Myeloma-Specific Oncolytic Adenovirus Induces Significant Tumour Oncolysis In Vitro and In Vivo and Prevents Cell Line Regrowth

Multiple myeloma is a largely incurable disease and despite current therapies achieving good initial responses, patients frequently relapse. Therefore, new approaches are required that not only reduce the tumour load, but also prevent the growth of residual disease. One such approach is the use of oncolytic viruses. We developed an oncolytic adenovirus that utilizes transcriptional control of E1A under the myeloma-specific promoter CS1 (ADCE1A). We hypothesised that ADCE1A would be myeloma-specific, inducing tumour oncolysis and preventing tumour regrowth.

A panel of myeloma cell lines (JJN3, L-363, OPM-2, U-266, RMPI-8226, NCI-H929, KMS-11, KMS-12-BM) were assessed for expression of CS1 by flow cytometry (FC). The same panel were treated with ADCE1A and cell death was monitored after 72 hours using FC and propidium iodide staining. Apoptosis was assessed following ADCE1A infection using annexin V staining and analysed by FC at 6 and 24 hours post treatment. CD138⁺ plasma cells from bone marrow aspirates were obtained from myeloma and plasma cell leukaemia patients and from the peripheral blood from healthy donors. The CD138⁺ and CD138^- populations from these samples were treated with ADCE1A and cell death was monitored after 4 days using FC and propidium iodide staining. Myeloma cell regrowth was assessed after bortezomib (0.56-2.81nM) or bortezomib in combination with ADCE1A treatment using cell counts. Viral efficacy was tested in a xenograft model of myeloma, where 5 weeks after tumour cell injection (106 U266 cells intravenously), mice were treated with ADCE1A (1x107 pfu, 2x/wk) or control (PBS) for 3 weeks. Tumour burden was measured ex vivo in bone marrow flushes of the long bones by FC.

CS1 was expressed in all myeloma cell lines, except KMS-12-BM. ADCE1A infected, replicated and caused oncolysis in JJN-3, L-363, OPM-2, U-266, RPMI-8226, NCI-H929, and KMS-11 myeloma cell lines. However, KMS-12-BM had the lowest sensitivity to ADCE1A. This correlated with CS1 expression, as CS1 was not expressed at the protein level in this cell line. Apoptosis, as detected by annexin V staining, was found to be a cell death mechanism involved in ADCE1A oncolysis. Importantly, ADCE1A induced oncolysis in primary patient malignant CD138⁺ plasma cells, but not in the non-malignant CD138^- bone marrow mononuclear population from these patients. Additionally, ADCE1A had no effect on cell death in non-malignant CD138⁺ plasma cells and non-malignant CD138^- peripheral blood mononuclear population from healthy donors. ADCE1A prevented regrowth of myeloma cell lines following treatment with bortezomib in vitro. In the U266 xenograft model, tumour load was significantly reduced (p<0.05) compared to control treated mice.

In summary, ADCE1A has potential clinical efficacy as shown by preclinical models and patient tumour samples. Additionally, ADCE1A was able to stop tumour cell regrowth after chemotherapy in vitro, therefore, the use of oncolytic adenoviruses to target minimal residual disease may be a novel yet promising approach for the treatment of myeloma.