Vector design for expression of O6-methylguanine-DNA methyltransferase in hematopoietic cells.
Enhancing DNA repair activity of hematopoietic cells by stably integrating gene vectors that express O(6)-methylguanine-DNA-methyltransferase (MGMT) is of major interest for innovative approaches in tumor chemotherapy and for the control of hematopoietic chimerism in the treatment of multiple other acquired or inherited disorders. Crucial determinants of this selection principle are the stringency of treatment with O(6)-alkylating agents and the level of transgenic MGMT expression. Attempts to generate clinically useful MGMT vectors focus on the design of potent expression cassettes, an important component of which is formed by enhancer sequences that are active in primitive as well as more differentiated hematopoietic cells. However, recent studies have revealed that vectors harboring strong enhancer sequences are more likely to induce adverse events related to insertional mutagenesis. Safety-improved vectors that maintain high levels of MGMT expression may be constructed based on the following principles: choice of enhancer-promoter sequences with relatively mild long-distance effects despite a high transcription rate, improved RNA processing (export, stability and translation), and protein design. The need for optimizing MGMT protein design is supported by recent observations suggesting that the P140K mutant of MGMT, developed to be resistant to inhibitors such as O(6)-benzylguanine, may confer a selective disadvantage when expressed at high levels. Here, we provide a review of the literature exploring MGMT expression vectors for bone marrow chemoprotection, and describe experimental evidence suggesting that high expression of MGMT P140K induces a selective disadvantage in the absence of alkylating agents. We conclude that the appropriate design of expression vectors and MGMT protein features will be crucial for the long-term prospects of this promising selection principle.
Schambach
Axel
A
Department of Experimental Hematology, Hannover Medical School, Hannover, Germany.
Baum
Christopher
C
D019943:Amino Acid Substitution; D000818:Animals; D015254:DNA Modification Methylases / Q000235:genetics; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000235:genetics; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000201:enzymology*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009369:Neoplasms / Q000628:therapy; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics
D011994:Recombinant Proteins; D025521:Tumor Suppressor Proteins; D015254:DNA Modification Methylases; C515491:MGMT protein, human; D019853:O(6)-Methylguanine-DNA Methyltransferase; D045643:DNA Repair Enzymes