Enhanced in vitro invasiveness and drug resistance with altered gene expression patterns in a human lung carcinoma cell line after pulse selection with anticancer drugs

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Abstract
The human lung carcinoma cell line DLKP was exposed to sequential pulses of 10 commonly used chemotherapeutic drugs (VP-16, vincristine, taxotere, mitoxantrone, 5-fluorouracil, methotrexate, CCNU, BCNU, cisplatin and chlorambucil); resulting cell lines exhibited resistance to the selecting agents (ranging approx. 1.5- to 36-fold) and, in some cases, cross-resistance to methotrexate (approx. 1.4- to 22-fold), vincristine (1.6- to 262-fold), doxorubicin (Adriamycin, approx. 1.1- to 33-fold) and taxotere (approx. 1.1- to 36-fold). Several of the variants displayed collateral sensitivity to cisplatin. A marked increase in in vitro invasiveness and motility was observed with variants pulsed with mitoxantrone, 5-fluorouracil, methotrexate, BCNU, cisplatin and chlorambucil. There was no significant change in invasiveness of cells pulsed with VP-16, vincristine, taxotere or CCNU. All of the pulse-selected variants showed elevated levels of MDR-1/P-gp protein by Western blot analysis, although mdr-1 mRNA levels were not increased (except for DLKP-taxotere). In DLKP-taxotere, MRP1 protein levels were also greatly elevated, but mrp1 mRNA levels remained unchanged. BCRP was upregulated in DLKP-mitoxantrone at both the mRNA and protein levels. Gelatin zymography, Western blot and RT-PCR showed that DLKP and its variants secreted MMPs 2, 9 and 13. MMP inhibition assays suggested that MMP-2 plays a more important role than MMPs 9 and 13 in cell invasion of these DLKP drug-resistant variants in vitro. These results indicate that drug exposure may induce not only resistance but also invasiveness in cancer cells.