Deciphering the molecular pathways of apoptosis using purified fractions from leaf extract of Basella alba through studying the regulation of apoptosis related genes.

Apoptosis plays a pivotal role in the exclusion of abnormal cells without any ruin of surrounding healthy cells. Generally, it occurs through an orderly and autonomously process which is controlled by proper function of various genes. Therefore, the current experiments detect the expression level/pattern of those genes to confirm the involvement of extrinsic and intrinsic pathway using Basella alba leaf (BAL). Several fractions after gel filtration chromatography of BAL extract have been pooled to evaluates its apoptosis induction potentiality on Ehrlich's Ascites Carcinoma (EAC) cells through conducting a number of bio-assays such as cell growth inhibition assay, fluorescence and optical microscopy, DNA fragmentation assay and gene expression analysis etc. The pooled fractions of BAL showed 12-56% inhibitory effect on EAC cell line at the concentration range of 25-400 µg/ml that was determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay. They also exhibited excellent cell growth inhibition at in vivo and in vitro condition when treated with 10, 20 and 40 mg/kg day. After administration of six consequent days, significant morphological features of apoptosis were observed in EAC cells under both fluorescence and optical microscope which was further supported by DNA fragmentation assay. The polymerase chain reaction amplification of bax, bcl-2 (B-cell lymphoma 2), p53, tumor necrosis factor-α, Fas, NF-kß (Nuclear factor-Kappa-B), PARP-1 (Poly (ADP-ribose) polymerase), Cyt-c cas-8, cas-9 and cas-3 revealed that the experimental sample able to induce apoptosis in both extrinsic and intrinsic pathways through altering the gene expression. The current findings suggest that sample from BAL occupy wonderful competence to induce cell apoptosis and become an ideal resource for cancer treatment.

Indole glucosinolates exhibit anti-inflammatory effects on Ehrlich ascites carcinoma cells through modulation of inflammatory markers and miRNAs.

BACKGROUND: Nuclear factor-κB (NF-κB) has been identified as the major link between inflammation and cancer. Natural agents that inhibit this pathway are essential in attenuating inflammation induced by cancer or chemotherapeutic drugs. High intake of Brassicaceae vegetables has been determined to modulate essential pathways related to chronic diseases. In this study, we investigated the anti-proliferative and anti-inflammatory effects of the indole glucosinolates; indole-3-carbinol (I3C) and its metabolite 3,3-diindolylmethane (DIM) on the inflammatory biomarkers and miRNAs controlling the NF-κB pathway. METHODS AND RESULTS: In our study, we inoculated Ehrlich ascites carcinoma (EAC) cells in female albino mice, which increased their packed cell volume and induced a significant increase in the levels of several cytokines and inflammatory biomarkers (NF-κB IL-6, IL-1b, TNF-α, and NO). A significant elevation in inflammatory-medicated miRNAs (miR-31 and miR-21) was also noted. Treatment with 5-fluorouracil (5-FU) significantly reduced packed cell volume and viable cell count. However, it was accompanied by a significant increase in the levels of inflammatory markers and expression of miR-31 and miR-21. Nevertheless, although treatment with indoles (I3C and DIM) significantly reduced the packed cell volume and viable cell count, their prominent effect was the marked reduction of all inflammatory biomarkers compared to both the EAC untreated group and the EAC group treated with 5-FU. Moreover, the anti-inflammatory effect of I3C or DIM was accompanied by a significant decrease in the expression of miR-31 and miR-21. CONCLUSION: Our findings have; therefore, revealed that I3C and DIM have strong anti-inflammatory effects, implying that their use as a co-treatment with chemotherapeutic drugs can effectively improve the anti-tumor effect of chemotherapeutic drugs.

The effect of antioxidants in Ehrlich Ascites Cancer.

A fundamental goal in molecular oncology is to unravel the underlying mechanisms which cause the cell transformation. In line with this approach, genome-wide functional screening approaches have revealed exciting insights into heterogeneous nature of cancer. Rapidly expanding horizons of research have unraveled myriad of pathways which play instrumental role in carcinogenesis and metastasis. Oxidative stress has also been reported to be significantly involved in cancer onset and progression. In line with this approach, oxidative stress modulating chemicals have always been sharply divided into antioxidants and oxidative stress-inducing agents. Conceptual and experimental advancements have enabled us to critically analyze full potential of these two different groups of chemicals in cancer chemoprevention. Different antioxidants are currently being analyzed in different phases of clinical trials. Although it has been reported in the literature that antioxidant supplements reduce tumor cells in some tumors or cause volume reduction in solid tumor sizes, there is no definite consensus. Therefore, an antioxidant supplement guideline based on more detailed clinical research and as a result of these is needed to achieve the best care for cancer patients and to avoid risky treatments for cancer patients.

Thymoquinone upregulates miR-125a-5p, attenuates STAT3 activation, and potentiates doxorubicin antitumor activity in murine solid Ehrlich carcinoma.

In breast cancer, there has been evidence of atypical activation of signal transduction and activators of transcription 3 (STAT3). Thymoquinone (TQ) exerts its anti-neoplastic effect through diverse mechanisms, including STAT3 inhibition. The tumor suppressor, microRNA-125a-5p was reported to be downregulated in various breast cancer cells. Therefore, we investigated the influence of TQ and/or doxorubicin on microRNA-125a-5p and its correlation with STAT3 activation as well as tumor growth in mice bearing solid Ehrlich tumors. We found that TQ markedly suppressed inducible and constitutive phosphorylation of STAT3 in tumor tissue without affecting STAT5. Moreover, it attenuated tumor growth, downregulated STAT3 downstream target proteins, and increased the apoptotic activities of caspase-3 and -9. Interestingly, TQ-elicited synergism of doxorubicin anti-neoplastic activity was coupled with upregulation of tumoral microRNA-125a-5p. Taken together, the current findings raise the potential of TQ as a promising chemomodulatory adjuvant to augment mammary carcinoma sensitivity to doxorubicin.

Molecular changes associated with the anticancer effect of sulforaphane against Ehrlich solid tumour in mice.

The anticancer effect of sulforaphane (SFN) is mediated by several signalling pathways. However, little is known regarding the underlying mechanism in Ehrlich solid tumours (ESTs) in mice. This study was conducted to determine molecular changes associated with the anticancer effect of SFN and to compare its preventive (cotreatment) and therapeutic (posttreatment) effects. Ehrlich (murine mammary adenocarcinoma) solid tumour was selected and changes in the gene expression were determined in tumour tissues by the real-time polymerase chain reaction. The results showed that SFN increased the expression of the oxidative stress gene NrF2 and its downstream targets (HO1 and CAT). Conversely, SFN administration decreased the expression of the epigenesis-related genes (HDAC1 and DNMT1) and inflammation-related genes (TNFa, NFkB and Cox2). Overall, SFN cotreatment presented notable molecular changes than the posttreatment strategy. These data suggest that molecular changes associated with the anticancer effects of SFN against EST involved induction of oxidative stress, inhibition of inflammation and epigenetic modifications.

An Unusual Pathway of Mitoptosis Found in Ehrlich Carcinoma Cells.

An integrated microscopic study of the destruction of mouse Ehrlich ascites carcinoma (EAC) cells under starvation conditions has been carried out. It has been found that, in addition to apoptosis, necrosis, and apoptotic necrosis, already known for EAC, cell destruction can also occur through mitochondrial autolysis, which is proposed to be considered a new kind of mitoptosis. A mitoptosis in EAC is characterized by the appearance of many autolyzing mitochondria, the fusion of which leads to rupture of the cell membrane and the ejection of the nucleus from the cell. It is assumed that the polymorphism of EAC destruction patterns is explained by the different physiological state of the cells, which determines the "choice" of the cell death mechanism. This situation poses a challenge for researchers to develop complex inducers with the ability to stimulate all possible types of cancer cell death.

Salvia lachnostachys Benth has antitumor and chemopreventive effects against solid Ehrlich carcinoma.

Salvia lachnostachys is an herbaceous plant with anti-inflammatory, analgesic and cytotoxic properties. This study investigated the antitumor effect of an ethanolic extract of Salvia lachnostachys leaves (EES) in a solid Ehrlich carcinoma model. Ehrlich cells were inoculated subcutaneously in the right pelvic member (2 × 106 cells) in female Swiss mice. The animals were treated with vehicle (10 mL kg-1, p.o.), EES (30 and 100 mg kg-1, p.o.), or methotrexate (2.5 mg kg-1, i.p.) for 21 days (early treatment) or 14 days (late treatment) after tumor inoculation, or 10 days before tumor inoculation and continued for 21 days after tumor inoculation (chemopreventive treatment). The acute toxicity test was performed according OECD guidelines Late treatment with EES had no antitumor effect. Early treatment with 100 mg kg-1 EES prevented tumor development, increased tumor necrosis factor-α (TNF-α) levels and decreased tumor superoxide dismutase (SOD) activity, interleukin-10 (IL-10) levels and Cyclin D1 expression, and tumor cell necrosis was observed. Chemopreventive treatment with EES for 10 and 31 days prevented tumor development in the same manner. EES treatment for 31 days decreased hepatic and tumor SOD activity, tumor IL-10 levels and Cyclin D1 expression, and increased tumor reduced glutathione, N-acetylglucosaminidase, reactive oxygen species, lipid peroxidation, TNF-α levels and Nrf2 expression. No toxicity was observed in the acute toxicity assay. In conclusion, EES had an antitumor effect by inhibiting Cyclin D1 expression and increasing inflammation with early and chemopreventive treatment. Modulation of the antioxidant system also contribute for the antitumor effects of EES.

Metformin augments doxorubicin cytotoxicity in mammary carcinoma through activation of adenosine monophosphate protein kinase pathway.

Since the incidence of breast cancer increases dramatically all over the world, the search for effective treatment is an urgent need. Metformin has demonstrated anti-tumorigenic effect both in vivo and in vitro in different cancer types. This work was designed to examine on molecular level the mode of action of metformin in mice bearing solid Ehrlich carcinoma and to evaluate the use of metformin in conjunction with doxorubicin as a combined therapy against solid Ehrlich carcinoma. Ehrlich ascites carcinoma cells were inoculated in 60 female mice as a model of breast cancer. The mice were divided into four equal groups: Control tumor, metformin, doxorubicin, and co-treatment. Metformin (15 mg/kg) and doxorubicin (4 mg/kg) were given intraperitoneally (i.p.) for four cycles every 5 days starting on day 12 of inoculation. The anti-tumorigenic effect of metformin was mediated by enhancement of adenosine monophosphate protein kinase activity and elevation of P53 protein as well as the suppression of nuclear factor-kappa B, DNA contents, and cyclin D1 gene expression. Metformin and doxorubicin mono-treatments exhibited opposing action regarding cyclin D1 gene expression, phosphorylated adenosine monophosphate protein kinase, and nuclear factor-kappa B levels. Co-treatment markedly decreased tumor volume, increased survival rate, and improved other parameters compared to doxorubicin group. In parallel, the histopathological findings demonstrated enhanced apoptosis and absence of necrosis in tumor tissue of co-treatment group. Metformin proved chemotherapeutic effect which could be mediated by the activation of adenosine monophosphate protein kinase and related pathways. Combining metformin and doxorubicin, which exhibited different mechanisms of action, produced greater efficacy as anticancer therapeutic regimen.

Assessment of DNA damage in Ehlrich carcinoma after treatment with doxorubicin encapsulated in nanoscales thermosensitive liposomes in combination with localized hyperthermia.

Nanoscales thermosensitive liposomes (TSL) composed of synthetic lipids (dipalmitoylphosphatidylcholine, and distearoylphosphatidylcholine), were used for doxorubicin encapsulation with 70% encapsulated efficiency. The liposomes were characterized by dynamic light scattering, transmission electron microscopy and turbidity method. Additionally, the liposomes exhibited a significant release of doxorubicin (Dox) by 60% within 5 min at 42°C. To assess the therapeutic efficacy of Dox in combination with hyperthermia, Dox free and encapsulated TSL were administered directly to Ehrlich tumor bearing mice at 1 mg/kg dose. Immediately after the drug administration, hyperthermia was applied to mention the temperature inside the tumor site at 42°C either for 5 min and 30 min. The results indicate a significant increase in the percent of apoptotic and necrotic cells in the treated group. Moreover, disrupts the integrity and the amount of intact DNA in tumor cells. In conclusion, Dox and hyperthermia may serve as a useful targeted drug delivery system for management of Ehrlich carcinoma.

Integrin ß1, Osmosensing, and Chemoresistance in Mouse Ehrlich Carcinoma Cells.

BACKGROUND/AIMS: Altered expression of the integrin family of cell adhesion receptors has been associated with initiation, progression, and metastasis of solid tumors as well as in the development of chemoresistance. Here, we investigated the role of integrins, in particular integrin ß1, in cell volume regulation and drug-induced apoptosis in adherent and non-adherent Ehrlich ascites cell lines. METHODS: Adhesion phenotypes were verified by colorimetric cell-adhesion-assay. Quantitative real-time PCR and western blot were used to compare expression levels of integrin subunits. Small interfering RNA was used to silence integrin ß1 expression. Regulatory volume decrease (RVD) after cell swelling was studied with calcein-fluorescence-self-quenching and Coulter counter analysis. Taurine efflux was estimated with tracer technique. Caspase assay was used to determine apoptosis. RESULTS: We show that adherent cells have stronger fibronectin binding and a significantly increased expression of integrin α5, αv, and ß1 at mRNA and protein level, compared to non-adherent cells. Knockdown of integrin ß1 reduced RVD of the adherent but not of the non-adherent cells. Efflux of taurine was unaffected. In contrast to non-adherent, adherent cells exhibited chemoresistance to chemotherapeutic drugs (cisplatin and gemcitabine). However, knockdown of integrin ß1 promoted cisplatin-induced caspase activity in adherent cells. CONCLUSION: Our data identifies integrin ß1 as a part of the osmosensing machinery and regulator of cisplatin resistance in adherent Ehrlich cells.

Effect of hesperidin on mice bearing Ehrlich solid carcinoma maintained on doxorubicin.

Doxorubicin (DOX) is widely used in cancer therapy of many carcinomas types. Unfortunately, DOX is not sufficiently effective in many cases, and increasing the dosage of it is limited due to its systemic toxicity. A citrus flavonoid hesperidin (HES) is proved to be potent antioxidant and protective agent against many diseases including cancer. In this context, the objective of this study was to examine effect of HES along with DOX on solid Ehrlich carcinoma (SEC) in mice. Forty male mice were divided into four equal groups (n = 10): control SEC, DOX, HES, and DOX + HES. HES (50 mg/kg body weight orally) was given day after day for 16 days along with DOX (4 mg/kg body weight i.p. injection) for 5 cycles every 4 days in ESC-inoculated mice. After 20 days, tumor volume, tumor weight, survival rate, tumor glutathione, nitric oxide content, and serum glutathione were determined. Tumor tissue was examined for histopathological and immunohistochemical study for p53 and VEGF. Tumor resistance for mdr1a gene was assessed in tumor tissue by RT-PCR. HES induced significant increase in tissue and serum glutathione with significant decrease in tumor volume and tumor weight. A possible role of HES to modulate gene expression of mdr1a in tumor tissue was established. In addition, HES alleviated the histopathological changes with significant decrease in p53 and VEGF expression. The use of HES as adjuvant therapy with DOX would enhance the therapeutic efficacy and alleviate the resistance to DOX in treatment of solid tumors.

[Genetic features of nitric oxide generating systems predetermine the body's resistance to the development of carcinoma].

Predisposition to tumors is often determined by how effectively the genotype of an individual forms an immune defense. An important factor of such protection is macrophage NO. We assumed that the body's vulnerability to the development of tumors may depend from the characteristics of the NO generating systems. The content of NO in the tumor changed by ITU, inhibitor of iNOS, c-PTIO, traps and SNP, donor NO. Production of macrophage NO were evaluated by nitrites in the culture media. iNOS was assessed using the Western blot analysis. Phenotype of macrophages was assessed using cytometry for CD labels. Life span of mice C57BL/6N with Ehrlich tumor was 25% greater than that of the C57BL/6J. Reducing the content of NO in the tumor reduced life expectancy of high-resistance to tumor subline C57BL/6N at 23%. Increase of NO increased life expectancy of low-resistance subline C57BL/6J at 26%. Macrophages of C57BL/6N were 1.5 times higher contents of iNOS and NO production, as compared with macrophages of C57BL/6J. CD phenotype markers determined the macrophage phenotype C57BL/6N as M1 and C57BL/6J mice macrophage phenotype as M2. Thus, the body's vulnerability to the development of tumors may depend from the characteristics of the NO generating systems. C57BL/6J, unlike C57BL/6N does not synthesize NNT (nicotinamide nucleotide transhydrogenase) and have differences in the single nucleotide polymorphism (SNP). The important role of NO in the resistance to Carcinoma, NNT and SNP deserve attention in the development of new methods of antitumor therapy.

Dextran-functionalized magnetic fluid mediating magnetohyperthermia for treatment of Ehrlich-solid-tumor-bearing mice: toxicological and histopathological evaluations.

Dextran-functionalized maghemite fluid (DexMF) has been tested to treat Ehrlich-solid-tumor-bearing mice, evidencing its potential use in mediating magnetohyperthermia in breast cancer treatment. However, although magnetic nanoparticles tend to accumulate in tumor tissues, part of the nanomaterial can reach the blood stream, and then the organism. The aim of this study was to investigate the acute systemic effects of the intratumoral injection of DexMF mediating magnetohyperthermia in the treatment of an advanced clinical Ehrlich-solid-tumor, assessed through histopathological analyses of liver, kidneys, heart and spleen, comet assay, micronucleus test, hemogram, and serum levels of bilirubin, aspartate aminotransferase, alanine aminotransferase, gamma glutamyl transferase, alkaline phosphatase, creatinine, and urea. The tumor's histopathology and morphometry were used to assess its aggressiveness and regression. DexMF mediating hyperthermia was effective in containing tumor aggressiveness and in inducing tumor regression, besides showing no toxic effects. Its physical characteristics also suggest that it is safe to use in other biomedical applications.

[The role of genetic peculiarityes (of organisms) in the resistance to neoplastic processes in August line and Wistar population rats].

The probability of development of the Ehrlich's ascites carcinoma in young August and Wistar rats was investigated. The Ehrlich's carcinoma strain was derived in mice in the N.N. Blokhin Russian Cancer Research Center. The tumor was transplanted into rats intraperitonially. It was shown that the transplanted murine carcinomas did not arouse tumors in rats, but caused pathologic effects: abrupt growth impairment and partial loss in the August rats while in the Wistar rats the growth impairment was slight and there was no loss. Thus, the first, there was no tumor growth in rats and the second, the indicated effects of the murine tumor transplantation were more dramatic in the August rats than thouse in the Wistar rats.

Bis(acridine-9-carboxylate)-nitro-europium(III) dihydrate complex a new apoptotic agent through Flk-1 down regulation, caspase-3 activation and oligonucleosomes DNA fragmentation.

New bis(acridine-9-carboxylate)-nitro-europium(III) dihydrate complex was synthesized and characterized. In vivo anti-angiogenic activities of bis(acridine-9-carboxylate)-nitro-europium(III) dihydrate complex against Ehrlich ascites carcinoma (EAC) cells are described. The newly synthesized complex resulted in inhibition of proliferation of EAC cells and ascites formation. The anti-tumor effect was found to be through anti-angiogenic activity as evident by the reduction of microvessel density in EAC solid tumors. The anti-angiogenic effect is mediated through down-regulation of VEGF receptor type-2 (Flk-1). The complex was also found to significantly increase the level of caspase-3 in laboratory animals compared to the acridine ligand and to the control group. This was also consistent with the DNA fragmentation detected by capillary electrophoresis that proved the apoptotic effect of the new complex. Our complex exhibited anti-angiogenic and apoptotic activity in vivo, a thing that makes it a potential effective chemotherapeutic agent. The interaction of calf thymus DNA (ct-DNA) with bis(acridine-9-carboxylate)-nitro-europium(III) dihydrate complex has been investigated using fluorescence technique. A competitive experiment of the europium(III)-acridine complex with ethidium bromide (EB) to bind DNA revealed that interaction between the europium(III)-acridine and DNA was via intercalation. The interaction of the synthesized complex with tyrosine kinases was also studied using molecular docking simulation to further substantiate its mode of action.

MicroRNA expression profiles associated with development of drug resistance in Ehrlich ascites tumor cells.

Multidrug resistance (MDR) poses a major obstacle to successful chemotherapeutic treatment of cancer, and often involves multiple genes, which may be regulated post-transcriptionally by microRNAs (miRNAs). The purpose of the present study was therefore to identify any resistance-associated changes in miRNA expression in a sensitive and five increasingly drug-resistant Ehrlich ascites tumor (EAT) cell lines, representing different steps in the development of resistance. We used an LNA-enhanced microarray platform to study the global miRNA expression profiles in the six murine EAT cell lines, and identified growth-, hypoxia-, and resistance-specific miRNA patterns. Among the differentially expressed miRNAs, we found the two clusters miR-183∼miR-96∼miR-182 and miR-200b∼miR-200a∼miR-429 as well as miR-141 to be consistently upregulated in the MDR cell lines, while miR-125b-5p and the two clusters miR-30d∼miR-30b and miR-23b∼miR-27b∼miR-24-1 were downregulated in most of the resistant EAT cells. Several of the target genes for these miRNAs-including Zeb1/Zeb2 and members of the Fox gene family-could contribute to the drug-resistant phenotype, although we did not find that the degree of resistance was directly correlated to any specific changes in miRNA expression. Probably, the observed miRNA expression patterns reflect the underlying genomic instability of the tumor cells, and further studies are needed to explore how the highly complex regulatory miRNA networks contribute to the development of MDR.

A N-acetyl-D-galactosamine-binding lectin from Amaranthus gangeticus seeds inhibits biofilm formation and Ehrlich ascites carcinoma cell growth in vivo in mice.

AGL, a 15-kDa lectin from Amaranthus gangeticus seeds was isolated using ion-exchange and gel filtration chromatography. AGL contained 8.55% of neutral sugar and became specifically inhibited by N-acetyl-D-galactosamine. Hemagglutination activity of the lectin was maximum over the pH range of 4.0-6.0 and temperatures of 30-60 °C though it lost the activity when treated with urea and EDTA. With an LC50 value of 250 µg/ml, AGL showed mild toxicity against Artemia nauplii. It inhibited the growth of pathogenic bacteria like Shigella boydii, Shigella dysenteriae and Staphylococcus aureus when treated for 8 and 16 h, respectively, but lost the antibacterial activity during a 24 h treatment. AGL could not inhibit the growth of Escherichia coli and mitogenic growth (7.0-9.0%) was observed instead. AGL inhibited 37.14%, 65.71% and 82.85% of biofilm formation of Escherichia coli at the concentrations of 250, 500 and 1000 µg/ml, respectively. Marked inhibition of the proliferation of Ehrlich ascites carcinoma cells was determined when treated with various doses of AGL. AGL inhibited 65.89% and 81.25% of the in vivo growth of EAC cells in mice at the doses of 2.0 and 4.0 mg/kg/day, respectively. Significant alteration of the expression of apoptosis related genes Fas, NF-kB and MAPK were observed.

Anti-cancer activity of two novel heterocyclic compounds through modulation of VEGFR and miR-122 in mice bearing Ehrlich ascites carcinoma.

Metastasis in breast cancer is a leading cause of mortality among women in many countries. This study investigated the anti-cancer role of benzoimidazoquinazoline and benzimidazotriazin; two novel compounds that were designed, synthesized, structurally elucidated, and biologically evaluated as potent anti-angiogenic agents that act through inhibition of vascular endothelial growth factor receptor-2 (VEGFR2). Breast cancer was induced by inoculation of Ehrlich Ascites Carcinoma (EAC) cells. Seventy swiss albino mice were randomly divided into 7 groups, 10 animals each: (1) normal, (2) control EAC group, (3) cisplatin treated group, (4&5) benzoimidazoquinazoline treated (5 mg/kg and 10 mg/kg), (6&7) benzimidazotriazin treated (5 mg/kg and 10 mg/kg). The expression of miR-122 was assessed in the tumor tissue by quantitative PCR, and the VEGF level was determined in serum by ELISA. VEGFR2 and cluster of differentiation (CD)34 were assessed by immunohistochemistry. Serum ALT, AST, creatinine, and urea were measured. Treatment with benzoimidazoquinazoline and benzimidazotriazin decreased tumor weight and serum levels of VEGF, and down-regulated expression of VEGFR2 and CD34 in the tumor tissue. miR-122 was upregulated, particularly in the benzimidazotriazin (10 mg/kg) group. Relative to cisplatin, the novel compounds were less toxic to kidneys. Benzoimidazoquinazoline and benzimidazotriazin are promising anti-cancer agents that act through inhibition of angiogenesis and thus provide a new strategy for advancement of chemotherapy.

Dual-targeted therapeutic strategy combining CSC-DC-based vaccine and cisplatin overcomes chemo-resistance in experimental mice model.

BACKGROUND AND PURPOSE: Emerging evidence suggests that one of the main reasons of chemotherapy treatment failure is the development of multi-drug resistance (MDR) associated with cancer stem cells (CSCs). Our aim is to identify a therapeutic strategy based on MDR-reversing agents. MATERIALS AND METHODS: CSC-enriched Ehrlich carcinoma (EC) cell cultures were prepared by drug-resistant selection method using different concentrations of cisplatin (CIS). Cell cultures following drug exposure were analyzed by flow cytometry for CSC surface markers CD44+/CD24-. We isolated murine bone marrow-derived dendritic cells (DCs) and then used them to prepare CSC-DC vaccine by pulsation with CSC-enriched lysate. DCs were examined by flow cytometry for phenotypic markers. Solid Ehrlich carcinoma bearing mice were injected with the CSC-DC vaccine in conjunction with repeated low doses of CIS. Tumor growth inhibition was evaluated and tumor tissues were excised and analyzed by real-time PCR to determine the relative gene expression levels of MDR and Bcl-2. Histopathological features of tumor tissues excised were examined. RESULTS AND CONCLUSION: Co-treatment with CSC-DC and CIS resulted in a significant tumor growth inhibition. Furthermore, the greatest response of downregulation of MDR and Bcl-2 relative gene expression were achieved in the same group. In parallel, the histopathological observations demonstrated enhanced apoptosis and absence of mitotic figures in tumor tissues of the co-treatment group. Dual targeting of resistant cancer cells using CSC-DC vaccine along with cisplatin represents a promising therapeutic strategy that could suppress tumor growth, circumvent MDR, and increase the efficacy of conventional chemotherapies.

Therapeutic effect of Arthrocnemum machrostachyum methanolic extract on Ehrlich solid tumor in mice.

BACKGROUND: The anti-cancer effect of the halophyte Arthrocnemum indicum, a member of Arthrocnemum family of salt-tolerant plants, was evaluated against colorectal cancer cell, CaCo2. However, the anti-cancer effect of another halophyte Arthrocnemum machrostachyum was not investigated yet. Herein, the anticancer effect of A. machrostachyum methanolic extract (AME) was evaluated against Ehrlich solid tumor (EST) in mice and the potential mechanism of action was also studied. METHODS: Male Swiss albino mice (n = 28) were randomly divided into 4 groups (n = 7/group). Group 1 (negative control group); group 2 (EST) injected intramuscularly by 0.2 mL Ehrlich ascitic carcinoma (2 × 106 cells); and groups 3 and 4 injected intratumorally with AME (180 and 360 mg/kg body weight, respectively) at D12 trice weekly for 2 weeks. Gene expression, protein expression, DNA damage, and TNFa level in tumors were determined by real-time PCR, western blot, comet assay, and Elisa, respectively. RESULTS: Treatment with AME induced anti-tumor effects against EST as indicated by 1) notable reduction in tumor size; 2) elevation in tissue necrosis and apoptosis, as confirmed histologically; 3) increased DNA fragmentation; 4) decreased expression of the apoptotic genes (p53, Bax and caspase 3), and increased expression of the anti-apoptotic marker Bcl2; 5) significantly upregulated cell cycle regulatory genes Cdc2 and connexin26, and; 6) decreased TNFa levels in tumor tissues. Interestingly, a high dose of AME exhibited a more potent anti-tumor effect against EST. CONCLUSION: These findings indicate that AME has a potent antitumor effect against EST and could be used as an adjuvant to anticancer drugs to combat tumor, but after application of further confirmatory clinical trials.