A comparison of the human and mouse protein corona profiles of functionalized SiO2 nanocarriers.

Nanoparticles are a promising cancer therapy for their use as drug carriers given their versatile functionalization with polyethylene glycol and proteins that can be recognized by overexpressed receptors in tumor cells. However, it has been suggested that in biological fluids, proteins cover nanoparticles, which gives the proteins a biological identity that could be responsible for unexpected biological responses: the so-called protein corona. A relevant biological event that is usually ignored in protein-corona formation is the interspecies differences in protein binding, which can be involved in the discrepancies observed in preclinical studies and the nanoparticle safety and efficiency. Hence, the aim of this study was to determine the differences between human and mouse plasma protein corona profiles in an active therapy model using silicon dioxide nanoparticles (SiO2 nanoparticles) functionalized with polyethylene glycol and transferrin. Functionalized SiO2 nanoparticles were made with a primary particle size of 25 nm and a transferrin content of 50 µg mg-1 of nanoparticles and were PEGylated with a cross-linker. The proteomic analysis by nanoliquid chromatography tandem-mass spectrometry (nanoLC-MS/MS) showed interspecies differences. The most abundant proteins found in the human protein corona profile were immunoglobulins, actin cytoplasmic 1, hemoglobin subunit beta, serotransferrin, ficolin-3, complement C3, and apolipoprotein A-1. Meanwhile, the mouse protein corona adsorbed the serine protease inhibitor A3K, serotransferrin, alpha-1-antitrypsin 1-2, hemoglobin subunit beta, and fibrinogen gamma and beta chains. These protein-corona profile differences in the functionalized SiO2 nanoparticles indicate that biological responses observed in in vivo models could not be translated to clinical use and must be considered in the interpretation of preclinical trials in order to design more efficient and safer nanomedicines.

hMLH1 promoter methylation is an early event in oral cancer.

Promoter methylation is believed to inactivate the expression of hMLH1. This process has been implicated in the tumorigenesis of oral squamous cell carcinoma (OSCC). Thus, the aim of this study was to determine the profile of hMLH1 methylation and protein expression in OSCC. The matched case-control study included 50 OSCC cases and 200 controls, with a median of age 64 (Q1-Q3 54-71) years. Protein expression was determined by immunohistochemical staining, and hMLH1 gene promoter methylation was analyzed by methylation-specific polymerase chain reaction (MSP). A conditional logistic regression model for risk factors was built for OSCC cases and matched controls. Promoter methylation of hMLH1 was detected in 38 (76%) OSCC cases, but in none of the control samples. Of the 38 OSCC samples with promoter methylation, 12 (32%) were negative for hMLH1 protein, and corresponded to early clinical stages (10 in stage II and 2 in stage I). All 12 unmethylated samples showed positive stain for hMLH1. Multiple logistic regression analysis showed an OR of 16.54 (IC 95%: 1.69-161.68, p=0.016) for methylation of the hMLH1 gene and early stages of OSCC, adjusting by gender and tobacco use. This study showed a high frequency of hMLH1 promoter methylation that occurred in most of the early stage cases and in about half of the late stage cases. It is proposed that hMLH1 promoter methylation is an early event that is maintained during tumor progression.

Mutational analysis of K-ras and Ras protein expression in larynx squamous cell carcinoma.

The ras gene family (H, K and N-ras) encodes the Ras protein, a GTPase-activating protein that regulates several signal transduction pathways including cellular proliferation and differentiation. Mutations in codons 12, 13 and 61 of the ras genes constitute one of the most frequent alterations in human cancer. In the Western Hemisphere, a low frequency of mutations in these genes has been observed in head and neck carcinomas; a higher frequency has been found in countries such as India and Taiwan. Increased protein expression is a relatively frequent event in larynx carcinomas. This study was aimed to evaluate the participation of the k-ras gene and Ras expression in 20 Mexican patients with larynx squamous carcinoma, 2 with dysplasia and 4 with normal mucosa. Samples (of 26 patients) were embedded in paraffin and immunohistochemical analysis was performed for the Ras protein, as well as amplification of the k-ras gene exon 1 (108 bp) by laser capture microdissection. Then, DNA extraction, PCR and sequencing were performed looking for possible mutation in codons 12 and 13. All patients with larynx carcinoma were men, median age 62 years. Eighty-five percent of the patients had risk factors such as smoking and/or alcohol consumption, 25% were in clinical stages I and II, and 75% in stages III and IV; 45% of the patients presented tumor recurrence or persistence. In this study, no mutations were found in codons 12 or 13 of the k-ras gene; however, protein expression was observed in 95% of the samples and a higher expression of the protein was associated with tumor recurrence or persistence, although this was not statistically significant. Unexpectedly, well-differentiated carcinomas and dysplasias presented an increase in protein expression. These results suggest that ras may be involved in early stages of larynx carcinogenesis and may be activated by other mechanisms different from mutations, such as epigenetic events.