Inhibition of tumor cell growth by adenine is mediated by apoptosis induction and cell cycle S phase arrest.

Gekko swinhonis has a long standing history in Chinese traditional medicine recognized for its application in treating patients with terminal cancer.In order to discover novel anticancer drugs with high anti-tumor efficacy and low toxicity to normal cells, we aim to investigate the anti-tumor components from Gekko swinhonis. Four nucleosides from the extracted samples were enriched, namely adenosine, guanosine, thymidine and inosine. We evaluated the antitumor effect of the four nucleosides and found that adenosine possessed the strongest anti-tumor effect. Besides, adenine could inhibit the growth of Bel-7402 and Hela cells in a dose and time dependent manner, but not normal human cervical keratinocytes. Bel-7402 and Hela cells had undergone apoptosis 48 hours after treatment as evidenced by morphologic changes under TEM, while adenine blocked cell cycle of tumor cells at S phase and subsequently causing cell cycle exit and promoting apoptosis. Moreover, the pharmacokinetics of adenine was stable in cell culture medium for up to 72 hours. Combining its potency with stability, we conclude adenine makes a promising candidate for an anti-tumor drug.

Identification and functional analysis of acute myeloid leukemia susceptibility associated single nucleotide polymorphisms at non-protein coding regions of RUNX1.

Little is known about the susceptibility to acute myeloid leukemia. We aim to search non-protein coding regions of key hematopoiesis transcription factors for genetic variations associated with acute myeloid leukemia susceptibility. We genotyped SNPs of RUNX1 P1 promoter, P2 promoter, +23 enhancer, intron 5.2 enhancer, PU.1 promoter, CEBPA promoter, and CEBPE promoter from acute myeloid leukemia patients and healthy controls. Rs2249650 and rs2268276 at RUNX1 intron 5.2 enhancer were found to be associated with acute myeloid leukemia susceptibility. Artificial reporters containing different rs2249650 and rs2268276 alleles showed differential activities in the K562 cell line, a human immortalized myeloid leukemia line. Rs2249650 contributes to reporter activities more than rs2268276. Gel shift assay is consistent with the luciferase assay. Supershift assay indicated that one potential binding protein was PU.1. To sum up, rs2268276 and especially rs2249650 may be qualified as new acute myeloid leukemia susceptibility-associated SNPs.

Proliferative and survival effects of PUMA promote angiogenesis.

The p53 upregulated modulator of apoptosis (PUMA) is known as an essential apoptosis inducer. Here, we report the seemingly paradoxical finding that PUMA is a proangiogenic factor critically required for the proliferation and survival of vascular and microglia cells. Strikingly, Puma deficiency by genetic deletion or small hairpin RNA knockdown inhibited developmental and pathological angiogenesis and reduced microglia numbers in vivo, whereas Puma gene delivery increased angiogenesis and cell survival. Mechanistically, we revealed that PUMA plays a critical role in regulating autophagy by modulating Erk activation and intracellular calcium level. Our findings revealed an unexpected function of PUMA in promoting angiogenesis and warrant more careful investigations into the therapeutic potential of PUMA in treating cancer and degenerative diseases.

PDGF-CC blockade inhibits pathological angiogenesis by acting on multiple cellular and molecular targets.

The importance of identifying VEGF-independent pathways in pathological angiogenesis is increasingly recognized as a result of the emerging drug resistance to anti-VEGF therapies. PDGF-CC is the third member of the PDGF family discovered after more than two decades of studies on PDGF-AA and PDGF-BB. The biological function of PDGF-CC and the underlying cellular and molecular mechanisms remain largely unexplored. Here, using different animal models, we report that PDGF-CC inhibition by neutralizing antibody, shRNA, or genetic deletion suppressed both choroidal and retinal neovascularization. Importantly, we revealed that PDGF-CC targeting acted not only on multiple cell types important for pathological angiogenesis, such as vascular mural and endothelial cells, macrophages, choroidal fibroblasts and retinal pigment epithelial cells, but also on the expression of other important angiogenic genes, such as PDGF-BB and PDGF receptors. At a molecular level, we found that PDGF-CC regulated glycogen synthase kinase (GSK)-3beta phosphorylation and expression both in vitro and in vivo. Activation of GSK3beta impaired PDGF-CC-induced angiogenesis, and inhibition of GSK3beta abolished the antiangiogenic effect of PDGF-CC blockade. Thus, we identified PDGF-CC as an important candidate target gene for antiangiogenic therapy, and PDGF-CC inhibition may be of therapeutic value in treating neovascular diseases.

Survival effect of PDGF-CC rescues neurons from apoptosis in both brain and retina by regulating GSK3beta phosphorylation.

Platelet-derived growth factor CC (PDGF-CC) is the third member of the PDGF family discovered after more than two decades of studies on the original members of the family, PDGF-AA and PDGF-BB. The biological function of PDGF-CC remains largely to be explored. We report a novel finding that PDGF-CC is a potent neuroprotective factor that acts by modulating glycogen synthase kinase 3beta (GSK3beta) activity. In several different animal models of neuronal injury, such as axotomy-induced neuronal death, neurotoxin-induced neuronal injury, 6-hydroxydopamine-induced Parkinson's dopaminergic neuronal death, and ischemia-induced stroke, PDGF-CC protein or gene delivery protected different types of neurons from apoptosis in both the retina and brain. On the other hand, loss-of-function assays using PDGF-C null mice, neutralizing antibody, or short hairpin RNA showed that PDGF-CC deficiency/inhibition exacerbated neuronal death in different neuronal tissues in vivo. Mechanistically, we revealed that the neuroprotective effect of PDGF-CC was achieved by regulating GSK3beta phosphorylation and expression. Our data demonstrate that PDGF-CC is critically required for neuronal survival and may potentially be used to treat neurodegenerative diseases. Inhibition of the PDGF-CC-PDGF receptor pathway for different clinical purposes should be conducted with caution to preserve normal neuronal functions.