Mutations resulting in the formation of hyperactive complement convertases support cytocidal effect of anti-CD20 immunotherapeutics

Abstract

Anti-CD20 monoclonal antibodies (mAbs) rituximab and ofatumumab are potent activators of the classical complement pathway, and have been approved for the treatment of B-cell malignancies. However, complement exhaustion and overexpression of complement inhibitors by cancer cells diminish their therapeutic potential. The strategies of targeting membrane complement inhibitors by function-blocking antibodies and the supplementation with fresh frozen plasma have been proposed to overcome tumour cell resistance. We present a novel approach, which utilizes gain-of-function variants of complement factor B (FB), a component of alternative C3/C5 convertases, which augment mAb-activated reactions through a positive feedback mechanism called an amplification loop. If complement concentration is limited, an addition of quadruple gain-of-function FB mutant p.D279G p.F286L p.K323E p.Y363A (or selected single mutants) results in significantly increased complement-mediated lysis of ofatumumab-resistant tumour cells, as well as the complete lysis of moderately sensitive cells. Importantly, this effect cannot be achieved by further increasing ofatumumab concentration. Potentiation of cytotoxic effect towards moderately sensitive cells was less apparent at physiological serum concentration. However, an addition of hyperactive FB could compensate the loss of cytotoxic potential of serum collected from the NHL and CLL patients after infusion of rituximab. Residual levels of rituximab in such sera, in combination with added FB, were able to efficiently lyse tumour cells. We suggest that the administration of gain-of-function variants of FB can restore cytotoxic potential of complement-exhausted serum and maximize the therapeutic effect of circulating anti-CD20 mAbs. Electronic supplementary material The online version of this article (10.1007/s00262-019-02304-0) contains supplementary material, which is available to authorized users.

The Role of Complement in the Mechanism of Action of Therapeutic Anti-Cancer mAbs

“…It is clear that manipulation of the complement cascade, through the design of anti-tumor mAbs with increased ability to activate complement [ 44 , 59 , 96 ], or with mAbs that hyperactivate the complement cascade [ 97 ] or block soluble or membrane bound inhibitors [ 63 , 84 , 91 ], may all be feasible strategies to enhance the CDC activity of mAbs for cancer immunotherapy. However, there is still the need for the demonstration that each of these strategies has efficacy in vivo.…”

Hydrogen deuterium exchange mass spectrometry identifies the dominant paratope in CD20 antigen binding to the NCD1.2 monoclonal antibody

“…Future mutagenesis of the CDR3 (V H ) domain, the Fc domain, as well as ‘canine-izing’ the variable framework domains, might optimize the immunochemical properties of the recombinant NCD1.2 antibody. In addition, as complement mutants are emerging that might augment the human anti-CD20 monoclonal antibody [ 53 ], canine cancers might form a physiological preclinical testbed for translating such novel combined synthetic biologics for use in human lymphoma.…”

Complement system in Anti-CD20 mAb therapy for cancer: A mini-review

“…[7][8][9][10] Manipulating complement cascade can serve as an effective strategy to increase the anti-cancer efficacy of anti-CD20 mAbs. 1,2,[11][12][13][14] In innate immunity, the complement system is a key component. The activation of the complement cascade is strictly restricted by different regulators in order to protect host tissues from being attacked, though, it can be detected on the surface of cancer cells and the fluids of cancer patients.…”

“…Lymphoma is a group of cancers that start in the lymph system. Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) are two main types of lymphoma, yet there also exist additional 1 , 2 subclasses of lymphoma based on lymphoid lineage cells that expand and undergo malignant transformation. 3 , 4 Lymphoma can occur in all age groups, yet HL is more common among people aged 15–39 years and 75 years or older than other age groups while the rates of NHL grow higher as people get old.…”

“… 7 – 10 Manipulating complement cascade can serve as an effective strategy to increase the anti-cancer efficacy of anti-CD20 mAbs. 1 , 2 , 11 – 14 …”