Fatemeh Ahmadipour

Receptor tyrosine kinase (c-Kit) inhibitors: a potential therapeutic target in cancer cells

c-Kit, a receptor tyrosine kinase, is involved in intracellular signaling, and the mutated form of c-Kit plays a crucial role in occurrence of some cancers. The function of c-Kit has led to the concept that inhibiting c-Kit kinase activity can be a target for cancer therapy. The promising results of inhibition of c-Kit for treatment of cancers have been observed in some cancers such as gastrointestinal stromal tumor, acute myeloid leukemia, melanoma, and other tumors, and these results have encouraged attempts toward improvement of using c-Kit as a capable target for cancer therapy. This paper presents the findings of previous studies regarding c-Kit as a receptor tyrosine kinase and an oncogene, as well as its gene targets and signaling pathways in normal and cancer cells. The c-Kit gene location, protein structure, and the role of c-Kit in normal cell have been discussed. Comprehending the molecular mechanism underlying c-Kit-mediated tumorogenesis is consequently essential and may lead to the identification of future novel drug targets. The potential mechanisms by which c-Kit induces cellular transformation have been described. This study aims to elucidate the function of c-Kit for future cancer therapy. In addition, it has c-Kit inhibitor drug properties and their functions have been listed in tables and demonstrated in schematic pictures. This review also has collected previous studies that targeted c-Kit as a novel strategy for cancer therapy. This paper further emphasizes the advantages of this approach, as well as the limitations that must be addressed in the future. Finally, although c-Kit is an attractive target for cancer therapy, based on the outcomes of treatment of patients with c-Kit inhibitors, it is unlikely that Kit inhibitors alone can lead to cure. It seems that c-Kit mutations alone are not sufficient for tumorogenesis, but do play a crucial role in cancer occurrence.

Koenimbin, a natural dietary compound of Murraya koenigii (L) Spreng: inhibition of MCF7 breast cancer cells and targeting of derived MCF7 breast cancer stem cells (CD44(+)/CD24(-/low)): an in vitro study.

Background Inhibition of breast cancer stem cells has been shown to be an effective therapeutic strategy for cancer prevention. The aims of this work were to evaluate the efficacy of koenimbin, isolated from Murraya koenigii (L) Spreng, in the inhibition of MCF7 breast cancer cells and to target MCF7 breast cancer stem cells through apoptosis in vitro. Methods Koenimbin-induced cell viability was evaluated using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Nuclear condensation, cell permeability, mitochondrial membrane potential, and cytochrome c release were observed using high-content screening. Cell cycle arrest was examined using flow cytometry, while human apoptosis proteome profiler assays were used to investigate the mechanism of apoptosis. Protein expression levels of Bax, Bcl2, and heat shock protein 70 were confirmed using Western blotting. Caspase-7, caspase-8, and caspase-9 levels were measured, and nuclear factor kappa B (NF-κB) activity was assessed using a high-content screening assay. Aldefluor™ and mammosphere formation assays were used to evaluate the effect of koenimbin on MCF7 breast cancer stem cells in vitro. The Wnt/β-catenin signaling pathway was investigated using Western blotting. Results Koenimbin-induced apoptosis in MCF7 cells was mediated by cell death-transducing signals regulating the mitochondrial membrane potential by downregulating Bcl2 and upregulating Bax, due to cytochrome c release from the mitochondria to the cytosol. Koenimbin induced significant (P<0.05) sub-G0 phase arrest in breast cancer cells. Cytochrome c release triggered caspase-9 activation, which then activated caspase-7, leading to apoptotic changes. This form of apoptosis is closely associated with the intrinsic pathway and inhibition of NF-κB translocation from the cytoplasm to the nucleus. Koenimbin significantly (P<0.05) decreased the aldehyde dehydrogenase-positive cell population in MCF7 cancer stem cells and significantly (P<0.01) decreased the size and number of MCF7 cancer stem cells in primary, secondary, and tertiary mammospheres in vitro. Koenimbin also significantly (P<0.05) downregulated the Wnt/β-catenin self-renewal pathway. Conclusion Koenimbin has potential for future chemoprevention studies, and may lead to the discovery of further cancer management strategies by reducing cancer resistance and recurrence and improving patient survival.

PASS-Predicted Hepatoprotective Activity of Caesalpinia sappan in Thioacetamide-Induced Liver Fibrosis in Rats

The antifibrotic effects of traditional medicinal herb Caesalpinia sappan (CS) extract on liver fibrosis induced by thioacetamide (TAA) and the expression of transforming growth factor β1 (TGF-β1), α-smooth muscle actin (αSMA), and proliferating cell nuclear antigen (PCNA) in rats were studied. A computer-aided prediction of antioxidant and hepatoprotective activities was primarily performed with the Prediction Activity Spectra of the Substance (PASS) Program. Liver fibrosis was induced in male Sprague Dawley rats by TAA administration (0.03% w/v) in drinking water for a period of 12 weeks. Rats were divided into seven groups: control, TAA, Silymarin (SY), and CS 300 mg/kg body weight and 100 mg/kg groups. The effect of CS on liver fibrogenesis was determined by Massons trichrome staining, immunohistochemical analysis, and western blotting. In vivo determination of hepatic antioxidant activities, cytochrome P450 2E1 (CYP2E1), and matrix metalloproteinases (MPPS) was employed. CS treatment had significantly increased hepatic antioxidant enzymes activity in the TAA-treated rats. Liver fibrosis was greatly alleviated in rats when treated with CS extract. CS treatment was noted to normalize the expression of TGF-β1, αSMA, PCNA, MMPs, and TIMP1 proteins. PASS-predicted plant activity could efficiently guide in selecting a promising pharmaceutical lead with high accuracy and required antioxidant and hepatoprotective properties.

Two Novel Tyrosinase (TYR) Gene Mutations with Pathogenic Impact on Oculocutaneous Albinism Type 1 (OCA1)

Oculocutaneous albinism (OCA) is a heterogeneous group of autosomal recessive disorders resulting from mutations of the tyrosinase (TYR) gene and presents with either complete or partial absence of pigment in the skin, hair and eyes due to a defect in an enzyme involved in the production of melanin. In this study, mutations in the TYR gene of 30 unrelated Iranian OCA1 patients and 100 healthy individuals were examined using PCR-sequencing. Additionally, in order to predict the possible effects of new mutations on the structure and function of tyrosinase, these mutations were analyzed by SIFT, PolyPhen and I-Mutant 2 software. Here, two new pathogenic p.C89S and p.H180R mutations were detected in two OCA1 patients. Moreover, the R402Q and S192Y variants, which are common non-pathogenic polymorphisms, were detected in 17.5% and 35% of the patients, respectively. The outcome of this study has extended the genotypic spectrum of OCA1 patients, which paves the way for more efficient carrier detection and genetic counseling.

α-Mangostin from Cratoxylum arborescens demonstrates apoptogenesis in MCF-7 with regulation of NF-κB and Hsp70 protein modulation in vitro, and tumor reduction in vivo.

Cratoxylum arborescens is an equatorial plant belonging to the family Guttiferae. In the current study, α-Mangostin (AM) was isolated and its cell death mechanism was studied. HCS was undertaken to detect the nuclear condensation, mitochondrial membrane potential, cell permeability, and the release of cytochrome c. An investigation for reactive oxygen species formation was conducted using fluorescent analysis. To determine the mechanism of cell death, human apoptosis proteome profiler assay was conducted. In addition, using immunofluorescence and immunoblotting, the levels of Bcl-2-associated X protein (Bax) and B-cell lymphoma (Bcl)-2 proteins were also tested. Caspaces such as 3/7, 8, and 9 were assessed during treatment. Using HCS and Western blot, the contribution of nuclear factor kappa-B (NF-κB) was investigated. AM had showed a selective cytotoxicity toward the cancer cells with no toxicity toward the normal cells even at 30 μg/mL, thereby indicating that AM has the attributes to induce cell death in tumor cells. The treatment of MCF-7 cells with AM prompted apoptosis with cell death-transducing signals. This regulated the mitochondrial membrane potential by down-regulation of Bcl-2 and up-regulation of Bax, thereby causing the release of cytochrome c from the mitochondria into the cytosol. The liberation of cytochrome c activated caspace-9, which, in turn, activated the downstream executioner caspace-3/7 with the cleaved poly (ADP-ribose) polymerase protein, thereby leading to apoptotic alterations. Increase of caspace 8 had showed the involvement of an extrinsic pathway. This type of apoptosis was suggested to occur through both extrinsic and intrinsic pathways and prevention of translocation of NF-κB from the cytoplasm to the nucleus. Our results revealed AM prompt apoptosis of MCF-7 cells through NF-κB, Bax/Bcl-2 and heat shock protein 70 modulation with the contribution of caspaces. Moreover, ingestion of AM at (30 and 60 mg/kg) significantly reduced tumor size in an animal model of breast cancer. Our results suggest that AM is a potentially useful agent for the treatment of breast cancer.

Synthesis, structural characterization, and anticancer activity of a monobenzyltin compound against MCF-7 breast cancer cells

A new monoorganotin Schiff base compound, [N-(3,5-dichloro-2-oxidobenzylidene)-4-chlorobenzyhydrazidato](o-methylbenzyl)aquatin(IV) chloride, (compound C1), was synthesized, and its structural features were investigated by spectroscopic techniques and single-crystal X-ray diffractometry. Compound C1 was exposed to several human cancer cell lines, including breast adenocarcinoma cell lines MCF-7 and MDA-MB-231, ovarian adenocarcinoma cell lines Skov3 and Caov3, and prostate cancer cell line PC3, in order to examine its cytotoxic effect for different forms of cancer. Human hepatic cell line WRL-68 was used as a normal cell line. We concentrated on the MCF-7 cell line to detect possible underlying mechanism involvement of compound C1. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed the strongest cytotoxicity of compound C1 against MCF-7 cells, with a half maximal inhibitory concentration (IC50) value of 2.5±0.50 μg/mL after 48 hours treatment. The IC50 value was >30 μg/mL in WRL-68 cells. Induced antiproliferative activity of compound C1 for MCF-7 cells was further confirmed by lactate dehydrogenase, reactive oxygen species, acridine orange/propidium iodide staining, and DNA fragmentation assays. A significant increase of lactate dehydrogenase release in treated cells was observed via fluorescence analysis. Luminescent analysis showed significant growth in intracellular reactive oxygen species production after treatment. Morphological changes of necrosis and early and late apoptosis stages were observed in treated cells after staining with acridine orange/propidium iodide. DNA fragmentation was observed as a characteristic of apoptosis in treated cells. Results of the present study obviously reveal potential cytotoxic effects of compound C1 against human breast cancer MCF-7 cells.

Four Novel p.N385K, p.V36A, c.1033–1034insT and c.1417–1418delCT Mutations in the Sphingomyelin Phosphodiesterase 1 (SMPD1) Gene in Patients with Types A and B Niemann-Pick Disease (NPD)

Background: Types A and B Niemann-Pick disease (NPD) are autosomal-recessive lysosomal storage disorders caused by the deficient activity of acid sphingomyelinase due to mutations in the sphingomyelin phosphodiesterase 1 (SMPD1) gene. Methods: In order to determine the prevalence and distribution of SMPD1 gene mutations, the genomic DNA of 15 unrelated Iranian patients with types A and B NPD was examined using PCR, DNA sequencing and bioinformatics analysis. Results: Of 8 patients with the p.G508R mutation, 5 patients were homozygous, while the other 3 were heterozygous. One patient was heterozygous for both the p.N385K and p.G508R mutations. Another patient was heterozygous for both the p.A487V and p.G508R mutations. Two patients (one homozygous and one heterozygous) showed the p.V36A mutation. One patient was homozygous for the c.1033–1034insT mutation. One patient was homozygous for the c.573delT mutation, and 1 patient was homozygous for the c.1417–1418delCT mutation. Additionally, bioinformatics analysis indicated that two new p.V36A and p.N385K mutations decreased the acid sphingomyelinase (ASM) protein stability, which might be evidence to suggest the pathogenicity of these mutations. Conclusion: with detection of these new mutations, the genotypic spectrum of types A and B NPD is extended, facilitating the definition of disease-related mutations. However, more research is essential to confirm the pathogenic effect of these mutations.

Prevalence of the UGT1A1*6 (c.211G>A) Polymorphism and Prediction of Irinotecan Toxicity in Iranian Populations of Different Ethnicities

Background: Pharmacogenetic studies on irinotecan treatment in patients with metastatic colorectal cancer have indicated that genetic polymorphisms in UGT1A1*6 can lead to decreased enzyme activity and accumulation of the toxic metabolite SN-38. Here, we compared the prevalence of UGT1A1*6 in an Iranian population of different ethnicities with those of other populations. Materials and Methods: A total of 300 healthy people of different ethnic groups including Persian, Azari, Lure, Kurdish, Arab, Baluch and Caspian in the Iranian population were enrolled. Genotyping of the UGT1A1*6 alleles (G/G, A/G, A/A) was performed by polymerase chain reaction-restriction fragment length polymorphism and direct genomic DNA sequencing. Result: The most predictive genotype among the Iranian ethnic groups, especially Persian, was the G/G genotype (wild-type genotype). The frequency of the A/G genotype among the Persian, Azari, Lure, Kurdish, Arab, Baluch and Caspian ethnicities were 15.69% (n = 27), 11.11% (n = 8), 5.88% (n = 1), 9.09% (n = 1), 10% (n = 1), 20% (n = 1) and 0% (n = 0), respectively. Only one person with Persian ethnicity was homozygous for the mutation in UGT1A1*6 (0.58%). Additionally, the frequency of the A and G alleles in Iranians was 6.83 and 93.16%, respectively. Conclusion: The identification of the UGT1A1*6 alleles is necessary among the different Iranian ethnic groups before irinotecan therapy, suggesting that genotyping would be helpful for clinicians to optimize chemotherapy or identify individuals at risk of adverse drug reactions before clinical trials.

Oral consumption of α-linolenic acid increases serum BDNF levels in healthy adult humans.

Background aimsDietary omega-6 and omega-3 fatty acids have remarkable impacts on the levels of DHA in the brain and retina. Low levels of DHA in plasma and blood hamper visual and neural development in children and cause dementia and cognitive decline in adults. The level of brain-derived neurotrophic factors (BDNF) changes with dietary omega-3 fatty acid intake. BDNF is known for its effects on promoting neurogenesis and neuronal survival.MethodsIn this study, we examined the effect of the oral consumption of α-Linolenic acid (ALA) on blood levels of BDNF and Malondialdehyde (MDA) in healthy adult humans. 30 healthy volunteers, 15 men and 15 women, were selected randomly. Each individual served as his or her own control. Before consuming the Flaxseed oil capsules, 5cc blood from each individual was sampled in order to measure the plasma levels of BDNF and MDA as baseline controls. During the experiment, each individual was given 3 oral capsules of flaxseed oil, containing 500mg of alpha linolenic acid, daily for one week. Then, plasma levels of BDNF and MDA were tested.ResultsThe plasma levels of BDNF and MDA significantly (P < 0.05) increased in individuals who received the oral capsules of ALA. Plasma levels of BDNF increased more in the women in comparison with the men.ConclusionALA treatment could be a feasible approach to reduce size of infarcts in stroke patients. Thus, ALA could be used in adjunction with routine stroke therapies to minimize brain lesions caused by stroke.

Prevalence of the CYP2D6*10 (C100T), *4 (G1846A), and *14 (G1758A) alleles among Iranians of different ethnicities.

The presence of polymorphisms in the CYP2D6 gene may modulate enzyme level and activity, thereby affecting individual responses to pharmacological treatment. Here, we compared the prevalence of the CYP2D6*10, *4, and 14* alleles in an Iranian population of different ethnicities with those of other populations. Allele and genotype frequency distributions of CYP2D6*10 variants and predicted phenotypes including extensive metabolizers, intermediate metabolizers, and poor metabolizers were analysed in blood samples of 300 unrelated healthy individuals in an Iranian population using polymerase chain reaction (PCR)-restriction fragment length polymorphism, PCR-single-strand conformation polymorphism, and direct genomic DNA sequencing. The CYP2D6*4 (G1846A) and *14 (G1758A) allelic frequencies were not detected in different ethnicities, demonstrating the absence of a significant contribution of these alleles in Iranian populations. However, the T/T, C/T, and C/C genotype frequencies of the CYP2D6*10 allele were significantly different (P<0.01) in all Iranian ethnic groups. Additionally, the frequency of the homozygous T/T variant of the CYP2D6*10 allele was significantly high in the Lure (P<0.017) and low in the Kurd (P<0.002) ethnicities. The frequency of the T/T variant of the CYP2D6*10 allele in central Iran was the highest (P<0.001), while the south of Iran had the lowest frequency (P<0.001). The frequency of the C/T variant of the CYP2D6*10 allele was significantly a bit high (P<0.001) in females compare to males, while the frequencies of the T/T variant in females is similar to males, which are 24.4% and 24.3%, respectively. In contrast to absence of the CYP2D6*4 (G1846A) and *14 (G1758A) alleles in Iranian populations of different ethnicities, the prediction of the CYP2D6*10 allele is required in drug research and routine treatment, where the information would be helpful for clinicians to optimize therapy or identify persons at risk of adverse drug reactions before clinical trials. Approximately 39.3% of subjects (24.3% homozygous T/T CYP2D6*10 as poor metabolizers and 15% heterozygous C/T CYP2D6*10 as intermediate metabolizers) had this allele; therefore, the harmful effects of drugs are relatively common among Iranians.