Commentary

Resistance to cisplatin: How to deal with the problem?

Drug targeting may contribute to overcoming resistance to chemotherapy and to reducing side effects. Here, by conjugating a nitrogenated base (NB) to the side chain of a bile acid (BA) moiety, we have synthesized and evaluated six novel compounds, designated BANB-1 to -6, with potential cytostatic activity and vectoriality toward enterohepatic tumors. These compounds were purified by liquid chromatography and their purity was checked by TLC and HPLC before being chemically characterized using IR, ¹H/¹³C NMR and FAB-MS. Using several cell lines – HepG2 (human hepatoblastoma), LS 174T and Caco-2 (human colon adenocarcinoma), Hepa 1-6 (mouse hepatoma), McA-RH7777 (rat hepatoma), CCRF S-180 II (mouse sarcoma) and CHO (Chinese hamster ovary) – their effect on cell viability was measured with the formazan test after drug exposure for 6 h (cytotoxic effect) or 72 h (cytostatic effect). A weak cytostatic effect of BANB-1, BANB-2 and BANB-3 was detected even in CHO cells stably transfected with rat bile acid transporters (Ntcp and Oatp1/1a1). In contrast, BANB-4, BANB-5 and BANB-6, similarly to cisplatin, showed strong cytostatic effects, together with mild non-specific toxicity. BANB-6 was effective even against Hepa 1-6/R cells, which were partly resistant to cisplatin. Treatment with BANB-6, but not cisplatin, was able to prolong the life span of Nude mice bearing tumors formed by Hepa 1-6/R cells orthotopically implanted in the liver. In conclusion, our results support the hypothesis that cytostatic bile acid derivatives such as BANB-6 may offer a useful pharmacological strategy for the treatment of tumors of the enterohepatic circuit.

The mammalian 5′- to 3′-exonuclease/endonuclease, called RAD2 homologue 1 or flap endonuclease 1, has a unique cleavage activity, dependent on specific substrate structure. On a primer-template, in which the primer has an unannealed 5′-tail, endonucleolytic cleavage near the annealing point releases the tail intact. Entering at the 5′-end, the nuclease tracks along the entire tail to the point of cleavage. Genetic analyses suggest that this nuclease removes DNA adducts in vivo (Sommers, C. H., Miller, E. J., Dujon, B., Prakash, S., and Prakash, L. (1995) J. Biol. Chem. 270, 4193-4196). Micrococcal nuclease footprinting shows that after tracking the nuclease protects a region of the tail 25 nucleotides long, adjacent to the cleavage site.

Substrates with adducts at specific locations were used to assess the mechanism of RAD2 homologue 1 nuclease tracking and its ability to cleave modified DNA. Either a conventional cis-diamminedichloroplatinum (II) (CDDP) or a bulky CDDP derivative was placed within or beyond the region protected by the nuclease. The nuclease cleaved the tail of both substrates. In contrast, a CDDP adduct just adjacent to the expected cleavage point was inhibitory. A CDDP adduct at the very 5′-end of the tail was also cleaved. The nuclease could remove tails containing adducts on the sugar-phosphate backbone. Apparently, the nuclease is designed to slide over various types of damage on single stranded DNA and then cut past the damaged site.

$\text{Purpose}$: Human glioma cell lines resistant (U373MG^CP) and sensitive (U373MG) to cisplatin were used to evaluate the effect of cisplatin as a sensitizer to low dose rate irradiation (LDRI).

$\text{Methods and Materials}$: A cisplatin resistant glioma cell line U373MG^CP was developed by chronic exposure of parental U373MG cells to cisplatin. Plateau phase cells were treated with cisplatin, high dose rate (HDR) irradiation (1.12 Gy/min), LDRI (0.0088 Gy/min), or cisplatin concurrent with LDRI. Cell survival was determined by the colony forming assay.

$\text{Results}$: Both cell lines showed increased resistance to radiation at LDR compared with HDR, with Dose Modifying Factors (DMF at 10% survival level) of 1.7 for U373MG and 2.5 for U373MG^CP. The increased LDR sparing effect in the cisplatin resistant U373MG^CP cells indicates increased repair proficiency. The resistant cell line showed a fourfold increase in resistance to cisplatin cytotoxicity at the 10% survival level compared with the parental U373MG cells. Cisplatin enhanced the response of both cell lines to LDRI. The DMFs were 1.2, 1.2, and 1.7, respectively, for the sensitive U373MG cell line given 1 μg/ml, and the resistant cell line given 3 or 6 μg/ml cisplatin treatments concurrent with LDRI.

$\text{Conclusions}$: These data show that cisplatin can be an effective sensitizer to LDRI in both cisplatin resistant and sensitive glioma cell lines. However, in the resistant cell line, higher concentrations of cisplatin were necessary to achieve the same level of sensitization as in the sensitive cell line.

Twenty patients with documented progression after two or three cisplatin-based regimens for advanced epithelial ovarian carcinoma were treated with high-dose folinic acid (200 mg/m²), 5-fluorouracil bolus (400 mg/m²) and continuous infusion (600 mg/m²) for two consecutive days every two weeks. One clinically complete and two partial responses were observed in 16 evaluable patients, with 5 remaining stable. Median survival was 9 months. Toxicity was mild. This combination achieved a 19%(95% confidence interval 4%-46%) response rate in heavily pretreated cisplatin-resistant patients.

The ovary is among the more complex organs of the body and its functions are achieved by numerous cell types. All of these cell types have some tendency to undergo malignant transformation, but the vast majority of ovarian cancers are believed to be the result of malignant transformation of the ovarian surface epithelium. The concept that most ovarian cancer arises from this modified peritoneal mesothelium is credited to Sir Spencer Wells in 1872. Ovarian cancer is the most frequently fatal gynecologic malignancy, and approximately 20,000 cases per year are diagnosed in the United States. Progress in understanding the biology of this disease, including factors involved in its etiology, progression, and tendency to change from a relatively chemotherapy-sensitive tumor to one with marked drug resistance, has been slow. In this review, the complex features of the normal ovarian surface epithelial cells are considered in relation to the etiology and progression of the disease. The hypothesis that incessant or repetitious ovulation contributes to the initiation of the disease is explored in detail based on experimental data, epidemiologic information, and the potential for antioncogene inactivation in this interesting cell type. Lastly, based on the experimental data available, potential mechanisms of resistance to platinum, the cornerstone of aggressive ovarian cancer therapy, are discussed, as are approaches to overcoming drug resistance. It is hoped that the reader will be left with the feeling that the pace of our understanding of the biology of ovarian cancer is increasing at such a rate that answers to the questions of etiology and why chemotherapy often fails will be known in the foreseeable future.

A major problem in cytostatic treatment of malignant tumors is the development of chemoresistant cell clones. An increased understanding of chemoresistance related mechanisms, improved methods for the detection and localization of resistant cell populations including predictive conclusions on the effectiveness of cytostatic drugs would contribute to the advancement of anti-tumor strategies. This paper reviews current concepts suggested for the development of cellular resistance to natural product drugs (anthracyclines, Vinca alkaloids, epipodophyllotoxines, antibiotics; so-called multidrug resistance substances), alkylating agents (nitrosureas, busul fan and mitomycin C), heavy metal compounds (cisplatin) and antifolates (5-fluorouracil, methotrexate) and describes the role of drug transporting and binding proteins (P-170-glycoprotein), detoxifying enzymes (glutathion-S-transferase, dihydrofolate reductase), DNA repair enzymes (topoisomerase I and II, polymerase alpha and beta), and genomic alterations (amplification, double minutes and homogeneous staining regions) due to resistance. It is focussed on the employment of morphological methods (light microscopy, immunocytochemistry, electron microscopy, fluorescence analysis, in situ hybridization, computer aided morphometric analysis) which will help to detect resistant cell clones in tumor biopsies. First correlations between histological data and clinical course will be reported. In the future, the morphological determination of chemoresistance may play an important role in applied functional tumor pathology.