A panel of human cell-based artificial APC enables the expansion of long-lived antigen-specific CD4+ T cells restricted by prevalent HLA-DR alleles

Abstract

Many preclinical experiments have attested to the critical role of CD4(+) T cell help in CD8(+) cytotoxic T lymphocyte (CTL)-mediated immunity. Recent clinical trials have demonstrated that reinfusion of CD4(+) T cells can induce responses in infectious diseases and cancer. However, few standardized and versatile systems exist to expand antigen-specific CD4(+) T(h) for clinical use. K562 is a human erythroleukemic cell line, which lacks expression of HLA class I and class II, invariant chain and HLA-DM but expresses adhesion molecules such as intercellular adhesion molecule-1 and leukocyte function-associated antigen-3. With this unique immunologic phenotype, K562 has been tested in clinical trials of cancer immunotherapy. Previously, we created a K562-based artificial antigen-presenting cell (aAPC) that generates ex vivo long-lived HLA-A2-restricted CD8(+) CTL with a central/effector memory phenotype armed with potent effector function. We successfully generated a clinical version of this aAPC and conducted a clinical trial where large numbers of anti-tumor CTL are reinfused to cancer patients. In this article, we shifted focus to CD4(+) T cells and developed a panel of novel K562-derived aAPC, where each expresses a different single HLA-DR allele, invariant chain, HLA-DM, CD80, CD83 and CD64; takes up soluble protein by endocytosis and processes and presents CD4(+) T-cell peptides. Using this aAPC, we were able to determine novel DR-restricted CD4(+) T-cell epitopes and expand long-lived CD4(+) T-cells specific for multiple antigens without growing bystander Foxp3(+) regulatory T cells. Our results suggest that K562-based aAPC may serve as a translatable platform to generate both antigen-specific CD8(+) CTL and CD4(+) T(h).

HLA-DP84Gly constitutively presents endogenous peptides generated by the class I antigen processing pathway

“…All DR-expressing T2 cells similarly produced and presented CLIP regardless of the DR allele tested, as previously reported (Supplementary Fig. 1a)21. T2 cells transduced with DP5 or DP8 also presented CLIP.…”

“…In the interest of demonstrating the reproducibility of these results, additional cell lines were examined. K562 cells lack the expression of HLA-class II, Ii and HLA-DM202122. Similar findings were obtained using a series of K562-derived transfectants expressing DR or DP alleles in conjunction with Ii (Supplementary Fig.…”

Key Residues at Third CDR3β Position Impact Structure and Antigen Recognition of Human Invariant NK TCRs

“…IL-2 ELISPOT assays were conducted as described elsewhere (33–36). Briefly, PVDF plates (Millipore, Etobicoke, ON) were coated with capture mAb (R&D Systems, Minneapolis, MN).…”

Specific Roles of Each TCR Hemichain in Generating Functional Chain-Centric TCR

“…Biotinylated HLA-A2/peptide monomers were purchased from ProImmune, multimerized in-house using SA-PE and SA-APC, and utilized to stain antigen-specific T cells as described previously (20, 37, 44, 45). A2/HIV multimer was always used as a control.…”