Experimental therapy of doxorubicin resistant human uveal melanoma with targeted cytotoxic luteinizing hormone-releasing hormone analog (AN-152)

Background: Cytotoxic analogs of LHRH (luteinizing hormone-releasing hormone) can be successfully used for the treatment of hormone-dependent cancers such as prostatic, ovarian, endometrial, but our knowledge about their effect on hormone-independent cancers such as human uveal melanoma (UM) is limited. Previously, we have demonstrated that 46% of UM express full-length LHRH receptors. This finding has led us to further examine the mechanism of action of LHRH receptor based targeted therapies in this malignancy. Aims: In the present study we investigated the cellular uptake of doxorubicin (DOX) and cytotoxic LHRH analog AN-152 (AEZS-108, zoptarelin doxorubicin) on human UM cell lines (OCM3) and its DOX resistant form OCM3_DOX320 by confocal laser scanning microscopy. The LHRH receptor expression was characterized by RT-PCR and immunocytochemistry. Results: We were able to establish a new, stable and DOX resistant human UM cell line OCM3_DOX320. Our results demonstrated the expression of splice variants and isoforms of receptor for LHRH in OCM3 UM cell line and its doxorubicin resistant form OCM3_DOX320. It has been revealed by MTT assay that AN-152 inhibited cell proliferation in a dose dependent manner in OCM3_DOX320 cells. Furthermore, receptor-mediated uptake of AN-152 was demonstrated using confocal laser scanning microscopy in both cell line. Conclusions: Our results suggest that the antiproliferative effect of AN-152 can be detected even if only LHRH receptor isoforms are expressed. Our study also demonstrates the LHRH receptor-mediated uptake of AN-152 in DOX resistant OCM3_DOX320 cells. Our experiments provide new insights into a potential targeted therapy of UM and give further details about the accumulation of AN-152 in hormone-independent DOX-resistant cells expressing splice variants of the LHRH receptors.