Hematogenous Micrometastases in Osteosarcoma Patients

Abstract

Bone marrow and peripheral blood samples from 60 patients with suspected bone sarcoma were examined for the presence and number of micrometastatic osteosarcoma cells by a sensitive immunomagnetic detection assay, using in parallel two osteosarcoma-associated antibodies. Forty-nine of the patients had osteosarcoma, and of these, as many as 31 (63%) had tumor cells in bone marrow, in many cases with a high number of cells. Only four (8%) were positive also in blood. None of 38 control bone marrow samples were positive, including 11from patients with suspected bone sarcoma at time of sampling who later were found not to have osteosarcoma. Fifteen of 28 patients without overt metastases at primary diagnosis (54%) were positive, 12 of whom had localized high-grade primary tumors in the extremity. Four of these have relapsed compared with none of 10 negative patients. In the group of 22 patients with extremity localized nonmetastatic osteosarcoma, information was available on the histologic response to preoperative chemotherapy in15 patients. None of the three patients in the bone marrow^negative group who had a poor response to chemotherapy have relapsed, whereas two of the four poor responders in the bone marrow^positive cohort are deadof disease. Among12 patients with overt metastasis at primary diagnosis, 11 (92%) were positive in bone marrow with a very high number of osteosarcoma cells. The immunomagnetically isolated cells were further characterized by the use of fluorescent latex microparticles with surface-bound antibodies targeting different membrane markers. Moreover, in cases with numerous osteosarcoma cells in bone marrow attempts to grow the selected cells in vitro were successful in two of eight attempts, and in two of five cases, isolated cells produced tumors with osteosarcoma characteristics in nude mice. In conclusion, already at primary diagnosis, a very high fraction of osteosarcoma patients had malignant cells in bone marrow, and a correlation between the presence of tumor cells, clinical stage, and disease progression was found. The data show the clinical potential of this immunomagnetic method. Attempts to subgroup osteosarcoma patients for more individualized treatment based on the presence of micrometastatic cells should be studied in a larger cohort of patients.

Osteosarcoma

“…Support for this hypothesis in osteosarcoma includes the identification of osteosarcoma cells in the bone marrow of patients. (34) Following the rationale of this hypothetical model, cells would disseminate from the primary tumor early during tumor formation to the bone marrow. Metastatic cells then would persist during the period of dormancy in the bone marrow and then emerge subsequently and colonize distant secondary sites at the break in dormancy.…”

Osteoblast-induced EGFR/ERBB2 signaling in androgen-sensitive prostate carcinoma cells characterized by multiplex kinase activity profiling

“…Primary, polyclonal Immunomagnetic cell separation MOC-31 (IQ Corporation BV, Groningen, The Netherlands) is an IgG1 class antibody that binds to the EPCAM antigen, which is consistently expressed in most epithelial cells [9]. The high-affinity monoclonal antibody 9.2.27, which was originally developed against melanoma [10], recognizes an epitope on the high molecular weight melanoma-associated antigen and has also been shown to bind some subgroups of sarcoma, including osteosarcoma [11,12]. The antibodies were conjugated to superparamagnetic particles coated with polyclonal sheep-antimouse IgG particles (Dynabeads; Dynal A.S., Oslo, Norway).…”

“…The antibodies were conjugated to superparamagnetic particles coated with polyclonal sheep-antimouse IgG particles (Dynabeads; Dynal A.S., Oslo, Norway). At the end of experimental incubations, cocultures of LNCaP cells with OHS cells and of LNCaP cells with differentiated mesenchymal stem cells were detached, and the resulting single cell suspensions were subjected to immunomagnetic target cell isolation, essentially as previously described [12][13][14][15]. The positive cell fractions were examined by light microscopy for the principal presence of cells with C5 immunobeads bound to their surface (bead rosettes).…”

“…The substrate phosphorylation state generated by such LNCaP cells overlapped the phosphopeptide signatures identified in the paracrine LNCaP entities (Table 2). We have previously evaluated the immunomagnetic cell separation method for selective isolation of target cells and demonstrated that target cell populations are highly enriched [12][13][14][15]. But because kinase activity profiling might be sensitive to the possible presence of contaminants, we screened for phosphopeptide signatures generated by immunoselected as well as monocultured OHS cells.…”

Inhibition of protein phosphatase 2A with the small molecule LB100 overcomes cell cycle arrest in osteosarcoma after cisplatin treatment

“…1 Despite total surgical resection of the primary lesion, approximately 80% of patients go on to develop metastases, particularly to the lungs. 2 The addition of systemic chemotherapy has improved 5-year survivals to 70%, 3 but long-term survival beyond 10 y remains a challenge, as metastatic disease burden worsens. 1,4 Understanding the mechanisms involved in chemotherapeutic resistance, leading to metastases may uncover new therapeutic targets.…”