Time to take HPV infection in colorectal cancer patients more seriously.

Background: The association between viral infections and colorectal cancer (CRC) remains an enigma in cancer research. Certain types of Human Papillomaviruses (hr-HPVs), known for their oncogenic properties, have been observed in particular CRC biopsies, further adding to the enigma surrounding this association. Materials and methods: This cross-sectional study was conducted on 40 confirmed cases of CRC adenocarcinoma. The presence and genotyping of HPV DNA in colorectal fresh tissue and urine samples was assessed using an HPV DNA hybridization kit. A subset of serum samples from both CRC cases and healthy volunteers was randomly chosen and subjected to western blot to investigate the presence of HPV16 E6/E7 oncoproteins carried by exosomes. Results: It was observed that 26/40 HPV-positive CRC patients demonstrated 7 times more chance to develop colorectal cancer when compared to those 8/40 normal tissue (odds ratio [OR] = 7.4; confidence interval [CI] 95% = 0.483156-0.793718; p < 0.001). Of 26 HPV-positive CRC patients, 14 urine samples were also showed HPV DNA positivity (p = 0.013). High-risk HPV16 was the most prevalent genotype detected in both 24/40 tumor and 12/40 urine samples (p < 0.001). The tumor sample of a male was HPV45, while another male's urine sample was HPV31. A female CRC patient had HPV83 in tumor and HPV56 in urine. Here, was the first detection of HPV83 in a CRC patient. Notably among 20 randomly selected serum exosome samples, one serum sample concurrently tested positive for both HPV16 E6 and E7 oncoproteins, and one sample tested positive for HPV16 E7 oncoprotein. Conclusion: High risk HPV DNA detection in CRC urine samples supports non-invasive screening tools. Detection of HPV16 E6 and E7 oncoproteins in exosomes from serum samples shows potential for non-invasive diagnostics. HPV's potential role in CRC development is also underscored. HPV vaccination should be implemented in low- and middle-income countries to prevent cancer.

Genetically engineered E. coli invade epithelial cells and transfer their genetic cargo into the cells: an approach to a gene delivery system.

PURPOSE: Despite advances in gene therapy, the lack of safe and efficient gene delivery systems limited the clinical effectiveness of gene therapy. Due to the inherent potential of bacteria, they can be considered as a good option for the gene transfer system. This study aimed to create a genetically engineered bacterium capable of entering epithelial cells and transferring its genetic cargo to the cell's cytoplasm, eventually expressing the gene of interest in the cell. METHODS: The invasin (inv) gene from Yersinia pseudotuberculosis and the listeriolysin (hlyA) gene from Listeria monocytogenes were isolated by PCR assay and inserted into a pACYCDuet-1 vector. The recombinant plasmid was then transformed into E. coli strain BL21. Subsequently, pEGFP-C1 plasmids containing a CMV promoter were transformed into the engineered bacteria. Finally, the engineered bacteria containing the reporter genes were incubated with the HeLa and LNCaP cell lines. Fluorescence microscopy, flow cytometry, and TEM were used to monitor bacterial entry into the cells and gene expression. We used native E. coli strain BL21 as a control. RESULTS: A fluorescence microscope showed that, in contrast to the control group, the manipulated E. coli were able to penetrate the cells and transport the plasmid pEGFP-C1 to the target cells. Flow cytometry also showed fluorescence intensity of 54.7% in HeLa cells and 71% in LNCaP cells, respectively. In addition, electron micrographs revealed the presence of bacteria in the cell endosomes and in the cytoplasm of the cells. CONCLUSION: This study shows that genetically engineered E. coli can enter cells, transport cargo into cells, and induce gene expression in the target cell. In addition, flow cytometry shows that the gene transfer efficiency was sufficient for protein expression.