Entire CD3ε, δ, and γ humanized mouse to evaluate human CD3–mediated therapeutics

Abstract

T cell–mediated immunotherapy is an attractive strategy for treatment in various disease areas. In this therapeutic approach, the CD3 complex is one of the key molecules to modulate T cell functions; however, in many cases, we cannot evaluate the drug candidates in animal experiments because the therapeutics, usually monoclonal antibodies specific to human CD3, cannot react to mouse endogenous Cd3. Although immunodeficient mice transfused with human hematopoietic stem or precursor cells, known as humanized mice, are available for these studies, mice humanized in this manner are not completely immune competent. In this study we have succeeded in establishing a novel mouse strain in which all the three components of the Cd3 complex — Cd3ε, Cd3δ, and Cd3γ — are replaced by their human counterparts, CD3E, CD3D, and CD3G. Basic immunological assessments have confirmed that this strain of human CD3 EDG–replaced mice are entirely immune competent, and we have also demonstrated that a bispecific antibody that simultaneously binds to human CD3 and a tumor-associated antigen (e.g. ERBB2 or GPC3) can be evaluated in human CD3 EDG–replaced mice engrafted with tumors. Our mouse model provides a novel means to evaluate the in vivo efficacy of human CD3–mediated therapy.

CD3-Bispecific Antibody Therapy Turns Solid Tumors into Inflammatory Sites but Does Not Install Protective Memory

“…Even the more elegant models in this research field, in which human CD3E is transgenically expressed in mouse T cells and human TAA are expressed in mouse tumor cell lines under heterologous promoters, fall short, due to artificial expression levels of the CD3 and TAA (21,22). Indeed, the expression levels and frequency of CD3E in these transgenic mouse models are lower than those in wild-type mice (24). Recently, a triple knock-in mouse was reported in which human CD3E, -D, and -G genes replace the mouse homologous genes CD3e, -d, and -g. In this knock-in mouse, T-cell numbers and CD3 expression levels are comparable to those in wild-type mice and treatment with CD3 bsAb binding to human CD3 can representatively be evaluated (23,24).…”

“…Indeed, the expression levels and frequency of CD3E in these transgenic mouse models are lower than those in wild-type mice (24). Recently, a triple knock-in mouse was reported in which human CD3E, -D, and -G genes replace the mouse homologous genes CD3e, -d, and -g. In this knock-in mouse, T-cell numbers and CD3 expression levels are comparable to those in wild-type mice and treatment with CD3 bsAb binding to human CD3 can representatively be evaluated (23,24). In our syngeneic mouse model, we report a strong increase of innate immune cells, such as inflammatory macrophages and natural killer cells, and a fast intratumoral activation of CD4 þ and CD8 þ T-cell subsets (Figs.…”

“…Recently, transgenic mice that coexpress human CD3e (hCD3e) on their T cells, in addition to mouse CD3e, were exploited in this research field, enabling the use of endogenous T cells in immune-competent mice (21,22). Although hCD3e transgenic mice harbor a relatively normal immune system and allow analysis of immune-tumor cell interaction, they still lack a natural distribution of the selected TAA in mouse tissues, hampering examination of "on-target, off-tumor" effects via target expression in host tissues (21)(22)(23)(24). Moreover, induction of neutralizing anti-human Ig antibodies prevents longterm evaluation in these mouse models.…”

LCK as a Potential Therapeutic Target for Acute Rejection after Kidney Transplantation: A Bioinformatics Clue

“…In the cocitation network, CD3D, CD3E, and CD3G, which are compositions of CD3 complex of TCR, interacted with each other to affect the assembly of TCR membrane complex and disturb T-cell responsiveness [ 27 ], especially CD3E [ 28 ]. After TCR engagement, the phosphorylation of CD3 immunoreceptor tyrosine-based activation motifs of CD3E in CD3 complex is combined with activated LCK, which is also called Src kinase lymphocyte kinase [ 29 ].…”

Anti-GPRC5D/CD3 Bispecific T-Cell–Redirecting Antibody for the Treatment of Multiple Myeloma

“…Expression vectors transfected into FreeStyle293 cells were expressed as recombinant IgGs, which were then purified from culture supernatants using rProtein A-Sepharose (GE Healthcare). GPRC5D TRABs were generated by Fab-arm exchange as described previously (16).…”