title: Persistent mismatch repair deficiency following targeted correction of hMLH1.
abstract: The use of gene therapy to correct mutated or lost gene function for the treatment of human cancers has been an active, yet problematic area of biomedical research. Many technical difficulties, including efficient tissue-specific delivery, integration site specificity and general toxicity, are being addressed. Little is known, however, about the genetic and phenotypic stability that accompanies a successful gene-specific targeting event in a cancer cell. This question was addressed following the creation of a colon cancer cell line in which a mutated hMLH1 gene was corrected via targeted homologous recombination. This correction resulted in the expression of wild-type hMLH1 protein, restoration of the hPMS2 protein and mismatch repair (MMR) proficiency. One of two hMLH1-corrected clones, however, was found to retain defects in MMR activity. These cells continued to express the corrected hMLH1 protein, but had lost expression of another MMR protein, hMSH6. DNA sequence analysis of the hMSH6 gene revealed biallelic expansions of a cytosine repeat region in exon 5 that result in frameshifts leading to premature stop codons. These findings suggest that, similar to acquired drug resistance, the presence of genetically heterogeneous cancer cell populations or acquisition of compensatory mutations can result in 'resistance' to gene replacement therapy.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Maryland Greenebaum Cancer Center, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: M B
authors.initials: MB
authors.lastname: Weiss
authors.forename: M I
authors.initials: MI
authors.lastname: Vitolo
authors.forename: K
authors.initials: K
authors.lastname: Baerenfaller
authors.forename: G
authors.initials: G
authors.lastname: Marra
authors.forename: B H
authors.initials: BH
authors.lastname: Park
authors.forename: K E
authors.initials: KE
authors.lastname: Bachman
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D020137:Base Pair Mismatch; D001483:Base Sequence; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics*; D045744:Cell Line, Tumor; D017931:DNA Primers; D004260:DNA Repair; D006801:Humans; D000070957:MutL Protein Homolog 1; D009687:Nuclear Proteins / Q000235:genetics*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D017931:DNA Primers; C501486:MLH1 protein, human; D009687:Nuclear Proteins; D000070957:MutL Protein Homolog 1