Directed-Complement Activation as a Novel Immunotherapeutic Approach for HER2-Breast Cancer.

PURPOSE

Directing selective complement activation towards tumor cells is an attractive strategy to promote their elimination. We have generated complement-activating multimeric immunotherapeutic complexes (CoMiX), stimulating either the alternative pathway (via Factor H Related protein 4 (FHR4)) or the classical pathway (via triple Fc dimers) on HER2-expressing tumor cells.

METHODS

We used the C-terminal α-chain multimerizing scaffold of the C4 binding protein (C4bp) to generate CoMiX-FHR4 as well as CoMiX-Fc with 2 different anti-HER2 VH, VH(T) and VH(P), recognizing trastuzumab- or pertuzumab-competing epitopes, respectively. The different CoMiX were compared in vitro for C3b and C5b9 depositions, complement-dependent cytotoxicity (CDC), and their ability to activate NK cells and phagocytosis by macrophages. We further explored their therapeutic efficacy on human BT474 tumor xenografts established in nude mice.

RESULTS

CoMiX-FHR4/VH(T) and -FHR4/VH(P) lead to the highest C3b and C5b9 depositions and CDC on BT474 tumor cells (p<0.0001), both individually and in combinations with their CoMiX-Fc counterparts, surpassing the low complement activating capacity of trastuzumab and pertuzumab. All CoMiX induced BT474 cell death and phagocytosis of tumor cells by macrophages while CoMiX-Fc also stimulated NK cell activation. In human BT474 xenografts sensitive to trastuzumab, CoMiX induced a massive C3b deposition 6 hours after injection. CoMiX-FHR4 reduced the tumor volume compared to controls (p< 0.05) but to a lesser extent than trastuzumab (p< 0.001) while CoMiX-VH(P)/Fc led to a tumor volume reduction similar to pertuzumab. Combinations of two CoMiX-FHR4 or two CoMiX-Fc were more potent, similarly to the combination of trastuzumab and pertuzumab, leading to increased NK cell infiltration in xenografts. Importantly, CoMiX-FHR4 was still active against trastuzumab-resistant xenografts, delaying tumor growth and inducing a large NK cell infiltration.

CONCLUSION

We showed here that directed complement activation on tumor cells is an alternative to therapeutic antibodies for future combination therapies upon resistance to standard-of-care treatment.