Abstract
The doxorubicin-selected multidrug resistant small cell lung cancer cell line, H69AR, is cross-resistant to the Vinca alkaloids and epipodophyllotoxins, but does not overexpress P-glycoprotein, a 170 kDa plasma membrane efflux pump usually associated with this type of resistance. Monoclonal antibodies were raised against the H69AR cell line and one of these, MAb 3.186, recognises a peptide epitope on a 36 kDa phosphorylated protein that is membrane associated, but not presented on the external surface of H69AR cells (Mirski & Cole, 1991). In the present study, in vitro translation and molecular cloning techniques were used to determine the relative levels of mRNA corresponding to the 3.186 antigen. In addition, a cDNA clone containing an insert of approximately 1.4 kb was obtained by screening an H69AR cDNA library with 125I-MAb 3.186. Fragments of this cloned DNA hybridised to a single mRNA species of approximately 1.6 kb that was 5 to 6-fold elevated in H69AR cells. Partial DNA sequencing and restriction endonuclease mapping revealed identity of the cloned DNA with p36, a member of the annexin/lipocortin family of Ca2+ and phospholipid binding proteins.
Highlights
The H69AR cell line was derived by selection of the small cell lung cancer (SCLC) cell line, H69, in doxorubicin (DOX) (Cole, 1986; Mirski et al, 1987) and is cross-resistant to other anthracyclines, the Vinca alkaloids and the epipodophyllotoxins (Mirski et al, 1987; Cole, 1990)
The relative levels of mRNA corresponding to the 3.186 antigen were compared by immunoprecipitation of equivalent amounts of "5S-methionine labelled in vitro translation products of RNA from both H69 and H69AR cells
Densitometry of autoradiographs showed that 5 to 6-fold more of the 36 kDa antigen was immunoprecipitated from the translation products of H69AR mRNA relative to H69 (Figure 1)
Summary
The H69AR cell line was derived by selection of the small cell lung cancer (SCLC) cell line, H69, in doxorubicin (DOX) (Cole, 1986; Mirski et al, 1987) and is cross-resistant to other anthracyclines, the Vinca alkaloids and the epipodophyllotoxins (Mirski et al, 1987; Cole, 1990) This pattern of cross-resistance is typical of cells that express elevated levels of P-glycoprotein, a plasma membrane glycoprotein that confers multidrug resistance by enhancing drug efflux Given the evidence that overexpression of P-glycoprotein in SCLC is an infrequent occurrence (Lai et al, 1989; Noonan et al, 1990; Brambilla et al, 1991), the investigation of drug resistance in H69AR cells is important because of its potential clinical relevance
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