The Demethylating Agent 5-Aza Reduces the Growth, Invasiveness, and Clonogenicity of Uveal and Cutaneous Melanoma

Abstract

Citation: Rajaii F, Asnaghi L, Enke R, Merbs SL, Handa JT, Eberhart CG. The demethylating agent 5-Aza reduces the growth, invasiveness, and clonogenicity of uveal and cutaneous melanoma. Invest Ophthalmol Vis Sci. 2014;55:6178-6186. DOI:10.1167/ iovs.14-13933 PURPOSE. Uveal melanoma is the most common primary intraocular malignancy in adults. Although local disease can be controlled with radiation therapy or enucleation, many cases are complicated by metastases, which account for the significant mortality from this disease. To date, no chemotherapeutic regimens effectively treat local or metastatic disease. Epigenetic silencing of tumor suppressor genes has been shown to be an important factor in the growth and metastasis of many cancers. One form of epigenetic alteration is DNA methylation, which often occurs at promoter elements resulting in the silencing of target gene transcription. METHODS. We used 5-aza-20 -deoxycytidine (5-Aza), a well characterized demethylating agent that is US Food and Drug Administration approved to decrease DNA methylation in multiple uveal and cutaneous melanoma cell lines.RESULTS. Demethylation of melanoma cell lines using 5-Aza causes significant decreases in growth, invasion, and clonogenicity. Treatment of melanoma cells with combined 5-Aza therapy and irradiation showed an even more pronounced effect on cell viability. In addition, treatment with 5-Aza decreased the number of metastases from the eye to the lung in a murine cutaneous melanoma xenograft model. CONCLUSIONS.We demonstrate in vitro and in vivo that demethylating agents such as 5-Aza may be promising chemotherapeutic agents for treating melanoma and decreasing progression to metastatic disease. These results provide proof of concept for an exciting potential therapy to reduce mortality from this disease. Future work will focus on identifying pathways that mediate these changes.Keywords: demethylation, epigenetics, melanoma, metastasis, uveal melanoma U veal melanoma is the most common primary intraocular malignancy in adults, with an age-adjusted incidence of 5.1 cases per million.1 Advances in local therapy have allowed preservation of the patient's eye and vision in many cases; however, the 5-year survival rate of 81.6% has remained unchanged from 1976 to 2008, underscoring the need to develop effective therapies for this deadly disease. 1 Epigenetic mechanisms controlling gene expression have long been known to have a role in cancer development. [2][3][4] Changes in DNA methylation serve as early markers of tumor formation, and DNA methylation at CpG islands in promoters has been shown to decrease expression of tumor suppressor genes in many malignancies, including uveal melanoma. 5,6 In addition, demethylating agents have been used to decrease tumor formation in animal models of cancer and are US Food and Drug Administration (FDA) approved as treatment of myelodysplastic syndrome.7 However, the functional effects of altered DNA methylation in uveal melanoma are unclear.To determine the role of DNA methylatio...

Novel Methylation Patterns Predict Outcome in Uveal Melanoma

“…While large-scale epigenetic changes have been documented in UM tumors, and these changes have been significantly related to prognosis, there are currently no approved treatments using these agents in UM. In vitro, treatment with epigenetic modifying drugs has been shown to reduce growth and invasiveness in UM cell lines, and decitabine (a DNMT inhibitor) in combination with MEK inhibition has been shown to suppress growth in UM cells [ 39 , 40 ]. Additionally, decitabine has been used safely in clinical trials via hepatic arterial infusion in patients with unresectable liver metastases (NCT02316028), which is promising in the context of high risk UM patients [ 41 ].…”

Genetic and epigenetic insights into uveal melanoma

“…The demethylating agent 5‐Aza—also in combination with radiation—has been shown to decrease growth and invasion in uveal (OCM1, OCM3, 92.1) and cutaneous melanoma cell cultures. In a murine cutaneous melanoma intraocular xenograft model, the number of lung metastases was reduced if the cells were exposed to 5‐Aza prior to subretinal injection . The same study also reported a subsequent decrease in LINE‐1 methylation in cell cultures after 5‐Aza treatment.…”

“…In a murine cutaneous melanoma intraocular xenograft model, the number of lung metastases was reduced if the cells were exposed to 5-Aza prior to subretinal injection. 118 The same study also reported a subsequent decrease in LINE-1 methylation in cell cultures after 5-Aza treatment.…”

Ocular melanoma-when you have seen one, you have not seen them all: a clinical outcome study from the Surveillance, Epidemiology and End Results (SEER) database (1973–2012)

“…Epigenetic drugs, including DNA methyltransferase (DNMT) inhibitors and HDAC inhibitors, have been shown to have anticancer properties and are beginning to be investigated for ocular tumors with hopes of restoring normal control of neoplastic genomes 42,43. Vidaza (5-azacytidine), a DNMT inhibitor, has been shown to reduce ocular metastasis to the lung in murine xenograft models 42,44. Tenovin-6, which inhibits the class 3 HDAC sirtuin 1 and 2, has shown promise in eliminating UM tumor cells and cancer stem cells 45…”