Comparative analysis of three functional predictive assays in lymphocytes of patients with breast and gynaecological cancer treated by radiotherapy.

PURPOSE: There is a need to develop predictive tests that would allow identifying cancer patients with a high risk of developing side effects to radiotherapy. We compared the predictive value of three functional assays: the G(0) aberration assay, the G(2) aberration assay and the alkaline comet assay in lymphocytes of breast cancer and gynaecological cancer patients. MATERIAL AND METHODS: Peripheral blood was collected from 35 patients with breast cancer and 34 patients with gynaecological cancer before the onset of therapy. Chromosomal aberrations were scored in lymphocytes irradiated in the G(0) or G(2) phase of the cell cycle. DNA repair kinetics was performed with the alkaline comet assay following irradiation of unstimulated lymphocytes. The results were compared with the severity of early and late side effects to radiotherapy. RESULTS: No correlation was observed between the results of the assays and the severity of side effects. Moreover, each assay identified different patients as radiosensitive. CONCLUSIONS: There is no simple correlation between the in vitro sensitivity of lymphocytes and the risk of developing early and late side effects.

DNA interstrand crosslinks are induced in cells prelabelled with 5-bromo-2'-deoxyuridine and exposed to UVC radiation.

It has been observed previously that 5-bromo-2'-deoxyuridine (BrdU) potentiates the effect of UVC radiation on the level of sister chromatid exchanges. It is not known which type of DNA damage is responsible for this enhancing effect and we have proposed this to be the DNA interstrand crosslink (ICL) which, theoretically, may arise in cells that are labelled with BrdU for one round of replication and exposed to UVC radiation. The aim of the present investigation was to verify if ICLs are indeed formed during this irradiation scenario. CHO-K1 cells were prelabelled with BrdU and exposed to UVC. ICLs were detected by a modified version of the comet assay that relies on the reduction of induced DNA migration in the agarose gel. Carboplatin was used as a positive control. We found that BrdU+UVC treatment indeed results in a reduction of the damage induced by gamma-radiation. Furthermore, we observed that CL-V4B cells exposed to BrdU+UVC, but not to UVC alone, showed a very high level of chromosomal damage. These cells have a deficient Rad51C paralog that renders them extremely sensitive towards ICLs. Interestingly, the cytogenetic results did not correlate with cell survival, where it was found that the CL-V4B cells tolerate BrdU+UVC better than the wild type cells. The possible reasons are discussed. Taken together our results indicate that ICLs are formed in DNA that was prelabelled with BrdU and exposed to UVC radiation.

Cytogenetic damage in lymphocytes of patients undergoing therapy for small cell lung cancer and ovarian carcinoma.

The level of cytogenetic damage in peripheral blood lymphocytes of patients undergoing chemotherapy has been analyzed incisively 20 years ago. The results showed that the highest level of cytogenetic damage was observed at the end of therapy. In recent years, the doses of anticancer drugs were intensified thanks to the discovery of colony stimulating factors. Therefore, it was interesting to analyze the kinetics of micronuclei formation in lymphocytes of patients undergoing modern chemotherapy. The frequencies of micronuclei were measured in lymphocytes of 6 patients with small cell lung cancer treated with a combination of cisplatin and etoposide and 7 patients with ovarian carcinoma treated with a combination of taxol and cisplatin. 3 patients with lung cancer received radiotherapy in addition to chemotherapy. Micronuclei were analyzed in lymphocytes collected before the start of therapy and 1 day before each following cycle of chemotherapy. The micronucleus frequencies were compared with the kinetics of leukocyte counts. The micronucleus frequencies showed an interindividual variability. On average, the frequencies of micronuclei increased during the first half of therapy and declined thereafter, reaching, in some patients with ovarian carcinoma, values below the pre-treatment level. Leukocyte counts decreased strongly at the beginning of therapy with an upward trend at the end. We suggest that the decline of micronuclei was due to repopulation of lymphocytes and acquired drug resistance.