pep005 and Melanoma

Skin cancer is a broad term used to describe a number of different malignant indications of the skin. Skin cancers mostly comprise of the keratinocyte cancers [Basal Cell Carcinoma (BCC) and cutaneous Squamous Cell Carcinoma (SCC)], and melanoma. Surgical excision of these malignancies has been the preferred treatment of patients for decades. However, the decision to perform surgery can be affected by various considerations, including co-morbidities of the patient, the anatomical site of the lesion and potential intolerance for repeated excisions. Topical treatment of skin cancer may therefore be more appropriate in certain instances. Topical treatment potentially allows for higher drug levels at the tumor site, and may result in less overall toxicity than systemic agents. This review will specifically address the current agents used in topical treatment of skin cancers, and introduce emerging treatments from the natural product field that may also find utility in these indications.

Topics: Administration, Cutaneous; Antineoplastic Agents; Carcinoma, Basal Cell; Clinical Trials as Topic; Diterpenes; Drug Delivery Systems; Fluorouracil; Humans; Hutchinson's Melanotic Freckle; Imiquimod; Keratosis, Actinic; Melanoma; Neoplasms, Squamous Cell; Photochemotherapy; Retinoids; Skin; Skin Neoplasms; Ultraviolet Rays

Taking into consideration that the immune system plays a very important role in the development of melanoma and non-melanoma skin cancers, which have a high prevalence in immunosuppressed patients and after prolonged ultraviolet radiation, the interest in developing novel therapies, in particular targeting the inflammation in cancer, has increased in the past years. The latest data suggest that therapies such as imiquimod (IMQ), ingenol mebutate (IM), 5-fluorouracil (5-FU), retinoids, and nonsteroidal anti-inflammatory drugs (NSAIDs) have been used with success in the topical treatment of some cancers. Herein, we review the topical treatment targeting the inflammation in skin cancer and the mechanisms involved in these processes. Currently, various associations have shown a superior success rate than monotherapy, such as systemic acitretin and topical IMQ, topical 5-FU with tretinoin cream, or IMQ with checkpoint inhibitor cytotoxic T lymphocyte antigen 4. Novel therapies targeting Toll-like receptor-7 (TLR-7) with higher selectivity than IMQ are also of great interest.

Topics: Administration, Topical; Aminoquinolines; Animals; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Cytokines; Diterpenes; Fluorouracil; Humans; Imiquimod; Inflammation; Melanoma; Neoplasm Metastasis; Skin; Skin Neoplasms; Toll-Like Receptor 7

BACKGROUND A recent focus in skin cancer prevention intervenes though modulating molecular links between inflammation and cell growth signaling, such as NF-κB. This study elucidates the effect of a non-tumor promoting phorbol ester, ingenol-3-angelate (I3A), on the growth of human melanoma cells and on the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin inflammation and 7,12-Dimethylbenz(a)anthracene (DMBA)-induced skin carcinoma in mice. MATERIAL AND METHODS Cell viability was assessed by MTT assay, cell proliferation by clonogenic assay, apoptosis and cell cycle arrest was analyzed by flow cytometry, protein expression was studied by IHC and Western blotting, and gene expression by qPCR. RESULTS I3A suppressed the survival and proliferation of human melanoma cells with estimated IC50 values around 38 and 46 μM for A2058 and HT144 cell, respectively. I3A activated the protein levels of PKCδ and PKCε, which induced apoptosis by activating caspase-9 and caspace-3 followed by lowering of mitochondrial membrane potential and enhancing DNA fragmentation. I3A induced G1 phase cell cycle arrest as well as G2/M phase arrest in both cell lines. I3A inhibited the levels of NFκB p65 protein as well as phosphorylation of p65 and its nuclear translocation. I3A suppressed the gene expression of NF-κB, COX-2 and iNOS. I3A inhibited TPA-induced inflammation and epidermal hyperplasia in female ICR mice by downregulating NF-κB and iNOS. I3A suppressed the growth of skin tumor in DMBA-induced mice in dose-dependent manner. CONCLUSIONS The mechanism of I3A induces apoptosis in human melanoma cells and suppresses skin inflammation and carcinoma via downregulation of NF-κB-iNOS-COX-2 signaling.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Apoptosis; Apoptosis Regulatory Proteins; Carcinogenesis; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclooxygenase 2; Diterpenes; Down-Regulation; Epidermis; Female; Hyperplasia; Inflammation; Melanoma; Mice, Inbred ICR; NF-kappa B; Signal Transduction; Skin Neoplasms; Tetradecanoylphorbol Acetate

Ingenol mebutate is approved for the topical treatment of actinic keratoses and may ultimately also find utility in treating skin cancers. Here we show that relapse rates of subcutaneous B16 melanoma tumours treated topically with ingenol mebutate were not significantly different in C57BL/6 and Rag1-/- mice, suggesting B and T cells do not play a major role in the anti-cancer efficacy of ingenol mebutate. Relapse rates were, however, significantly increased in MyD88-/- mice and in C57BL/6 mice treated with the anti-IL-1 agent, anakinra. Ingenol mebutate treatment induces a pronounced infiltration of neutrophils, which have been shown to have anti-cancer activity in mice. Herein we provide evidence that IL-1 promotes neutrophil recruitment to the tumour, decreases apoptosis of infiltrating neutrophils and increases neutrophil tumour killing activity. These studies suggest IL-1, via its action on neutrophils, promotes the anti-cancer efficacy of ingenol mebutate, with ingenol mebutate treatment causing both IL-1β induction and IL-1α released from keratinocytes.

Topics: Animals; Antineoplastic Agents; Diterpenes; Female; Gene Deletion; Immunity, Cellular; Interleukin 1 Receptor Antagonist Protein; Interleukin-1; Melanoma; Mice, Inbred C57BL; Myeloid Differentiation Factor 88; Neoplasm Recurrence, Local

Ingenol mebutate is the active ingredient in Picato® a new drug for the treatment of actinic keratosis. A number of derivatives related to ingenol mebutate were prepared by chemical synthesis from ingenol with the purpose of investigating the SAR and potency in assays relating to pro-inflammatory effects (induction of PMN oxidative burst and keratinocyte cytokine release), the potential of cell death induction, as well as the chemical stability. By modifications of the ingenol scaffold several prerequisites for activity were identified. The chemical stability of the compounds could be linked to an acyl migration mechanism. We were able to find analogues of ingenol mebutate with comparable in vitro properties. Some key features for potent and more stable ingenol derivatives have been identified.

Topics: Antineoplastic Agents; Apoptosis; Cell Line; Diterpenes; Humans; Interleukin-8; Keratinocytes; Keratosis, Actinic; Leukocytes, Mononuclear; Melanoma; Oxidative Stress; Reactive Oxygen Species; Skin Neoplasms; Structure-Activity Relationship; Tumor Necrosis Factor-alpha

The sap of Euphorbia peplus, commonly know as 'petty spurge', 'radium weed' or 'milkweed' has been used for centuries as a traditional treatment for skin conditions, including warts, corns and cancers of the skin. Documentation of its use by medical professionals to treat basal cell carcinoma (BCC) dates from the early 19 century. Individuals who participated in a 1988 survey of home treatments for cancer indicated the sap of E. peplus was an effective cure for actinic lesions leading the investigators to suggest that this potential utility should be further explored in controlled clinical trials. The fractionation of the sap E. peplus using solvents of varying polarity yielded several macrocyclic diterpenes, many of which were found to have cytotoxic activity or the ability to influence cellular differentiation. Ultimately, ingenol 3-angelate (I3A) of PEP005, emerged as a promising potential new anti-cancer treatment. Here we report the proceedings from the First International Conference on PEP005, covering the exciting potential of PEP005 as the therapeutic agent for the treatment of skin cancer, leukemia and bladder cancer.

Topics: Administration, Topical; Animals; Antineoplastic Agents, Phytogenic; Diterpenes; Esters; Humans; Leukemia; Melanoma; Mice; Neoplasms; Skin Neoplasms; Urinary Bladder Neoplasms

The diterpene ester PEP005 is a novel anticancer agent that activates protein kinase C (PKC) and induces cell death in melanoma at high doses. We now describe the in vitro cytostatic effects of PEP005 and the diterpene ester phorbol 12-myristate 13-acetate, observed in 20% of human melanoma cell lines. Primary cultures of normal human neonatal fibroblasts were resistant to growth arrest, indicating a potential for tumor selectivity. Sensitive cell lines were induced to senesce and exhibited a G(1) and G(2)-M arrest. There was sustained expression of p21(WAF1/CIP1), irreversible dephosphorylation of the retinoblastoma protein, and transcriptional silencing of E2F-responsive genes in sensitive cell lines. Activation of mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK) kinase (MEK) 1/2 by PKC was required for diterpene ester-induced senescence. Expression profiling revealed that the MAP kinase inhibitor HREV107 was expressed at a higher transcript level in resistant compared with sensitive cell lines. We propose that activation of PKC overstimulates the RAS/RAF/MEK/ERK pathway, resulting in molecular changes leading to the senescent phenotype.

Topics: Cell Cycle; Cell Line, Tumor; Cellular Senescence; Diterpenes; E2F1 Transcription Factor; Enzyme Activation; Esters; Gene Silencing; Humans; Isoenzymes; MAP Kinase Signaling System; Melanoma; Phosphorylation; Protein Kinase C; Retinoblastoma Protein; Tetradecanoylphorbol Acetate; Transcription, Genetic

Ingenol 3-angelate (PEP005), one of the active ingredients in an extract from Euphorbia peplus, was shown in preclinical studies to have activity against human melanoma xenografts in nude mice. In the present study, we have tested its ability to induce the apoptosis of melanoma cells in vitro in the absence or presence of tumor necrosis factor-related apoptosis inducing ligand (TRAIL). The results showed that at relatively high concentrations (100 microg/mL), PEP005 killed melanoma cells mainly by induction of necrosis. In 20% of cell lines, evidence of apoptosis was observed. Apoptosis was caspase-dependent and associated with changes in mitochondrial membrane potential that were not inhibitable by overexpression of Bcl-2 or inhibition of caspases but were blocked by inhibition of protein kinase C (PKC). Low concentrations (1 or 10 microg/mL) of PEP005 either increased or decreased TRAIL-induced apoptosis in a cell line-dependent manner. These changes in TRAIL-induced apoptosis seemed to be due to activation of PKC and varying levels of PKC isoenzymes in different melanoma cell lines. PEP005-mediated enhancement of apoptosis seemed to be associated with low expression of the PKCepsilon isoform. These results indicate that PEP005 may enhance or inhibit sensitivity of melanoma to treatments associated with TRAIL-induced apoptosis depending on the PKC isoform content of melanoma cells.

Topics: Apoptosis; Apoptosis Regulatory Proteins; Caspases; Diterpenes; Enzyme Activation; Esters; Gene Expression Regulation, Neoplastic; Humans; Melanoma; Membrane Glycoproteins; Membrane Potentials; Mitochondria; Protein Kinase C; Protein Kinase C-epsilon; Proto-Oncogene Proteins c-bcl-2; Skin Neoplasms; TNF-Related Apoptosis-Inducing Ligand; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha