Several different cancer studies have indicated that lymphocyte mutagen sensitivity is a marker of DNA repair deficiency and increased cancer risk. We have used a mutagen sensitivity assay (MSA) measuring gamma-radiation-induced chromosomal aberrations in freshly cultured lymphocytes and assessed breast cancer risk in African-American women. Concurrently, we conducted duplicate cultures in the presence of caffeine, which overrides G(2) arrest in cultured cells, decreases time to DNA repair, and hence increases the aberration rate. In comparison with the non-caffeine-treated cells, we are conceptually segregating the contribution of DNA repair and time for DNA repair as individual susceptibility phenotypes. Blood samples were obtained from 61 cases and 86 controls at Howard University Hospital. Two sets of whole-blood cultures were established and gamma-irradiated (1 Gy) at 67 hours, one of which was treated with caffeine (1 mg/mL). Thereafter, cultures were processed for obtaining metaphase spreads. Fifty metaphases were screened for chromatid breaks. The mean breaks per cell (MBPC) for cases (0.34 +/- 0.15) was significantly greater than for controls (0.24 +/- 0.12; P < 0.0001). Using the 75th percentile value of controls as a cutoff to define mutagen sensitivity, the sensitive individuals had an odds ratio of 4.5 (95% confidence intervals, 2.2-9.1) for breast cancer compared with individuals that were not sensitive. The adjusted odds ratio was 3.3 (95% confidence intervals, 0.147-73.917), which was statistically significant but was limited by the small number of subjects. The results for caffeine co-culture were not predictive of breast cancer (MBPC: cases, 1.6 +/- 0.9 versus controls, 1.5 +/- 0.8; P = 0.8663). Comparing the MBPC for caffeine and non-caffeine cultures, there was a correlation in controls (n = 79; Spearman r = 0.4286; P < 0.0001), but not in cases (n = 58; Spearman r = 0.06609; P = 0.6221). This study indicates that the MSA phenotype is a risk factor for breast cancer in African-American women, with a significant effect observable even in small studies. The use of caffeine did not enhance the predictivity of MSA, but the correlation with non-caffeine cultures in controls indicates that the MSA phenotype is due to both DNA repair and G(2) arrest capacity.