Cancer therapy in mice using a pure population of CD8+ T cell specific to the AH1 tumor rejection antigen

There is a growing interest in the use of patient-derived T cells for the treatment of various types of malignancies. The expansion of a polyclonal and polyspecific population of tumor-reactive T cells, with a subsequent infusion into the same donor patient, has been implemented, sometimes with positive results. It is not known, however, whether a set of T cells with a single antigen specificity may be sufficient for an effective therapy. To gain more insights in this matter, we used naturally occurring T cells recognizing a retroviral peptide (AH1), which is endogenous in many tumor cell lines of BALB/c origin and which serves as potent tumor rejection antigen. We were able to isolate and expand this rare population of T cells to numbers suitable for therapy experiments in mice (i.e., up to 30 × 106 cells/mouse). After the expansion process, T cells efficiently killed antigen-positive tumor cells in vitro and demonstrated tumor growth inhibition in two syngeneic murine models of cancer. However, AH1-specific T cells failed to induce complete regressions of established tumors. The incomplete activity was associated with a failure of injected T cells to survive in vivo, as only a very limited amount of T cells was found in tumor or secondary lymphoid organs 72 h after injection. These data suggest that future therapeutic strategies based on autologous T cells may require the potentiation of tumor-homing and survival properties of cancer-specific T cells.

Identification and expression distribution of esophageal carcinoma autoantigens.

e16520 Background: To identify the autoantigen protein molecules with autoserum in the tissues from the esophageal carcinoma (EC) patients, analyze autoantigen expression distribution in EC tissues, so as to provide basis for the molecular pathogenesis and clinical medication of EC. Methods: 69 cases of EC patients tissues and serum and 81 cases of healthy people serum were collected, serological proteome analysis (SERPA) was modified with sequential extraction of subcellular protein fractions to identify esophageal oncopathgensis stages autoantigen with autoserum in the tissues from the EC patients. Another 93 cases of EC patients tissue were collected, immunohistochemical and western blot method were used to detect expression distribution of EC autoantigens in esophageal carcinoma tissue, para-carcinoma tissue and normal tissue. Results: Autoantigens CK13, CK16, CaD, ACTG2, tumor rejection antigen (gp96) 1 variant, heat shock protein gp96 precursor were identified, among wihich, CK16, CaD, ACTG2, tumor rejection antigen (gp96)1 variant, heat shock protein gp96 precursor were firstly reported as EC autoantigens. Expression of autoantigens CK16, CaD, ACTG2 were increased in EC carcinoma tissue than para-carcinoma tissue and normal tissue, while CK13 were decreased. Positive expression level of CK16 in normal tissue, para-carcinoma tissue and cancer tissue of EC patients was 0.0076±0.0033, 0.0158±0.0065, 0.0356±0.0165 respectively, CaD was 0.0085±0.0048, 0.0107±0.0056, 0.0177±0.0103 respectively, ACTG2 was 0.0091±0.0039, 0.0136±0.0043, 0.0214±0.0110 respectively, and CK13 was 0.2053±0.0311, 0.1633±0.0280, 0.0412±0.0239 respectively. Conclusions: 6 EC autoantigens were identified, and 5 were first reported. Autoantigens CK13, CK16, CaD and ACTG2 were expressed in EC patients carcinoma tissue, which can be the potential biomarkers of esophageal carcinoma. This study provides new basis for the EC molecular mechanism and development of molecular drugs.

Immunotherapy of CT26 murine tumors is characterized by an oligoclonal response against the AH1 tumor rejection antigen

The possibility to cure immunocompetent mice bearing murine CT26 colorectal tumors using cytokine-based therapeutics allows to study the tumor rejection process at a molecular level. Following treatment with L19-mIL12 or F8-mTNF, two antibody fusion proteins which preferentially concentrate a murine cytokine payload at the tumor site, CT26 tumors could be cured in a process that crucially relies on CD8+ T cells. In both settings, the AH1 peptide (derived from the gp70 envelop protein of murine leukemia virus) acted as the main tumor rejection antigen and ~50% of CD8+ T cells in the tumor mass are AH1-specific after therapy. In order to characterize the clonality of the T cell response after successful antibody-cytokine immunotherapy, we isolated CD8+ T cells from tumors and submitted them to T cell receptor (TCR) sequencing. As expected, different TCR sequences were found in different mice, as these molecules originate from a stochastic rearrangement process. CD8+ T cells featuring the ten most abundant TCR sequences represented more than 60% of total CD8+ T cell clones in the tumor mass, but less than 10% in the spleen. Looking at sorted CD8+ T cells from individual animals, AH1-specific TCRs were consistently found among the most abundant sequences. Collectively, these data suggest that the antitumor response driven by two different antibody-cytokine fusions proceeds through an oligoclonal expansion and activation of tumor-infiltrating CD8+ T cells.