Chemically induced increases and decreases in the rate of expansion of a CAG*CTG triplet repeat

Abstract

Somatic mosaicism of repeat length is prominent in repeat expansion disorders such as Huntington disease and myotonic dystrophy. Somatic mosaicism is age-dependent, tissue-specific and expansion-biased, and likely contributes toward the tissue-specificity and progressive nature of the symptoms. We propose that therapies targeted at somatic repeat expansion may have general utility in these disorders. Specifically, suppression of somatic expansion would be expected to be therapeutic, whilst reversion of the expanded mutant repeat to within the normal range would be predicted to be curative. However, the effects of genotoxic agents on the mutational properties of specific nuclear genes are notoriously difficult to define. Nonetheless, we have determined that chronic exposure over a three month period to a number of genotoxic agents can alter the rate of triplet repeat expansion in whole populations of mammalian cells. Interestingly, high doses of caffeine increased the rate of expansion by approximately 60%. More importantly, cytosine arabinoside, ethidium bromide, 5-azacytidine and aspirin all significantly reduced the rate of expansion by from 35 to 75%. These data establish that drug induced suppression of somatic expansion is possible. These data also suggest that highly unstable expanded simple sequence repeats may act as sensitive reporters of genotoxic assault in the soma.

Figure 1

Chemical treatment and expanded CAG·CTG repeat dynamics. (A) The autoradiographs shown are representative SP-PCR analyses of DNA samples extracted from replicate D2763Kc2 cells cultured for up to 130 days with the drugs indicated (see Materials and Methods and Table 1 for details) and the progenitor culture from which all cells were derived at day zero. Evidence for clonal growth in the hydrogen peroxide and rhodamine 6G treated cultures is indicated with arrows. (B) The box plots show the degree of repeat length variation observed in treated and control cells at the time points indicated. The top and bottom of the boxes correspond to the third (Q3) and first quartiles (Q1), respectively and the line across the box displays the median. The lines extending from the top and the bottom of the boxes, include values that fall inside the lower and upper limits: Q1-1.5(Q3-Q1) and Q3+1.5(Q3-Q1), respectively. Statistically significant differences (*, P < 0.05) in the median rates of expansion were observed for some treatments relative to both time and PD control cultures.