Oncogenic alterations of metabolism associated with resistance to chemotherapy.

Metabolic reprogramming in cancer cells is a strategy to meet high proliferation rates, invasion, and metastasis. Also, several researchers indicated that the cellular metabolism changed during the resistance to chemotherapy. Since glycolytic enzymes play a prominent role in these alterations, the ability to reduce resistance to chemotherapy drugs is promising for cancer patients. Oscillating gene expression of these enzymes was involved in the proliferation, invasion, and metastasis of cancer cells. This review discussed the roles of some glycolytic enzymes associated with cancer progression and resistance to chemotherapy in the various cancer types.

Nanostructured multifunctional stimuli-responsive glycopolypeptide-based copolymers for biomedical applications.

Inspired by natural resources, such as peptides and carbohydrates, glycopolypeptide biopolymer has recently emerged as a new form of biopolymer being recruited in various biomedical applications. Glycopolypeptides with well-defined secondary structures and pendant glycosides on the polypeptide backbone have sparked lots of research interest and they have an innate ability to self-assemble in diverse structures. The nanostructures of glycopolypeptides have also opened up new perspectives in biomedical applications due to their stable three-dimensional structures, high drug loading efficiency, excellent biocompatibility, and biodegradability. Although the development of glycopolypeptide-based nanocarriers is well-studied, their clinical translation is still limited. The present review highlights the preparation and characterization strategies related to glycopolypeptides-based copolymers, followed by a comprehensive discussion on their biomedical applications with a specific focus on drug delivery by various stimuli-responsive (e.g., pH, redox, conduction, and sugar) nanostructures, as well as their beneficial usage in diagnosis and regenerative medicine.

Design, construction, and expression of recombinant human interferon beta gene in CHO-s cell line using EBV-based expression system.

BACKGROUND AND PURPOSE: Codon optimization has been considered as a powerful strategy to increase the expression level of protein therapeutics in mammalian cells. As an empirical approach to study the effects of the codon usage and GC content on heterologous gene expression in suspension adapted Chinese hamster ovary (CHO-s) cells, we redesigned the recombinant human interferon beta (rhIFN- ß) gene based on the codon preference of the CHO cell in a way to increase the GC content in the third position of each codon. EXPERIMENTAL APPROACH: The nucleotide sequence of the codon-optimized rhIFN-ß was synthesized in parallel with the wild-type and expressed transiently in CHO-s cells using Epstein-Bar virus (EBV)-based expression system. The protein expression of the rhIFN-ß by codon-optimized and wild-type genes were quantified using ELISA test. FINDINGS / RESULTS: The results indicated a 2.8-fold increase in the expression level of the biologically active form of the rhIFN-ß by codon-optimized sequence. CONCLUSION AND IMPLICATIONS: These results shed light on the capability of codon optimization to create a stable CHO cell for scaling up the production of recombinant therapeutics such as rhIFN-ß.

Phototherapy alters the oncogenic metabolic activity of breast cancer cells.

BACKGROUND: Metabolic reprogramming in cancer cells is a strategy to attain a high proliferation rate, invasion, and metastasis. In this study, the effects of phototherapy at different wavelengths were investigated on the metabolic activity of breast cancer cells. METHODS: The states of the MCF7 cells proliferation and viability were measured by the MTT assay. Glucose consumption and the lactate formation in the LED-irradiated cells culture were analyzed by biochemical assay kits. The Amino acid concentration in the culture media of the MCF7 cells was analyzed using HPLC. Moreover, the gene expression of some glycolytic, TCA cycle and pentose phosphate cycleenzymes were assessed by real time PCR. RESULTS: Phototherapy at wavelength of 435 nm decreased the cell viability by 23 % when the energy dose was 17.5 J/cm2 compared to the control group. The expression of the LDHA and GLS was up-regulated in 629 nm-treated cells while the expression of these genes was down-regulated in the MCF7 cells irradiated at 435 nm in comparison with the control group. Consequently, the glucose consumption and the lactate formation were diminished respectively by 22 % and 15 % in the 435 nm-irradiated cells while the glucose consumption and the lactate formation were increased in the 629 nm-irradiated cells by 112 % and 107 % in comparison with the control group. In addition, the analysis of the glutamine concentration by the HPLC indicated that the blue light irradiation decreased the glutamine consumption while the red light increased it in comparison with the control group.

Synergistic effect of phototherapy and chemotherapy on bladder cancer cells.

Drug resistance as an important barrier to cancer treatment, has a close relation with alteration of cancer metabolism. Therefore, in this study the synergistic effect of phototherapy and chemotherapy were investigated on the bladder cancer cells viability. The cytotoxicity effect of blue light irradiation was measured by the MTT assay. Glucose consumption, lactate and ammonium formation were analyzed in the blue LED-irradiated cancer cells culture. Also, the expression of some genes involved in apoptosis and epithelial-mesenchymal transition was assessed using real-time PCR in comparison with the control group. The analysis of the results indicated that blue light irradiation inhibited the cell viability in a dose-dependent manner. Blue light irradiation decreased the cell viability by 7% and 19% (p < .05) in 5637 cells at doses of 8.7 J/cm2 and 17.5 J/cm2 in comparison with the control group respectively. Glucose consumption, lactate and ammonium formation diminished in the blue LED-irradiated 5637 cells in both doses. The real time PCR results indicated that the expression of Bax increased in blue light-irradiated cells. In addition, the cell cycle analysis showed that blue light irradiation arrested the bladder cancer in the G1 phase. Also, the effect of combination therapy on cancer cells was investigated in presence of blue light irradiation and cisplatin. The obtained results of the MTT assay indicated that blue light irradiation enhance the cytotoxicity effect of cisplatin on bladder cancer cells.

Improving the Growth Rate of Human Adipose-Derived Mesenchymal Stem Cells in Alginate/Gelatin Versus Alginate Hydrogels.

BACKGROUND: Expansion and differentiation of stem cells relies on the soluble materials as well as the physical conditions of their microenvironment. Several methods have been studied in attempt to enhance the growth and differentiation rates of different adult stem cells extracted from different sources. OBJECTIVES: The purpose was to improve the three-dimensional (3D) culture condition of the semi-permeable polymeric beads for encapsulation of the human adipose-derived mesenchymal stem cells (hADSCs) by modifying the ratio of the alginate-gelatin composition. MATERIALS AND METHODS: Following isolation and characterization of hADSCs by flow cytometry and their functional differentiation, encapsulation in the alginate and alginate/gelatin compositions were performed. Moreover, the stability, swelling, size frequency, growth kinetics, and cytotoxicity of the beads were measured to meet proper condition in the designed experimental and control culture conditions. Finally, the growth rates of the cells in different experimental groups and control were measured and analyzed statistically. RESULTS: Viability decreased in 2 and 3 percent alginate once compared to 1% alginate in beads (p≤0.05). Moreover swelling of the beads in the alginate/gelatin compositions (50:50 and 70:30) were higher than the pure alginate beads (p≤0.05). Finally, the cell growth rate in alginate/gelatin (50:50) beads was significantly higher than alginate and alginate/gelatin (70:30) beads (p≤0.05). CONCLUSIONS: These findings suggested for the first time that the composite of alginate/gelatin beads with the ratio of 50:50 might provide a suitable culture condition for the encapsulation and in vitro expansion of the hADSCs.

A novel in vitro model for cancer stem cell culture using ectopically expressed piwil2 stable cell line.

OBJECTIVE: Piwil2, a member of Ago/Piwi gene family containing Piwi and PAZ domains, has been shown to be ectopically expressed in different cancer cells, especially its remarkable expression in cancer stem cells (CSCs), and is also known to be essential for germ line stem cell self-renewal in various organisms. The hypothesis that CSC may hold the key to the central problem of clinical oncology and tumor relapse leads to more anticancer treatment studies. Due to emerging controversies and extreme difficulties in studying of CSC, like the cells using in vivo models, more attempts have expended to establish different in vitro models. However, the progress was slow owing to the problems associated with establishing proper CSC cultures in vitro. To overcome these difficulties, we prompted to establish a novel stable cell line over-expressing Piwil2 to develop a potential proper in vitro CSC model. MATERIALS AND METHODS: In this experimental study, mouse embryonic fibroblasts (MEFs) were isolated and electroporated with a construct containing Piwil2 cDNA under the control of the cytomegalovirus promoter (CMV). Stable transfectants were selected, and the established MEF-Piwil2 cell line was characterized and designated as CSC-like cells using molecular markers. Functional assays, including proliferation, migration, and invasion assays were performed using characterized CSC like cells in serum-free medium. Additionally, MEF-Piwil2 cell density and viability were measured by direct and indirect methods in normoxic and hypoxic conditions. RESULTS: The results of reverse transcriptase-polymerase chain reaction (RT-PCR), western blot, and immunocytochemistry revealed an overexpression for Piwil2 in the transfected Piwil2 cells both in the RNA and protein levels. Furthermore, analysis of the kinetic and stoichiometric parameters demonstrated that the specific growth rate and the yield of lactate per glucose were significantly higher in the MEF-Piwil2 group compared to the MEF cells (ANOVA, p< 0.05). Also, analysis of functional assays including migration and invasion assays demonstrated a significantly higher number of migrated and invaded cells in the MEF-Piwil2 compared to that of the MEF cells (ANOVA, p< 0.05). The MEF-Piwil2 cells tolerated hypoxia mimetic conditions (CoCl2 ) with more than 95% viability. CONCLUSION: According to the molecular and functional studies, it has been realized that Piwil2 plays a key role(s) in tumor initiation, progression and metastasis. Therefore, Piwil2 can be used not only as a common biomarker for tumor, but also as a target for the development of new anticancer drug. Finally, the main outcome of our study was the establishment of a novel CSC-like in vitro model which is expected to be utilized in understanding the complex roles played by CSC in tumor maintenance, metastasis, therapy resistance or cancer relapse.