Abstract
We evaluated the MDR1 expression levels in 77 osteosarcomas and investigated whether MDR1 mRNA expression in osteosarcomas varies with location within the tumour, following chemotherapy, or after metastasis. We modified the semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) assay to determine accurately the levels of MDR1 mRNA expression in clinical specimens. We show that specimens collected from multiple locations in six tumours revealed very little variation in MDR1 expression suggesting that the levels of MDR1 in these tumours do not vary greatly with location within the tumour mass. In a comparison of pre and post-chemotherapy specimens it was found that MDR1 levels did not change appreciably following chemotherapy in 16 of 20 cases. In addition, in eight of ten specimens obtained before and after metastasis, the amount of MDR1 mRNA was found to remain relatively constant despite metastatic spread. Thus, many osteosarcomas exhibited intrinsic expression of MDR1 mRNA before multidrug regimens which invariably included doxorubicin and, in most cases, MDR1 expression was not induced following chemotherapeutic treatment. Our results suggest that some osteosarcoma patients may have primary tumours which are resistant to doxorubicin. These individuals may benefit from different chemotherapeutic regimens, e.g. the addition of MDR reversal agents.
Highlights
In the present study we describe a refined reverse transcription-polymerase chain reaction (RT-PCR) assay capable of sensitive detection and quantitation of low-level MDR1 expression in tumour specimens
We investigated whether MDR1 expression changes with progression by analysing specimens from biopsy through recurrence with metastasis
The semiquantitative reverse transcriptase (RT)-PCR assay of Noonan et al (1990) was modified to quantitate the levels of MDR1 mRNA observed in osteosarcomas
Summary
KB3-1 parental drug-sensitive and KB8 and KB8-5 drugresistance carcinoma cell lines were obtained from MM Gottesman (Akiyama et al, 1985). KB8 represents the first step in selection from KB3-1 cells and exhibits 2-fold resistance to colchicine and a 6- to 10-fold increase in MDR1 mRNA. KB8-5 cells have 40-fold increased expression of MDR1 without gene amplification and are four times more resistant to colchicine than KB3-1 cells. All osteosarcoma specimens were obtained from patients who presented free of metastatic disease. Specimens were collected at both biopsy and resection, as well as from various locations within the tumour. Specimens were collected from lung metastasis in the event of recurrent systemic disease. Specimens were obtained immediately after removal and flash-frozen in liquid nitrogen
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