Opium use and risk of lung cancer: A multicenter case-control study in Iran.

Opium use was recently classified as a human carcinogen for lung cancer by the International Agency for Research on Cancer. We conducted a large, multicenter case-control study evaluating the association between opium use and the risk of lung cancer. We recruited 627 cases and 3477 controls from May 2017 to July 2020. We used unconditional logistic regression analyses to estimate the odds ratios (OR) and 95% confidence intervals (CI) and measured the association between opium use and the risk of lung cancer. The ORs were adjusted for the residential place, age, gender, socioeconomic status, cigarettes, and water pipe smoking. We found a 3.6-fold risk of lung cancer for regular opium users compared to never users (95% CI: 2.9, 4.6). There was a strong dose-response association between a cumulative count of opium use and lung cancer risk. The OR for regular opium use was higher for small cell carcinoma than in other histology (8.3, 95% CI: 4.8, 14.4). The OR of developing lung cancer among opium users was higher in females (7.4, 95% CI: 3.8, 14.5) than in males (3.3, 95% CI: 2.6, 4.2). The OR for users of both opium and tobacco was 13.4 (95% CI: 10.2, 17.7) compared to nonusers of anything. The risk of developing lung cancer is higher in regular opium users, and these results strengthen the conclusions on the carcinogenicity of opium. The association is stronger for small cell carcinoma cases than in other histology.

Opium use and risk of colorectal cancer: a multi-center case-referent study in Iran.

BACKGROUND: Opium use has been associated with an increased risk of cancers of the lung, oesophagus, and pancreas, and it was recently classified by the International Agency for Cancer Research as carcinogenic to humans. It is not clear whether opium also increases the risk of colorectal cancer (CRC). The aim of our study was to assess the association between various metrics of opium use and the risk of CRC. METHODS: This case-referent study from seven provinces in Iran comprised 848 CRC cases and 3215 referents. Data on opium use (duration, amount, frequency) and potential confounders were collected by trained interviewers. Multivariable unconditional logistic regression models were used to measure odds ratios (OR) adjusted for age, gender, province, marital status, family history of CRC-linked cancers, consumption of red meat, fruits and vegetables, body shape, occupational physical activity, and socioeconomic status. RESULTS: Regular opium consumption was not associated with the risk of CRC (OR 0.9, 95% confidence interval, CI: 0.7, 1.2) compared to subjects who never used opium. However, frequent opium use more than twice a day was associated with an increased risk of CRC compared to non-users of opium (OR: 2.0, 95% CI: 1.1, 3.8; p for quadratic trend 0.008). CONCLUSION: There seems to be no overall association between opium use and CRC, but the risk of CRC might be increased among persons who use opium many times a day.

Mutations in the S gene of hepatitis B virus in three generations of patients with chronic hepatitis B.

Failure to neutralize HBsAg and subsequent escape from the host immune system may be caused by HBsAg mutations, particularly in the "a" determinant, which alters the antigenicity of the protein. The purpose of this study was to examine the frequency of S gene mutations in three generations of HBV cases in northeastern Iran. In this study, 90 patients with chronic HBV were assigned to three groups according to the inclusion criteria. The plasma were utilized to extract viral DNA, and the PCR was applied. Direct sequencing and alignment were performed on the S gene, using reference sequence. The results indicated that all HBV genomes were categorized as the genotype D/ayw2. Among 79 point mutations detected, 36.8% were silent, and 56.2% were missense. In the S region, mutations were observed in 88.9% of CHB subjects studied. In the three-generation group, 21.5% of mutations were in the "a" determinant, and 2.6%, 19.5%, and 87.0% of these mutations were observed in antigenic epitopes of CTLs, CD4+, and B cells, respectively. In addition, 56.7% of mutations occurred at Major Hydrophilic Region. S143L and G145R mutations which the most prevalent in the three-generation (36.7%, 20%), and two-generation (42.5%, 20%) groups, related to the failure of HBsAg detection, vaccine, and immunotherapy escape. The findings showed that most of the mutations were concentrated in the B cell epitope. Most CHB cases from the three-generation, especially grandmothers, had HBV S gene mutations and subsequent amino acid mutations, suggesting that these mutations may be critical for pathogenesis and vaccine evasion.

Opium use and the risk of head and neck squamous cell carcinoma.

Scant evidence exists to support the association of opium use with head and neck cancer, limited to the larynx and oral cavity. In a multicenter case-control study-Iran Opium and Cancer study, we recruited 633 cases of head and neck squamous cell carcinoma (HNSCC) (254 lip and oral cavity, 54 pharynx, 327 larynx and 28 other subsites within the head and neck) and 3065 frequency-matched controls from April 2016 to April 2019. Odds ratios (ORs) for opium use and 95% confidence intervals (95% CIs) were obtained using mixed-effects logistic regression because of heterogeneity among centers. The adjusted OR (95% CI) for regular opium use was 3.76 (2.96-4.79) for all HNSCC combined. Strong dose-response effects were observed by frequency or amount of use, and duration of use. Regular opium uses significantly increased the risk of HNSCC of the pharynx, larynx and other subsites within the head and neck with OR (95% CI) of 2.90 (1.40-6.02), 6.55 (4.69-9.13) and 5.95 (2.41-14.71), respectively. The observed associations were significant even among never tobacco smokers (including cigarette and water-pipe smoking). Moreover, by the multiplicative interaction scale, the effect of opium use could be varied by cigarette smoking on HNSCC, 8.16 (6.20-10.74). For the first time, the current study showed opium users have an increased risk of several anatomic subsites of HNSCC.

Functional variation (Q63R) in the cannabinoid CB2 receptor may affect the severity of COVID-19: a human study and molecular docking.

Evidence supports a role of host genetic diversity in the clinical course of coronavirus disease 2019 (COVID-19). Variation in the cannabinoid CB2 receptor gene (CNR2) could affect the regulatory action of endocannabinoids on the immune system, resulting in an increased risk of various inflammatory diseases. The present study investigated the relationship between the CNR2-Q63R variant and COVID-19 severity. A total of 200 Iranian COVID-19 patients were enrolled in the study and genotyped using a TaqMan assay. The co-dominant, dominant, recessive, over-dominant, and additive inheritance models were analyzed using SNPStats software. In silico molecular docking was also performed to simulate the effects of the Q63R variation on CB2 binding with a ligand and with the G-protein. A significant difference in the Q63R allele and genotype distribution was found between expired and discharged COVID-19 patients in co-dominant, recessive, and additive inheritance models. The molecular docking results showed that the predicted structure of mutant CB2 (63R type) could not bind to the G-protein in the correct position. The data indicated that the Q63R variation in the CNR2 gene may affect the severity of COVID-19. Identification of genes related to susceptibility and severity of COVID-19 may lead to specific targets for drug repurposing or development.

Opium use reporting error in case-control studies: neighborhood controls versus hospital visitor controls.

Background: There are relatively scant data to determine whether hospital visitors could serve as a proper source of controls in case-control studies of illicit drug use. The aim of this study was to evaluate using neighborhood versus hospital visitor controls in reporting opium use. Methods: We used data from 2 independent case-control studies of cancer in Iran. In the first study, controls were selected from neighborhoods of the patients. For the second one, controls were selected from among hospital visitors. In the latter study, hospital visitors were companions of the patients or others visiting the hospital for reasons other than disease treatment. We used stata (version 12; Stata Corp( for all analyses and with a significance level of 0.05. Results: Data from 616 of neighborhood controls and 414 of hospital visitor controls were analyzed. Opium point prevalence among men was significantly higher in hospital visitors than neighborhood controls (43.3% vs 32.2%; P = 0.047), while the prevalence of cigarette smoking was very similar in both control groups (46.3% vs 47.2%; P =.847). Using a logistic regression analysis, in an unadjusted analysis, neighborhood controls were less likely to report opium use in both genders, with (unadjusted OR = 0.77; 95% CI: 0.59,1). After adjusting for potential confounders, the differences of opium use between the 2 control groups became more pronounced (Adjusted OR = 0.26; 95% CI: 0.10, 0.69). Conclusion: Because of the similarity of reporting cigarette smoking among neighborhood controls but substantially lower reporting of opium use among them, we concluded that neighborhood controls underreport opium use-a sensitive question- and that using neighborhood control biases the findings in case-control studies. Hospital visitor controls may be more appropriate than neighborhood controls for case-control studies of illicit drugs.

Effects of microRNAs polymorphism in cancer progression.

MicroRNAs (miRNAs) are known as a new class of small RNAs (18-25 nucleotides) that regulate gene expression at multiple levels from transcription to translation. Considering the important role of miRNAs in cell proliferation, differentiation, and apoptosis, any variations in their expression can contribute to various anomalies, such as tumorigenesis. Single-nucleotide polymorphisms (SNPs) have received much attention as potential genetic markers for diseases due to their advantage of being present at a high frequency in the human genome. SNPs can occur in different parts of the miRNA genes (primary, precursor, and mature) which result in pathological conditions. In this study, recent findings related to the effects of SNPs in miRNAs on their biogenesis and functions and their role in cancer development and progression are discussed. This review was performed using PubMed to search for related reports. The identified effects may be useful for clinical decision-making and providing important new information about the pathophysiology of miRNAs.

Increased expression of miR-146a, miR-10b, and miR-21 in cancer stem-like gastro-spheres.

BACKGROUND: Gastric cancer remains one of the leading causes of cancer-associated mortalities globally. Accumulating evidence support the presence of gastric cancer stem cells (CSCs) and their role in the pathogenesis and therapeutic challenges of gastric cancer. MicroRNAs (miRNAs) may be influenced by the cellular differentiative state and as critical regulators of the cellular fate in development and cancer, can modulate the behavior of CSCs too. Here, we aimed to investigate the expression relevance of three prognostic miRNAs (miR-21, miR-10b, and miR-146a) in CSCs of AGS and MKN-45 gastric cancer cell lines. METHODS: Serial sphere-forming assay in serum-free culture medium was used to enrich the cellular population with stem-like properties. Gastro-spheres were characterized by evaluating the stemness gene expression, clonogenicity, and resistance to docetaxel and cisplatin in comparison with their parental cells. The expression level of miRNAs in gastro-spheres and their parental cells was measured using quantitative reverse transcription polymerase chain reaction. RESULTS: Gastro-spheres from both cell lines exhibit stem-like properties: upregulated stemness associated genes (P < 0.05), more colonogenicity and more resistance to docetaxel (P < 0.05). MKN-45 gastro-spheres exhibited upregulated expression of miR-21 (1.8-folds), miR-10b (1.34-folds) and miR-146a (4.8-folds; P < 0.05) compared with the parental cells. AGS-derived gastro-spheres showed upregulation of miR-21 (4.7-folds; P < 0.01), miR-10b (15.2-folds; P < 0.001) and miR-146a (39.3-folds; P < 0.05). CONCLUSION: Our data exhibited upregulation of miR-21, miR-10b, and miR-146a in the stem-like gastro-spheres; however; their function in gastric CSCs remains to be verified by further experiments.

Inhibition of H1N1 influenza virus infection by zinc oxide nanoparticles: another emerging application of nanomedicine.

BACKGROUND: Currently available anti-influenza drugs are often associated with limitations such as toxicity and the appearance of drug-resistant strains. Therefore, there is a pressing need for the development of novel, safe and more efficient antiviral agents. In this study, we evaluated the antiviral activity of zinc oxide nanoparticles (ZnO-NPs) and PEGylated zinc oxide nanoparticles against H1N1 influenza virus. METHODS: The nanoparticles were characterized using the inductively coupled plasma mass spectrometry, x-ray diffraction analysis, and electron microscopy. MTT assay was applied to assess the cytotoxicity of the nanoparticles, and anti-influenza activity was determined by TCID50 and quantitative Real-Time PCR assays. To study the inhibitory impact of nanoparticles on the expression of viral antigens, an indirect immunofluorescence assay was also performed. RESULTS: Post-exposure of influenza virus with PEGylated ZnO-NPs and bare ZnO-NPs at the highest non-toxic concentrations could be led to 2.8 and 1.2 log10 TCID50 reduction in virus titer when compared to the virus control, respectively (P < 0.0001). At the highest non-toxic concentrations, the PEGylated and unPEGylated ZnO-NPs led to inhibition rates of 94.6 and 52.2%, respectively, which were calculated based on the viral loads. There was a substantial decrease in fluorescence emission intensity in viral-infected cell treated with PEGylated ZnO-NPs compared to the positive control. CONCLUSIONS: Taken together, our study indicated that PEGylated ZnO-NPs could be a novel, effective, and promising antiviral agent against H1N1 influenza virus infection, and future studies can be designed to explore the exact antiviral mechanism of these nanoparticles.

M51R and Delta-M51 matrix protein of the vesicular stomatitis virus induce apoptosis in colorectal cancer cells.

Colorectal cancer (CRC) is the third most common cancer in both men and women. Oncolytic viral-based therapy methods seem to be promising for CRC treatment. Vesicular stomatitis virus (VSV) is considered as a potent candidate in viral therapy for several tumors. VSV particles with mutated matrix (M) protein are capable of initiating cell death cascades while not being harmful to the immune system. In the current study, the effects of the VSV M-protein was investigated on the apoptosis of the colorectal cancer SW480 cell. Wild-type, M51R, and ΔM51 mutants VSV M-protein genes were cloned into the PCDNA3.1 vector and transfected into the SW480 cells. The results of the MTT assay, Western blotting, and Caspase 3, 8, and 9 measurement, illustrated that both wild and M51R mutant M-proteins can destroy the SW480 colorectal cancer cells. DAPI/TUNEL double-staining reconfirmed the apoptotic effects of the M-protein expression. The ΔM51 mutant M-protein is effective likewise M51R, somehow it can be considered as a safer substitution.

Association of socioeconomic status with consumption of cigarettes, illicit drugs, and alcohol.

Introduction and Aims: Socioeconomic Status (SES) is considered as one of the important factors associated with use of various drugs. The present study aimed to investigate the effect of SES on cigarette smoking, alcohol use, drug use, and passive exposure to opium and cigarette smoke. Design and Methods: In this study, which is part of a multicenter case-control study, the research hypothesis was checked among controls who had referred to hospitals. Data were collected through a questionnaire and laboratory tests to determine the actual consumers of opium and other illicit drugs. Then, the data were analyzed using STATA 13. Result: This study was performed on 364 individuals within the age range of 30 to 75 years. More than 55% of the participants had a history of life-time consumption of cigarettes and hookah as well as alcohol and drugs. The results revealed an inverse relationship between SES and life-time consumption of hookah and alcohol. Furthermore, individuals with higher SES were more likely to deny their drug use. Discussion and Conclusions: The results revealed little robust evidence supporting the assumption that SES level can have an important effect on illicit drug use. On the other hand, the participants' characteristics could have a prominent effect on precise evaluation of the relationship between SES and drug use. Further multicenter studies are needed with samples diversified in terms of age and ethnicity to identify these confounding relationships.

The First Characterization of HIV-1 Subtypes and Drug Resistance Mutations among Antiretrovirally Treated Patients in Kermanshah, Iran.

BACKGROUND: Insufficient therapy during HIV-1 replication can promote the emergence of drug-resistant strains, reduce the effectiveness of antiretroviral treatment (ART), and increase the likelihood of the onward transmission of drug-resistant viruses. We characterized, for the first time, the prevalence of HIV-1 subtypes and drug resistance mutations in a western region of Iran. METHODS: This study was conducted among 122 patients on ART at a major referral center in Kermanshah, Iran. Nested PCR was performed using RT gene-specific primers from the pol gene. Sequencing was followed by amplification and purification of the desired sequence. Subtypes and mutations were determined using the Stanford HIV Drug Resistance Database. RESULTS: Most patients (92.6%) had subtype CRF 35-AD; 7.4% had subtype B. In total, 36.1% of the patients had at least 1 mutation associated with resistance RT inhibitors. The greatest rates of high-level resistance were observed for nevirapine (21.3%) and efavirenz (19.7%). CONCLUSIONS: Our results showed a high prevalence of drug resistance mutations in strains isolated from patients on treatment. At our center, we therefore recommend that genotyping be performed. This would allow the physician to prescribe appropriate drugs, reduce treatment costs, and increase the longevity and quality of life of patients.

Frequent pUL27 Variations in HIV-Infected Patients.

OBJECTIVE: Drug-resistant isolates of human cytomegalovirus (HCMV) have led to the development of new anti-HCMV drugs. Maribavir (MBV) is a novel inhibitor of the HCMV viral kinase. Resistance to MBV is mapped to gene UL27, a viral nuclear protein. In this study, we investigated UL27 polymorphisms in MBV-naive HIV-positive and HCMV congenitally infected clinical samples. METHODS: DNA was extracted from 20 CMV-positive HIV (9/20) and congenitally infected (11/20) patients and used for UL27 polymerase chain reaction amplification. Sanger sequencing and multiple sequence alignment of products was performed. RESULTS: K90 was the most prevalent polymorphism in both HIV-positive and congenitally infected patients. Polymorphisms Q54, D123, and R107 (10%) were seen in more than one sample. There were significantly more polymorphisms in the HIV-positive samples (p = 0.038). CONCLUSION: HCMV pUL27 is highly variable in adult immunocompromised HIV-positive patients.

Antiretroviral drug resistance mutations in naïve and experienced patients in Shiraz, Iran, 2014.

Resistance to antiretroviral agents is a significant concern in the clinical management of HIV-infected individuals, particularly in areas of the world where treatment options are limited. In this study, we aimed to identify HIV drug-resistance-associated mutations in 40 drug-naïve patients and 62 patients under antiretroviral therapy (ART) referred to the Shiraz HIV/AIDS Research Center - the first such data available for the south of Iran. HIV reverse transcriptase and protease genes were amplified and sequenced to determine subtypes and antiretroviral- resistance-associated mutations (RAMs). Subtype CRF35-AD recombinant was the most prevalent in all patients (98 of 102, 96 %), followed by subtype A1, and subtype B (one each, 2 %). Among the 40 ART-naïve patients, two mutations associated with nucleoside reverse transcriptase inhibitor (NRTI) resistance (two with Y115F and T215I) and three associated with non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance (two with G190S and Y181C, four with V179T) were found. Among ART-experienced patients, four mutations associated with resistance to NRTI, four with NNRTI, and five with protease inhibitors (PI) were found. Twenty patients with high levels of resistance were already on second-line therapy. We document for the first time in this region of Iran high levels of ART resistance to multiple drugs. Our findings call for more vigilant systematic ART resistance surveillance, increased resistance testing, careful management of patients with existing regimens, and strong advocacy for expansion of available drugs in Iran.

Antitumor effect of therapeutic HPV DNA vaccines with chitosan-based nanodelivery systems.

BACKGROUND: Cervical cancer is the second-most-common cause of malignancies in women worldwide, and the oncogenic activity of the human papilloma virus types (HPV) E7 protein has a crucial role in anogenital tumors. In this study, we have designed a therapeutic vaccine based on chitosan nanodelivery systems to deliver HPV-16 E7 DNA vaccine, considered as a tumor specific antigen for immunotherapy of HPV-associated cervical cancer. We have developed a Nano-chitosan (NCS) as a carrier system for intramuscular administration using a recombinant DNA vaccine expressing HPV-16 E7 (NCS-DNA E7 vaccine). NCS were characterized in vitro for their gene transfection ability. RESULTS: The transfection of CS-pEGFP NPs was efficient in CHO cells and the expression of green fluorescent proteins was well observed. In addition, NCS-DNA E7 vaccine induced the strongest E7-specific CD8+ T cell and interferon γ responses in C57BL/6 mice. Mice vaccinated with NCS-DNA E7 vaccine were able to generate potent protective and therapeutic antitumor effects against challenge with E7-expressing tumor cell line, TC-1. CONCLUSIONS: The strong therapeutic effect induced by the Chitosan-based nanodelivery suggest that nanoparticles may be an efficient carrier to improve the immunogenicity of DNA vaccination upon intramuscular administration and the platform could be further exploited as a potential cancer vaccine candidate in humans.

Enhanced cell immune responses to hepatitis C virus core by novel heterologous DNA prime/lambda nanoparticles boost in mice.

Hepatitis C virus (HCV) is a worldwide problem which does not have an effective vaccine and more than 170 million people worldwide are chronically infected by HCV. T cell responses are associated with spontaneous clearance of HCV infection. We report here the development of recombinant Lambda bacteriophage nanoparticles encoding HCV Core antigen. The aim of this study was to investigate the antigen-specific immune responses triggered in mice by different prime-boost combinations of DNA and Lambda phage nanoparticles encoding the HCV Core. The homologous prime/boost with recombinant Lambda nanoparticles induced higher levels of cellular and humoral immune response than the DNA vaccines. However, a heterologous prime/boost of HCV Core protein, using DNA vaccine priming followed by Lambda boost, induced highest level of lymphocyte proliferation, CD8 lymphocytes with cytotoxic function, and shifting the immune response toward a T helper (Th1) pattern and in overall improved immunity. Our study provides a new, safe, and effective vaccine for the prime-boost regimen which augments robust immunity and highlights novel promising strategies in HCV vaccine development.

Molecular identification of neuraminidase gene mutations in influenza A/H1N1 and A/H3N2 isolates of Mazandaran province, north of Iran.

OBJECTIVES: The neuraminidase (NA) mutations causing resistance to NA inhibitors (NAIs) mostly compromise the fitness of influenza viruses. Considering the importance of these mutations, constant monitoring of the effectiveness of available drugs is critical. This study aimed to identify NA mutations in the influenza A/H1N1 and A/H3N2 subtypes in the samples of Mazandaran, Iran from 2016 to 2020. METHODS: In this cross-sectional study, 20 influenza A/H1N1 and 20 influenza A/H3N2 samples were included in the study. After design of appropriate primers for NA gene, all samples subjected to RT-PCR and electrophoresis. Then the PCR product was sequenced to determine the mutations. RESULTS: In the present study, no oseltamivir resistance-related mutations were detected. Still, NA gene showed variations compared to the vaccine strains. In A/H1N1, a total of 43 mutations were detected. Similarly, in A/H3N2, a total of 66 mutations were observed. In all isolates of H1N1, N200S, N248D and I321V mutations were detected in the antigenic site of NA protein, which can affect vaccine incompatibility and virus escape from the host's immune system. Also, H150R mutation was observed in the NA active site of H3N2, which is the cause of agglutination by NA protein. Also, S245N mutation was identified as a new N-Glycosylation site of H3N2 subtype. CONCLUSIONS: The study of NA gene sequences revealed no oseltamivir resistance mutations. In H1N1 isolates, ca. 97% identities and in the H3N2 subtype, 96% identities were observed compared to reference isolate of 2009, which indicates the importance of constant monitoring of the emergence of the drug resistance mutations.

Association of HLADQ-B1 polymorphisms in three generations of chronic hepatitis B patients.

The presence of polymorphisms in the human leukocyte antigen (HLA)-DQB1 gene, along with its expression, has been demonstrated to be correlated with spontaneous clearance and susceptibility to HBV infection. The present study aimed to evaluate the possible role of genetic polymorphisms in HLA-DQB1 in three generations of patients with chronic hepatitis B (CHB). Based on the inclusion criteria, 90 CHB patients, 18 individuals recovered from HBV infection, and 40 healthy subjects were chosen. The DNA contents of the whole blood samples were extracted in order to perform HLA-DQB1 typing by the PCR technique. Besides whole blood samples, sera were applied to measure liver function tests (LFTs), as well as the titers of anti-HDV and anti-HCV. Also, in all CHB patients were measured liver stiffness (LSM) by Fibro Scan. The results of HLA-DQB1 polymorphisms (rs2856718 and rs7453920) demonstrated that the majority of polymorphisms in CHB patients were HLA-DQB1*03, HLA-DQB1*05, HLA-DQB1*04:01 and HLA-DQB1*03:01 that associated with HBV persistence and chronicity. Among the patients who showed these polymorphisms, the mean±SD, LSM was 4±1.57 KPa and most of them, F grade was reported as F2, which was a sign of disease progression towards chronicity. HLA polymorphisms imputation revealed that HLA-DQB1*06:04 (3.4%, P-Value= 0.2) was detected only in healthy subjects as protective polymorphism, while the allele HLA-DQB1*03:03 was reported in both healthy subjects (P-Value= 0.06) and recovered patients (P-Value= 0.1) as suppressor of CHB formation. The allele HLA-DQB1*05:02 was found in both healthy subjects (3.4%) and CHB patients (4.5%) which was associated with risk to liver cirrhosis (P-Value= 0, OR: 0.002 0.95CI: 0.000-0.15). HLA polymorphism analysis indicated that 17.39% of patients who were seropositive for anti-HCV carried the HLA-DQB1*03:01. HBV resistance or infection risk could be assessed by DBQ1 typing. The existence of polymorphisms in HLA gene could influence the clearance (HLA-DQB1*03:03) or susceptibility and persistence of infection (HLA-DQB1*03, HLA-DQB1*05, HLA-DQB1*04:01 and HLA-DQB1*03:01). These results have the potential to improve personalized therapy and prognosis for HBV infection.

NSP4 as adjuvant for immunogenicity and design of effective therapeutic HPV16 E6/E7/L1 DNA vaccine in tumor-bearing and healthy C57BL/6 mice.

INTRODUCTION: In humans, approximately 5% of all cancers are attributable to HPV infection. Prophylactic vaccines can inhibit viral migration and persistence. However, further studies are still required to develop such treatments. To achieve this goal, we designed a therapeutic HPV DNA vaccine encoding a construct of E6/E7/L1 and used NSP4 antigen as an adjuvant to assess the efficiency of this construct in generating antigen-specific antitumor immune responses. MATERIALS AND METHODS: Sixty female C57BL/6 mice (6-8 weeks old) were purchased from the Institute Pasteur of Iran. Through a subcutaneous (s.c) injection of a suspension of 100 µl PBS containing 106 TC-1 cells/mouse in the back side, 30 of them became cancerous, while 30 of them were healthy control mice. To amplify E6/E7/L1-pcDNA3 and NSP4-pcDNA3, the competent cells of DH5α and to generate a tumor, TC-1 cell line was used. Mice were then immunized with the HPV DNA vaccine. Cell proliferation was assessed by MTT assay. Finally, cytokine responses (IL-4, IL-12, IFN- γ) were measured in the supernatant of mice spleen cells. RESULT: Mice receiving the NSP4/E6-E7-L1 vaccine had the highest stimulatory index compared to other groups, although it was not statistically significant. Interleukin 4/12 and IFN-γ production were significantly higher in E6-E7-L1 / NSP4 group and E6-E7-L1 group compared to other groups (P < 0.05). Among different groups, E6/E7/L1 + NSP4 group was able to slow down the tumor growth process, but it was not significant (p > 0.05). Among the aforementioned cytokines, IFN-γ and IL-12 are among the cytokines that stimulate the Th1 pathway and IL-4 cytokine stimulates the Th2 pathway and B lymphocytes. CONCLUSION: Our data revealed that the present vaccine can reduce tumor size, and cytokine measurement showed that it stimulates innate and acquired immune responses, thus it can be a therapeutic vaccine in the tumor-bearing mice model.

A New Insight Into p53-Inhibiting Genes in Epstein­Barr Virus-Associated Gastric Adenocarcinoma

Background: The p53 mutation is uncommon in Epstein­Barr virus-linked gastric carcinoma, but its suppression occurs through mechanisms such as ubiquitin specific peptidase 7 (USP7) inhibitions via Epstein­Barr virus nuclear antigen-1 (EBNA1) activity. This study aimed to evaluate the effect of EBNA1 on p53-inhibiting gene expression and the impact of USP7 inhibition on p53 suppression. Methods: MKN-45 cells were transfected with the EBNA1 plasmid. A stable EBNA1 expression cell line was developed through selection based on hygromycin B resistance. Murine double minute (MDM)4, MDM2, sirtuin (SIRT)3, histone deacetylase (HDAC)1, proteasome 26S subunit, Non-ATPase (PSMD)10, USP7, and p53 expression were checked using real-time PCR. Also, cells containing EBNA1 or control plasmid were treated with GNE-6776, and the expression of the interested genes and cell survival were assessed. Results: MDM4, MDM2, and PSMD10 were significantly upregulated in the MKN-45 cell line following EBNA1 transfection. Morphological changes were observed in the cells harboring EBNA1 after 20 days. In the control cells, USP7 inhibition significantly upregulated the HDAC1, PSMD10, MDM4, and MDM2 genes after 24 h, but downregulated these genes after four days. In the EBNA1-harboring cells, MDM2, MDM4, and PSMD10 genes were significantly upregulated after 24 h, and this effect was sustained for all genes except for MDM4, even after four days. Furthermore, USP7 inhibition induced apoptosis in both cell groups. Conclusion: EBNA1 enhances the expression of p53-inhibiting genes. Two events­p53 protein overexpression and apoptosis activation­followed the suppression of the USP7 protein and provided evidence for its possible function. The significance of the EBNA1-USP7 interaction in p53 suppression warrants additional investigation and possibly reconsideration.