Drug Resistance Gene Therapy to Augment Allogeneic Transplantation for the Treatment of Leukemia Following Relapse

Abstract 2050

Relapse remains a significant problem after allogeneic hematopoietic cell transplantation (HCT) for high-risk hematopoietic malignancies. Unfortunately treatment options for recurrent disease after HCT have remained limited and are often ineffective. In addition, chemotherapy to treat relapse after HCT results in myelosuppression and thus infectious complications. Here we investigated a novel concept of conferring donor cell resistance to the chemotherapy thus preventing or minimizing these complications.

To model this approach we used a mutant version of the chemotherapeutic resistance gene methylguanine-methyltransferase (MGMT) that are insensitive to O6-benzylguanine (O6BG), an agent that can be used to sensitize unmodified/tumor cells prior to alkylating agents. Overexpression of MGMT mediates resistance to hematopoietic stem cells (HSCs) to alkylating agents like Carmustine (BCNU) and methylating agents like Temozolomide (TMZ). Chemotherapeutic effects can be further enhanced with O6BG, which blocks the endogens MGMT, a mechanism that can be exploited with the O6BG resistant human MGMT mutant MGMT(P140K) or the equivalent dog mutant MGMT(P144K). We have previously demonstrated in dog and nonhuman primate large animal models that MGMT(P140K) gene transfer to HSCs provided multilineage hematopoietic chemoprotection and allowed for dose intense chemotherapy.

Using a DLA-identical transplantation setting in the dog we established donor chimerism following non-myeloablative conditioning with 300 cGy total body irradiation and infusion of whole bone marrow. To model the treatment of a hypothetical relapse 10 weeks after the initial HCT, the animals received chemotherapy with O6BG/TMZ and the infusion of MGMT(P144K) gene-modified cells from the same donor. Dogs then receive several cycles of chemotherapy, consisting of a combination of O6BG and TMZ or BCNU. Donor chimerism was assessed by analyzing the variable number of tandem repeats (VNTR) and gene marking was analysed with a quantitative real-time PCR.

We have treated 2 dogs thus far. Gene marked cells increased ∼100-fold up to 11% after two rounds of O6BG and TMZ. The overall chimerism increased by 10% up to 90% during selection. After a third round of chemotherapy for dog 2 we observed protection from myelosuppression indicated by a drop of platelet counts to only 200,000/μl which compared to historical treatments would have resulted in pronounced myelosuppression.

Here we successfully developed a clinical relevant dog model to test a new treatment option for patients with a a recurrent hematologic malignancy after HCT that combines aggressive alkylating agent chemotherapy and drug resistant allogeneic HCT. We demonstrate that allogeneic transplantation of genetic modified HSCs into a recipient that previously received a HCT is feasible and well tolerated. MGMT(P144K) modified cells engraft in the recipient and provide efficient chemoprotection of hematopoiesis following aggressive chemotherapy that would likely be required to achieve a response in patients with recurrent disease This strategy should the protection of donor cells using MGMT(P140K) and/or cytidine deaminase modified HSCs that confers protection to alkylating agents, methylating agents, or antimetabolites and and thus allow high dose and/or dose dense chemotherapy for patients with recurrent hematologic malignancy.