cytochrome-c-t and Mastocytosis

Gain-of-function mutations of membrane receptor tyrosine kinase KIT, especially gatekeeper D816V point mutation in KIT, render kinase autoactivation, disease progression, and poor prognosis. D816V KIT is found in approximately 80% of the patients with systemic mastocytosis, and is resistant to the first and second generations of tyrosine kinase inhibitors (TKI). The purpose of this investigation was aimed at exploring whether ponatinib (AP24534), a novel effective TKI against T315I Bcr-Abl, was active against D816V KIT. We discovered that ponatinib abrogated the phosphorylation of KIT harboring either V560G (sensitive to imatinib) or D816V mutation (resistant to imatinib) and the downstream signaling transduction. Ponatinib inhibited the growth of D816V KIT-expressing cells in culture and nude mouse xenografted tumor. Ponatinib triggered apoptosis by inducing the release of cytochrome c and AIF, downregulation of Mcl-1. Furthermore, ponatinib abrogated the phosphorylation of β-catenin at the site Y654, suppressed the translocation of β-catenin, and inhibited the transcription and DNA binding of TCF and the expression of its targets (e.g., AXIN2, c-MYC, and CCND1). Moreover, ponatinib was highly active against xenografted D816V KIT tumors in nude mice and significantly prolonged the survival of mice with aggressive systemic mastocytosis or mast cell leukemia by impeding the expansion and infiltration of mast cells with imatinib-resistant D814Y KIT. Our findings warrant a clinical trial of ponatinib in patients with systemic mastocytosis harboring D816V KIT.

Topics: Animals; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; beta Catenin; Cell Line; Cell Proliferation; Cytochromes c; Disease Models, Animal; Drug Resistance; Gene Expression Regulation; Gene Silencing; Humans; Imidazoles; Male; Mast Cells; Mastocytosis; Mice; Myeloid Cell Leukemia Sequence 1 Protein; Phosphorylation; Point Mutation; Protein Binding; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-kit; Pyridazines; Signal Transduction; TCF Transcription Factors; Wnt Signaling Pathway; Xenograft Model Antitumor Assays

As diverse pruritic cutaneous diseases respond to ultraviolet treatment, we have examined whether ultraviolet light is capable of inducing apoptosis in mast cells. Human mast cell line 1 (HMC1) derived from a patient with malignant mastocytosis and purified skin mast cells were irradiated with single doses of ultraviolet B or ultraviolet A1, or pretreated with 8-methoxypsoralen prior to ultraviolet A1 exposure. After 0 to 48 h of incubation, the percentage of apoptotic and dead cells was assessed. In HMC1 cells, morphologic features of apoptosis were further evaluated by electron microscopy. All ultraviolet treatment induced apoptosis of HMC1 cells in a time- and dose-dependent manner. Apoptosis was associated with activation of caspase-3, release of cytochrome C, cleavage of poly(ADP-ribose)-polymerase, and nuclear accumulation of p53. In contrast, resting skin mast cells were resistant to ultraviolet light induced apoptosis. After incubation with stem cell factor and interleukin-4 for 2 wk, however, slowly proliferating skin mast cells also underwent apoptosis in response to ultraviolet light. In conclusion, these data demonstrate that ultraviolet light directly affects mast cells, but mainly aims at the proliferating mast cells as found in mastocytosis and mast cell dependent pruritic diseases, where increased numbers are observed due to the recruitment mast cell precursors from the blood.

Topics: Apoptosis; Caspase 3; Caspases; Cell Differentiation; Cell Division; Cell Line, Tumor; Cytochromes c; Humans; Leukemia; Mast Cells; Mastocytosis; Poly(ADP-ribose) Polymerases; Skin; Tumor Suppressor Protein p53; Ultraviolet Rays