kn-93 and Melanoma

Acral melanoma is a predominant and aggressive subtype of melanoma in non-Caucasian populations. There is a lack of genotype-driven therapies for over 50% of patients. TRPM1 (transient receptor potential melastatin 1), a nonspecific cation channel, is mainly expressed in retinal bipolar neurons and skin. Nonetheless, the function of TRPM1 in melanoma progression is poorly understood.. We investigated the association between TRPM1 and acral melanoma progression and revealed the molecular mechanisms by which TRPM1 promotes tumor progression and malignancy.. TRPM1 expression and CaMKII phosphorylation in tumor specimens were tested by immunohistochemistry analysis and scored by two independent investigators. The functions of TRPM1 and CaMKII were assessed using loss-of-function and gain-of-function approaches and examined by western blotting, colony formation, cell migration and invasion, and xenograft tumor growth assays. The effects of a CaMKII inhibitor, KN93, were evaluated using both in vitro cell and in vivo xenograft mouse models.

Topics: Animals; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Humans; Melanoma; Melanoma, Cutaneous Malignant; Mice; Neoplastic Processes; Proto-Oncogene Proteins c-akt; TRPM Cation Channels

Topics: Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Line, Tumor; Cell Proliferation; Humans; Melanoma; Membrane Proteins; Phosphorylation; Signal Transduction

1. The purpose of the present study was to investigate the molecular mechanisms that control tumour cell resistance and to search for molecules that could overcome Fas ligand (FasL) or CH-11 resistance in certain tumours, including glioma and melanoma. 2. Twelve tumour cell lines were examined for their sensitivity to CH-11-induced apoptosis and then two of each of the CH-11-sensitive and -resistant tumour cell lines were analysed for Fas-mediated death-inducing signalling complex (DISC). The calmodulin kinase II (CaMKII) inhibitor KN-93 and the chemotherapeutic drug cisplatin were used to treat resistant cells; the effects of these two drugs on CH-11-resistant tumour cells were investigated. 3. In CH-11-sensitive tumour cells, apoptosis-initiating caspase 8 and caspase 10 were recruited to the DISC, where they became activated through autocatalytic cleavage, leading to apoptosis through cleavage of downstream substrates, such as caspase 3 and DNA fragmentation factor 45. 4. In CH-11-resistant cells, cellular Fas-associated death domain-like interleukin-1b-converting enzyme inhibitory protein (c-FLIP) proteins were recruited to the DISC, resulting in inhibition of caspase 8 and caspase 10 cleavage. The c-protein expression and phosphorylation of FLIP and CaMKII protein and enzyme activity were upregulated in resistant cells. Treatment of resistant cells with 100 micromol/L KN-93 and 10 microg/mL cisplatin downregulated c-FLIP expression, inhibited c-FLIP phosphorylation and rescued CH-11 sensitivity. 5. In conclusion, KN-93 and cisplatin inhibit c-FLIP protein expression and phosphorylation restores CH-11-induced apoptosis in tumour cells. tHe present study provides evidence for the use of a new combination therapeutic strategy in the treatment of malignant tumours.

Topics: Antibodies; Antineoplastic Agents; Apoptosis; Benzylamines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; CASP8 and FADD-Like Apoptosis Regulating Protein; Caspase 10; Caspase 8; Cell Line, Tumor; Cell Survival; Cisplatin; Down-Regulation; Drug Resistance, Neoplasm; Fas Ligand Protein; Glioma; Humans; Melanoma; Phosphorylation; Protein Kinase Inhibitors; Sulfonamides