TCR engineered T cell therapies for hematologic malignancies

Division Of Basic Sciences - Nci

Investigators

Abstract

Activity 1: TCR class I and II epitope discovery for myeloid malignancies (AML/MDS/MPN): The objective of this activity is to discover epitopes that can be targeted with T cell receptors. The project is pursued in collaboration with Dr. Josh Elias (Stanford/Chan Zuckerberg Biohub). Recurrent somatic mutations or chromosomal breakpoints shared across patients with frequencies of at least 2% in a given disease category were curated using the publicly accessible COSMIC database. A total of 47 such genetic abnormalities were chosen, and expression vectors for tandem minigenes (as described by Tran E et al. Science 2014) encoding these genetic abnormalities were manufactured. Then a panel of artificial antigen presenting cells expressing tandem minigene with one or two HLA class I or II alleles were engineered. A total of sixty cell lines were established, which cover 12 most frequent HLA class I and II alleles in the US population. These artificial cell lines were lysed, peptide-MHCs were pulled down, and the lysates are currently being analyzed by mass spectrometry by Dr. Elias' team. Activity 2: Pre-clinical development of HLA-A*02:01-restricted CD22TCR: We have discovered T cell receptor that recognizes HLA-A*02:01-restricted epitope of CD22, which is an antigen expressed broadly by mature B-cell malignancies and a portion of acute lymphoblastic leukemia. The CD22TCR demonstrates promising pre-clinical anti-leukemia/lymphoma activity against multiple cell lines in both CD22- and HLA-A2-restricted manner. Furthermore, the CD22TCR does not demonstrate severe cross-reactivity that would cause off-target toxicities on normal tissues. We will soon complete the pre-clinical safety dataset necessary to make a final decision about clinical translation. We are also interested in studying the biological differences between our CD22TCR and CD22-directed chimeric antigen receptor (CAR)-T cells. CD22CAR-T cells are remarkably effective therapies for acute lymphoblastic leukemia. However, leukemia that reduce the levels of cell-surface CD22 expression escape CAR-T cells, leading to disease relapses. As mentioned in the Goals and Objectives section, TCR and CAR have different antigen recognition modalities, the former recognizing the intracellularly processed peptides on an MHC with exquisite sensitivity while the latter binding directly to the cell-surface protein. Therefore, we hypothesized that our CD22TCR can overcome CD22CAR resistant diseases, and we are actively studying this concept in preclinical models. Additionally, we are working with Dr. Ling Zhang (Genitourinary Malignancies Branch, CCR, NCI) who has extensive expertise in the safe delivery of membrane anchored IL-12 in the context of T cell therapy; we are studying whether combining Dr. Zhang's IL-12 construct would improve our CD22TCR-T cell efficacy. The preliminary results confirm that the IL-12 expressed on the T-cell surface improves CD22TCR-T cell activities against leukemia/lymphoma. Activity 3: Establishment of TCR cross-reactivity screening strategy using the HLA-A*02:01-restricted CD20TCR: Anti-CD20 TCRs were isolated from HLA-A2 transgenic mouse. However, the TCR was found to cross-react with irrelevant protein from the human proteome. Based on this result, the TCR will not be translated into clinic because it may cause off-target toxicities. However, another aspect of this project was that we were able to establish an efficient and successful strategy to detect clinically relevant cross-reactivity and to identify the specific cross-reactive epitope. This strategy was validated across multiple TCRs against different epitopes. Activity 4: Pre-clinical development of HLA-A*02:01-restricted EBV LMP2 TCR: Epstein Barr virus is an oncogenic virus associated with lymphoid malignancies, post-transplant lymphoproliferative disease, and gastric cancer, and nasopharyngeal cancers. EBV-associated diseases frequently express one of latency proteins called LMP2. T cells transduced with the TCR specific to the HLA-A*02:01-restricted epitope of EBV LMP2 demonstrated appropriate specificity and high functional avidity against low concentration of the cognate epitope. However, the TCR-T cells demonstrated limited recognition of HLA-A*02:01+ and EBV+ LMP2+ lymphoma cell lines and EBV-transformed lymphoblastoid cell lines. Further investigation revealed that the epitope of this TCR to be TAP-independently-presented epitope. EBV is known to express TAP inhibitor (BNLF2a) during lytic cycle. We are currently studying our hypothesis that manipulation of EBV latency and lytic cycles with epigenetic modifiers and immunomodulatory agents would induce EBV-derived TAP inhibitor, thereby making the EBV-infected cells more susceptible to the TCR-T cells.