A CDNA Microarray Study Of Apoptosis-related Genes And Assessment Of ML-IAP Expression Status In Malignant Melanoma

Background: Malignant melanoma (MM) is highly resistant to various apoptosis-inducing therapeutic agents, indicating serious deregulation of apoptosis in MM. However, the complex anti-apoptotic mechanisms of MM are not clear, nor is the global expression status of apoptosis-related genes in MM. ML-IAP, a recently identified member of the intrinsic cellular inhibitor of apoptosis protein (IAP) family, is overexpressed in some melanoma cell lines and is a potential target for MM therapy. Nonetheless, the ML-IAP expression status in benign and malignant melanocytic lesions has yet to be elucidated. The present study aims to (1) systematically evaluate the global expression status of apoptosis-related genes and (2) to determine the expression rate of ML-IAP in MM and melanocytic nevus tissues. Materials and methods: (1) Human apoptosis cDNA microarray (HS-002, SuperArray) was employed for expression profiling of 96 apoptosis-related genes in 6 malignant melanoma and 3 dermal nevus samples. (2) ML-IAPcDNA probe for chromogenic in situ hybridization (CISH) was designed and prepared. The expression rate in 34 archived melanoma and 14 dermal nevus samples was assessed by immunohistochemistry (IHC), and by CISH and reverse transcription PCR (RT-PCR) assays.Result: Pathway-specific CDNA microarray data showed that (1) anti-apoptotic genes cIAPl, cIAP2, XIAP, and Apollon/Bruce, of the IAP family, and Bcl-2, Mcl-1, and Bcl-xL, and BCL2A1, of the Bcl-2 family were highly expressed in MM; (2) the initiator caspases, including caspase 8 and caspase 9, had low expression levels in MM; (3) some pro-apoptotic genes, including Bax, Bak, of the Bcl-2 family, and the effector caspase 3, were paradoxically expressed in a fairly high level in MM; (4) most of the TRAF, TNF and TNFR superfamily members were expressed at low levels in both nevus and melanoma.ML-IAP assays demonstrated that (1) expression of ML-IAP was detected in 47.6-70.6% of the melanomas, varying with detection methods, with the expression rate in melanoma much higher than that in melanocytic nevus (21.4~25.0%) (p<0.05); (2) no statistically significant difference was observed between primary and secondary melanomas (p>0.05); and (3) ML-IAP expression rates assessed by the three methods were in general agreement.Conclusions: (1) The high expression levels of the IAP and Bcl-2 family members, together with the low expression levels of initiator caspases (caspase 8 and 9), may have contributed significantly to the anti-apoptotic mechanisms in MM. However, some pro-apoptotic genes, such as Bax andthe effector caspase 3, were also expressed in fairly high levels in MM, the significance of which required further investigation. (2) ML-IAP positivity rate in archived melanoma tissues was 47.6-70.6%, which was much higher than that of dermal nevus (21.4-25.0%), indicating a significant role of ML-IAP in the pathogenesis of MM. (3) No significant difference of ML-IAP positivity rate was observed between primary and secondary melanomas, suggesting that ML-IAP was not involved in the development of metastatic potential of MM.