Cancer Chemother Pharmacol (1993) 33: 113-122 ancer hemotherapy and harmacology 9 Springer-Verlag 1993 Antitumor alkylating agents: in vitro cross-resistance and collateral sensitivity studies Emil Frei III, Sylvia A. Holden, Rene Gonin, David J. Waxman, Beverly A. Teicher Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA Received 22 February 1993/Accepted 2 July 1993 Abstract. Cell lines resistant to five antitumor alkylating agents (CDDP, PAM, 4-HC, HN2, and BCNU) were developed from five parental human tumor lines repre- sentative of solid tumors with a range of sensitivities to antitumor alkylating agents. The parental cell lines were SCC-25 squamous carcinoma of the head and neck, MCF-7 breast carcinoma, SW2 small-ceU lung cancer, SL6 non-small-cell lung carcinoma, and G3361 melanoma. Survival curves using colony formation as the endpoint were generated for each of the 25 cell lines to each of the five alkylating agents. Comparison of the drug concentra- tions that reduced the survival of the alkylating agent-re- sistant cell lines by 90% (IC90 values) with the IC90 values obtained for the corresponding parental cell lines was used as a measure of the resistance/sensitivity of the alkylating agent-resistant lines to each drug tested. Although cross-re- sistance among the alkylating agents was generally uncom- mon, several patterns of response emerged. Cross-resis- tance occurred in 27 of the 105 determinations and oc- curred most frequently in the cell lines in which resistance was developed to PAM (57%) or BCNU (38%). Cross-re- sistance to HN2 occurred most frequently. Collateral sensi- tivity was equally as common, occurring in 25 of the 105 determinations. Collateral sensitivity occurred most frequently in the cell lines made resistant to 4-HC. The 4-HC-resistant cell lines were most frequently collaterally sensitive to PAM and to BCNU. Cross-resistance This work was supported by NIH grant PO1-38493, by a grant from the Mathers Foundation, by a grant from Bristol-Myers-Squibb, Inc., Wal- lingford, Connecticut, and by ACS grant CH-487 Abbreviations: AAs, antitumor alkylating agents; CDDP, cisplatin, cis- diamminedichloroplatinum(II); HN2, nitrogen mustard; BCNU, N,N'- bis(2-chloroethyl)-N-nitrosourea; PAM, L-phenylalanine mustard, mel- phalan; 4-HC, 4-hydroxyperoxycyclophosphamide; CPA, cyclo- phosphamide; THIO, trimethyleneiminethiophosphoramide; DME, Dul- becco's modified Eagle's medium; ICg0, 90% inhibitory concentration; PBS, phosphate-buffered 0.9% saline; FBS, fetal bovine serum; GSH, glutathione; GST, glutathione-S-transferase Correspondence to: Beverly A. Teicher developed most frequently in the MCF-7 breast carcinoma and SCC-25 squamous-cell carcinoma cell lines, whereas collateral sensitivity developed most frequently in the SW2 small-cell lung cancer line and the G3361 melanoma cell line and least frequently in the MCF-7 breast carcinoma cell line and the SL6 non-small-cell lung cancer cell line. The implication of these findings for the development of strategies for clinical treatment are discussed. Introduction Antitumor alkylating agents (AAs) have been considered to be a relatively homogeneous class of compounds. AAs were described as radiomimetic agents and were consid- ered to be cross-resistant with each other, primarily on the basis of clinical impressions [43]. These generalizations were challenged by the classic data of Schabel et al. [35-37], which indicated that in tumor sublines of the L1210 murine leukemia system made resistant to specific AAs, cross-resistance among the AAs was the exception rather than the rule [39], and, more recently, by data ac- crued in several human tumor lines [17, 19, 20, 43-47, 51]. We have confirmed the findings of Schabel et al. in our studies in human tumor cell lines, which indicated that cross-resistance among, the AAs is indeed the exception rather than the rule [17, 19, 20, 37, 44-47, 51]. Chemi- cally, AAs are quite heterogeneous, leading to different mechanisms of resistance to specific agents within the class. Similarly, other determinants of drug action, such as plasma membrane transport and intracellular biotransfor- mation (activation or inactivation), vary substantially among AAs [21, 22, 32, 49]. The mechanisms of resistance to AAs are multiple and multifactorial, consistent with the general lack or, at best, low levels of cross-resistance among these drugs [17, 19, 20, 43, 45-47, 51]. There has been renewed interest in the AAs in recent years largely because, as a class, they are ideal agents for