Issn Print: 0960-8931
Publication Date: 2007/02/01
INV 22 Drug resistance mechanisms
Excerpt
Treatment of disseminated melanoma with chemotherapy has had limited success, since the tumors frequently show intrinsic resistance. Up to now, the most extensively used drug has been the DNA alkylating agent dacarbazine (DTIC), which generates O6-methylguanine cytotoxic lesions. Now an increasing number of patients are being treated with temozolomide (TMZ), an orally administered drug that has the same mechanism of action as DTIC. The DNA repair protein, O6-methylguanine-DNA methyltransferase (MGMT) has been shown to inhibit the killing of tumor cells by alkylating agents such as DTIC and TMZ by removing cytotoxic alkyl adducts from the O6-guanine base. Loss of MGMT expression in tumor cells is associated with hypermethylation of the MGMT CpG islands in the promoter, supposing that methylation of the MGMT promoter may predict sensitivity to chemotherapy. We are therefore investigating the role of MGMT hypermethylation in melanoma tumors and its potential relationship to clinical response to DTIC/TMZ-based chemotherapy and protein expression. The MGMT promoter methylation status was assessed, using both methylation-specific PCR (MSP) and pyrosequencing, in tumors from melanoma patients who had received DTIC/TMZ-based chemotherapy, 19 responders and 45 non-responders. The pyrosequencing analyses gave us the possibility to study the methylation status of, totally, 32 CpG sites within the promoter region and to compare the impact of methylation of specific CpGs on protein expression. There was a better association between MGMT promoter methylation and clinical response to treatment with DTIC/TMZ only than to DTIC/TMZ in combination with other drugs. In the cases where patients received DTIC/TMZ only, four of six (67%) responders had MGMT-methylated tumors compared with six (MSP) or seven (pyrosequencing) of 27 (22–26%) non-responders, which is consistent with the hypothesis that MGMT promoter methylation may predict the clinical response to DTIC/TMZ-based chemotherapy. A comparison between MGMT methylation and protein expression was performed showing a lack of or weak protein expression in 14/19 (74%) MGMT-methylated tumors compared with 16/37 (43%) unmethylated tumors. Some of the methylated tumors still show high protein expression, which may be due to differences in the number of methylated CpG sites indicating that the number of methylated CpGs have an impact on the inactivation of the gene. The limitation with the above-described study is that it focuses on only one putative drug resistance factor. There are also other possible candidate factors of importance for resistance to temozolomide and DTIC, such as other DNA repair systems (DNA mismatch repair) (Barvaux et al., 2004; Hunter et al., 2006) and genes involved in the regulation of apoptosis (bax, bcl-2, Apaf-1) (Barvaux et al., 2004; Gyo¨rffy et al., 2006; Soengas et al., 2001). In addition, expression patterns of some, at present, unidentified genes may be of importance for resistance to these drugs. It is therefore appropriate to perform large-scale expression analysis using microarray technology to screen for gene expression patterns, which may have an impact on the clinical outcome. The underlying mechanisms causing resistance is very complex and there is a need in the future to study patterns rather than single genes.
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