Biomimetic-gasdermin-protein-expressing nanoplatform mediates tumor-specific pyroptosis for cancer immunotherapy.

Pyroptosis, mediated by gasdermin proteins, has shown excellent efficacy in facilitating cancer immunotherapy. The strategies commonly used to induce pyroptosis suffer from a lack of tissue specificity, resulting in the nonselective activation of pyroptosis and consequent systemic toxicity. Moreover, pyroptosis activation usually depends on caspase, which can induce inflammation and metabolic disorders. In this study, inspired by the tumor-specific expression of SRY-box transcription factor 4 (Sox4) and matrix metalloproteinase 2 (MMP2), we constructed a doubly regulated plasmid, pGMD, that expresses a biomimetic gasdermin D (GSDMD) protein to induce the caspase-independent pyroptosis of tumor cells. To deliver pGMD to tumor cells, we used a hyaluronic acid (HA)-shelled calcium carbonate nanoplatform, H-CNP@pGMD, which effectively degrades in the acidic endosomal environment, releasing pGMD into the cytoplasm of tumor cells. Upon the initiation of Sox4, biomimetic GSDMD was expressed and cleaved by MMP2 to induce tumor-cell-specific pyroptosis. H-CNP@pGMD effectively inhibited tumor growth and induced strong immune memory effects, preventing tumor recurrence. We demonstrate that H-CNP@pGMD-induced biomimetic GSDMD expression and tumor-specific pyroptosis provide a novel approach to boost cancer immunotherapy.

Pharmacological approaches to promote cell death of latent HIV reservoirs.

PURPOSE OF REVIEW: HIV requires lifelong antiviral treatment due to the persistence of a reservoir of latently infected cells. Multiple strategies have been pursued to promote the death of infected cells. RECENT FINDINGS: Several groups have focused on multipronged approaches to induce apoptosis of infected cells. One approach is to combine latency reversal agents with proapoptotic compounds and cytotoxic T cells to first reactivate and then clear infected cells. Other strategies include using natural killer cells or chimeric antigen receptor cells to decrease the size of the reservoir.A novel strategy is to promote cell death by pyroptosis. This mechanism relies on the activation of the caspase recruitment domain-containing protein 8 (CARD8) inflammasome by the HIV protease and can be potentiated by nonnucleoside reverse transcriptase inhibitors. SUMMARY: The achievement of a clinically significant reduction in the size of the reservoir will likely require a combination strategy since none of the approaches pursued so far has been successful on its own in clinical trials. This discrepancy between promising in vitro findings and modest in vivo results highlights the hurdles of identifying a universally effective strategy given the wide heterogeneity of the HIV reservoirs in terms of tissue location, capability to undergo latency reversal and susceptibility to cell death.

Curcumin inhibits the development of colorectal cancer via regulating the USP4/LAMP3 pathway.

In this study, we aimed to explore the effects of curcumin on the progression of colorectal cancer and its underlying mechanisms involved. Cell proliferation, apoptosis and invasion were determined through CCK-8 assay, colony formation assay, EdU assay, flow cytometry, and transwell invasion assay, respectively. The protein expression of Bax, MMP2, USP4 and LAMP3 was measured using western blot. Pearson correlation coefficient was used to evaluate the relationship between USP4 and LAMP3. Co-IP was also conducted to determine the interaction between USP4 and LAMP3. Xenograft tumor model was established to explore the role of curcumin in colorectal cancer in vivo. IHC was utilized to measure the expression of Bax, MMP2, USP4 and LAMP3 in tumor tissues from mice. Curcumin significantly accelerated cell apoptosis, and inhibited cell proliferation and invasion in LoVo and HCT-116 cells. LAMP3 was augmented in colorectal cancer tissues and cells, and curcumin could reduce the expression of LAMP3. Curcumin decreased LAMP3 expression to exhibit the inhibition role in the progression of colorectal cancer. USP4 interacted with LAMP3, and positively regulated LAMP3 expression in colorectal cancer cells. LAMP3 overexpression could reverse the suppressive effects of USP4 knockdown on the development of colorectal cancer. Curcumin downregulated USP4 to impeded the progression of colorectal cancer via repressing LAMP3 expression. In addition, curcumin obviously restrained tumor growth in mice through downregulating USP4 and LAMP3 expression. These data indicated that curcumin exert the anti-tumor effects on the development of colorectal cancer through modulating the USP4/LAMP3 pathway.

MLLT11 siRNA Inhibits the Migration and Promotes the Apoptosis of MDA-MB-231 Breast Cancer Cells.

Breast cancer is considered the most prevalent malignancy due to its high incidence rate, recurrence, and metastasis in women that makes it one of the deadliest cancers. The current study aimed to predict the genes associated with the recurrence and metastasis of breast cancer and to validate their effect on MDA-MB-231 cells. Through the bioinformatics analysis, the transcription factor 7 cofactor (MLLT11) as the target gene was obtained. MLLT11-specific siRNA was synthesized and transfected into MDA-MB-231 cells. The results demonstrated that the siRNA significantly reduced the MLLT11 mRNA levels. Moreover, cell migration and invasion, as well as the protein levels of phosphatidylinositol 3-kinase (PI3K), AKT, matrix metalloproteinase (MMP) 2, and MMP9, were significantly lower in the groups treated with siRNA while the apoptosis was augmented. Collectively, MLLT11 siRNA elicited ameliorative properties on breast cancer cells, possibly via the inhibition of the PI3K/AKT signaling pathway.

Tamoxifen resistance induction results in the upregulation of ABCG2 expression and mitoxantrone resistance in MCF-7 breast cancer cells.

Cancer endocrine therapy can promote evolutionary dynamics and lead to changes in the gene expression profile of tumor cells. We aimed to assess the effect of tamoxifen (TAM)-resistance induction on ABCG2 pump mRNA, protein, and activity in ER + MCF-7 breast cancer cells. We also evaluated if the resistance to TAM leads to the cross-resistance toward mitoxantrone (MX), a well-known substrate of the ABCG2 pump. The ABCG2 mRNA and protein expression were compared in MCF-7 and its TAM-resistant derivative MCF-7/TAMR cells using RT-qPCR and western blot methods, respectively. Cross-resistance of MCF-7/TAMR cells toward MX was evaluated by the MTT method. Flow cytometry was applied to compare ABCG2 function between cell lines using MX accumulation assay. ABCG2 mRNA expression was also analyzed in tamoxifen-sensitive (TAM-S) and tamoxifen-resistant (TAM-R) breast tumor tissues. The levels of ABCG2 mRNA, protein, and activity were significantly higher in MCF-7/TAMR cells compared to TAM-sensitive MCF-7 cells. MX was also less toxic in MCF-7/TAMR compared to MCF-7 cells. ABCG2 was also upregulated in tissue samples obtained from TAM-R cancer patients compared to TAM-S patients. Prolonged exposure of ER + breast cancer cells to the active form of TAM and clonal evolution imposed by the selective pressure of the drug can lead to higher expression of the ABCG2 pump in the emerged TAM-resistant cells. Therefore, in choosing a sequential therapy for a patient who develops resistance to TAM, the possibility of the cross-resistance of the evolved tumor to chemotherapy drugs that are ABCG2 substrates should be considered. Prolonged exposure of MCF-7 breast cancer cells to tamoxifen can cause resistance to it and an increase in the expression of the ABCG2 mRNA and protein levels in the cells. Tamoxifen resistance can lead to cross-resistance to mitoxantrone.

The CARD8 inflammasome in HIV infection.

The biggest challenge to immune control of HIV infection is the rapid within-host viral evolution, which allows selection of viral variants that escape from T cell and antibody recognition. Thus, it is impossible to clear HIV infection without targeting "immutable" components of the virus. Unlike the adaptive immune system that recognizes cognate epitopes, the CARD8 inflammasome senses the essential enzymatic activity of the HIV-1 protease, which is immutable for the virus. Hence, all subtypes of HIV clinical isolates can be recognized by CARD8. In HIV-infected cells, the viral protease is expressed as a subunit of the viral Gag-Pol polyprotein and remains functionally inactive prior to viral budding. A class of anti-HIV drugs, the non-nucleoside reverse transcriptase inhibitors (NNRTIs), can promote Gag-pol dimerization and subsequent premature intracellular activation of the viral protease. NNRTI treatment triggers CARD8 inflammasome activation, which leads to pyroptosis of HIV-infected CD4+ T cells and macrophages. Targeting the CARD8 inflammasome can be a potent and broadly effective strategy for HIV eradication.

Cytotoxicity of the methanol extracts and compounds of Brucea antidysenterica (Simaroubaceae) towards multifactorial drug-resistant human cancer cell lines.

BACKGROUND: Cancer remains a global health concern and constitutes an important barrier to increasing life expectancy. Malignant cells rapidly develop drug resistance leading to many clinical therapeutic failures. The importance of medicinal plants as an alternative to classical drug discovery to fight cancer is well known. Brucea antidysenterica is an African medicinal plant traditionally used to treat cancer, dysentery, malaria, diarrhea, stomach aches, helminthic infections, fever, and asthma. The present work was designed to identify the cytotoxic constituents of Brucea antidysenterica on a broad range of cancer cell lines and to demonstrate the mode of induction of apoptosis of the most active samples. METHODS: Seven phytochemicals were isolated from the leaves (BAL) and stem (BAS) extract of Brucea antidysenterica by column chromatography and structurally elucidated using spectroscopic techniques. The antiproliferative effects of the crude extracts and compounds against 9 human cancer cell lines were evaluated by the resazurin reduction assay (RRA). The activity in cell lines was assessed by the Caspase-Glo assay. The cell cycle distribution, apoptosis via propidium iodide (PI) staining, mitochondrial membrane potential (MMP) through 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining, and the reactive oxygen species (ROS) via 2´,7´-dichlorodihydrofluoresceine diacetate (H2DCFH-DA) staining, were investigated by flow cytometry. RESULTS: Phytochemical studies of the botanicals (BAL and BAS) led to the isolation of seven compounds. BAL and its constituents 3, (3-(3-Methyl-1-oxo-2-butenyl))1H indole (1) and hydnocarpin (2), as well as the reference compound, doxorubicin, had antiproliferative activity against 9 cancer cell lines. The IC50 values varied from 17.42 µg/mL (against CCRF-CEM leukemia cells) to 38.70 µg/mL (against HCT116 p53-/- colon adenocarcinoma cells) for BAL, from 19.11 µM (against CCRF-CEM cells) to 47.50 µM (against MDA-MB-231-BCRP adenocarcinoma cells) for compound 1, and from 4.07 µM (against MDA-MB-231-pcDNA cells) to 11.44 µM (against HCT116 p53+/+ cells) for compound 2. Interestingly, hypersensitivity of resistant cancer cells to compound 2 was also observed. BAL and hydnocarpin induced apoptosis in CCRF-CEM cells mediated by caspase activation, the alteration of MMP, and increased ROS levels. CONCLUSION: BAL and its constituents, mostly compound 2, are potential antiproliferative products from Brucea antidysenterica. Other studies will be necessary in the perspective of the discovery of new antiproliferative agents to fight against resistance to anticancer drugs.

ATP-binding cassette efflux transporters and MDR in cancer.

Of the many multidrug resistance (MDR) mechanisms, the ATP binding cassette (ABC) transporters expelling drug molecules out of cells is a major culprit in limiting the efficacy of present-day anticancer drugs. The present review offers an updated information on structure, function, and regulatory mechanisms of major MDR related ABC transporters such as P-glycoprotein, MRP1, BCRP and effect of modulators on their functions. An attempt has been made to provide a focused information on different modulators of ABC transporters so as utilize them in clinical practice for amelioration of emerging MDR crisis in cancer treatment. Finally, the importance ABC transporters as therapeutic targets has been discussed in the light of future strategic planning for translating the ABC transporter inhibitors in clinical practice.

The Novel 5-Fluorouracil Loaded Ruthenium-based Nanocarriers Enhanced Anticancer and Apoptotic Efficiency while Reducing Multidrug Resistance in Colorectal Cancer Cells.

Recently, nanocarriers have been made to eliminate the disadvantages of chemotherapeutic agents by nanocarriers. Nanocarriers show their efficacy through their targeted and controlled release. In this study, 5-fluorouracil (5FU) was loaded into ruthenium (Ru)-based nanocarrier (5FU-RuNPs) for the first time to eliminate the disadvantages of 5FU, and its cytotoxic and apoptotic effects on HCT116 colorectal cancer cells were compared with free 5FU. 5FU-RuNPs with a size of approximately 100 nm showed a 2.61-fold higher cytotoxic effect compared to free 5FU. Apoptotic cells were detected by Hoechst/propidium iodide double staining, and the expression levels of BAX/Bcl-2 and p53 proteins, in which apoptosis occurred intrinsically, were revealed. In addition, 5FU-RuNPs was also found to reduce multidrug resistance (MDR) according to BCRP/ABCG2 gene expression levels. When all the results were evaluated, the fact that Ru-based nanocarriers alone did not cause cytotoxicity proved that they were ideal nanocarriers. Moreover, 5FU-RuNPs did not show any significant effect on the cell viability of normal human epithelial cell lines BEAS-2B. Consequently, the 5FU-RuNPs synthesized for the first time may be ideal candidates for cancer treatment because they can minimize the potential drawbacks of free 5FU.

Bisphenol A interacts with DLGAP5 and regulates IL-6/JAK2/STAT3 signaling pathway to promote tumorigenesis and progression of osteosarcoma.

OBJECTIVE: It has been suggested that Bisphenol A (BPA), a high-production-volume industrial chemical, can accelerate the development of various type of cancers. However, the effect of BPA on osteosarcoma and the underlying mechanisms are yet to be established. Therefore, in this study we tried to explore the carcinogenic effects of BPA on osteosarcoma and the underlying mechanism. METHODS: SaOs-2 cancer cell line was used to treat with BPA at the doses of 0.1, 1, 10 µM DGLAP5 knockdown and overexpression methods were constructed by using adenovirus mediated transfection, and the functional analysis of DGLAP5 was investigated to evaluate the carcinogenic effect of BPA on osteosarcoma through DLGAP5. Xenograft and metastatic mouse model were used to evaluate in vivo experiments. RESULTS: In this study, BPA at 10 µM promoted the proliferation, migration and invasion in vitro, and accelerate the progression and metastasis in vivo. Also, exposure to BPA was associated with poor survival of osteosarcoma in mice. In addition, we observed that BPA at 10 µM significantly increased the expression of DGLAP5 in osteosarcoma. Silencing DGLAP5 could reverse the effect of BPA on proliferation, migration and invasion. Mechanically, BPA promoted IL-6, JAK2, and STAT3 expression and promoted tumor progression in an IL-6-dependent manner through up-regulation of DLGAP5. CONCLUSION: Our findings demonstrated that BPA could promote the proliferation, migration, invasion of osteosarcoma cells and related to poor survival in a mouse model. DLGAP5 is one of the most critical targets of BPA to act as a carcinogen through IL-6/JAK2/STAT3 signaling pathway.

Organic Cation Transporters are Involved in Fluoxetine Transport Across the Blood-Brain Barrier In Vivo and In Vitro.

BACKGROUND: The research and development of drugs for the treatment of central nervous system diseases faces many challenges at present. One of the most important questions to be answered is, how does the drug cross the blood-brain barrier to get to the target site for pharmacological action. Fluoxetine is widely used in clinical antidepressant therapy. However, the mechanism by which fluoxetine passes through the BBB also remains unclear. Under physiological pH conditions, fluoxetine is an organic cation with a relatively small molecular weight (<500), which is in line with the substrate characteristics of organic cation transporters (OCTs). Therefore, this study aimed to investigate the interaction of fluoxetine with OCTs at the BBB and BBB-associated efflux transporters. This is of great significance for fluoxetine to better treat depression. Moreover, it can provide a theoretical basis for clinical drug combination. METHODS: In vitro BBB model was developed using human brain microvascular endothelial cells (hCMEC/D3), and the cellular accumulation was tested in the presence or absence of transporter inhibitors. In addition, an in vivo trial was performed in rats to investigate the effect of OCTs on the distribution of fluoxetine in the brain tissue. Fluoxetine concentration was determined by a validated UPLC-MS/MS method. RESULTS: The results showed that amantadine (an OCT1/2 inhibitor) and prazosin (an OCT1/3 inhibitor) significantly decreased the cellular accumulation of fluoxetine (P <.001). Moreover, we found that N-methylnicotinamide (an OCT2 inhibitor) significantly inhibited the cellular uptake of 100 and 500 ng/mL fluoxetine (P <.01 and P <.05 respectively). In contrast, corticosterone (an OCT3 inhibitor) only significantly inhibited the cellular uptake of 1000 ng/mL fluoxetine (P <.05). The P-glycoprotein (P-gp) inhibitor, verapamil, and the multidrug resistance associated proteins (MRPs) inhibitor, MK571, significantly decreased the cellular uptake of fluoxetine. However, intracellular accumulation of fluoxetine was not significantly changed when fluoxetine was incubated with the breast cancer resistance protein (BCRP) inhibitor Ko143. Furthermore, in vivo experiments proved that corticosterone and prazosin significantly inhibited the brain-plasma ratio of fluoxetine at 5.5 h and 12 h, respectively. CONCLUSION: OCTs might play a significant role in the transport of fluoxetine across the BBB. In addition, P-gp, BCRP, and MRPs seemed not to mediate the efflux transport of fluoxetine.

The multifaceted roles of gasdermins in cancer biology and oncologic therapies.

The involvement of the Gasdermin (GSDM) protein family in cancer and other pathologies is one of the hottest topics in biomedical research. There are six GSDMs in humans (GSDMA, B, C, D, GSDME/DFNA5 and PJVK/DFNB59) and, except PJVK, they can trigger cell death mostly by pyroptosis (a form of lytic and pro-inflammatory cell death) but also other mechanisms. The exact role of GSDMs in cancer is intricate, since depending on the biological context, these proteins have diverse cell-death dependent and independent functions, exhibit either pro-tumor or anti-tumor functions, and promote either sensitization or resistance to oncologic treatments. In this review we provide a comprehensive overview on the multifaceted roles of the GSDMs in cancer, and we critically discuss the possibilities of exploiting GSDM functions as determinants of anti-cancer treatment and as novel therapeutic targets, with special emphasis on innovative GSDM-directed nano-therapies. Finally, we discuss the issues to be resolved before GSDM-mediated oncologic therapies became a reality at the clinical level.

PLGA Nanoparticles Co-encapsulating NY-ESO-1 Peptides and IMM60 Induce Robust CD8 and CD4 T Cell and B Cell Responses.

Tumor-specific neoantigens can be highly immunogenic, but their identification for each patient and the production of personalized cancer vaccines can be time-consuming and prohibitively expensive. In contrast, tumor-associated antigens are widely expressed and suitable as an off the shelf immunotherapy. Here, we developed a PLGA-based nanoparticle vaccine that contains both the immunogenic cancer germline antigen NY-ESO-1 and an α-GalCer analog IMM60, as a novel iNKT cell agonist and dendritic cell transactivator. Three peptide sequences (85-111, 117-143, and 157-165) derived from immunodominant regions of NY-ESO-1 were selected. These peptides have a wide HLA coverage and were efficiently processed and presented by dendritic cells via various HLA subtypes. Co-delivery of IMM60 enhanced CD4 and CD8 T cell responses and antibody levels against NY-ESO-1 in vivo. Moreover, the nanoparticles have negligible systemic toxicity in high doses, and they could be produced according to GMP guidelines. Together, we demonstrated the feasibility of producing a PLGA-based nanovaccine containing immunogenic peptides and an iNKT cell agonist, that is activating DCs to induce antigen-specific T cell responses.

Human Amnion Chorion Membrane Allografts in the Treatment of Chronic Diabetic Foot Ulcers: A Literature Review.

OBJECTIVE: To discuss human amnion chorion (placental) membrane allograft (HACMA) use for the treatment of chronic diabetic foot ulcers (DFUs) and to evaluate the effectiveness, cost, and product waste of this therapy. DATA SOURCES: PubMed, Cochrane, and OVID databases. STUDY SELECTION: Twenty-four articles pertaining to HACMA and DFUs published from 2016 to 2020 were selected. DATA EXTRACTION: The data collected included type of wound care product, study design, study size, baseline size of DFU, cost, product wastage, number of applications, and wound healing outcomes. DATA SYNTHESIS: Human amnion chorion membrane allografts in the treatment of chronic DFUs have led to a reduction in healing time and increased the overall percentage of healing, making them more effective in treating DFUs compared with standard of care. These products are offered in multiple sizes with various shelf lives and methods of storage, making them accessible, easy to use, less wasteful, and lower in cost compared with other commercially available products. Promising evidence demonstrates that HACMAs are beneficial in treating complex, high-grade DFUs with exposed tendon or bone. CONCLUSIONS: Human amnion chorion membrane allografts are effective in treating chronic DFUs with a greater percentage of complete wound closure and a reduction in healing time versus standard of care.

TMEM16A/ANO1 calcium-activated chloride channel as a novel target for the treatment of human respiratory syncytial virus infection.

INTRODUCTION: Human respiratory syncytial virus (HRSV) is a common cause of respiratory tract infections (RTIs) globally and is one of the most fatal infectious diseases for infants in developing countries. Of those infected, 25%-40% aged ≤1 year develop severe lower RTIs leading to pneumonia and bronchiolitis, with ~10% requiring hospitalisation. Evidence also suggests that HRSV infection early in life is a major cause of adult asthma. There is no HRSV vaccine, and the only clinically approved treatment is immunoprophylaxis that is expensive and only moderately effective. New anti-HRSV therapeutic strategies are therefore urgently required. METHODS: It is now established that viruses require cellular ion channel functionality to infect cells. Here, we infected human lung epithelial cell lines and ex vivo human lung slices with HRSV in the presence of a defined panel of chloride (Cl-) channel modulators to investigate their role during the HRSV life-cycle. RESULTS: We demonstrate the requirement for TMEM16A, a calcium-activated Cl- channel, for HRSV infection. Time-of-addition assays revealed that the TMEM16A blockers inhibit HRSV at a postentry stage of the virus life-cycle, showing activity as a postexposure prophylaxis. Another important negative-sense RNA respiratory pathogen influenza virus was also inhibited by the TMEM16A-specific inhibitor T16Ainh-A01. DISCUSSION: These findings reveal TMEM16A as an exciting target for future host-directed antiviral therapeutics.

Influence of HLDF Differentiation Factor on Nonspecific Invasive Breast Carcinoma in vitro.

The study was carried out on samples of invasive breast carcinoma of no special type from 36 patients aged 48.0 to 62.8 years. The effect of HLDF on nonspecific invasive breast carcinoma was a decrease in the relative content of low-differentiated cells and an increase in the relative content of highly differentiated cells. HLDF did not have a cytotoxic effect leading to the death of low-differentiated cells but promoted promotes the acquisition of a higher degree of differentiation by them. A more pronounced effect of HLDF was observed in more aggressive metastasizing forms of neoplasia, which allows us to consider this differentiation factor as a candidate for use in the differentiation therapy of malignant neoplasms.

LIMD2 Regulates Key Steps of Metastasis Cascade in Papillary Thyroid Cancer Cells via MAPK Crosstalk.

Although papillary thyroid carcinoma (PTC) has a good prognosis, 20-90% of patients show metastasis to regional lymph nodes and 10-15% of patients show metastasis to distant sites. Metastatic disease represents the main clinical challenge that impacts survival rate. We previously showed that LIMD2 was a novel metastasis-associated gene. In this study, to interrogate the role of LIMD2 in cancer invasion and metastasis, we used CRISPR-mediated knockout (KO) of LIMD2 in PTC cells (BCPAP and TPC1). Western blot and high-content screening (HCS) analysis confirmed functional KO of LIMD2. LIMD2 KO reduced in vitro invasion and migration. Ultrastructural analyses showed that cell polarity and mitochondria function and morphology were restored in LIMD2 KO cells. To unveil the signals supervising these phenotypic changes, we employed phospho-protein array. Several members of the MAPK superfamily showed robust reduction in phosphorylation. A Venn diagram displayed the overlap of kinases with reduced phosphorylation in both cell lines and showed that they were able to initiate or sustain the epithelial-mesenchymal transition (EMT) and DNA damage checkpoint. Flow cytometry and HCS validation analyses further corroborated the phospho-protein array data. Collectively, our findings show that LIMD2 enhances phosphorylation of kinases associated with EMT and invasion. Through cooperation with different kinases, it contributes to the increased genomic instability that ultimately promotes PTC progression.

Chronic exposure of human glioblastoma tumors to low concentrations of a pesticide mixture induced multidrug resistance against chemotherapy agents.

Recent evidence indicates that chronic, low-dose exposure to mixtures of pesticides can cause adverse responses in a variety of cells, tissues and organs, although interactions between pesticides circulating in the blood and cancer cells remain largely unexplored. The aim of this study was to investigate the potential of a mixture of four pesticides to induce multidrug resistance against the chemotherapeutic agents cisplatin, 5-fluorouracil and temozolomide in the human U87 glioblastoma cell line, and to explore the molecular mechanisms underlying this resistance. We found that the repeated administration of the pesticide mixture (containing the insecticides chlorpyrifos-ethyl and deltamethrin, the fungicide metiram, and the herbicide glyphosate) induced a strong drug resistance in U87 cells. The resistance was durable and transferred to subsequent cell generations. In addition, we detected a significant over-expression of the ATP-binding cassette (ABC) membrane transporters P-gp/ABCB1 and BRCP/ABCG2 as well as a glutathione-S-transferase (GST)/M1-type cellular detoxification function, known to have important roles in multidrug resistance, thus providing molecular support for the acquired multidrug resistance phenotype and shedding light on the mechanism of resistance. We further determined that there was lower mortality in the resistant brain tumor cells and that the mitochondrial apoptosis pathway was activated at a lower rate after chemotherapy compared to non-resistant control cells. In addition, multidrug-resistant cells were found to have both higher motility and wound-healing properties, suggesting a greater metastatic potential. Our results suggest that the investigation of P-gp, BRCP and GST/M1 multidrug resistance gene expression and/or protein levels in biopsy specimens of brain tumor patients who were at risk of pesticide exposure could be beneficial in determining chemotherapy dose and prolonging patient survival.

Preparation of a new combination nanoemulsion-encapsulated MAGE1-MAGE3-MAGEn/HSP70 vaccine and study of its immunotherapeutic effect.

BACKGROUND: MAGE family genes have been studied as targets for tumor immunotherapy for a long time. Here, we combined MAGE1-, MAGE3- and MAGEn-derived peptides as a cancer vaccine and tested whether a new combination nanoemulsion-encapsulated vaccine could be used to inhibit the growth of tumor cells in humanized SCID mice. METHODS: The nanoemulsion-encapsulated complex protein vaccine (MAGE1, MAGE3, and MAGEn/HSP70 fusion protein; M1M3MnH) was prepared using a magnetic ultrasonic technique. After screening, human PBMCs were injected into SCID mice to mimic the human immune system. Then, the humanized SCID mice were challenged with M3-HHCC cells and immunized with nanoemulsion-encapsulated MAGE1-MAGE3-MAGEn/HSP70 [NE(M1M3MnH)] or M1M3MnH. The cellular immune responses were detected by IFN-γ ELISPOT and cytotoxicity assays. Therapeutic and tumor challenge experiments were also performed. RESULTS: The results showed that the immune responses elicited by NE(M1M3MnH) were apparently stronger than those elicited by M1M3MnH, NE(-) or PBS, suggesting that this novel nanoemulsion carrier induces potent antitumor immunity against the encapsulated antigens. The results of the therapeutic and tumor challenge experiments also indicated that the new vaccine had a definite effect on SCID mice bearing human hepatic cancer. CONCLUSION: Our study indicated that the combination of several tumor antigen-derived peptides may be a relatively good strategy for peptide-based cancer immunotherapy. These results suggest that the complex nanoemulsion vaccine could have broader applications for both therapy and prevention mediated by antitumor effects in the future.

Role of ATP-Binding Cassette Transporter Proteins in CNS Tumors: Resistance- Based Perspectives and Clinical Updates.

Despite gigantic advances in medical research and development, chemotherapeutic resistance remains a major challenge in complete remission of CNS tumors. The failure of complete eradication of CNS tumors has been correlated with the existence of several factors including overexpression of transporter proteins. To date, 49 ABC-transporter proteins (ABC-TPs) have been reported in humans, and the evidence of their strong association with chemotherapeutics' influx, dissemination, and efflux in CNS tumors, is growing. Research studies on CNS tumors are implicating ABC-TPs as diagnostic, prognostic and therapeutic biomarkers that may be utilised in preclinical and clinical studies. With the current advancements in cell biology, molecular analysis of genomic and transcriptomic interplay, and protein homology-based drug-transporters interaction, our research approaches are streamlining the roles of ABC-TPs in cancer and multidrug resistance. Potential inhibitors of ABC-TP for better clinical outcomes in CNS tumors have emerged. Elacridar has shown to enhance the chemo-sensitivity of Dasatanib and Imatinib in various glioma models. Tariquidar has improved the effectiveness of Temozolomide's in CNS tumors. Although these inhibitors have been effective in preclinical settings, their clinical outcomes have not been as significant in clinical trials. Thus, to have a better understanding of the molecular evaluations of ABC-TPs, as well as drug-interactions, further research is being pursued in research labs. Our lab aims to better comprehend the biological mechanisms involved in drug resistance and to explore novel strategies to increase the clinical effectiveness of anticancer chemotherapeutics, which will ultimately improve clinical outcomes.