Identification of semaphorin E as a non-MDR drug resistance gene of human cancers

To improve cancer chemotherapy, a better understanding of the molecular mechanisms of drug resistance is essential. To identify the molecules responsible for drug resistance that is unrelated to MDR1 or MRP gene products, a eukaryotic expression cDNA library of cis-diamminedichloroplatinum(II) (CDDP)-resistant ovarian cancer TYKnuR cells was introduced into Cos-7 cells. After repeated CDDP selection, cDNA homologous to murine semaphorin E was isolated from surviving cells. Human semaphorin E (H-sema E) was overexpressed in CDDP-resistant cell lines and was readily induced not only by diverse chemotherapeutic drugs but also by x-ray and UV irradiation. Transfection of H-sema E conferred a drug-resistant phenotype to CDDP-sensitive cells. In addition, the aberrant expression of H-sema E protein was detected immunohistochemically in 14 of 42 (33.3%) recurrent squamous cell carcinomas removed at autopsy after extensive radiochemotherapy. Recently, another member of the semaphorin family, CD100, was shown to significantly improve the viability of B lymphocytes. These results suggest the involvement of semaphorins in diverse cell survival mechanisms.

Heat effects on DNA repair after ionising radiation: hyperthermia commonly increases the number of non-repaired double-strand breaks and structural rearrangements

After ionising radiation double-strand breaks (dsb) are lethal if not repaired or misrepaired. Cell killing is greatly enhanced by hyperthermia and it is questioned here whether heat not only affects dsb repair capacity but also fidelity in a chromosomal context. dsb repair experiments were designed so as to mainly score non-homologous end joining, while homologous recombination was largely precluded. Human male G0 fibroblasts were either preheated (45°C, 20 min) or not before X-irradiation. dsb induction and repair were measured by conventional gel electrophoresis and an assay combining restriction digestion using a rare cutting enzyme (NotI) and Southern hybridisation, which detects large chromosomal rearrangements (>100 kb). dsb induction rate in an X-chromosomal NotI fragment was 4.8 × 10–3 dsb/Gy/Mb. Similar values were found for the genome overall and also when cells were preheated. After 50 Gy, fibroblasts were competent to largely restore the original restriction fragment size. Five per cent of dsb remained non-rejoined and 14% were misrejoined. Correct restitution of restriction fragments occurred preferably during the first hour but continued at a slow rate for 12–16 h. In addition, dsb appeared to misrejoin throughout the entire repair period. After hyperthermia the fractions of non-rejoined and misrejoined dsb were similarly increased to 13 and 51%, respectively. It is suggested that heat increases the probability of dsb being incorrectly rejoined but it is not likely to interfere with one dsb repair pathway in particular.