Efficiency and safety of O⁶-methylguanine DNA methyltransferase (MGMT(P140K))-mediated in vivo selection in a humanized mouse model.

Abstract

Efficient O⁶-methylguanine DNA methyltransferase (MGMT(P140K))-mediated myeloprotection and in vivo selection have been demonstrated in numerous animal models and most recently in a phase I clinical study in glioblastoma patients. However, this strategy may augment the genotoxic risk of integrating vectors because of chemotherapy-induced DNA damage and the proliferative stress exerted during the in vivo selection. Thus, to improve the safety of the procedure, we evaluated a self-inactivating lentiviral MGMT(P140K) vector for transduction of human cord blood-derived CD34⁺ cells followed by transplantation of the cells into NOD/LtSz-scid/Il2rγ⁻/⁻ mice. These experiments demonstrated significant and stable enrichment of MGMT(P140K) transgenic human cells in the murine peripheral blood and bone marrow. Clonal inventory analysis utilizing linear amplification-mediated polymerase chain reaction and high-throughput sequencing revealed a characteristic lentiviral integration profile. Among the bone marrow insertions retrieved, we observed considerable overlap to previous MGMT(P140K) preclinical models or the clinical study. However, no significant differences between our chemotherapy-treated and nontreated cohorts were observed. This also hold true when specific cancer gene databases and a functional annotation of hit genes by the Panther Database with respect to molecular function, biological process, or cellular component were assessed. Thus, in summary, our data demonstrate efficient and long-term in vivo selection without overt hematological abnormalities using the lentiviral MGMT(P140K) vector. Furthermore, the study introduces humanized mouse models as a novel tool for the pre-clinical assessment of human gene therapy related toxicity.

• Authors: Ruhi Phaltane, Reinhard Haemmerle, Michael Rothe, Ute Modlich, Thomas Moritz
• Journal: Human gene therapy (Hum Gene Ther) Vol. 25 Issue 2 Pg. 144-55 (Feb 2014) ISSN: 1557-7422 [Electronic] United States
• PMID: 24218991 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antineoplastic Agents
• O(6)-Methylguanine-DNA Methyltransferase

Topics

• Animals
• Antineoplastic Agents (administration & dosage, pharmacology)
• Gene Expression
• Gene Order
• Genetic Therapy
• Genetic Vectors (administration & dosage, genetics)
• Graft Survival (drug effects)
• Hematopoietic Stem Cell Transplantation
• Hematopoietic Stem Cells (drug effects, metabolism)
• Heterografts
• Humans
• Lentivirus (genetics)
• Mice
• O(6)-Methylguanine-DNA Methyltransferase (genetics, metabolism)
• Transduction, Genetic
• Transgenes
• Virus Integration