Generation of T Cells of Desired HLA Restriction Specific for Epitopes of a Self-Antigen, WT1, for the Adoptive Immunotherapy of WT1 Positive Malignancies Using A Panel of Artificial Antigen Presenting Cells Expressing Prevalent HLA Alleles.

Abstract 4086

Poster Board III-1021

We have recently established a panel of artificial antigen-presenting cells (AAPCs) each expressing a single HLA class I allele (such as A0201, A0301, A2402, B0702, C0401) and costimulatory molecules (B7.1, ICAMI, LFA3, b2-microglobulin). These cells were capable of eliciting T cells responses specific for immunogenic epitopes of the cytomegalovirus protein CMVpp65 presented by single HLA allele expressed by each AAPC (J.Immunol, 2009;183(4):2837-50). The initial studies with HLA-A0201 expressing AAPCs had suggested that they could sensitize the T cells against single immunogenic A0201-binding peptides derived from antigens such as S100 and telomerase. However, there are no data regarding the potentials of AAPCs expressing other HLA alleles to present epitopes of endogenous self antigens and stimulate specific T cells. We have recently defined a series of immunogenic epitopes for WT1 that can be presented by each of the HLA alleles represented in this panel, by an epitope mapping analysis of WT1 specific T cells sensitized in vitro with autologous dendritic cells(DC) loaded with the pool of overlapping 15-mer peptides spanning the sequence of WT1. In the present study we asked whether the established panel of AAPCs expressing different HLA alleles when loaded with the human WT1 peptide pool, could be used to generate T cells directed against this self antigen, whether the peptides recognized by these T cells would be identical to the peptides defined to be immunogenic when presented in the context of the same HLA alleles after stimulation with the autologous DC loaded with the same WT1 pool and further whether AAPCs could also be used to elicit responses against subdominant epitopes presented by an HLA allele expressed on AAPCs that were not elicited among T cells sensitized with peptide pool loaded autologous DC. The pool of 141 synthetic pentadecapeptides each overlapping the next by 11aa loaded on the autologous DC elicited responses in 80% of 14 normal individuals tested. Epitope mapping permitted identification immunodominant WT1 peptide sequences eliciting responses and their restricting HLA allele(s) as determined in a Cr51 release assay against the panel of WT1 peptide loaded EBV transformed B cells matching one of the HLA alleles of the T cell genotype.. AAPCs expressing single HLA allele shared by the same T cells and loaded with the same WT1 total peptide pool elicited responses in each normal individual. In each case the T cells recognized one of the same epitopes defined to be previously immunogenic in man when presented in the context of the HLA allele expressed by the AAPCs. At the same time, single peptide epitopes that were defined to be presented by, for example, HLA A0201 alleles after stimulation with WT1 total pool loaded DCs did not elicit WT1 specific T cells responses if they were loaded on AAPCs expressing another HLA allele such as B0702. In comparisons of T cells sensitized with autologous DCs versus AAPCs loaded with WT1 peptides pool we also found that while AAPCs expressing the HLA allele presenting the immunodominant epitope recognized by T cells sensitized with pool loaded autologous DC also elicited strong responses, AAPCs expressing HLA alleles shared by the responding T cells could also elicit responses against immunogenic but subdominant epitopes that were not generated after sensitization of the same T cells with WT1 peptide pool loaded autologous DCs. Thus, these studies suggest that the panel of murine derived AAPCs can effectively present self antigens, such as WT1, and permits in vitro sensitization and propagation of tumor reactive WT1 peptide specific T cells of desired HLA restriction for the adoptive immunotherapy of WT1⁺ malignancies.