, there are numerous applications of dendritic cell-based immunotherapy. Immunotherapy has been evaluated in several clinical trials suggesting the utility of this new modality for various malignancies. Many of the trials examined patients with advanced disease, accounting for the mixed results. However, there are few studies of dendritic cell-based immunotherapy for bone and soft tissue tumors, including osteosarcoma. We previously established a highly metastatic cell line, lung metastasis 8 (LM8), from a Dunn murine osteosarcoma. Lung metastasis 8 was obtained through eight cycles (each cycle consists of tail vein injection, preparation of lung metastasis, in vitro culture, and repeat tail vein injection) of in vivo selection according to a reported method that showed high metastatic potential to the lung Received: August 18, 2005 Revised: February 24, 2006; June 26, 2006 Accepted: July 10, 2006 From the Department of Biology, Department of Orthopedic Surgery, and Department of Pathology, Osaka Medical Center for Cancer & Cardiovascular Diseases, Osaka, Japan; and the Department of Orthopedic Surgery, Osaka University Graduate School of Medicine, Osaka, Japan. One of the authors (KI) has received funding from Grants-in-Aid from the Ministry of Education, Science, Sports, and Culture of Japan, and Takeda Science Foundation. Each author certifies that his or her institution has approved the animal protocol for this investigation and that all investigations were conducted in conformity with ethical principles of research. Correspondence to: Kazuyuki Itoh, MD, PhD, Department of Biology, Osaka Medical Center for Cancer and Cardiovascular Diseases, 1-3-3 Nakamichi, Higashinari-ku, Osaka, 537-8511, Japan. Phone: 81-6-6972-1181; Fax: 816-6973-5691; E-mail: itou-ka@mc.pref. osaka.jp. DOI: 10.1097/01.blo. 0000229347.02834. 5e CLINICAL ORTHOPAEDICS AND RELATED RESEARCH Number 453, pp. 318–327 © 2006 Lippincott Williams & Wilkins