Obinutuzumab-Based Drug-Free Macromolecular Therapeutics Synergizes with Topoisomerase Inhibitors

Drug-Free Macromolecular Therapeutics (DFMT) utilizes modified monoclonal antibodies (or antibody fragments) to generate antigen-crosslinking induced apoptosis in target cells. DFMT is a two-component system containing a morpholino oligonucleotide (MORF1) modified antibody (Ab-MORF1) and human serum albumin conjugated with multiple copies of complementary morpholino oligonucleotide (MORF2), (HSA-(MORF2)x ). The two components recognize each other via Watson-Crick base pairing complementation of their respective MORFs. One HSA-(MORF2)x molecule can hybridize with multiple Ab-MORF1 molecules on the cell surface, thus serving as the therapeutic crosslink-inducing mechanism of action. Herein, we examined various anti-neoplastic agents in combination with the anti-CD20 Obinutuzumab (OBN)-based DFMT system. We examined three different classes of chemotherapies: DNA alkylating agents; proliferation pathway inhibitors; and DNA replication inhibitors. We utilized Chou-Talalay combination index mathematics to determine which drugs engaged synergistically with OBN-based DFMT. We determined that OBN-based DFMT synergizes with topoisomerase inhibitors and DNA nucleotide analogues but is antagonistic with proliferation pathway inhibitors. We performed cell mechanism experiments to analyze points of synergism or antagonism by investigating Ca2+ influx, mitochondrial health, lysosomal stability, and cell cycle arrest. Finally, we demonstrated the synergistic drug combinatorial effects of OBN-based DFMT with etoposide in vivo using a human xenograft non-Hodgkin's lymphoma mouse model.