CD20-specific T cell receptors in gene therapy of leukemia and lymphoma

T-cell receptors constitute the key element of T cells that mediates recognition and killing of virally infected cells with a high degree of efficacy and specificity. This is the reason why T cells can cure influenza without side effects. The project utilizes these characteristics of T cells to kill cancer cells. Since cancer cells, unlike virus, are often ignored by the T cells, we utilize differences in tissue-type molecules to make the T cells respond effectively to self-proteins. This allows the induction of powerful T-cell responses to proteins that are normally, but selectively, expressed on the cell type from which the cancer cell originates. This constitutes a new immunological concept that we have demonstrated is highly efficient. By use of this strategy we have generated a number of T-cell receptors that recognize the B-cell specific protein CD20, expressed in lymphoma and leukemia. We will transfer the genes for such T-cell receptors to patient T cells to transform them into killer cells that efficiently recognize and kill lymphoma cells after re-infusion into the patient. During the project period, we have 1) cloned a number of CD20-specific T-cell receptors into vectors that allow genetic modification of T cells, 2) tested and identified which method and which vector that results in the most efficient gene transfer to T cells (modified with murine constant parts, extra cysteine bridges and codon optimization), 3) established expansion protocols for T cells, 4) tested and compared a large number of CD20-specific receptors for functionality following gene transfer into peripheral blood T cells. This has resulted in the identification of two candidate T-cell receptors that both mediate efficient killing of leukemia- and lymphoma cells (cell lines and patient cells), while sparing cells originating from a wide variety of tissues, and 5) established a mouse model for testing of efficacy of human gene-modified T cells that express CD20-specific T-cell receptors, and started to establish a mouse model for testing of safety.

The main objective of the project is to treat leukemia and lymphoma patients with gene therapy. We will conduct a clinical trial using a T cell receptor (TcR) targeted at CD20 that mediates highly efficient killing of primary human leukemia cells. T cell- mediated killing is independent of p53, as opposed to chemotherapy or monoclonal antibodies. Rather than seeking a cancer-specific response, we have developed an innovative approach to generate T cells that recognize peptides from cell type-restricted pro teins. To overcome tolerance to wild-type human proteins, T cells are exposed to peptides in the context of a foreign HLA-molecule. We have established that this is a highly efficient method to generate T cells that specifically recognize the B cell-restr icted antigen CD20 and that kill primary chronic lymphocytic leukemia cells with unprecedented efficiencies. From these cells we have now cloned a number of TcRs. The present application seeks funding for a phase I/II clinical trial that will include pati ents with small lymphocytic lymphoma or follicular lymphoma. The TcRs can be used to treat HLA-A*0201+ patients (50% of Caucasians). The primary end point is safety/tumor response rate. The preclinical phase involves optimization of the protocols for T ce ll expansion and large-scale gene transfer of CD20-specific TcRs to T cells. We will transfect the cells by electroporation with mRNA encoding the TcR. This method has been used in many clinical protocols at Radiumhospitalet, is documented to be safe and leads to transient expression. Research will be performed to maximize TcR expression, and to investigate if expression of molecules that direct T cell migration will improve homing to tumor areas. An innovative, stepwise trial design will ensure that info rmation is gathered along the way and that each patient will serve as his/her own control.