Murinization and H Chain Isotype Matching of the Anti-GITR Antibody DTA-1 Reduces Immunogenicity-Mediated Anaphylaxis in C57BL/6 Mice

Abstract

Recent advances in immuno-oncology have shown that the immune system can be activated to induce long-term, durable antitumor responses. For immuno-oncology drug development, immune activation is often explored using rat Abs in immunocompetent mouse models. Although these models can be used to show efficacy, antidrug immune responses to experimental protein-based therapeutics can arise. Immunogenicity of surrogate Abs may therefore represent an important obstacle to the evaluation of the antitumor efficacy of immunomodulator Abs in syngeneic models. A recent publication has shown that anti-glucocorticoid–induced TNFR family–related protein agonistic Ab DTA-1 (rat or murinized IgG2a) can induce the development of anaphylaxis in C57BL/6 mice upon repeated i.p. dosing because of an anti-idiotypic anti-drug Ab immune response. This study was undertaken to address the impact of the immunogenicity derived from the Fc and variable domains. To this end, chimerized (rat V domains/mouse constant regions) and murinized (95% mouse sequence) DTA-1–based surrogate Abs with a murine IgG2c H chain isotype were created. Chimerization and murinization of DTA-1 did not affect receptor binding and glucocorticoid-induced TNFR family–related protein–induced T cell agonistic properties. Similar in vivo antitumor efficacy and intratumoral CD8+/regulatory T cells were also observed. Finally, treatment of C57BL/6 mice with the chimerized and murinized DTA-1 Abs on a C57BL/6-matched IgG2c isotype resulted in reduced development and severity of anaphylaxis as measured by decline of body temperature, behavioral effects, serum IL-4, IgE, and anti-drug Ab levels. These results suggest that careful murinization and selection of a strain-matched H chain isotype are critical to generate ideal surrogate Abs for testing immuno-oncology mechanisms in vivo.

An anti-PD-1–GITR-L bispecific agonist induces GITR clustering-mediated T cell activation for cancer immunotherapy

“…Our anti-PD-1–GITR-L is highly efficacious in different anti-PD-1-resistant syngeneic models and has a distinct MoA in comparison to the combination of anti-PD-1 and anti-GITR antibodies (efficacious only in the highly immunogenic, anti-PD-1-responsive MC-38), where anti-GITR (rIgG2b, FcγR effector active) mainly works by induction of T reg depletion and anti-PD-1 induces PD-L1 inhibition due to prolonged saturation of PD-1 (refs. 51 , 52 ). The anti-PD-1–GITR-L is a PD-1-directed GITR-L that may enhance binding of GITR-L to PD-1 + CD8 + T cells due to higher levels of PD-1 expression in CD8 + versus CD4 + T cells.…”

HERA-GITRL activates T cells and promotes anti-tumor efficacy independent of FcγR-binding functionality

“…As an additional complication, the benchmark anti-mouse GITR antibody (clone DTA-1) is a rat IgG2b that causes severe anaphylaxis when repeatedly injected into mice [27]. Unfortunately, even “murinized” DTA-1 antibodies can cause anaphylaxis due to mouse strain differences in IgG2 expression [28].…”