How integrin phosphorylations regulate cell adhesion and signaling

Cell adhesion is essential for the formation of organs, cellular migration, and interaction with target cells and the extracellular matrix. Integrins are large protein alpha/13-chain heterodimers and form a major family of cell adhesion molecules. Recent research has dramatically increased our knowledge of how integrin phos-phorylations regulate integrin activity. Phosphorylations determine the signaling complexes formed on the cytoplasmic tails, regulating downstream signaling. alpha-Chain phosphorylation is necessary for inducing 13-chain phosphorylation in LFA-1, and the crosstalk from one integrin to another activating or inactivating its function is in part mediated by phosphorylation of 13-chains. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus receptor angiotensin-converting enzyme 2 (ACE2) and possible integrin coreceptors may crosstalk and induce a phosphorylation switch and autophagy.Peer reviewe

Regulation of Dynamic Cell Adhesion by Integrin-Integrin Crosstalk

Most cells express several integrins. The integrins are able to respond to various cellular functions and needs by modifying their own activation state, but in addition by their ability to regulate each other by activation or inhibition. This crosstalk or transdominant regulation is strictly controlled. The mechanisms resulting in integrin crosstalk are incompletely understood, but they often involve intracellular signalling routes also used by other cell surface receptors. Several studies show that the integrin cytoplasmic tails bind to a number of cytoskeletal and adaptor molecules in a regulated manner. Recent work has shown that phosphorylations of integrins and key intracellular molecules are of pivotal importance in integrin-cytoplasmic interactions, and these in turn affect integrin activity and crosstalk. The integrin β-chains play a central role in regulating crosstalk. In addition to Integrin-integrin crosstalk, crosstalk may also occur between integrins and related receptors, including other adhesion receptors, growth factor and SARS-CoV-2 receptors

Regulation of Dynamic Cell Adhesion by Integrin-Integrin Crosstalk

Most cells express several integrins. The integrins are able to respond to various cellular functions and needs by modifying their own activation state, but in addition by their ability to regulate each other by activation or inhibition. This crosstalk or transdominant regulation is strictly controlled. The mechanisms resulting in integrin crosstalk are incompletely understood, but they often involve intracellular signalling routes also used by other cell surface receptors. Several studies show that the integrin cytoplasmic tails bind to a number of cytoskeletal and adaptor molecules in a regulated manner. Recent work has shown that phosphorylations of integrins and key intracellular molecules are of pivotal importance in integrin-cytoplasmic interactions, and these in turn affect integrin activity and crosstalk. The integrin β-chains play a central role in regulating crosstalk. In addition to Integrin-integrin crosstalk, crosstalk may also occur between integrins and related receptors, including other adhesion receptors, growth factor and SARS-CoV-2 receptors

Patient-Derived Organoids for Precision Cancer Immunotherapy

Cancer immunotherapy has revolutionized the way tumors are treated. Nevertheless, efficient and robust testing platforms are still missing, including clinically relevant human ex vivo tumor assays that allow pretreatment testing of cancer therapies and selection of the most efficient and safe therapy for a specific patient. In the case of immunotherapy, this testing platform would require not only cancer cells, but also the tumor microenvironment, including immune cells. Here, we discuss the applications of patient-derived tumor organoid cultures and the possibilities in using complex immune-organoid cultures to provide preclinical testing platforms for precision cancer immunotherapy.Peer reviewe

Phosphorylation of the α-chain in the integrin LFA-1 enables β2-chain phosphorylation and α-actinin binding required for cell adhesion

The integrin leukocyte function-associated antigen-1 (LFA-1) plays a pivotal role in leukocyte adhesion and migration, but the mechanism(s) by which this integrin is regulated has remained incompletely understood. LFA-1 integrin activity requires phosphorylation of its 2-chain and interactions of its cytoplasmic tail with various cellular proteins. The -chain is constitutively phosphorylated and necessary for cellular adhesion, but how the -chain regulates adhesion has remained enigmatic. We now show that substitution of the -chain phosphorylation site (S1140A) in T cells inhibits the phosphorylation of the functionally important Thr-758 in the 2-chain, binding of -actinin and 14-3-3 protein, and expression of an integrin-activating epitope after treatment with the stromal cell-derived factor-1. The presence of this substitution resulted in a loss of cell adhesion and directional cell migration. Moreover, LFA-1 activation through the T-cell receptor in cells expressing the S1140A LFA-1 variant resulted in less Thr-758 phosphorylation, -actinin and talin binding, and cell adhesion. The finding that the LFA-1 -chain regulates adhesion through the -chain via specific phosphorylation at Ser-1140 in the -chain has not been previously reported and emphasizes that both chains are involved in the regulation of LFA-1 integrin activity.Peer reviewe

Geenien muokkaus uusilla tekniikoilla: kasvit, eläimet, mikrobit

CRISPR/Cas9 teknologian soveltaminen genomieditointiin on saanut valtavasti huomiota ja sitä on pidetty mullistavana teknologiana, jonka vaikutukset ja mahdollisuudet ovat paljon laajemmat kuin perinteisten geenimuuntelumenetelmien. Tässä julkaisussa, joka on valmisteltu geenitekniikan lautakunnan pyynnöstä keväällä 2018 valmistuneen muistion pohjalta, selvitetään esimerkkien avulla CRISPR/Cas9 teknologian erilaisia sovelluksia erityisesti kasvinjalostuksessa ja malarian torjunnassa. Mikrobeihin kohdistuvaa genomie-ditointia sivutaan lyhyesti. CRISPR/Cas9-teknologia mahdollistaa myös populaatioissa itseään levittävien geenielementtien (ns. geeniajurien) rakentamisen. Julkaisussa tarkas-tellaan myös genomieditointisovellusten suhdetta geenitekniikkalainsäädäntöön. Erityisen ongelman muodosti pitkään se, että oli epäselvää miten EU:n lainsäädäntöä sovelletaan uusiin tekniikoihin, jotka mahdollistavat kohdennetun mutageneesin tavalla, jota ei voi erottaa luonnon mutaatioista. Tilannetta on selkeyttänyt EU-tuomioistuimen (EUT) heinä-kuussa 2018 tekemä päätös, jossa se otti kantaa uusien mutageneesitekniikoiden juridiseen asemaan. Biotekniikan neuvottelukunta laatii terveyskysymyksistä erillisen selvityksen. </p

Peptides-Coated Oncolytic Vaccines for Cancer Personalized Medicine

Publisher Copyright: Copyright © 2022 Feola, Russo, Martins, Lopes, Vandermeulen, Fluhler, De Giorgi, Fusciello, Pesonen, Ylösmäki, Antignani, Chiaro, Hamdan, Feodoroff, Grönholm and Cerullo.Oncolytic Viruses (OVs) work through two main mechanisms of action: the direct lysis of the virus-infected cancer cells and the release of tumor antigens as a result of the viral burst. In this sc.enario, the OVs act as in situ cancer vaccines, since the immunogenicity of the virus is combined with tumor antigens, that direct the specificity of the anti-tumor adaptive immune response. However, this mechanism in some cases fails in eliciting a strong specific T cell response. One way to overcome this problem and enhance the priming efficiency is the production of genetically modified oncolytic viruses encoding one or more tumor antigens. To avoid the long and expensive process related to the engineering of the OVs, we have exploited an approach based on coating OVs (adenovirus and vaccinia virus) with tumor antigens. In this work, oncolytic viruses encoding tumor antigens and tumor antigen decorated adenoviral platform (PeptiCRAd) have been used as cancer vaccines and evaluated both for their prophylactic and therapeutic efficacy. We have first tested the oncolytic vaccines by exploiting the OVA model, moving then to TRP2, a more clinically relevant tumor antigen. Finally, both approaches have been investigated in tumor neo-antigens settings. Interestingly, both genetically modified oncolytic adenovirus and PeptiCRAd elicited T cells-specific anti-tumor responses. However, in vitro cross-representation experiments, showed an advantage of PeptiCRAd as regards the fast presentation of the model epitope SIINFEKL from OVA in an immunogenic rather than tolerogenic fashion. Here two approaches used as cancer oncolytic vaccines have been explored and characterized for their efficacy. Although the generation of specific anti-tumor T cells was elicited in both approaches, PeptiCRAd retains the advantage of being rapidly adaptable by coating the adenovirus with a different set of tumor antigens, which is crucial in personalized cancer vaccines clinical setting.Peer reviewe

A novel immunopeptidomic-based pipeline for the generation of personalized oncolytic cancer vaccines

Besides the isolation and identification of major histocompatibility complex I-restricted peptides from the surface of cancer cells, one of the challenges is eliciting an effective antitumor CD8+ T-cell-mediated response as part of therapeutic cancer vaccine. Therefore, the establishment of a solid pipeline for the downstream selection of clinically relevant peptides and the subsequent creation of therapeutic cancer vaccines are of utmost importance. Indeed, the use of peptides for eliciting specific antitumor adaptive immunity is hindered by two main limitations: the efficient selection of the most optimal candidate peptides and the use of a highly immunogenic platform to combine with the peptides to induce effective tumor-specific adaptive immune responses. Here, we describe for the first time a streamlined pipeline for the generation of personalized cancer vaccines starting from the isolation and selection of the most immunogenic peptide candidates expressed on the tumor cells and ending in the generation of efficient therapeutic oncolytic cancer vaccines. This immunopeptidomics-based pipeline was carefully validated in a murine colon tumor model CT26. Specifically, we used state-of-the-art immunoprecipitation and mass spectrometric methodologies to isolate > 8000 peptide targets from the CT26 tumor cell line. The selection of the target candidates was then based on two separate approaches: RNAseq analysis and HEX software. The latter is a tool previously developed by Jacopo, 2020, able to identify tumor antigens similar to pathogen antigens in order to exploit molecular mimicry and tumor pathogen cross-reactive T cells in cancer vaccine development. The generated list of candidates (26 in total) was further tested in a functional characterization assay using interferon-gamma enzyme-linked immunospot (ELISpot), reducing the number of candidates to six. These peptides were then tested in our previously described oncolytic cancer vaccine platform PeptiCRAd, a vaccine platform that combines an immunogenic oncolytic adenovirus (OAd) coated with tumor antigen peptides. In our work, PeptiCRAd was successfully used for the treatment of mice bearing CT26, controlling the primary malignant lesion and most importantly a secondary, nontreated, cancer lesion. These results confirmed the feasibility of applying the described pipeline for the selection of peptide candidates and generation of therapeutic oncolytic cancer vaccine, filling a gap in the field of cancer immunotherapy, and paving the way to translate our pipeline into human therapeutic approach.Peer reviewe

Controlled release of enhanced cross-hybrid IgGA Fc PD-L1 inhibitors using oncolytic adenoviruses

Immune checkpoint inhibitors have clinical success in prolonging the life of many cancer patients. However, only a minority of patients benefit from such therapy, calling for further improvements. Currently, most PD-L1 checkpoint inhibitors in the clinic do not elicit Fc effector mechanisms that would substantially increase their efficacy. To gain potency and circumvent off-target effects, we previously designed an oncolytic adenovirus (Ad-Cab) expressing an Fc fusion peptide against PD-L1 on a cross-hybrid immunoglobulin GA (IgGA) Fc. Ad-Cab elicited antibody effector mechanisms of IgG1 and IgA, which led to higher tumor killing compared with each isotype alone and with clinically approved PD-L1 checkpoint inhibitors. In this study, we further improved the therapy to increase the IgG1 Fc effector mechanisms of the IgGA Fc fusion peptide (Ad-Cab FT) by adding four somatic mutations that increase natural killer (NK) cell activation. Ad-Cab FT was shown to work better at lower concentrations compared with Ad-Cab in vitro and in vivo and to have better tumor- and myeloid-derived suppressor cell killing, likely because of higher NK cell activation. Additionally, the biodistribution of the Fc fusion peptide demonstrated targeted release in the tumor microenvironment with minimal or no leakage to the peripheral blood and organs in mice. These data demonstrate effective and safe use of Ad-Cab FT, bidding for further clinical investigation