We are reporting qualitative and quantitative changes of the extracellular matrix (ECM) and associated receptor proteomes, occurring during the transition from liver fibrosis and steatohepatitis to hepatocellular carcinoma (HCC). We compared two mouse models relevant to human HCC: PDGFC transgenic (Tg) and Pten null mice, models of disease progression from fibrosis and steatohepatitis to HCC. Using mass spectrometry, we identified in the liver of both models proteins for 26 collagen-encoding genes, providing the first evidence of expression at the protein level for 16 collagens. We also identified post-transcriptional protein variants for six collagens and lysine hydroxylation modifications for 14 collagens. Tumor-associated collagen proteomes were similar in both models with increased expression of collagens type IV, VI, VII, X, XIV, XV, XVI, and XVIII. Splice variants for Col4a2, Col6a2, Col6a3 were co-upregulated while only the short form of Col18a1 increased in the tumors. We also identified tumor specific increases of nidogen 1, decorin, perlecan, and of six laminin subunits. The changes in these non-collagenous ECM proteins were similar in both models with the exception of laminin β3, detected specifically in the Pten null tumors. Pdgfa and Pdgfc mRNA expression was increased in the Pten null liver, a possible mechanism for the similarity in ECM composition observed in the tumors of both models. In contrast and besides the strong up-regulation of integrin α5 protein observed in the liver tumors of both models, the expression of the six other integrins identified was specific to each model, with integrins α2b, α3, α6, and β1 up-regulated in Pten null tumors and integrins α8 and β5 up-regulated in the PDGFC Tg tumors. In conclusion, HCC–associated ECM proteins and ECM–integrin networks, common or specific to HCC subtypes, were identified, providing a unique foundation to using ECM composition for HCC classification, diagnosis, prevention, or treatment.

Pandemic threats of the H1N1 influenza virus have drawn attention to developing a universal vaccine against circulating and future strains of this virus. An immunoinformatics study was conducted to identify conserved peptides containing CD4+ and CD8+ T-cell epitopes from all the hemagglutinin (HA) and neuraminidase (NA) protein sequences available until February 2013 to cover the seasonal as well as the pandemic strains of the H1N1 virus. In the present study, six different immunoinformatics prediction programs were used in order to define the epitopes. Five conserved peptides of HA and six of NA protein were obtained that contained overlapping CD4+ and CD8+ T-cell epitopes. These identified peptides have a binding affinity for a large number of major histocompatibility complex (MHC) alleles. WHGSNRPWVSF of NA protein is a new peptide whose T-cell response has not been previously reported. Population coverage studies have shown that these peptide fragments have the capacity to induce a potent immune response among individuals from different populations around the world. Hence, these HA and NA peptides may be considered as interesting candidates for vaccine design.

Cell mediated immune response plays a key role in combating viral infection and thus identification of new vaccine targets manifesting T cell mediated response may serve as an ideal approach for influenza vaccine. The present study involves the application of an immunoinformatics-based consensus approach for epitope prediction (three epitope prediction tools each for CD4+ and CD8+ T cell epitopes) and molecular docking to identify peptide sequences containing T cell epitopes using the conserved sequences from all the Matrix 1 protein sequences of H1N1 virus available until April 2015. Three peptides comprising CD4+ and CD8+ T cell epitopes were obtained, which were not exactly reported in earlier studies. Population coverage study of these multiepitope peptides revealed that they are capable of inducing a potent immune response belonging to individuals from different populations and ethnicity distributed around the globe. Conservation study with other subtypes of influenza virus infecting humans (H2N2, H5N1, H7N9, and H3N2) revealed that these three peptides were conserved (>90%), with 100% identity in most of these strains. Hence, these peptides can impart immunity against H1N1 as well as other subtypes of influenza virus. A molecular docking study of the predicted peptides with class I and II human leukocyte antigen (HLA) molecules has shown that the majority of them have comparable binding energies to that of native peptides. Hence, these peptides from Matrix 1 protein of H1N1 appear to be promising candidates for universal vaccine design.

Influenza vaccine development is considered to be complicated and challenging. Constantly evolving influenza viruses require continuous global monitoring and reformulation of the vaccine strains. Peptides that are conserved among different strains and subtypes of influenza A virus are strongly considered to be attractive targets for development of cross protective influenza vaccines that stimulate cellular responses. In this study, three highly conserved (>90%) matrix 1 peptides that contain multiple T cell epitopes, ILGFVFTLTVPSERGLQRRRF (P 1), LIRHENRMVLASTTAKA (P 2) and LQAYQKRMGVQMQR (P 3), were assessed for their immunogenic potential in vitro by subjecting peripheral blood mononuclear cells from healthy volunteers to repetitive stimulation with these chemically synthesised peptides and measuring their IFN-γ concentrations, proliferation by ELISA, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, respectively. Seven samples were screened for immunogenicity of P 1 and P 2, and six for that of P 3. All six samples had positive responses (IFN-γ secretion) to P 3 stimulation, as did five and three for P 2 and P 1 respectively. In contrast, seven (P 1 and P 2) and four (P 3) samples showed proliferative response as compared with unstimulated cells. The encouraging immunogenic response generated by these highly conserved matrix 1 peptides indicates they are prospective candidates for development of broadly reactive influenza vaccines.

The concept of peptide-based vaccines against cancer has made noteworthy progress. Metadherin (MTDH) overexpression and its role in the development of diverse cancers make it an attractive target for cancer immunotherapy. In the current study, six different T cell epitope prediction tools were run to identify MTDH peptides with multiple immunogenic regions. Further, molecular docking was performed to assess HLA-peptide binding interactions. Nine and eleven peptides fragments containing multiple CD8 (+) and CD4 (+) T-cell epitopes, ranging from 9 to 20 amino acids, respectively, were obtained using a consensus immunoinformatics approach. The three peptides that were finally identified as having overlapping CD4 (+) and CD8 (+) T- cell epitopes are ARLREMLSVGLGFLRTELG, FLLGYGWAAACAGAR, YIDDEWSGLNGLSSADP. These peptides were found to not only have multiple T cell epitopes but also to have binding affinity with wide HLA molecules. A molecular docking study revealed that the predicted immunogenic peptides (with single or multiple T cell epitopes) of MTDH have comparable binding energies with naturally bound peptides for both HLA classes I and II. Thus, these peptides have the potential to induce immune responses that could be considered for developing synthetic peptide vaccines against multiple cancers.