In vivo emergence of a highly metastatic tumour cell line from a rat rhabdomyosarcoma after treatment with an alkylating agent

Abstract

Summary Rats bearing a transplanted nickel-induced rhabdomyosarcoma (RMS 9-4/0), treated with chlorozotocin (CZT), an alkylating agent, showed an amplified metastatic invasion of the lung (median of 165 lung tumour nodules, compared to 3 for untreated controls). A higher level of metastatic invasion (200 nodules) was reached spontaneously after the grafting of the S4T line, which was obtained by successive in vivo passages of RMS 9-4/0 cells in CZT treated rats. S4T tumour cells also invaded the liver and a considerable proportion of the lymph nodes. The NT4T line, obtained by successive in vivo passages in untreated rats, showed a lesser degree of enhancement of metastatic capacity (57 nodules). Both derived lines proved to be more aggressive than the parental, proliferated more rapidly, and were resistant to CZT toxicity. Only the non-treated lineage became more resistant to NK lysis. The S4T line lost its myogenic differentiation and was best described as a fibrohistiosarcoma, whereas NT4T did not. Chromosome analysis demonstrated a reduced range of chromosome number per cell in both lines. We conclude that both S4T and NT4T tumours became more metastatic than RMS 9-4/0 as the result of tumour progression through in vivo passages, and that in addition S4T acquired a spontaneously higher metastatic potential, similar to that which occurred in rats grafted with RMS 9-4/0 or NT4T tumours and treated by CZT. This suggests an inheritable mutation in the S4T line.A major problem of cancer therapy is the emergence of tumour subpopulations resistant to the treatment. The probability that a tumour will generate resistant variants could be related to genetic instability (Goldie & Coldman, 1984;Stephens et al., 1986 Davies, 1982). Repair mechanisms may allow mutated cells to survive, and increase both tumour drug resistance and phenotypic heterogeneity.We have described the enhancing effect of a nitrosourea, chlorozotocin 2-(3-(2-chloroethyl)-3-nitrosoureido)-D-glucopyranose (CZT), on the metastatic ability of RMS 9-4/0, a rat rhabdomyosarcoma (Poupon et al., 1984). We hypothesized that this action was due to the emergence of a subpopulation of the RMS 9-4/0 tumour, that was both resistant to CZT and highly metastatic. We failed to obtain a selection for CZT resistance in the primary tumour because the treated rats were rapidly killed by the growth of lung metastases. To overcome this, the treated primary tumour cells were injected into new recipients that were also treated and this cycle was repeated until a fully resistant population was obtained. The metastatic ability of these cells was then studied. A group of the tumour-bearing rats were not drug-treated and gave rise to the NT(x)T tumour lineage, where (x) designates the number of cycles performed. When rats were moribund the tumour was excised, finely minced with a scalpel, and then allowed to grow in the complete medium described above. At confluence, cells were injected s.c. into new animals. Part of the cells were frozen for further analyses. Materials and me...

Intrinsic or acquired drug resistance and metastasis: Are they linked phenotypes?

“…The magnitude of the increase in metastasis was similar to that observed by Teicher [19931 when studying the alkylating-agent resistant sublines of the EMT-6 mouse mammary tumor, as discussed earlier [Teicher et al, 19901. Poupon and her colleagues have also reported evidence that single chlorozotocin exposures of tumor-bearing animals can result in a similar "metastatic amplification" even if the growth of the primary tumor is suppressed [Pauwels-Vergely and Poupon, 1988;Poupon et al, 19841. However, in these cases it is difficult to determine whether the enhancing effect on metastasis is due t o druginduced damage of host tissues, leading to elevated tumor cell arrest andlor growth at the sites of damage, or to a direct effect on tumor cell genotype and phenotype [McMillan and Hart, 19871.…”

In vivo establishment and characterization of a paclitaxel-resistant human ovarian cancer cell line showing enhanced growth properties and drug-resistance only in vivo

“…2a). Although OM1/Tvivo acquired an enhanced in vivo growth property-as previous reports describing the in vivo established drug-resistant cell lines often show increased malignant potentials (Antoine et al 1988)-no invasion or metastasis was observed as in its parental OVMG1 (Table 1). In the case of our previously reported in vivo established cisplatin-resistant murine cancer cell lines, both invasive and metastatic properties were enhanced, compared with the in vivo-passaged control (drug sensitive) cell lines (Mitsumoto et al 1998).…”

“…Some of these problems might be overcome by in vivo establishment of drug-resistant cell lines which exhibit enhanced malignant potentials as well as drug-resistance (Antoine et al 1988). By the administration of cisplatin into tumor-bearing animals, we previously established, in vivo, drug-resistant murine cell lines showing enhanced invasive and metastatic properties, and reported that they were considered to be more faithful and useful models for simulating biological aggressiveness of clinically recurrent cancers rather than conventional ÔclassicÕ in vitro established drug-resistant cell lines (Mitsumoto et al 1998).…”

Comprehensive model for simultaneous MRI determination of perfusion and permeability using a blood-pool agent in rats rhabdomyosarcoma

“…Experiments were performed under anesthesia by means of intraperitoneal injection of a 50/50 mix of xylazine (10 mg/kg, Rompun, Bayer, Leverkusen, Germany) and ketamine (50 mg/kg, Imalgene, Bayer). The tumor model was a rhabdomyosarcoma [13] implanted subcutaneously bilaterally in the flanks of each animal (at the level of the heart) by means of injection of 0.2 ml cell S4MH (provided by M.F. Poupon, Institut Curie, Paris, France) diluted in 1 ml fetal calf serum.…”