Our Science – Kammula Website
Udai S. Kammula, M.D.
Project I: Translational Studies to Define and Deliver Optimal T Cells for the Adoptive Immunotherapy of Cancer
The adoptive transfer of autologous tumor-reactive lymphocytes derived from surgically resected tumor specimens (i.e., tumor infiltrating lymphocytes, TIL), in conjunction with a lymphodepleting conditioning regimen, has been shown to mediate significant tumor regression in some patients with metastatic melanoma. In addition to demonstrating proof of concept, these pilot studies have also defined the practical and therapeutic limitations to this approach. The isolation and expansion of reactive TIL has only been feasible from patients with metastatic melanoma. Furthermore, not all patients with melanoma are eligible for this type of immunotherapy either because resectable tumor is not available, the lymphocytes from the specimen do not expand sufficiently, or the lymphocytes that do proliferate do not exhibit sufficient tumor-specific reactivity. When reactive bulk TIL is generated, the composition is largely unknown and varies among treated patients. Thus, it has been difficult from these studies to define an 'optimal' population of lymphocytes suitable to consistently mediate tumor regression in melanoma patients. We hypothesized that the transfer and clinical evaluation of a consistent and homogeneous population of tumor-reactive lymphocytes would help to define lymphocytes with demonstrated therapeutic efficacy for use in future adoptive immunotherapy efforts. Further, we hypothesized that isolation of these lymphocytes from peripheral blood lymphocytes (PBL) would have several advantages. From a practical perspective, the use of a blood draw or leukapheresis would avoid the need for surgery and the potential for post-operative complications and delays to eventual systemic treatment. The broad repertoire of PBL might allow for the isolation of unique populations of tumor-reactive lymphocytes that are not commonly found in TIL. Finally, the use of PBL may serve as a generalized strategy to isolate tumor-reactive lymphocytes from patients with diverse histologies and, thus, expand the therapeutic relevance of this approach. To test these hypotheses, we set the following specific aims:
1. Develop a novel high-throughput methodology to rapidly detect, isolate, and expand low frequency, high avidity antigen-specific CD8+ T cells from PBL.
2. Characterize the phenotype, differentiation, and avidity of the tumor-reactive T cells isolated with this novel approach.
3. Develop clinical adoptive transfer trials of highly selected and homogeneous populations of tumor-specific lymphocytes and prospectively correlate their characteristics and quantity with tumor response in patients with advanced cancer.
Accomplishments and Future Plans
Since joining the Surgery Branch, our laboratory has successfully developed a novel high-throughput methodology to isolate and expand low frequency, high avidity antigen specific CD8+ T cells from PBL. This technique utilizes a highly sensitive PCR-based screening assay that can detect the reactivity of rare T cells in a bulk population. Using this approach, after approximately one week of culture, we have been able to prospectively detect T cells from the natural peripheral blood repertoire of melanoma patients that recognize the gp100154-162 epitope from the gp100 antigen, a process that would have been exceedingly difficult and time consuming with conventional repetitive in vitro stimulation techniques. These cells were then isolated and characterized to possess high avidity for the gp100154-162 epitope, an early effector memory phenotype with high surface expression of CD27, and were generated in the setting of limited antigen and interleukin-2 (IL-2) exposure. We have translated these findings to the design of a phase II clinical trial transferring gp100154-162 reactive lymphocytes to patients with metastatic melanoma. In parallel studies, we have applied this platform to the robust isolation of CD8+ T cells that recognize the cancer-testes antigen, NY-ESO, which is expressed in melanoma, breast cancer, prostate cancer, and to varying degrees in bladder, lung, hepatocellular, ovarian, and thyroid cancers. Future plans include the clinical transfer of lymphocytes that recognize the NY-ESO157-165 epitope, which would extend our current studies of adoptive immunotherapy to a variety of common malignancies.
Project II: Development of a Novel High-Throughput Reverse Immunology Approach to the Screening and Identification of Novel Tumor Antigen Epitopes
The identification of novel immunogenic epitopes presented by tumors in the context of common HLA molecules is critical to extending current immune-based therapies for cancer. The use of natural TIL to identify and clone the genes that encode for the antigens recognized by these TILs has had significant success in defining therapeutic targets against melanoma. However, the difficulty in generating reactive TIL from nearly all other solid tumors has limited this 'forward' immunology approach to antigen discovery. 'Reverse' immunology attempts to predict and identify immunogenic peptides from the sequence of a gene product that is believed to be specific and highly expressed on the tumor. This strategy typically uses computer-based HLA binding algorithms to help identify putative peptides that are then used to stimulate lymphocytes. The inherent weakness of this peptide 'mining' approach is the incredibly low probability of identifying a peptide that is naturally processed, presented, and sufficient to induce tumor lysis upon recognition. As a consequence, low-throughput attempts to screen limited numbers of peptides are typically unsuccessful. The aims of this project are:
1. Develop a novel high-throughput methodology to rapidly screen large numbers of predicted peptides from putative antigen targets by analyzing their ability to stimulate HLA matched human PBL.
2. Expand and characterize peptide-reactive lymphocytes to determine their ability to recognize natural tumor targets and antigen-expressing transfectants.
Accomplishments and Future Plans
We have applied the T cell detection methodology in Project I to rapidly screen reactivity against predicted peptides from mesothelin, a glycoprotein that is overexpressed on mesothelioma, pancreatic cancer, and ovarian cancer. We have identified two candidate peptides that we are pursuing for further evaluation. Our plans are to further optimize this approach to screen peptide libraries from other candidate antigens.
Project III: Development of Immunotherapy Strategies for Gastrointestinal Cancers with a Focus on Hepatocellular Cancer
Building upon evidence that immune manipulation can result in complete and durable regression in select patients with certain metastatic cancers, our efforts have focused on translating these observations into the development of effective immune therapies for hepatocellular cancer (HCC). Essential to this goal is the identification and evaluation of immunotherapeutic targets expressed by HCC. We have begun an active evaluation of putative HCC therapeutic targets, including the alpha-fetoprotein members of the cancer-testes antigen family (NY-ESO), and most recently glypican-3 (GPC-3). Our studies have found that GPC-3 mRNA is overexpressed in >90% of fresh HCC tumor specimens and is minimally expressed in normal tissues. Studies evaluating whether this glycoprotein is naturally recognized by T lymphocytes of the human immune system are underway, using traditional techniques involving TIL derived from surgically resected HCC specimens and also screening of peripheral blood mononuclear cells (PBMC). To facilitate the identification of low frequency and high avidity T cells that recognize novel tumor antigen epitopes, we have pioneered the use of high-throughput quantitative gene expression as a tool to rapidly screen for antigen-specific T cell reactivity. In pilot studies, we have found this promising technique to be a highly sensitive methodology to identify, stratify, and select melanoma antigen-reactive CD8+ T cells from the bulk PBMC of cancer patients. We are applying this method to the screening of potential HCC antigens, and are in the process of adapting this assay as a novel immune monitoring tool for future immunotherapy clinical trials. We have active pre-clinical studies evaluating the role of demethylating agents as a means to modulate the expression of tumor antigens on HCC to enhance their recognition by immune cells. Further pre-clinical efforts are aimed at evaluating the use of non-specific immune-stimulating agents such as IL-2 and anti-CTLA-4 antibody as possible adjunct therapies for the treatment of HCC. With growing evidence suggesting the importance of regulatory T cells in suppressing natural immune responses against tumors, including HCC, we have begun to evaluate their role in HCC and potential therapies that may abate their suppression.
This page was last updated on 5/2/2013.