Perturbation of naive TCR transgenic T cell functional responses and upstream activation events by anti-CD4 monoclonal antibodies

Abstract

It is well established that non‐depleting anti‐CD4 monoclonal antibodies (mAb) have potent immunomodulatory properties in vivo and as such can induce a profound state of tolerance. Receptor blockade, CD4 modulation, or the transmission of a negative signal have all been proposed to explain their effects, however their precise mechanism of action, particularly at the level of cellular signaling, remains obscure. Experiments were thus carried out to examine the underlying mechanisms of action of two non‐depleting anti‐mouse CD4 mAb, YTS177 and KT6, which differ in their ability to modulate CD4 expression. The effects of the mAb were examined on CD4+ T cells derived from D0.11.10 TCR transgenic mice. Functional studies revealed that both mAb could effectively block antigen‐driven proliferation and IL‐2 production but had only modest effects at higher peptide doses. Importantly, mAb pre‐treatment of T cells stimulated by sub‐optimal peptide seemed to induce an anergy‐like state making them unresponsive to subsequent re‐stimulation. Analysis of intracellular signaling demonstrated that certain key upstream events such as the phosphorylation of Zap‐70 and LAT were also blocked by mAb pretreatment which may be due to interference with stable T cell‐APC conjugate formation.

Anti-coreceptor therapy drives selective T cell egress by suppressing inflammation-dependent chemotactic cues

“…signaling (28,37), suggesting that coreceptor therapy mediates islet T cell egress by inhibiting TCR signal transduction and downstream effector functions. To initially test this model, the effect of YTS177 and YTS105 on TCR-induced intracellular Ca 2+ flux was studied in vitro.…”

Engagement of the CD4 Receptor Affects the Redistribution of Lck to the Immunological Synapse in Primary T Cells: Implications for T-Cell Activation during Human Immunodeficiency Virus Type 1 Infection

“…However, the gp120-treated cells exhibited defective proliferation and failed to upregulate activation markers such as HLA-DR and CD25 (33). While the two anti-CD4 clones we tested significantly inhibited Lck recruitment to the IS, we have previously shown that only anti-domain 1 MAb or AT-2-treated HIV-1 virions had a significant impact on downstream signaling events such as CD40L upregulation by activated CD4 ϩ T cells (11,18,69). Previous studies have shown that different anti-CD4 MAbs or gp120 could trigger activation of Lck, but they differed in their ability to activate NF-AT (4), proliferation, or interleukin-2 (IL-2) production (11,18,69).…”

Recombinant Anti-CD4 Antibody 13B8.2 Blocks Membrane-Proximal Events by Excluding the Zap70 Molecule and Downstream Targets SLP-76, PLCγ1, and Vav-1 from the CD4-Segregated Brij 98 Detergent-Resistant Raft Domains

“…To our knowledge, little data are available concerning a "physical" disconnection of the Ab-target CD4 molecule with membrane-proximal Zap70 kinase through dynamically oriented raft segregation. Other anti-CD4 Abs (30,(31)(32)(33) are known to block Zap70 phosphorylation in vitro and in vivo, but neither CD4/Zap70 differential patching by confocal microscopy nor CD4/Zap70 differential raft partitioning has been observed. Although no link between murine parental mAb 13B8.2 and Zap70 kinase has ever been established, down-regulation of LFA-1-dependent adhesion of Jurkat T cells to B cells, induced by murine parental mAb 13B8.2 cross-linking, requires colocalization of p56 Lck, LFA-1, and PI3K, but not Src homology 2 domain-containing phosphatase SHP-2, in Brij 58 GM3 ϩ -membrane rafts together with CD4 (34,35).…”

“…This phenomenon was also observed by Houtman et al (62), who demonstrated that early Zap70 phosphorylation showed maximal effect of 50% at 19 s following anti-CD3 T cell stimulation, occurred maximally at 30 s, and persisted throughout the 120-s experiment. Rapid (Ͻ5 min) and strong Ag-specific phosphorylation of Zap70 has been shown to be stable for 1 h and can be reduced by nondepleting anti-CD4 Abs KT6 and YTS177 (31). Ab-induced CD20 redistribution into Triton DRM occurs as early as 15 s, becomes maximal at 15 min, and remains stable until the end of the 1-h experiment (4).…”

“…Epitope specificity governing raft localization and further biological effects have mainly been defined for anti-CD20 reagents (5,6,16). It would be interesting to compare the epitope specificity of anti-CD4 Abs YNB.46 (30), KT6, YTS177, and YTA3.1 (31,32), all of which are known to induce inhibition of Zap70 phosphorylation, with the CDR3-like epitope specificity (residues Glu 87 and Asp 88 in the D1 domain of CD4) of Ab 13B8.2 (64), which reorganizes Zap70 outside the rafts and modulates its phosphorylation profile. Concerning the anti-CD4 Ab OKT4, directed against the D4 domain, we were not able, using our experimental procedure, to induce CD4 accumulation/retention into 37°C-extracted Brij 98 DRM upon OKT4 binding without anti-mouse hyper-cross-linking, whereas Fragoso et al (33) showed an increase in the amount of CD4 together with LAT and p56 Lck in 4°C-Triton DRM upon OKT4 cross-linking, followed by anti-mouse hyper-cross-linking.…”