Ali Farhadi

Introduction: Co-infection of high-risk human papillomaviruses (HR-HPVs) with human polyomaviruses, such as Merkel cell polyomavirus (MCPyV), may indicate a potential synergistic effect. This study focused on the role of the MCPyV sT protein, which shares similarities with HPV oncoproteins in disrupting cellular regulatory mechanisms. This research aimed to investigate the effect ...

Gastric cancer (GC) is the third leading cause of cancer‐related mortality worldwide. The Epstein–Barr virus (EBV) may play a role in certain cases of GC. Therefore, this study aimed to investigate the prevalence of EBV in the gastric tissue of both gastric cancer patients and non‐cancer patients in Shiraz, Iran. In this cross‐sectional study, 159 formalin‐fixed paraffin‐embedded (FFPE) tissues from gastric cancer patients and 137 from non‐cancer patients were assessed. All samples were assessed using PCR on the β‐globin gene. The nested PCR method was used to investigate the presence of the EBNA‐1‐EBV gene. The results were analysed using chi‐squared statistical tests. The mean age of the GC group and the control group was 62.08 ± 13.45 and 63.97 ± 9.13. In the GC and control groups, 33.33% (53/159) and 35.04% (48/137) were female. The results showed that in the cancer group, 1.88% (3/159) of tissue samples were positive for EBV, while this statistic was 22.62% (31/137) for non‐cancer samples (p < .0001). All EBV‐positive cancer patients were female with a mean age of 64.66 ± 7.37 while in the control group, 20.83% (10/48) of females and 23.59% (21/89) of males were positive for EBV, which was not statistically significant (p = .832). The low frequency of EBV infection in GC tissue might indicate the ‘hit and run’ mechanism of EBV in GC carcinogenesis. Additionally, the abundance of B lymphocytes in the inflamed samples of the healthy control group might influence the high frequency of EBV in this group. More investigations are recommended to verify these results.

Objective: Activating transcription factor 3 (ATF3) has attracted recent scientific attention as a novel mediator of tumor suppression, particularly within the context of cervical cancer (CC). Our prior research demonstrated that ATF3 overexpression ...

Purpose: Cervical cancer (CxCa), is mainly caused by high-risk human papillomaviruses (hrHPV), which disrupt the regulation of p53 and pRb, leading to uncontrolled cell growth and cancer progression. Co-infection with human polyomaviruses like MCPyV has been observed in some HPV-positive cases, suggesting a potential combined effect on tumor development. Cisplatin is commonly used for advanced CxCa, but resistance, often due to p53 disruption, remains a significant challenge. challenge. This study investigates if MCPyV sT oncoprotein, together with HPV-18 oncoproteins, affects the transcription of key genes, influencing cell proliferation and cisplatin resistance in CxCa. Methods: The sT gene was cloned into the pCMV6 vector and introduced into E. coli DH5α cells. HeLa cells were transfected with pCMV6-sT using Lipofectamine 3000. Transfection efficiency was assessed using fluorescence microscopy and flow cytometry. Protein expression was analyzed by SDS-PAGE and Western blotting. Cytotoxicity was measured with the MTT assay, gene expression was analyzed by RT-qPCR, Ki-67 staining was performed on cell blocks, and cisplatin-induced effects on cell proliferation and apoptosis were examined. Results: Cytotoxicity assays showed a significant increase in cell viability at 0.2 μg of sT plasmid after 72 hours (13.76%, P < 0.05). MCPyV sT expression significantly upregulated the mRNA levels of E1 (4.22-fold), E6/E7 (3.80-fold), and MMP1 (6-fold) (P < 0.001). Increased Ki-67 positivity, indicated enhanced cell proliferation. Additionally, sT expression reduced cisplatin-induced apoptosis, with fewer apoptotic cells observed in the sT + cisplatin group compared to the cisplatin-only group (25.9% vs. 38.3%, P < 0.05). Conclusion: The simultaneous presence of MCPyV sT and HPV oncoproteins enhances resistance to cisplatin-induced apoptosis in cervical cancer cells, necessitating further investigation into viral oncoprotein interactions to overcome therapeutic resistance.

Activating Transcription Factor 3 (ATF3) is known for its tumor-suppressive properties in cervical cancer, particularly through its role in stress response and interactions with human papillomavirus (HPV) oncogenes. This study investigates ATF3’s regulatory impact on metastasis-related genes, oxidative stress, and DNA damage in HPV-positive cervical cancer cells. HeLa and Ca Ski cell lines were transfected with ATF3-expressing vectors. Western blotting and quantitative reverse transcription polymerase chain reaction (RT-qPCR) were used to confirm ATF3 overexpression following transfection. ROS assays and Comet assays assessed the impact of ATF3 on oxidative stress and DNA damage, while RT-qPCR was used to evaluate changes in HPV E6/E7, SHARP1, and MMP1 gene expression. ATF3 overexpression led to elevated ROS levels (p < 0.02), resulting in oxidative DNA damage. These results demonstrate ATF3’s cytotoxic impact on cervical cancer cells through oxidative stress and DNA damage. Additionally, ATF3 overexpression significantly decreased MMP1 expression (p < 0.03), indicating a potential anti-metastatic effect, while SHARP1 and HPV E6/E7 expression levels were not significantly altered, indicating selective gene modulation by ATF3. These findings reveal that ATF3 contributes to tumor suppression in cervical cancer by modulating oxidative stress and DNA damage, selectively targeting genes involved in metastasis. These findings supports ATF3’s role in regulating key pathways in HPV-positive cervical cancer cells, providing a basis for further exploration of ATF3 as a target in therapeutic strategies aimed at improving outcomes in cervical cancer.

Cell-penetrating peptides (CPPs) are effective for delivering therapeutic molecules with minimal toxicity. This study focuses on the use of penetratin, a well-characterized CPP, to deliver a DNA vector encoding short hairpin RNA (shRNA) targeting the respiratory syncytial virus (RSV) F gene into infected cells. RSV is known to cause severe lower respiratory infections in infants and poses significant risks to immunocompromised individuals and the elderly. We evaluated the antiviral efficacy of the penetratin-shRNA complex by comparing its ability to inhibit RSV replication and induce apoptosis with ribavirin treatment. Penetratin-shRNA complexes were prepared at different ratios and analyzed using gel retardation assays, dynamic light scattering, and zeta potential measurements. The complexes were tested in HEp-2 and A549 cells for transfection efficiency, cytotoxicity, viral load, and apoptosis using plaque assays, real-time reverse transcription-polymerase chain reaction (RT-PCR), DNA fragmentation, propidium iodide staining, and caspase 3/7 activation assays. The gel shift assay determined that a 20:1 CPP-to-shRNA ratio was optimal for effective complexation, resulting in particles with a size of 164 nm and a zeta potential of 8.7 mV. Transfection efficiency in HEp-2 cells was highest at this ratio, reaching up to 93%. The penetratin-shRNA complex effectively silenced the RSV F gene, reduced viral titers, and decreased DNA fragmentation and apoptosis in infected cells. Penetratin effectively delivers shRNA targeting the RSV F gene, significantly reducing viral load and preventing apoptosis without toxicity. This approach surpasses Lipofectamine and shows potential for future therapeutic interventions, especially when combined with ribavirin, against RSV infection.

Background: It is believed that loss of p53 function plays a crucial role in the progression of well to poorly differentiated thyroid cancers including anaplastic thyroid carcinoma (ATC). Given the poor prognosis of ATC due to its strong therapeutic resistance, there is a need to establish new therapeutic targets to extend the survival of ATC patients. Activating transcription factor 3 (ATF3) can inhibit the oncogenic activity of mutant p53 and, as a result, contribute to tumor suppression in several TP53-mutated cancers. Herein, we demonstrate that the ectopic overexpression of ATF3 leads to the suppression of oncogenic mutant p53 activity in chemo-resistant 8305 C thyroid cancer cells harboring R273C p53 gene mutation. Methods: The biological behavior of 8305 C cells was assessed pre- and post-transfection with pCMV6-ATF3 plasmid using MTT assay, fluorescent microscopy, cell cycle, and annexin V/PI flow cytometric analysis. The effect of ectopic ATF3 overexpression on the cellular level of p53 was examined by western blotting assay. The mRNA expression levels of TP53, TAp63, ΔNp63, and SHARP1 were evaluated in ectopic ATF3-expressing cells compared to controls. Results: The overexpression of ATF3 in 8305 C thyroid cancer cells significantly decreased cell viability and induced apoptosis and cell cycle arrest in vitro. The immunoblotting of p53 protein revealed that ATF3 overexpression significantly increased the level of mutant p53 in 8305C cells compared to mock-transfected control cells. Additionally, elevated mRNA levels of TAp63 and SHARP1 and a decreased mRNA level of ΔNp63 were observed in PCMV6-AC-ATF3-transfected 8305 C cells with significant differences compared to the mock and untreated cells. Conclusion: In light of our findings, it is evident that therapeutic strategies aimed at increasing ATF3 expression or enhancing the interaction between ATF3 and mutant p53 can be a promising approach for the treatment of p53-mutated metastatic thyroid cancer.

Purpose: This research investigated the development of short hairpin RNA (shRNA) molecules designed to target specific regions of the human respiratory syncytial virus (HRSV) M and F genes. The study aimed to assess the therapeutic potential of these shRNAs and evaluate the effectiveness of Tat peptide-mediated delivery in enhancing their functionality. Methods: We acquired isolates from pediatric patients experiencing respiratory illness then cultured in HEp-2 cells. We constructed plasmids expressing shRNAs. Tat peptide as a facilitator for shRNA plasmid delivery was used. The cytotoxicity of ribavirin, shRNA constructs, and control agents was assessed using the MTT assay. The transfection efficiency of Tat peptide-mediated shRNA delivery with that of lipofectamine 3000™ were compared. Finally, real-time PCR was employed to quantify HRSV replication in the treated cells. Results: Tat peptide-mediated delivery of shRNA plasmids significantly suppressed the expression of the M and F genes of HRSV compared to lipofectamine 3000™. This suppression was evident in both short-term experiments and scenarios involving stable shRNA expression. Furthermore, the combination of ribavirin with shRNA treatment resulted in a substantial reduction in viral load. Notably, the most pronounced antiviral effect was observed when both shRNAs were employed simultaneously. Conclusion: Our findings suggest that Tat peptide-mediated delivery of shRNA plasmids holds significant potential for achieving stable suppression of HRSV genes. This approach warrants further investigation as a potential gene therapy strategy for HRSV. By demonstrating promising results in vitro, this study highlights the need for future in vivo studies to comprehensively evaluate the therapeutic potential of this approach in a clinical setting.