A Novel Murine Model of Human Disseminated Multiple Myeloma with A Subcutaneous Human Bone Chip from an Adult Donor Confers Proliferative Advantage and Is Suitable for the In Vivo Evaluation of Novel Drugs
Authors: Ocio Enrique M. , San-Segundo Laura , Blanco Juan , Hernández-Iglesias Teresa , Fernández-Lázaro Diego , Paíno Teresa , Garayoa Mercedes , Álava Enrique , Jiménez-Barredo Fernando , Díaz-Rodríguez Elena , Pandiella Atanasio , San Miguel J.F 
Published: October 2019

Abstract
Abstract 4077
Several murine models resembling multiple myeloma have been developed in the last years: the subcutaneous plasmocytoma model and the disseminated model, based on the iv or intracardiac injection of MM cells, have been widely used to test the efficacy of novel agents. Nevertheless, it is well known the crucial role that the interaction between MM plasma cells and the bone marrow microenvironment plays in MM pathogenesis as well as in the proliferation of MM cells and drug resistance; however, unfortunately, most murine models do not recapitulate these interactions. In order to overcome this caveat, other models such as the SCIDhu or the SCIDrab have been developed. However, in these models, the induced microenvironmental milleu is derived from a fetus and a rabbit respectively, which can be relatively different from that found in an adult patient with MM. We have developed a novel model of a human disseminated MM with the presence of human adult microenvironment.
For this purpose, CB17-SCID mice, which lack B and T lymphocytes, were anesthetized and a human bone chip obtained from a healthy adult patient undergoing a hip replacement due to a non-malignant fracture, was subcutaneously implanted in the back of the mice. In the same day 3×106 MM1S-Luc cells were iv injected through the tail vein. Two groups were also included as controls in the study, one with MM1S-Luc cells without the bone chip, and the other with only the human bone chip (without MM cells). Luciferase activity was measured three times per week using a Xenogen-IVIS 50 system after ip injection of 0,15 mg/Kg luciferin. Mice were sacrificed when they developed signs associated with advanced disease, such as body weight loss or back limb paralysis.
Although the group of mice receiving MM1S-Luc without bone, did develop a disseminated myeloma, this was much more precocious in the group of mice with a human bone subcutaneous implant. In fact, in mice with human bone and MM1S-Luc cells, the luciferase activity was detectable a median of 9 days after the injection of the cells whereas it was necessary 16 days for obtaining the same effect in mice without the bone chip. This is pointing out that the bone chip in these mice induces a favourable environment for myeloma growth and could be appropriate model to study the interactions of MM cells with human stroma. The increase of tumour growth rate also correlated with a decrease in survival. Median survival (CI 95%) of 64 (47-80) days for mice without bone vs 48 (45-50) days for mice with human bone (Log Rank=0.004). Regarding the sites of infiltration, luminescence images showed a disseminated involvement in all skeletal regions, and this was confirmed by histopathology, as there was a general infiltration of spine, femora, tibia and cranium of these mice. By contrast we did not find infiltration of extra-skeletal regions such as heart, lung or spleen in any mice and only unclear infiltrates were observed in the kidney and liver of some mice.
In order to validate the suitability of this model for testing the in vivo antimyeloma activity of novel drugs in the presence of a human microenvironment, 6 mice bearing this novel model were randomized to receive: bortezomib (0.5 mg/Kg ip five days per week) + dexamethasone (0.5 mg/Kg 2 days per week) or vehicle control. A second group of the same number of mice with MM1S-Luc but without the bone chip were randomized in the same way and used as control. Bortezomib + dex significantly decreased tumour growth in both groups of mice without any clear differences in activity between both models, suggesting that this combination overcomes the influence of the microenvironment.
In summary we have developed a murine model of MM that may allows us to 1) gain insights into the role of the BM microenvironment interactions in MM pathogenesis. 2) Evaluate the in vivo activity of different anti-MM agents in the presence of human microenvironment. 3) Potentially, this model could also contribute to analyze the effect of different agents in the myeloma bone disease.

Disclosures:
No relevant conflicts of interest to declare.