Alphafoetoprotein uptake by cloned cell lines derived from a nickel-induced rat rhabdomyosarcoma.

Rat, mouse, pig and chicken alphafoetoproteins (AFP), rat serum albumin and egg albumin, or their fluoresceinated conjugates were added to cultures of several cloned cell lines isolated from a nickel-induced rat rhabdomyosarcoma. The intracellular uptake of assayed proteins was revealed by the indirect immunoperoxidase technique and/or by direct fluorescence microscopy. All the clones examined bound AFP, and all but one internalized the protein. The protein localized in the membrane and the cytoplasm, as well as along straight processes interconnecting cells. Nuclei were always AFP negative. The protein uptake of fluoresceinated conjugates of AFP and serumalbumin was already visible 15 min after incubation and progressed with time to reach a plateau 4-5 h later. Ultrastructural radioautographs of cells incubated with [3H]-AFP (rat) showed protein accumulation in several organelles and particularly in lipid droplets. Parallel to these observations, the intracellular presence of AFP within myofibrillar structures was demonstrated in tongue sections of rat foetuses and neonates. The results presented here provide experimental evidence of the reappearance in cloned cell lines derived from a primary rhabdomyosarcoma of a property pertaining to foetal striated muscle

Sensitivity to CPT-11 of xenografted human colorectal cancers as a function of microsatellite instability and p53 status

Biological parameters influencing the response of human colorectal cancers (CRCs) to CPT-11, a topoisomerase 1 (top1) inhibitor, were investigated using a panel of nine CRCs xenografted into nude mice. CRC xenografts differed in their p53 status (wt or mut) and in their microsatellite instability phenotype (MSI+when altered). Five CRC xenografts were established from clinical samples. All five had a functional p53, two were MSI+and three were MSI–. Tumour-bearing nude mice were treated intraperitonealy (i.p.) with CPT-11. At 10 mg kg–1of CPT-11, four injections at 4-day intervals, four of the five xenografts responded to CPT-11 (growth delay of up to 10 days); the non-responder tumour was MSI−. At 40 mg kg−1of CPT-11, six injections at 4-day intervals, the five CRCs displayed variable but marked responses with complete regressions. In order to assess the role of p53 status in CPT-11 response, four CRC lines were used. HT29 cell line was MSI−/ Ala273-mutp53, its subclone HT29A3 being transfected by wtp53. LoVo cell line was MSI+/ wtp53, its subclone X17LoVo dominantly expressed Ala273-mutp53 after transfection. LoVo tumours (MSI+/ mutp53) were more sensitive than X17LoVo (MSI+/ mutp53. HT 29 tumours (MSI−Imutp53), were refractory to CPT-11 while HT29A3 tumours (MSI−/ wtp53) were sensitive, showing that wtp53 improves the drug-response in these MSI−tumours. Levels of mRNA expression of top1, fasR, TP53 and mdr1 were semi-quantified by reverse transcription polymerase chain reaction. None of these parameters correlated with CPT-11 response. Taken together, these observations indicate that MSI and p53 alterations could be associated with different CPT-11 sensitivities; MSI phenotype moderately influences the CPT-11 sensitivity, MSI+being more sensitive than MSI−CRC freshly obtained from patients, mutp53 status being associated with a poor response to CPT-11. © 2000 Cancer Research Campaig