Expression and Targeting of the Apoptosis Inhibitor, Survivin, in Human Melanoma

Abstract

The newly described apoptosis inhibitor survivin is expressed in many human cancers and appears to play a critical part in both apoptosis regulation and cell cycle progression. Its potential role in malignant melanoma is unknown. In a panel of 30 malignant melanomas, survivin was strongly expressed in all cases (15 of 15) of metastatic malignant melanomas and 13 of 15 cases of invasive malignant melanomas by immunohistochemistry. In invasive malignant melanomas, survivin was also expressed in the in-situ component of the lesion. Survivin expression was found in all cases (11 of 11) of nevi, but not in melanocytes in sections of normal skin. The apoptosis inhibitor bcl-2 was expressed in 26 of 30 cases, but generally at lower levels than that of infiltrating lymphocytes. The mitotic index, as assessed by MIB-1 staining, was consistently higher in metastatic than invasive malignant melanomas. Assessment of apoptotic index by in situ end-labeling revealed extremely low rates of apoptosis in most malignant melanomas. Survivin expression by western blotting was detected in four human metastatic malignant melanoma cell lines but not in cultured normal human melanocytes. Transfection of both YUSAC-2 and LOX malignant melanoma cells with green fluorescence protein-conjugated survivin anti-sense or green fluorescence protein-conjugated survivin dominant negative mutant (Cys84Ala) [corrected] resulted in increased apoptosis in the absence of other genotoxic stimuli. Two-color flow cytometry confirmed that YUSAC-2 cells transfected with survivin anti-sense expressed less endogenous survivin and exhibited an increased fraction of cells with sub-G1 DNA content. These data demonstrate that apoptosis inhibition by survivin may participate in the onset and progression of malignant melanomas, and suggest that therapeutic targeting of survivin may be beneficial in patients with recurrent or metastatic disease.

Survivin knockdown by short hairpin RNA abrogates the growth of human hepatocellular carcinoma xenografts in nude mice

“…7,8,25,24 We postulated that distinct functional properties of survivin have different roles in antiapoptosis and cell mitosis. The strategy of dominant-negative mutants can only abrogate one function of survivin at one time, whereas siRNA can interrupt all the functions of survivin simultaneously and this may reinforce the inhibitory effect on the functions of survivin.…”

“…1 Attenuation of survivin expression or function by using antisense oligonucleotides, ribozymes, small interfering RNAs (siRNAs), or dominant-negative mutants has been shown to induce apoptosis or mitotic catastrophe in multi types of cancer. [5][6][7][8][9][10] The use of RNA interference (RNAi) has grown at a remarkable rate since the first discovery of RNAi in Caenorhabditis elegans. 11 Subsequently, RNAi was shown to occur in mammalian cells in response to doublestranded siRNAs of B21 nt in length that serve as the effector molecules of sequence-specific gene silencing.…”

Decreased proliferation of human melanoma cell lines caused by antisense RNA against translation factor eIF-4A1

“…For melanoma, clinical studies using antisense oligonucleotides against the antiapoptotic Bcl-2 are on the way aiming the sensitisation of tumour cells against chemotherapy (Jansen et al, 2000). Several other genes were targeted in melanoma cells by in vitro studies, as the anti-apoptotic factor survivin (Grossman et al, 1999), the transcription factor c-myc (Citro et al, 1998) and the growth factor pleiotrophin (Satyamoorthy et al, 2000).…”

Survivin expression in oral squamous cell carcinoma

“…Immunohistochemical staining was carried out with a rabbit polyclonal antibody supplied by NOVUS (NOVUS Biologicals, Littleton, CO, USA) raised against full-length recombinant survivin characterised in previous studies (Grossman et al, 1999a;Grossman et al, 2001), using the standard streptavidin -biotin -peroxidase complex technique using the Dako LSAB kit (DAKO A/S, Carpinteria, CA, USA) after antigen retrieval by pressure cooking. Briefly, deparaffined sections were immersed in a 10 À3 M sodium citrate buffer (pH 6.0) after bringing the solution to a boil in a pressure cooker, and then heated two times for 3 min each at a 10-min interval while keeping the pressure indicator valve rising.…”