A Clinically Relevant Zebrafish in Vivo Model of Human Multiple Myeloma (MM) to Study Disease Biology and Preclinical Therapeutic Efficacy

Abstract 4078

Although a number of murine models of human MM have been developed and the SCID-hu murine model allows for growth of primary MM cells, these models have significant limitations including need for insertion of human bones, generation of limited number of mice with human MM Growth and over 2 months lag period, highlighting the need for a model that allows growth of human MM cells rapidly and in larger number of animals. We here describe a zebrafish model which can sustain human patient MM cell growth and be able to test efficacy of therapeutic agents. In this model we have utilized Casper fish, which are transparent and hence ideal for in vivo observation of tumor, Moreover, their 48 hours post fertilization (hpf) stage embryos are immune deficient allowing growth of xenogeneic human MM cells. We have tested growth of both MM cell lines as well as primary patient cells. About 50–200 cells from MM cell lines or CD138⁺ plasma cells from MM patient bone marrow samples labeled by CM-Dil were injected into the abdominal perivitelline space of 48 hpf stage Casper fish larvae and observed tumor progression, including tumor size and cell invasion by fluorescence microscopy at 24, 48 and 72 hrs post injection. MM cells were confirmed by immunohistochemistry. We observe over 80% larvae demonstrating MM cell survival for up to 9 days. As we require very small number of cells we were able to inject and observe growth of primary cells from all the patients producing over 50 larvae, We next treated the MM cells in vivo by adding first common anti-MM agents into fish water through 24 hrs to 72 hrs post injection (hpi) and observed change in tumor cell survival by fluorescence microscopy again at 24, 48 and 72 hours post treatment (hpt). We have here evaluated response of MM cell lines and MM primary patient cells to single or combination drug treatment, and defined response. We have observed response of the traditional and novel agents as predicted. For example, we have confirmed the response of dexamethasone to MM1.S and MM1.R cell lines which are sensitive or resistant to dexemathasone in this model. We observed that 100 nM dexemathasone treatment led to observation of MM cell survival, and found response ratio of MM1.S and MM1.R xenograft tumors to were 37.5% vs 87.5% fish at 24 hpt and 26.7% vs 87.5% at 48 hpt respectively. We have confirmed efficacy of bortezomib as well as lenalidomide in this model using MM cell lines as well as observed efficacy against primary MM cells. Importantly, we have been able to confirm similar drug resistance profile as patients; for example cells from a patient with bortezomib resistance survived bortezomib treatment in this model. Any combination treatment produced higher response than single drug treatment indicating that MM xenograft zebrafish model can predict drug response accurately. The zebrafish larvaes tolerated all the treatment at the dose utilized well. In conclusion, we have characterized a highly-reproducible zebrafish model of human myeloma that supports primary myeloma cell growth using very small number of cells and can allow generation of large number of fish with evalution of drug activity in a very short time of 3–5 days. This model provides a powerful tool to evaluate preclinical efficacy of novel agents and combinations and may provide opportunity to evaluate personalized therapy.