Recombinant immunotoxin (RIT) therapy is limited in patients by neutralizing antibody responses. Most patients with normal immune systems make neutralizing antibodies after one cycle (three doses) of RIT, preventing repeated dosing. Furthermore, some patients have preexisting antibodies from environmental exposure to Pseudomonas exotoxin (PE), the component of the RIT that elicits the neutralizing antibody response. Bortezomib is an FDA-approved proteasome inhibitor which selectively targets and kills plasma cells which are necessary for a neutralizing antibody response. Bortezomib, in combination with methotrexate, rituximab, and cyclophosphamide, was shown to reduce antibody levels >64-fold following enzyme replacement therapy in the clinical setting of Pompe disease. We hypothesized bortezomib may abrogate the neutralizing antibody levels, making dosing of RIT possible in mice already immune to RIT.

We immunized BALB/c mice with multiple doses of SS1P, a RIT whose antibody portion targets mesothelin. Antibody titers were determined by immune-capture ELISA. Mice with elevated levels were separated into groups to receive bortezomib (1mg/kg i.v., twice a week for 5.5 weeks) or saline control. Bortezomib treatment lowered the starting antibody titer in mice previously immunized with RIT by an average of 50%, while antibody titers in control mice increased by an average of 20%. Despite success in lowering titers, bortezomib did not abrogate titer in any mice.

To further diminish preexisting antibodies, we performed a separate experiment in which mice with preexisting anti-SS1P antibodies received bortezomib with concomitant pentostatin and cyclophosphamide (P/C) therapy. P/C is an immune-depleting regimen already shown to be effective in preventing the onset of new neutralizing antibodies. Combination treatment lowered the starting antibody titer in mice previously immunized with RIT by an average of 73%, while antibody titers in control mice increased by an average of 80%.

We have shown that bortezomib reduces, but does not eliminate, antibody titer in mice with preexisting antibodies to RIT. Addition of the immune-depleting P/C regimen further diminishes antibody titers. We are investigating remaining plasma cell populations as a potential source of remaining antibody. Ongoing experiments also include incorporating other FDA-approved agents into our study.

Citation Format: Michael L. Manning, Emily Mason-Osann, Masanori Onda, Ira Pastan. Bortezomib reduces preexisting antibodies to recombinant immunotoxins in mice. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2586. doi:10.1158/1538-7445.AM2014-2586