A New Recombinant Fibronectin/Cadherin Protein-Hydrophobically Modified Glycol Chitosan/Paclitaxel Layer-By-Layer Self-Assembly Strategy for the Postoperative Therapy of Osteosarcoma and Correlated Bone Injury.

Osteosarcoma is one of the most aggressive cancers which greatly threatens the health of adolescents and surgery is difficult to resect the whole piece of tumor tissue. The residual tumor cells might proliferate at the tumor site and invade into the blood circulation, leading to tumor recurrence and metastasis. Besides, the invasion of tumor cells could also lead to bone injury. We designed a recombinant fibronectin-cadherin fusion protein/hydrophobically modified glycol chitosan-PTX nanoparticles (rFN-CDH/HGC-PTX) layer-by-layer self-assembly polymer based on biphasic calcium phosphate ceramic (BCP) (BCP-PEI-(rFN/CDH-PTX/HGC)n-rFN/CDH). The SEM, FTIR, XPS and contact angle experiments proved the successful synthesis of the polymer. The chemotherapy drug PTX and bone-repairing-related rFN/CDH fusion protein could be stably released within one week and the in vitro experiments exhibited the efficacy of the polymer to kill residual tumor cells and promote the proliferation of osteoblast, confirming that our polymer was a superior material for postoperative osteosarcoma therapy.

Synthesis and Biological Evaluation of Spirocyclic Chromane Derivatives as a Potential Treatment of Prostate Cancer.

As a significant co-activator involved in cell cycle and cell growth, differentiation and development, p300/CBP has shown extraordinary potential target in cancer therapy. Herein we designed new compounds from the lead compound A-485 based on molecular dynamic simulations. A series of new spirocyclic chroman derivatives was prepared, characterized and proven to be a potential treatment of prostate cancer. The most potent compound B16 inhibited the proliferation of enzalutamide-resistant 22Rv1 cells with an IC50 value of 96 nM. Furthermore, compounds B16-P2 displayed favorable overall pharmacokinetic profiles, and better tumor growth inhibition than A-485 in an in vivo xenograft model.

Smad4 deficiency substitutes Cdkn2b but not Cdkn2a downregulation in pancreatic cancer following induction of genetic events in adult mice.

BACKGROUND: Minor progress in pancreatic cancer treatment and prognosis implies that more reliable animal models are urgently needed to decipher its molecular mechanisms and preclinical research. We recently reported a genetically engineered adult mouse model where Cdkn2b downregulation was required together with Cdkn2a downregulation to inactivate the Rb pathway. Besides, the role of Smad4, which is mutated more frequently than Cdkn2b in human pancreatic cancer, was determined critical on the development of the pancreas tumor by some reports. However, the impact of Smad4 deficiency in combination with PDAC-relevant mutations, such as Cdkn2a when induced in adult pancreas has not been completely elucidated in mice. METHODS: Lentiviral delivered oncogene/tumor suppressors in adult pancreas. The development of pancreatic cancer was monitored. Hematoxylin and eosin staining and immunofluorescence were performed for pathological identification of the pancreatic cancer. Real-time polymerase chain reaction, immunofluorescence and western blot were used to test gene expression. RESULTS: Loss of Smad4 could cooperate with alterations of KRAS, Trp53, and Cdkn2a to induce pancreatic cancer in adult mice. The role of Smad4 was mainly in downregulating the expression of Cdkn2b and further inducing phosphorylation of the Rb1 protein. CONCLUSIONS: These findings show an essential role of Smad4 deficiency in pancreatic ductal adenocarcinoma (PDAC) formation. This model better recapitulates the adult onset, clonal origin, and genetic alterations in human PDAC and can be simply generated on a large-scale.

Biogenic silver/silver chloride nanoparticles inhibit human glioblastoma stem cells growth in vitro and Ehrlich ascites carcinoma cell growth in vivo.

The importance of biogenic silver/silver chloride nanoparticles has become increasing day by day. In the present study, silver/silver chloride nanoparticles (Ag/AgCl-NPs) were synthesized from Kaempferia rotunda tuberous rhizome extract to evaluate the antiproliferative activity against human glioblastoma stem cells (GSCs) in vitro and Ehrlich ascites carcinoma (EAC) cells in vivo in mice. Synthesis of nanoparticles was confirmed by colour change and UV-visible spectrum and characterized by TEM, XRD, TGA, AFM and FTIR. K rotunda and recently synthesized Zizyphus mauritiana fruit extract-mediated Ag/AgCl-NPs inhibited 77.2% and 71% of GSCs growth at 32 µg/mL concentration with the IC50 values of 6.8 and 10.4 µg/mL, respectively. Cell morphological studies and caspase-3 immunofluorescence assay revealed that both biogenic nanoparticles induced apoptosis in GSCs. Expression levels of several genes were checked by real-time PCR after treatment with K rotunda tuberous rhizome-mediated Ag/AgCl-NPs. PARP, EGFR, NOTCH2 and STAT3 gene expression were decreased with the increase of NFκB, TLR9, IL1, TNFα, IKK and p21 gene that would be the cause of induction of apoptosis in GSCs. The cell cycle arrest at G2 /M phase was confirmed by flow cytometric assay. Both nanoparticles were injected intraperitoneally to rapidly growing EAC cells for 5 consecutive days. Approximately, 32.3% and 55% EAC cells growth were inhibited by K rotunda tuberous rhizome-mediated Ag/AgCl-NPs at 6 and 12 mg/kg/day doses, respectively while only 20% cell growth inhibition was monitored at 12 mg/kg/day dose of Z mauritiana-mediated Ag/AgCl-NPs. From the above results, it can be concluded that presently synthesized nanoparticles would be a potent anticancer agent.

Targeting ß2 subunit of Na+/K+-ATPase induces glioblastoma cell apoptosis through elevation of intracellular Ca2.

Glioblastoma (GBM) is the most frequent brain cancer with poor prognosis and few therapies and urgently requires effective treatments. Na+/K+-ATPase is considered as a target for GBM therapy and development of anticancer drugs. Cardiac glycosides bind the Na+/K+-ATPase α subunit to inhibit enzymatic activity and are promising candidates for anticancer drug development including GBM. However, the comparatively higher doses required for effective anticancer actions cause severe cardiotoxicity. Selectively targeting the ATPase Na+/K+ transporting subunit beta 2 (ATP1B2) that is not expressed in the heart might avoid the cardiotoxicity. However, the effect of targeting ATP1B2 in GBM remains unknown. In this study, we found that high ATP1B2 expression is significantly associated with poor prognosis of patients with GBM. ATP1B2 silencing in GBM cells resulted in remarkably cell cycle arrest at the G2/M phase and apoptosis with concomitant increase in intracellular Ca2+ and activation of p38 kinase, similar to Na+/K+-ATPase inhibition by the classic cardiac glycoside digoxin. ATP1B2 is expressed higher in glioblastoma stem-like cells (GSCs) than in GBM cells and its downregulation induces apoptosis of GSCs. Furthermore, inducible ATP1B2 knockdown significantly inhibit tumor growth in vivo. Our data suggest ATP1B2 has potential as a therapeutic target for GBM.

A novel pancreatic cancer model originated from transformation of acinar cells in adult tree shrew, a primate-like animal.

Pancreatic cancer is one of the most lethal common cancers. The cell of origin of pancreatic ductal adenocarcinoma (PDAC) has been controversial, and recent evidence suggested acinar cells as the most probable candidate. However, the genetic alterations driving the transformation of pancreatic acinar cells in fully mature animals remain to be deciphered. In this study, lentivirus was used as a tool to introduce genetic engineering in tree shrew pancreatic acinar cells to explore the driver mutation essential for malignant transformation, establishing a novel tree shrew PDAC model, because we found that lentivirus could selectively infect acinar cells in tree shrew pancreas. Combination of oncogenic KRASG12D expression and inactivation of tumor suppressor genes Tp53, Cdkn2a and Cdkn2b could induce pancreatic cancer with full penetrance. Silencing of Cdkn2b is indispensable for Rb1 phosphorylation and tumor induction. Tree shrew PDAC possesses the main histological and molecular features of human PDAC. The gene expression profile of tree shrew PDAC was more similar to human disease than a mouse model. In conclusion, we established a novel pancreatic cancer model in tree shrew and identified driver mutations indispensable for PDAC induction from acinar cells in mature adults, demonstrating the essential roles of Cdkn2b in the induction of PDAC originating from adult acinar cells. Tree shrew could thus provide a better choice than mouse for a PDAC model derived from acinar cells in fully mature animals.

HUWE1 controls the development of non-small cell lung cancer through down-regulation of p53.

Lung cancer is the most frequent cancer type and the leading cause of tumor-associated deaths worldwide. TP53 is an important tumor suppressor gene and is frequently inactivated in lung cancer. E3 ligases targeting p53, such as MDM2, are involved in the development of lung cancer. The E3 ligase HUWE1, which targets many tumor-associated proteins including p53, has been reported to be highly expressed in lung cancer; however, its role in lung tumorigenesis is unclear. Methods: The expression of HUWE1 and p53 in lung cancer cells was modulated and the phenotypes were assessed by performing soft agar colony forming assays, cell cycle analysis, BrdU incorporation assays, and xenograft tumor growth assays. The effect on tumorigenesis in genetically-engineered mice was also analyzed. The mechanism through which HUWE1 sustained lung cancer cell malignancy was confirmed by western blotting. HUWE1 expression in clinical lung cancer was identified by immunohistochemistry and validated by analyzing lung adenocarcinoma and lung squamous carcinoma samples from the Cancer Genome Atlas (TCGA) database. Finally, we assessed the association between HUWE1 expression and patient outcome using online survival analysis software including survival information from the caBIG, GEO, and TCGA database. Results: Inactivation of HUWE1 in a human lung cancer cell line inhibited proliferation, colony-forming capacity, and tumorigenicity. Mechanistically, this phenotype was driven by increased p53, which was due to attenuated proteasomal degradation by HUWE1. Up-regulation of p53 inhibited cancer cell malignancy, mainly through the induction of p21 expression and the down-regulation of HIF1α. Huwe1 deletion completely abolished the development of EGFRVIII-induced lung cancer in Huwe1 conditional knockout mice. Furthermore, survival analysis of lung cancer patients showed that increased HUWE1 expression is significantly associated with worse prognosis. Conclusion: Our data suggest that HUWE1 plays a critical role in lung cancer and that the HUWE1-p53 axis might be a potential target for lung cancer therapy.

Aptamer TY04 inhibits the growth of multiple myeloma cells via cell cycle arrest.

The aptamer TY04 is a single-stranded DNA. However, its biological function has not been elucidated. Here, we found that TY04 specifically bound to multiple myeloma cells MM.1S, and some membrane proteins on the surface of MM.1S cells constituted the target molecules of TY04. TY04 inhibited the growth of multiple myeloma cell lines, induced cell cycle arrest in mitosis, and resulted in a significant accumulation of binucleated cells. Following TY04 treatment, a concomitant increase in CDK1 and cyclin B1 expression occurred. In addition, TY04 treatment also resulted in a significant downregulation of γ-tubulin. Considering the unique advantages of aptamers, TY04 shows great potential as a drug candidate to treat multiple myeloma.

Lycorine induces cell-cycle arrest in the G0/G1 phase in K562 cells via HDAC inhibition.

BACKGROUND: Lycorine, a natural alkaloid extracted from Amaryllidaceae, has shown various pharmacological effects. Recent studies have focused on the potential antitumor activity of lycorine. In our previous study, we found that lycorine decrease the cell viability of leukemia HL-60 cells and multiple myeloma KM3 cells and induces cell apoptosis. However, the effect and molecular mechanism of lycorine on human chronic myelocytic leukemia cells has yet to be determined. METHODS: Human chronic myelocytic leukemia cells K562 were treated with lycorine. Cell viability was monitored using the method of CCK-8. The histone deacetylase (HDAC) enzymatic activity was detected by HDAC colorimetric assay, and the cell cycle was analyzed by flow cytometry. The expression of cell-cycle related proteins were identified using Western blot. RESULTS: In the present study, we further revealed that lycorine can inhibit the proliferation of K562 cells. Analysis of HDAC activity showed that lycroine decreases HDAC enzymatic activities in K562 cells in a dose-dependent manner. Inhibition of HDAC activity has been associated with cell-cycle arrest and growth inhibition. We evaluated the cell cycle distribution after lycorine treatment and found that lycorine causes cell-cycle arrest in the G0/G1 phase. To investigate the mechanism behind this cell cycle arrest, G1-related proteins were assayed by Western blot. After lycorine treatment, cyclin D1 and cyclin-dependent kinase 4 expressions were inhibited and retinoblastoma protein phosphorylation was reduced. Lycorine treatment also significantly upregulated the expression of p53 and its target gene product, p21. CONCLUSIONS: These results suggest that inhibition of HDAC activity is responsible for at least part of the induction of cell-cycle arrest in the G0/G1 phase by lycorine and provide a mechanistic framework for further exploring the use of lycorine as a novel antitumor agent.

Mechanism and kinetics of apatite formation on nanocrystalline TiO2 coatings: a quartz crystal microbalance study.

Apatite (Ca5(PO4)3OH) has long been considered as an excellent biomaterial to promote bone repairs and implant. Apatite formation induced by negatively charged nanocrystalline TiO2 coatings soaked in simulated body fluid (SBF) was investigated using in situ quartz crystal microbalance (QCM), scanning electron microscopy (SEM), Fourier-transformed infrared spectroscopy (FTIR), X-ray diffraction (XRD) and energy-dispersive X-ray spectroscopy (EDX) techniques, and factors affecting its formation such as pH, size of TiO2 particles and thickness of TiO2 coatings, were discussed in detail. Two different stages were clearly observed in the process of apatite precipitation, indicating two different kinetic processes. At the first stage, the calcium ions in SBF were initially attracted to the negatively charged TiO2 surface, and then the calcium titanate formed at the interface combined with phosphate ions, consequently forming apatite nuclei. After the nucleation, the calcium ions, phosphate ions and other minor ions (i.e. CO3(2-) and Mg2+) in supersaturated SBF deposited spontaneously on the original apatite coatings to form apatite precipitates. In terms of the in situ frequency shifts, the growth-rate constants of apatite (K1 and K2) were estimated, respectively, at two different stages, and the results were (1.96+/-0.14)x10(-3)s(-1) and (1.28+/-0.10)x10(-4)s(-1), respectively, in 1.5 SBF solution. It was found that the reaction rate at the first stage is obviously higher than that at the second stage.

Piezoelectric urea biosensor based on immobilization of urease onto nanoporous alumina membranes.

The urease was immobilized onto nanoporous alumina membranes prepared by the two-step anodization method, and a novel piezoelectric urea sensing system with separated porous alumina/urease electrode has been developed through measuring the conductivity change of immobilized urease/urea reaction. The process of urease immobilization was optimized and the performance of the developed urea biosensor was evaluated. The obtained urea biosensor presented high-selectivity monitoring of urea, better reproducibility (S.D.=0.02, n=6), shorter response time (30s), wider linear range (0.5 microM to 3mM), lower detection limit (0.2 microM) and good long-term storage stability (with about 76% of the enzymatic activity retained after 30 days). The clinical analysis of the urea biosensor confirmed the feasibility of urea detection in urine samples.