H69AR is a multidrug-resistant human small-cell lung carcinoma cell line that was selected in doxorubicin and has previously been shown to be cross-resistant to a variety of natural-product-type anticancer drugs. H69AR is unlike many other multidrug-resistant cell lines in that it does not overexpress P-glycoprotein. In the present study, the drug sensitivity and cross-resistance patterns of H69AR cells were further characterized. A total of 15 drugs belonging to a number of chemical classes were screened. These compounds included anthracyclines, DNA binders (anthrapyrazoles, benzothiopyranoindazoles, and pyrazoloacridines), and lipophilic antifolates. The alkylating agent melphalan and the antimetabolite cytosine arabinofuranoside (Ara-C) were also tested. In general, the drug sensitivity and cross-resistance profiles of H69AR cells were consistent with those reported by others using other drug-resistant cell lines. However, there were several unexpected instances of cross-resistance. Thus, the H69AR cell line was more resistant than its parent cell line to the potent 3'-deamino-3'-(3-cyano-4-morpholinyl) doxorubicin, bisantrene, the pyrazoloacridine PD 114541, Ara-C, and melphalan. In addition, no cross-resistance to the four lipophilic antifolates tested, including trimetrexate, was found. The absence of a consistent pattern among the various drug-resistant cell lines indicates that assumptions about the efficacy of anticancer drugs in multidrug resistance should be made with caution.