Antibody Targeting to a Class I MHC-Peptide Epitope Promotes Tumor Cell Death

Abstract

Therapeutic mAbs that target tumor-associated Ags on the surface of malignant cells have proven to be an effective and specific option for the treatment of certain cancers. However, many of these protein markers of carcinogenesis are not expressed on the cells’ surface. Instead these tumor-associated Ags are processed into peptides that are presented at the cell surface, in the context of MHC class I molecules, where they become targets for T cells. To tap this vast source of tumor Ags, we generated a murine IgG2a mAb, 3.2G1, endowed with TCR-like binding specificity for peptide-HLA-A*0201 (HLA-A2) complex and designated this class of Ab as TCR mimics (TCRm). The 3.2G1 TCRm recognizes the GVL peptide (GVLPALPQV) from human chorionic gonadotropin β presented by the peptide-HLA-A*0201 complex. When used in immunofluorescent staining reactions using GVL peptide-loaded T2 cells, the 3.2G1 TCRm specifically stained the cells in a peptide and Ab concentration-dependent manner. Staining intensity correlated with the extent of cell lysis by complement-dependent cytotoxicity (CDC), and a peptide concentration-dependent threshold level existed for the CDC reaction. Staining of human tumor lines demonstrated that 3.2G1 TCRm was able to recognize endogenously processed peptide and that the breast cancer cell line MDA-MB-231 highly expressed the target epitope. The 3.2G1 TCRm-mediated CDC and Ab-dependent cellular cytotoxicity of a human breast carcinoma line in vitro and inhibited in vivo tumor implantation and growth in nude mice. These results provide validation for the development of novel TCRm therapeutic reagents that specifically target and kill tumors via recognition and binding to MHC-peptide epitopes.

TCR Mimic Monoclonal Antibody Targets a Specific Peptide/HLA Class I Complex and Significantly Impedes Tumor Growth In Vivo Using Breast Cancer Models

“…We showed previously the IgG2a Ab RL4B (3.2G1) TCRm prevented tumor growth in a prophylactic model of human breast cancer (41). Further, the RL4B TCRm demonstrated CDC and ADCC activity against tumor cells in vitro.…”

“…Many methods rely on using bacteriophage display technology to generate antipeptide/MHC class I Abs from nonimmunized and immunized libraries (36,38,39). Our approach combines immunization with synthetic immunogen and high throughput screening techniques to generate antipeptide/MHC Abs we have dubbed TCR mimic (TCRm) Abs (40,41). The majority of the Abs generated by our laboratory using this process have shown high binding affinity and fine binding specificity, and they can be used for the detection and quantitation of peptide/MHC class I molecules (32,33).…”

TCR Mimic Monoclonal Antibodies Induce Apoptosis of Tumor Cells via Immune Effector-Independent Mechanisms

“…The BB7.2 (anti-HLA-A2 mAb)-and W6/32 (pan-MHC class I mAb)-expressing mouse hybridoma cell lines were purchased from the American Type Culture Collection. RL4B and RL6A TCRm were described previously (5,10). RL21A TCRm (mouse IgG2a) is specific for peptide FLSELTQQL derived from protein phage migration inhibitory protein in the context of HLA-A2.…”

“…The RL6A recognizes peptide YLLPAIVHI from human p68 RNA helicase and presented by HLA-A*0201 (10). Although it has been shown that these mAbs induce complement-dependent cytotoxicity (CDC) and Ab-dependent cellular cytotoxicity (ADCC) in vitro (5,10), it remains to be elucidated how these Abs kill tumor cells in vivo.…”

“…Intact intracellular proteins are shielded from the immune system recognition; however, peptides originating from these proteins are presented on the surface of all nucleated cells, including tumor cells, by MHC class I molecules. Thus, mAbs that recognize these complexes can become a new class of therapeutics for targeting tumor cells (5)(6)(7). We termed these Abs TCR mimic mAbs (TCRm), as T cells, similar to TCRm, recognize peptides presented by MHC class I molecules through the TCR engagement.…”