603 research outputs found
Impact of enzymatic and microbial bioprocessing on antioxidant properties of hemp (Cannabis sativa L.)
Although the hemp seed boasts high nutritional and functional potential, its use in food preparations is still underestimated due to scarce technological properties and the presence of several anti-nutritional factors. Here, an optimization of a biotechnological protocol aimed at improving the antioxidant properties and the protein digestibility of the whole hemp seed has been proposed. Processing based on the use of commercial food grade enzymes and ad hoc selected lactic acid bacteria was tested and the phenolic and protein profiles were investigated through an integrated approach including selective extraction, purification, and identification of the potentially active compounds. The influence of the bioprocessing on the antioxidant activity of the hemp was evaluated both in vitro and on human keratinocytes. The lactic acid bacteria fermentation was the best method to significantly improve the antioxidant potential of the hemp through intense proteolysis which led to both the release of bioactive peptides and the increase in the protein digestibility. Moreover, changes in the phenolic profile allowed a significant protective effect against oxidative stress measured on the human keratinocyte cell line
Bioprocessing of Brewers’ Spent Grain Enhances Its Antioxidant Activity: Characterization of Phenolic Compounds and Bioactive Peptides
Brewers’ spent grain (BSG) is the major by-product of the brewing industry which remain largely unutilized despite its nutritional quality. In this study, the effects of fermentation on BSG antioxidant potential were analyzed. A biotechnological protocol including the use of xylanase followed by fermentation with Lactiplantibacillus plantarum (Lactobacillus plantarum) PU1, PRO17, and H46 was used. Bioprocessed BSG exhibited enhanced antioxidant potential, characterized by high radical scavenging activity, long-term inhibition of linoleic acid oxidation and protective effect toward oxidative stress on human keratinocytes NCTC 2544. Immunolabelling and confocal laser microscopy showed that xylanase caused an extensive cell wall arabinoxylan disruption, contributing to the release of bound phenols molecules, thus available to further conversion through lactic acid bacteria metabolism. To clarify the role of fermentation on the antioxidant BSG potential, phenols were selectively extracted and characterized through HPLC-MS techniques. Novel antioxidant peptides were purified and identified in the most active bioprocessed BSG
Characterization of an immunologically conserved epitope from hepatitis C virus E2 glycoprotein recognized by HLA-A2 restricted cytotoxic T lymphocytes
BACKGROUND/AIMS:
Identification of epitopes recognized by cytotoxic T lymphocytes (CTL) in hepatitis C virus (HCV) proteins is of importance because they can be used for vaccination, treatment of infection or monitoring of immune responses. Our purpose was to characterize new CTL epitopes in HCV structural proteins.
METHODS:
Peptides were synthesized and tested in HLA-A2 binding assays. Binder peptides were used to stimulate peripheral blood mononuclear cells from HCV+ patients and controls, and activity measured in chromium release and ELISPOT assays.
RESULTS:
Twenty binder peptides were found, and stimulation of HCV+ patient cells with nine peptides showing high binding ability led to the growth of CD8+ CTL recognizing peptide E2(614-622) in association with HLA-A2. Peptide E2(614-622) was recognized by 30% of HLA-A2+ patients with chronic HCV infection, but no responses were observed in control groups. Five peptides derived from region E2(614-622) from 26 different viral isolates bound to HLA-A2 molecules, and all of them but one, containing Phe at position 622, were recognized by E2(614-622) specific CTL.
CONCLUSIONS:
These results show that peptide E2(614-622) belongs to a highly conserved region of HCV E2, and might be a good candidate to induce anti-HCV CTL responses in HLA-A2+ subjects
Active immunotherapy in the treatment of haematological neoplasias
Abstract
The continuous search for therapeutic approaches that improve the conventional treatments of neoplasms, together with an improved understanding of the immune system, has led in recent years to the development of Immunotherapy. Basically, a distinction can be made between two forms of immunotherapy: passive immunotherapy, which consists in the transfer of antibodies or cells previously generated in vitro that are directed against the tumour, and active immunotherapy, which attempts to activate in vivo the immune system and induce it to elaborate a specific response against the tumor antibodies. Hematological neoplasms, specifically some B lymphomas, express in their membrane an immunoglobulin that is considered a specific antigen of the tumour, which is why these diseases have become the ideal target for immunotherapy treatments. There are many alternatives, ranging from protein vaccines, which have already shown clinical benefits, to those of the second generation, which make use of the new techniques of molecular biology to increase the efficacy of the vaccines and obtain their production in a quicker and less costly way, but with which there are not yet definitive clinical results
Acquired potential N-glycosylation sites within the tumor-specific immunoglobulin heavy chains of B-cell malignancies
Background and Objectives. Among B-cell malignancies, follicular lymphomas (FL)
more frequently show acquired, potential N-glycosylation sites (AGS) within tumor-specific
immunoglobulin. The aim of this study was to extend this observation and to evaluate
the pattern of presentation of AGS within five different forms of B-cell lymphoma.
Design and Methods. We sequenced the tumor-specific immunoglobulin heavy chain
variable region fragment, including complementarity-determining regions 2 and 3, of
forty-seven consecutive patients with a B-cell malignancy enrolled in idiotype vaccine
clinical trials. This sequencing approach is known to allow the identification of most AGS.
We then statistically analyzed differences in presentation pattern, in terms of tumor histology,
immunoglobulin isotype, AGS location and amino acid composition.
Results. All twenty-four FL cases presented with at least one AGS, whereas the vast
majority of four B-cell lymphoma types other than FL did not. The non- FL group of tumors
included four cases of Burkitt’s lymphoma, six of diffuse large cell lymphoma, seven mantle
cell lymphomas and six small lymphocytic lymphomas. Most IgM-bearing follicular
lymphoma cases featured their AGS within complementarity-determining region 2, as
opposed to those bearing an IgG, which mostly displayed the AGS within complementarity-
determining region 3. The vast majority of AGS located within either complementarity-
determining region ended with a serine residue, whereas those located within framework
regions mostly featured threonine as the last amino acid residue.
Interpretation and Conclusions. In our series, all cases of FL had AGS within their
tumor-specific immunoglobulin heavy chain variable regions. In contrast, most B-cell
malignancies other than FL did not. Further studies are warranted in order to establish
the possible meaning of these findings in terms of disease pathogenesis, their diagnostic
value in doubtful cases and their potential implications for immunotherapy
Optimization of universal allogeneic CAR-T cells combining CRISPR and transposon-based technologies for treatment of acute myeloid leukemia
Despite the potential of CAR-T therapies for hematological malignancies, their efficacy in patients with relapse and refractory Acute Myeloid Leukemia has been limited. The aim of our study has been to develop and manufacture a CAR-T cell product that addresses some of the current limitations. We initially compared the phenotype of T cells from AML patients and healthy young and elderly controls. This analysis showed that T cells from AML patients displayed a predominantly effector phenotype, with increased expression of activation (CD69 and HLA-DR) and exhaustion markers (PD1 and LAG3), in contrast to the enriched memory phenotype observed in healthy donors. This differentiated and more exhausted phenotype was also observed, and corroborated by transcriptomic analyses, in CAR-T cells from AML patients engineered with an optimized CAR construct targeting CD33, resulting in a decreased in vivo antitumoral efficacy evaluated in xenograft AML models. To overcome some of these limitations we have combined CRISPR-based genome editing technologies with virus-free gene-transfer strategies using Sleeping Beauty transposons, to generate CAR-T cells depleted of HLA-I and TCR complexes (HLA-IKO/TCRKO CAR-T cells) for allogeneic approaches. Our optimized protocol allows one-step generation of edited CAR-T cells that show a similar phenotypic profile to non-edited CAR-T cells, with equivalent in vitro and in vivo antitumoral efficacy. Moreover, genomic analysis of edited CAR-T cells revealed a safe integration profile of the vector, with no preferences for specific genomic regions, with highly specific editing of the HLA-I and TCR, without significant off-target sites. Finally, the production of edited CAR-T cells at a larger scale allowed the generation and selection of enough HLA-IKO/TCRKO CAR-T cells that would be compatible with clinical applications. In summary, our results demonstrate that CAR-T cells from AML patients, although functional, present phenotypic and functional features that could compromise their antitumoral efficacy, compared to CAR-T cells from healthy donors. The combination of CRISPR technologies with transposon-based delivery strategies allows the generation of HLA-IKO/TCRKO CAR-T cells, compatible with allogeneic approaches, that would represent a promising option for AML treatment
A randomized phase II clinical trial of dendritic cell vaccination following complete resection of colon cancer liver metastasis
Surgically resectable synchronic and metachronic liver metastases of colon cancer have high risk of relapse in spite
of standard-of-care neoadjuvant and adjuvant chemotherapy regimens. Dendritic cell vaccines loaded with autologous
tumor lysates were tested for their potential to avoid or delay disease relapses (NCT01348256). Patients with surgically
amenable liver metastasis of colon adenocarcinoma (n = 19) were included and underwent neoadjuvant chemotherapy,
surgery and adjuvant chemotherapy. Fifteen patients with disease-free resection margins were randomized 1:1 to receive
two courses of four daily doses of dendritic cell intradermal vaccinations versus observation. The trial had been originally
designed to include 56 patients but was curtailed due to budgetary restrictions. Follow-up of the patients indicates a
clear tendency to fewer and later relapses in the vaccine arm (median disease free survival –DFS-) 25.26 months, 95% CI 8.
74-n.r) versus observation arm (median DFS 9.53 months, 95% CI 5.32–18.88)
Immunological Biomarkers of Fatal COVID-19: A Study of 868 Patients
Information on the immunopathobiology of coronavirus disease 2019 (COVID-19) is rapidly increasing; however, there remains a need to identify immune features predictive of fatal outcome. This large-scale study characterized immune responses to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection using multidimensional flow cytometry, with the aim of identifying high-risk immune biomarkers. Holistic and unbiased analyses of 17 immune cell-types were conducted on 1,075 peripheral blood samples obtained from 868 COVID-19 patients and on samples from 24 patients presenting with non-SARS-CoV-2 infections and 36 healthy donors. Immune profiles of COVID-19 patients were significantly different from those of age-matched healthy donors but generally similar to those of patients with non-SARS-CoV-2 infections. Unsupervised clustering analysis revealed three immunotypes during SARS-CoV-2 infection; immunotype 1 (14% of patients) was characterized by significantly lower percentages of all immune cell-types except neutrophils and circulating plasma cells, and was significantly associated with severe disease. Reduced B-cell percentage was most strongly associated with risk of death. On multivariate analysis incorporating age and comorbidities, B-cell and non-classical monocyte percentages were independent prognostic factors for survival in training (n=513) and validation (n=355) cohorts. Therefore, reduced percentages of B-cells and non-classical monocytes are high-risk immune biomarkers for risk-stratification of COVID-19 patients
Pasado, presente y futuro de la vacunación anti-idiotipo
Cancer vaccines are conceived as therapeutic tools, in contrast to the prophylactic vaccines used to fight against infectious diseases. Among the most potent therapeutic vaccines, anti-idiotype vaccination is directed against the tumor idiotype, the only well-characterized tumor antigen displayed in neoplastic B-cells. Anti-idiotype vaccines have demonstrated clinical benefit against follicular lymphoma and are currently being evaluated in two different phase III clinical trials. Additional emerging strategies, which include the use of dendritic cells and the production of vaccines via molecular means will surely allow us to draw important conclusions concerning the treatment of cancer patients
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