Cell Journal (Yakhteh)
Volume:19 Issue: 1, Spring 2017

Cancer cells have recently been shown to activate hundreds of normally silent tissue-restricted genes, including a specific subset associated with cancer progression and poor prognosis. Within these genes, a class of testis-specific genes designed as cancer/testis, attracted special attention because of their oncogenic roles as well as their potential use in immunotherapy. Here we focus on one of these genes encoding the testis-specific member of the bromodomain and extra-terminal (BET) family, known as BRDT. Aberrant activation of BRDT was first detected in lung cancers. In this study, we report that the frequency of BRDT’s aberrant activation in lung cancer varies according to the histological subtypes and in contrast with other cancer/testis genes, it is rarely expressed in other solid tumours. The functional characterization of BRDT in its physiological setting in male germ cells is now painting a clear portrait of its normal activity and also suggests possible underlying oncogenic activities, when the gene is ectopically activated in cancers. Also, these functional studies of BRDT point to specific anti-cancer therapeutic strategies that could be used to “high-jack” BRDT’s action and turn it against cancer cells, which express this gene. Finally, BRDT’s expression could be used as a biomarker for cell sensitivity to BET bromodomain inhibitors, which have become newly available as anti-cancer drugs.

Epigenetic and genetic alterations are two mechanisms participating in leukemia, which can inactivate genes involved in leukemia pathogenesis or progression. The purpose of this review was to introduce various inactivated genes and evaluate their possible role in leukemia pathogenesis and prognosis. By searching the mesh words “Gene, Silencing AND Leukemia” in PubMed website, relevant English articles dealt with human subjects as of 2000 were included in this study. Gene inactivation in leukemia is largely mediated by promoter’s hypermethylation of gene involving in cellular functions such as cell cycle, apoptosis, and gene transcription. Inactivated genes, such as ASPP1, TP53, IKZF1 and P15, may correlate with poor prognosis in acute lymphoid leukemia (ALL), chronic lymphoid leukemia (CLL), chronic myelogenous leukemia (CML) and acute myeloid leukemia (AML), respectively. Gene inactivation may play a considerable role in leukemia pathogenesis and prognosis, which can be considered as complementary diagnostic tests to differentiate different leukemia types, determine leukemia prognosis, and also detect response to therapy. In general, this review showed some genes inactivated only in leukemia (with differences between B-ALL, T-ALL, CLL, AML and CML). These differences could be of interest as an additional tool to better categorize leukemia types. Furthermore; based on inactivated genes, a diverse classification of Leukemias could represent a powerful method to address a targeted therapy of the patients, in order to minimize side effects of conventional therapies and to enhance new drug strategies.

Multiple Myeloma (MM) is a heterogeneous cytogenetic disorder in which clonal plasma cells proliferate in the bone marrow (BM) and cause bone destruction. The BM microenvironment plays a crucial role in pathogenesis of this disease, and mesenchymal stem cells (MSCs) are one of the key players. Herein, we propose to investigate the expressions of hsa-MIR-204, runt-related transcription factor 2 (RUNX2), peroxisome proliferator-activated receptor gamma (PPARγ), and B-cell lymphoma 2 (BCL2) as factors involved in osteogenesis, adipogenesis, and MSC survival in BM-MSCs from MM patients and normal individuals.
In this experimental study, we isolated MSCs from BM aspirates of MM patients and healthy donors. Total RNA were extracted before and after co-culture with L363 myeloma cells. Gene expressions of RUNX2, PPARγ, BCL2, and hsa-MIR-204 were assessed by quantitive real time polymerase chain reaction (qRT-PCR).
Higher levels of RUNX2, PPARγ, and hsa-MIR-204 expressions existed in MM- MSCs compared to normally derived (ND)-MSCs. BCL2 expression decreased in MM- MSCs. We observed different results in the co-culture model.
In general, the MM-MSCs gene expression profile differed compared to ND- MSCs. Upregulation of RUNX2, PPARγ, and hsa-MIR-204 in MM-MSCs compared to ND- MSCs would result in formation of bone defects. Downregulation of BCL2 would lead to MM-MSC cell death.

There is a positive correlation between higher serum phytoestrogen concentrations and lower risk of breast cancer. The activation of telomerase is crucial for the growth of cancer cells; therefore, the aim of this study was to examine the effects of enterolactone (ENL) and enterodiol (END) on this enzyme.
In this experimental study, we performed the viability assay to determine the effects of different concentrations of ENL and END on cell viability, and the effective concentrations of these two compounds on cell growth. We used western blot analysis to evaluate human telomerase reverse transcriptase catalytic subunit (hTERT) expression and polymerase chain reaction (PCR)-ELISA based on the telomeric repeat amplification protocol (TRAP) assay for telomerase activity.
Both ENL and END, at 100 μM concentrations, significantly (P High concentration of ENL decreased the viability of MCF-7 breast cancer cells and inhibited the expression and activity of telomerase in these cells. Although END could reduce breast cancer cell viability, it did not have any effect on telomerase expression and activity.

This study attempted to identify altered metabolism and pathways related to non-Hodgkin’s lymphoma (NHL) and myeloma patients.
In this retrospective study, we collected plasma samples from 11 patients-6 healthy controls with no evidence of any blood cancers and 5 patients with either multiple myeloma (n=3) or NHL (n=2) during the preliminary study period. Samples were analyzed using quadrupole time-of-flight liquid chromatography mass spectrometry (LC-MS). Significant features generated after statistical analyses were used for metabolomics and pathway analysis.
Data after false discovery rate (FDR) adjustment at q=0.05 of features showed 136 for positive and 350 significant features for negative ionization mode in NHL patients as well as 262 for positive and 98 features for negative ionization mode in myeloma patients. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis determined that pathways such as steroid hormone biosynthesis, ABC transporters, and arginine and proline metabolism were affected in NHL patients. In myeloma patients, pyrimidine metabolism, carbon metabolism, and bile secretion pathways were potentially affected by the disease.
The results have shown tremendous differences in the metabolites of healthy individuals compared to myeloma and lymphoma patients. Validation through quantitative metabolomics is encouraged, especially for the metabolites with significantly expression in blood cancer patients.

In this study we prepared a novel formulation of liposomal doxorubicin (L- DOX). The drug dose was optimized by analyses of cellular uptake and cell viability of osteosarcoma (OS) cell lines upon exposure to nanoliposomes that contained varying DOX concentrations. We intended to reduce the cytotoxicity of DOX and improve characteristics of the nanosystems.
In this experimental study, we prepared liposomes by the pH gradient hydration method. Various characterization tests that included dynamic light scattering (DLS), cryogenic transmission electron microscopy (Cryo-TEM) imaging, and UV- Vis spectrophotometry were employed to evaluate the quality of the nanocarriers. In addition, the CyQUANT® assay and fluorescence microscope imaging were used on various OS cell lines (MG-63, U2-OS, SaOS-2, SaOS-LM7) and Human primary osteoblasts cells, as novel methods to determine cell viability and in vitro transfection efficacy.
We observed an entrapment efficiency of 84% for DOX within the optimized liposomal formulation (L-DOX) that had a liposomal diameter of 96 nm. Less than 37% of DOX released after 48 hours and L-DOX could be stored stably for 14 days. L-DOX increased DOX toxicity by 1.8-4.6 times for the OS cell lines and only 1.3 times for Human primary osteoblasts cells compared to free DOX, which confirmed a higher sensitivity of the OS cell lines versus Human primary osteoblasts cells for L-DOX. We deduced that L- DOX passed more freely through the cell membrane compared to free DOX.
We successfully synthesized a stealth L-DOX that contained natural phospholipid by the pH gradient method, which could encapsulate DOX with 84% efficiency. The resulting nanoparticles were round, with a suitable particle size, and stable for 14 days. These nanoparticles allowed for adequately controlled DOX release, increased cell permeability compared to free DOX, and increased tumor cell death. L-DOX provided a novel, more effective therapy for OS treatment.

Forkhead box (FOX) proteins are important regulators of the epithelial-to-mesenchymal transition (EMT), which is the main mechanism of cancer metastasis. Different studies have shown their potential involvement in progression of cancer in different tissues such as breast, ovary and colorectum. In this study, we aimed to analyze the expression of genes encoding two FOX proteins in gastric adenocarcinoma.
In this experimental case-control study, the expression of FOXC2 and FOXQ1 was examined in 31 gastric adenocarcinoma tumors and 31 normal adjacent gastric tissues by reverse transcription polymerase chain reaction (PCR).
The expression of both genes was significantly up-regulated in gastric adenocarcinoma tumors compared with the normal tissues (P We show that up-regulation of FOXC2 and FOXQ1 are likely to be involved in the progression of gastric adenocarcinoma.

The genetic variants of the long non-coding RNA ANRIL (an antisense noncoding RNA in the INK4 locus) as well as its expression have been shown to be associated with several human diseases including cancers. The aim of this study was to examine the association of ANRIL variants with breast cancer susceptibility in Iranian patients.
In this case-control study, we genotyped rs1333045, rs4977574, rs1333048 and rs10757278 single nucleotide polymorphisms (SNPs) in 122 breast can- cer patients as well as in 200 normal age-matched subjects by tetra-primer amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR).
The TT genotype at rs1333045 was significantly over-represented among pa- tients (P=0.038) but did not remain significant after multiple-testing correction. In addi- tion, among all observed haplotypes (with SNP order of rs1333045, rs1333048 rs4977574 and rs10757278), four haplotypes were shown to be associated with breast cancer risk. However, after multiple testing corrections, TCGA was the only haplotype which remained significant.
These results suggest that breast cancer risk is significantly associated with ANRIL variants. Future work analyzing the expression of different associated ANRIL haplotypes would further shed light on the role of ANRIL in this disease.

This study aimed to determine the effect of 13.56 MHz radiofrequency (RF) capacitive hyperthermia (HT) on radiosensivity of human prostate cancer cells pre and post X-ray radiation treatment (RT).
In this experimental study, the human prostate cancer cell line DU145 was cultured as 300 µm diameter spheroids. We divided the spheroids into group I: control, group II: HT at 43˚C for 30 minutes (HT), group III: 4 Gy irradiation with 6 MV X-ray [RT (6 MV)], group IV: 4 Gy irradiation with 15 MV X-ray [RT (15 MV)], group V: HT (6 MV), group VI: HT (15 MV), group VII: RT (6 MV), and group VIII: RT (15 MV). The alkaline comet assay was used to assess DNA damages in terms of tail moment (TM). Thermal enhancement factor (TEF) was obtained for the different treatment combinations.
Mean TM increased with increasing photon energy. Group II had significantly greater TM compared to group I. Groups III and IV also had significantly higher TM compared to group I. Significant differences in TM existed between groups V, VII, and III (P Our results suggest that HT applied before RT leads to higher radiosensivity compared to after RT. HT at 43˚C for 30 minutes added to 6 MV X-ray causes higher enhancement of radiation compared to 15 MV X-ray.

This study evaluated enhanced induced DNA damages and apoptosis of a spheroid culture of DU145 prostate cancer cells treated by a combination of radiofrequency hyperthermia (RF HT) with radiation treatment (RT) from an external radiotherapy machine compared to RT alone.
In this experimental study, DU145 cells were cultured as spheroids until they reached 300 µm in diameter. We exposed these cultures to either: RF HT for 90 minutes at 43˚C originated from a Celsius TCS system, RF HT followed by RT at doses of 2 Gy or 4 Gy (15 MV energy) with 15-minute interval, or RT alone at the above mentioned doses. The trypan blue exclusion assay, alkaline comet assay, and annexin V/PI flow cytometry were performed to measure cell viability, the amount of DNA damage in an individual cell as the tail moment, and percentage of induced cell apoptosis in response to treatments explained.
We calculated the thermal enhancement factor (TEF) for the combined treatment regime. RF HT followed by the 4 Gy dose of RT resulted in minimum viability (85.33 ± 1.30%), the highest tail moment (1.98 ± 0.18), and highest percentage of apoptotic cells (64.48 ± 3.40%) compared to the other treatments. The results of the TEF assay were 2.54 from the comet assay and 2.33 according to flow cytometry.
The present data suggest that combined treatment of mega voltage X-rays and RF HT can result in significant radiosensitization of prostate cancer cells.

Colorectal cancer (CRC) is the second leading cause of cancer death in occidental countries. Chronic inflammatory bowel disease (crohn’s disease and ulcerative colitis) is associated with an increased risk for CRC development. The aim of this work was to investigate the relationship between inflammatory status and absorption of nutrients with a role in CRC pathogenesis.
In this experimental study, we evaluated the in vitro effect of tumour necrosis factor-alpha (TNF-α), interferon-γ (IF-γ), and acetylsalicylic acid on 14C-butyrate (14C- BT), 3H-folic acid (3H-FA) uptake, and on proliferation, viability and differentiation of Caco-2 and IEC-6 cells in culture.
The proinflammatory cytokines TNF-α and INF-γ were found to decrease uptake of a low concentration of 14C-BT (10 µM) by Caco-2 (tumoral) and IEC-6 (normal) intestinal epithelial cell lines. However, the effect of TNF-α and INF-γ in IEC-6 cells is most probably related to a cytotoxic and antiproliferative impact. In contrast, INF-γ increases uptake of a high concentration (10 mM) of 14C-BT in Caco-2 cells. The anticarcinogenic effect of BT (10 mM) in these cells is not affected by the presence of this cytokine. On the other hand, acetylsalicylic acid stimulates 14C-BT uptake by Caco-2 cells and potentiates its antiproliferative effect. Finally, both TNF-α and INF-γ cause a significant decrease in 3H-FA uptake by Caco-2 cells.
The inflammatory status has an impact upon cellular uptake of BT and FA, two nutrients with a role in CRC pathogenesis. Moreover, the anti-inflammatory acetylsalicylic acid potentiates the anticarcinogenic effect of BT in Caco-2 cells by increasing its cellular uptake.

We studied effect of high glucose levels on coronary artery endothelial cell proliferation and human colon cancer cell proliferation. To examine the long-term effect of glucose exposure on cell growth, cells were cultured for 14 days in the absence or presence of 183 mg/dL D-glucose addition in the culture medium. Short effect of elevated glucose levels was examined by addition of 183 mg/dL D-glucose addition in the culture medium for just one hour per day followed by changing the culture to standard medium (5.5 mM D-glucose) during the next 23-hours period. Cell proliferation was estimated by 2,3-Bis (2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carbox-anilide (XTT) assay and phosphor-Erk western blot analysis. We found that coronary artery endothelial cell proliferation was significantly increased in the culture medium with the acute one-hour addition of 183 mg/dL D-glucose compared to the absence or chronic presence of 183 mg/dL D-glucose addition in the culture medium. In contrast, colon cancer cell proliferation was significantly increased in the continuous presence of 183 mg/dL D-glucose addition in the culture medium compared to the acute one-hour addition of glucose. The extent of Erk2 phosphorylation paralleled with the relative changes in cellular proliferation in both cell types. Taken together, these results suggested that continuous or transient high level of glucose exposure differentially effects coronary artery endothelial and human colon cancer cell proliferation.

The detection of KRAS and BRAF mutations is a crucial step for the correct therapeutic approach and predicting the epidermal growth factor receptor (EGFR)-targeted therapy resistance of colorectal carcinomas. The concomitant KRAS and BRAF mutations occur rarely in the colorectal cancers (CRCs) with the prevalence of less than 0.001% of the cases. In patients with KRAS-mutant tumors, BRAF mutations should not regularly be tested unless the patient is participating in a clinical trial enriching for the presence of KRAS or BRAF-mutated tumor. The current report demonstrates a case with advanced adenocarcinoma of the colon showing the coexistence of KRAS and BRAF mutations and may have profound clinical implications for disease progression and therapeutic responses.