Overexpression of GSTA2 protects against cell cycle arrest and apoptosis induced by the DNA inter‐strand crosslinking nitrogen mustard, mechlorethamine

Abstract

The effectiveness of bifunctional alkylating nitrogen mustard compounds in chemotherapy is related to their ability to form DNA inter-strand crosslinks. Patients exposed to DNA inter-strand crosslinking (ICL) agents subsequently experience an elevated incidence of myelodysplastic syndromes (MDS) and MDS related acute myeloid leukemia. Fanconi's anemia (FA) patients are deficient in the repair of crosslink DNA damage and they experience a high incidence of MDS. These observations indicate that hematopoietic cells are specific target for the transforming effects of DNA crosslinking damage. Changes in transcript levels were characterized in human hematopoietic cells occurring in response to the nitrogen mustard, mechlorethamine (HN2), but not in response to monofunctional analogs. Only modest changes in a few gene transcripts were detected in HL60 cells exposed to levels of HN2 tittered to maximal dose that caused growth suppression with minimal cell death and allowed eventual resumption of normal cell growth. Under conditions of transient growth suppression, a subset of glutathione-S-transferase (GST) isoenzyme genes was consistently upregulated three to fourfold by HN2, but not by monofunctional analogs. Subsequent efforts to confirm the changes detected by microarray analyses revealed an unexpected dependence on treatment conditions. The GST alpha class A2 subfamily member transcripts were upregulated 24 h after a 1 h exposure to HN2 that caused an extensive, but transient block in late S/G2 cell cycle phase, but were minimally altered with continuous exposure. The 1-h exposure to HN2 caused a transient late S/G2 cell cycle arrest in both the HL-60 cell line and the Colo 320HSR human colon cancer cell line. Overexpression of GSTA2 by transient transfection protected Colo 320HSR cells against both cycle arrest and apoptosis following exposure to HN2. Overexpression of GSTA2 in Colo 320HSR cells induced after exposure to HN2 did not alter cycle arrest or apoptosis. The results indicate that human GSTA2 facilitates the protection of cells from HN2 damage and not repair. Our results are consistent with the possibility that GSTA2 polymorphisms, variable isoenzyme expression, and variable induced expression may be factors in the pathogenesis of MDS.

Intercalation‐enhanced “Click” Crosslinking of DNA

“…TheAML cell lines were somewhat more sensitive towards the single agents,with maximal tolerated doses of 3 mm of POM-AmdU and up to 10 mm of DiMOC ( Figure S8, SI). These results revealed cell-type specific effects consistent with those previously reported for the formation of DNA-DNAi nterstrand crosslinks in AML [26] and HeLa [27] cell cultures. Interestingly,a ctive and complete cell killing by the combination of 4 + 5 was observed in HeLa cell cultures (Figure 2a), whereas OCI-AML2 (Figure 2b)a nd OCI-AML3 ( Figure S8c,S I) exhibited am aximal 50 % reduction in the number of viable cells,i ndicative of cell cycle arrest.…”

“…Interestingly, active and complete cell killing by the combination of 4 + 5 was observed in HeLa cell cultures (Figure a), whereas OCI‐AML2 (Figure b) and OCI‐AML3 (Figure S8c, SI) exhibited a maximal 50 % reduction in the number of viable cells, indicative of cell cycle arrest. These results revealed cell‐type specific effects consistent with those previously reported for the formation of DNA‐DNA interstrand crosslinks in AML and HeLa cell cultures.…”

Nitrogen mustard hydrochloride-induced acute respiratory failure and myelosuppression: A case report

“…Nitrogen mustard has a high affinity for DNA guanine residues and forms adducts and crosslinks with DNA, RNA and proteins; therefore, nitrogen mustard is potentially mutagenic and carcinogenic ( 14 ). Overexpression of the glutathione-S-transferase (GST) subfamily member GSTA2 has been found to protect cells against nitrogen mustard-induced cell cycle arrest and apoptosis ( 15 ). Recently, Inturi et al ( 16 ) studied the mechanism of DNA repair following nitrogen mustard-induced double-strand breaks.…”

Anticancer Drug Metabolism: Chemotherapy Resistance and New Therapeutic Approaches

“…By contrast, the overexpression of GSTP1 in human renal UOK130 tumor cells was accompanied by a decreased sensitivity to cisplatin, melphalan and chlorambucil (Wang et al, 2007). Similarly, overexpression of Alpha class GST has been correlated with the resistance to alkylating agents in Colo 320HSR cells (Xie et al, 2005) and to doxorubicin in MCF-7 human breast cancer and small cell lung cancer (H69) cell lines (Sharma et al, 2006;Wang et al, 1999).…”