Programmed Necrosis Inducers for Cancer Treatment / 中国医学科学院学报

Programmed necrosis,a mode of cell death independent of Caspase,is mainly mediated by receptor-interacting protein kinase-1 (RIPK1),receptor-interacting protein kinase-3 (RIPK3),and mixed lineage kinase domain-like protein (MLKL).Studies have demonstrated that programmed necrosis has the dual role of promoting and inhibiting tumor growth and thus we can control the development of tumor by regulating programmed necrosis.The drugs capable of inducing programmed necrosis show potential anti-tumor activity.In addition,inducing programmed necrosis is an effective way to overcome tumor resistance to apoptosis.This paper summarized the mechanisms of programmed necrosis and its relationship with tumors.We focused on the antitumor activity of programmed necrosis inducers including natural products,chemotherapeutic drugs,death receptor ligands,kinase inhibitors,inorganic salts,metal complexes,and metal nanoparticles.These agents will provide new therapeutic candidates for the treatment of tumors,especially the tumors acquiring resistance to apoptosis.

Synthesis and anti-angiogenic activity of a novel microtubule inhibitor IMB5046 / 药学学报

IMB5046 is a newly discovered nitrobenzoate functioning as a microtubule inhibitor. Here we report its synthesis and in vitro anti-angiogenic activity. IMB5046 was synthesized by conjugation of 2-morpholin-4-yl-5-nitrobenzoic acid with 4-(methylthio)benzyl alcohol via two-step reactions. The structure of the end product was verified using 1H NMR and HR-MS spectroscopy. The effect of these compounds on cell proliferation was determined using MTT assay, and their impact on cytoskeleton was investigated using fluorescence assay. Flow cytometry was performed to examine the effect of IMB5046 on cell cycle. Cell wound scratch assay and Transwell assay were performed to examine cell migration. Endothelial tube formation assay was used to evaluate the anti-angiogenic activity of IMB5046. The results indicated that IMB5046 induced endothelial cell contraction and microtubule depolymerization, and inhibited the proliferation of endothelial cells and tumor cells, while two raw materials showed no obvious effects. IMB5046 arrested cell cycle at G2/M phase, even at low-cytotoxic concentrations it significantly inhibited the motility of endothelial cells. IMB5046 inhibited the tube formation of endothelial cells according to the number of tubes and junctions. In conclusion, IMB5046 is a promising microtubule-targeting drug with anti-angiogenic activity.

Isolation and culture of mouse spermatogonial stem cells and determination of the related markers / 中国医学科学院学报

<p><b>OBJECTIVE</b>To establish a simple and highly effective isolation and culture system of mouse spermatogonial stem cells(SSCs)and detect the expression of stem cell-related markers in the isolated cells.</p><p><b>METHODS</b>The structures of seminiferous tubules of neonatal(6-8 days of age)and adult(26-28 weeks)DBA/2 mice were compared using histochemical examination. Testes of neonatal mice were selected for preparing primary cells. The digestive efficiency of different enzymes was compared. SSCs were isolated according to the different binding abilities of testicle somatic cells and SSCs to gelatin matrix. The effects of different base culture media such as StemPro34 and α-MEM,gelatin,and serum on the SSCs binding activity and growth were studied. The cell morphology was observed during the culture process. Immunofluorescence was used to detect the expression of SSCs and cancer stem cells(CSCs)-related markers in SSCs.</p><p><b>RESULTS</b>The content of SSCs in the testes of neonatal mice was relatively higher than that in adult mice. Trypsin showed the highest digestive efficiency. In StemPro34 supplemented with 1% fetal bovine serum and on the gelatin matrix,testicular somatic cells could bind with the plate efficiently. Spermatogonial cells grew well when using mitomycin C-treated testicular somatic cells as feeder cells and showed typical characteristic of SSCs. After 13 days of culture,spermatogonial cells formed cell clusters. Immunofluorescence assay showed that SSCs markers glial cell line-derived neurotrophic factor(GDNF)family receptor α1(GFRα1)and VASA protein were highly expressed in the cell clusters. CSCs marker CD44 was expressed in the As,Apr,Aal and the inner cells of the cell clusters,while seldom expressed in the somatic cells.</p><p><b>CONCLUSIONS</b>An isolation and culture system of SSCs derived from DBA/2 mice was established. CD44 is highly expressed in the early stage of spermatogonial cell development.</p>

Inhibition of tumor cell invasion and induction of apoptosis by ubenimex / 药学学报

This study is to investigate the effects of ubenimex on tumor cell invasion and apoptosis, dose relationship and mechanism. Immunofluorescence staining was performed to detect the expression of CD13 in HT-1080 cells. MTT assay was used to analyze the effect of ubenimex on cell proliferation. Annexin V-EGFP/PI was used to detect apoptotic cells by flow cytometry. Cell cycle was analyzed using flow cytometry. Ala-pNA was used as substrate to evaluate the effect of ubenimex on the aminopeptidase activity. Transwell assay was used to analyze the effect of ubenimex on cell invasion and migration ability. Western blotting was used to detect the expression level of CD13. MMP activity was analyzed using gelatin zymography. The results showed that ubenimex at high concentration inhibited the proliferation of HT-1080 cells (IC50: 3.8 mg x mL(-1)), and induced cell apoptosis. Cell cycle was blocked at G1 phase. Ubenimex at low concentration inhibited the aminopeptidase activity of HT-1080 cells (IC50: 8.3 microg x mL(-1)) and inhibited cell invasion, but it had no effects on the cell migration and proliferation. Ubenimex had no effects on CD13 expression and MMP activity. In conclusion, ubenimex at low concentration can inhibit the invasion ability of tumor cells by directly inhibiting the aminopeptidase activity; ubenimex at high concentration can inhibit the proliferation of tumor cells and induce cell apoptosis by a CD13-independent pathway.