University of Pittsburgh Archives | Alliance for Cancer Gene Therapy

Pancreatic cancer is a major public health problem with approximately 57,600 new cases in 2020. Five-year survival is less than 9% for all stages, with surgical resection the only potentially curative therapy. Nonetheless, a clear majority of patients are not candidates for surgery due to locally advanced disease or metastatic spread, and ultimately derive little to no survival benefit from radiation or chemotherapy.

Advances in cancer immunotherapy have revolutionized cancer care, providing hope in the form of durable cancer regression and long-term survival. In patients with treatment refractory metastatic melanoma, immunotherapy in the form of adoptive transfer with tumor infiltrating immune cells (TIL therapy) has yielded practice changing response rates, with complete remission in 10-20% of patients.

Unfortunately, an immune response to pancreatic cancer has been elusive in most clinical trials, and thus identifying methods to enhance the immune response to cancer has been the focus of our research program in the last two decades. Recent transient responses to TIL therapy in pancreatic cancer obtained at the NCI Surgery Branch and our institution have revitalized our interest in improving TIL therapy outcomes.

A tumor’s ability to evade the immune system can be attributed to an immunosuppressive tumor microenvironment and downregulation or loss of the major histocompatibility complex (MHC), which is required by immune cells to recognize malignant cells. Conventional immunotherapy approaches, including TIL therapy, rely on activation and tumor specific recognition of T cells with alpha-beta T cell receptors that require intact MHC expression. However, with 60% of primary and 90% of metastatic pancreatic tumors exhibiting mutated or reduced expression of MHC molecules, new immunotherapy approaches are necessary to elicit improved treatment responses. Gamma Delta (gd) T cells are an understudied tissue resident immune subset that can recognize a broad range of antigens without the presence of MHC molecules.

Gamma Delta T cells comprise as much as 40% of tumor infiltrating lymphocytes in pancreatic cancer and display potent antitumor immunity. Utilizing the resources of our world-renowned pancreatic cancer treatment center and established cell therapy production facility, we aim to evaluate the efficacy of gd TIL therapy for patients with metastatic pancreatic cancer. After developing a clinical grade cell manufacturing protocol to grow gd TIL and compare tumor specific reactivity with alpha beta TIL, we aim to conduct a pilot Phase I/II clinical trial assessing the safety and preliminary treatment efficacy of gd TIL therapy. We will also identify MHC independent TCRs from gd TIL for further development as universal gene therapies.

Ultimately, this study will define the therapeutic potential of tumor infiltrating gd T cells, helping to advance pancreatic cancer into a disease that our own immune system, ‘the best doctor’ can control.

Significant strides have been made in the treatment of solid tumors using viruses designed to attack and kill tumors (Oncolytic Viruses: OV) following direct injection into a detectable tumor mass. Tumor cell killing releases tumor-related proteins capable of inducing anti-tumor immunity, potentially eliminating similar tumor masses throughout the body.

The first OV to achieve FDA approval is an oncolytic herpes simplex virus (oHSV) whose use in treatment of late-stage melanoma achieved a significant “cure” rate (~50%) when administered in combination with antibodies that enhance tumor rejection. Here we propose to create substantially improved oHSV vectors that will be extraordinarily safe (tumor targeted) and highly resistant to pre-existing anti-HSV immunity common in the human population. The advanced vector design will allow intravenous administration of a “heat-seeking missile” that targets metastatic cancer for destruction and consequently induces protective immunity against relapse.

Recent clinical successes have revealed that the immune system can be successfully harnessed to fight cancer. Various strategies are utilized, including enhancing a patient’s ‘natural’ response to cancer as well as ‘redirecting’ a patient’s immune cells (‘T cells’) to the tumor using genetic engineering. While these T cell therapies have had major success in leukemias, they have not yet shown promise in the treatment of solid tumors.

T cells require an enormous amount of fuel to perform their tumor-killing functions. However, we have recently shown that in solid tumors, cancer cells evade immune responses in part by depriving the T cell of the ability to generate energy and depleting the local environment of nutrients.

In this Alliance for Cancer Gene Therapy funded research program, we will utilize genetic engineering to metabolically ‘reprogram’ tumor-specific T cells. Using this technology, they will become more fit to fight cancer for an extended period of time. We will test these T cells in animal models and translate these findings into human T cells as well. The goal is to generate super-soldier type T cells, those that can be both redirected to the tumor site, but also bolstered metabolically to support long-term and durable responses.

This Alliance for Cancer Gene Therapy Research Fellow is funded in part by Swim Across America.

Our group is exploring the use of gene therapy techniques to engineer viruses to stimulate the immune system to fight pancreatic cancer. In this proposal we will modify a vaccinia virus (similar to the virus used to immunize against smallpox) to produce hormones that attract a certain kind of tumor fighting immune cell called T cells to the sites of tumor.

We have already administered a similar virus to patients and found that it is able to be delivered through the blood stream safely, travel to sites of tumor and infect the tumor cells. We have also shown that a special way to stimulate the immune system to generate tumor fighting T cells called aDC1 is effective in patients with a very aggressive kind of brain tumor.

In this study we will combine these two approaches (stimulate T cells with aDC1 and attract them to the tumor with the vaccinia virus) in patient who have pancreatic cancer. There are several clinical trials examining immunotherapy for pancreatic cancer in progress at this time, we believe our approach will be a significant improvement on current trials. 

Probably the greatest limitation to the application of gene therapy for the treatment of cancer remains the difficulty in delivering a therapeutic gene efficiently and selectively to its tumor target. 

We have used oncolytic viruses, viruses that have been modified so that they selectively replicate in tumors and not normal tissues, to deliver a variety of genes to tumors. These viruses, based on vaccinia virus can be delivered intravenously into the blood stream, and even though they will then infect many different tissues (including the tumor), the engineered selectivity means they are cleared from all non-tumor tissues. The small amount of virus that does make it to the tumor will rapidly and selectively amplify as the virus replicates and spreads through the tumor. We have also used certain immune cell therapies that are attracted to the tumor, as delivery vehicles to carry the viruses to the tumor, so increasing the amount of virus that gets to the tumor and reducing the amount that goes to other organs. 

We have found that if we express therapeutic genes from these viruses (such as cytokines, that attract the host immune response to help destroy the tumor), we are able to increase their therapeutic benefit. However, because the expression of the cytokines results in destruction of the infected cells, we also reduce the amount of viral replication that occurs. This means that the virus does not get a chance to replicate properly, so reducing its initial delivery and spread within the tumor, and leading to clearance of the virus before it is capable of achieving its own therapeutic potential. 

We have therefore taken an approach that was developed by our collaborators that allows us to control the stability, and so the function, of any protein, and applied this to our oncolytic viral therapies.

By attaching a small sequence onto any protein, we can flag the protein for rapid destruction by the cell’s degradation pathways. However, if we apply a small molecule that binds to this sequence and shields it, then we stabilize the protein and its function is restored. This allows us to rapidly, reversibly and safely control the levels of functional protein. In this way we can block cytokine function for a period of time (while the virus is cleared from normal cells and amplifies itself to high levels within the tumor; or while the virus is being delivered within an immune cell to the tumor), and so enhance initial delivery and establishment within the tumor. If we then stabilize the cytokine, there is increased production exclusively within the tumor, and so we can safely increase anti-tumor effects.

We propose to use this system to control the expression of different genes expressed from oncolytic vaccinia viruses, and will use the system to control the virus replication itself. As a result, we can improve the delivery, safety and effectiveness of these (and other) gene therapy approaches, using a system that can be rapidly moved into the clinic. 

Lung cancer remains one of the greatest public health threats, despite advancement in the understanding of molecular genetics. We have been accumulating preliminary data and developing reagents for this project for the past two years, and we are encouraged by the early results of using PUMA, a protein, as a novel target to selectively encourage apoptosis, cell death, in lung cancer cells. With this funding, we will be able to expand the research into animal model trials, and it is our hope that these efforts will allow us to examine the feasibility of moving PUMA gene therapy towards clinical trials.