Adenosine triphosphate acts as a paracrine signaling molecule to reduce the motility of T cells.

Organization of immune responses requires exchange of information between cells. This is achieved through either direct cell-cell contacts and establishment of temporary synapses or the release of soluble factors, such as cytokines and chemokines. Here we show a novel form of cell-to-cell communication based on adenosine triphosphate (ATP). ATP released by stimulated T cells induces P2X4/P2X7-mediated calcium waves in the neighboring lymphocytes. Our data obtained in lymph node slices suggest that, during T-cell priming, ATP acts as a paracrine messenger to reduce the motility of lymphocytes and that this may be relevant to allow optimal tissue scanning by T cells.

The challenge for a European network of biobanks for rare diseases taken up by RD-Connect.

Access to biological materials is a key prerequisite for scientific research in any medical field and in particular for research into rare diseases (RDs), for which obtaining high-quality samples and the related clinical data remains a major hurdle. RD biobanks play a pivotal role in making such materials and data available to the scientific community. In order to increase the effectiveness of RD biobanks, three major challenges need to be met: maximise access to rare biological samples stored in RD biobanks spread globally by the international scientific community, promote networking among such biobanks to share quality standards and procedures and allow collaboration with RD registries and databases, and finally adopt an efficient management model compliant with legal and ethical issues and ensuring biobank sustainability. The European program RD-Connect, funded under the FP7 program, addresses all of these issues through an articulated action plan aimed at building a network of European RD biobanks. Ultimately, RD-Connect will offer access to precious, quality-controlled biological samples from RD patients through an online, searchable, dynamic catalogue in the context of an integrated platform that links RD patient registries to biobanks and to clinical bioinformatics data for RD research.

667C>T and 1298A>C polymorphisms of MTHFR do not predict response to methotrexate in patients with gestational trophoblastic neoplasia.

OBJECTIVE: Approximately one third of patients treated with methotrexate for gestational trophoblastic neoplasia (GTN) following a molar pregnancy are reported to develop resistance to methotrexate and need to change to different chemotherapeutic agents. Previous studies, in other clinical settings, have suggested that polymorphisms in key folate metabolising enzymes such as 5,10-methylenetetrahydrofolate reductase (MTHFR) influence both toxicity and efficacy of methotrexate. Our objective was to investigate the impact of two common functional MTHFR polymorphisms, 677C>T and 1298A>C, on the efficacy of methotrexate in women treated for GTN following a molar pregnancy. METHODS: DNA from 121 women treated with methotrexate for GTN was genotyped for the 677C>T and 1298A>C polymorphisms using TaqMan SNP Genotyping Assays. In 64 cases these polymorphisms were also genotyped in the antecedent molar pregnancy, using DNA extracted from archival blocks of tissue. Response to methotrexate was evaluated with reference to serial human chorionic gonadotrophin (hCG) levels in patient serum. RESULTS: No significant association was found between the genotype of the patient, or presence of the variant allele, and clinical response to methotrexate therapy for either the 677C>T or the 1298A>C SNP. No significant association was found between the genotypes of the molar tissue and response to methotrexate. In molar tissue there was a significant reduction in the expected number with the 677TT genotype suggesting the 677C>T SNP may identify a subgroup of molar pregnancies less likely to progress to GTN. CONCLUSION: Neither the genotype for the 677C>T SNP or the 1298A>C SNP in MTHFR predict the therapeutic outcomes of women treated with single agent methotrexate for GTN.

Perspectives, Expectations, and Concerns of European Patient Advocates on Advanced Therapy Medicinal Products.

This paper presents the results of a qualitative study based on semi-structured interviews of 10 expert patient advocates on several different issues around Advanced Therapy Medicinal Products (ATMPs). The interviews were conducted between February and May 2020 based on a guideline with a list of 8 topics that covered concerns about safety and ethics, access problems and limitations, pricing of ATMPs and educational needs for patient communities. Overall, the interviewees expressed a high degree of convergence of opinions on most of the topics and especially on the identification of the reasons for concern. Conversely, when asked about possible solutions, quite a wide range of solutions were proposed, although with many common points. However, it highlights that the debate is still in its infancy and that there are not yet consolidated positions across the whole community. A general concern emerging from all the interviews is the potential limitation of access to approved ATMPs, both due to the high prices and to the geographical concentration of treatment centers. However, patients recognize the value of a model with a limited number of specialized clinical centers administering these therapies. On the ethical side, patients do not show particular concern as long as ATMPs and the underlying technology is used to treat severe diseases. Finally, patients are asking for both more education on ATMPs as well as for a more continuous involvement of patient representatives in the whole "life-cycle" of a new ATMP, from the development phase to the authorization, from the definition of the reimbursement scheme to the collection of Real Word Data on safety and long-term efficacy of the treatment.

Governance of Access in Biobanking: The Case of Telethon Network of Genetic Biobanks.

The discussion concerning the measure of the quality of a biobank should focus not only on the number of stored samples and their quality but also on the assessment of their access arrangements and governance. This article aims at contributing to the ongoing debate on samples and data access governance in biobanking by presenting the case of the Telethon Network of Genetic Biobanks (TNGB). We attempt to contribute to the need for clear and available access criteria and harmonization in access arrangements to maximize the influence of biobanks in the progress of biomedical research. We reviewed all the sample requests submitted to the TNGB from 2008 to 2020, focusing on those rejected by the Access Committee and the reasons behind the rejections. The analysis of the reasons behind the rejected requests allowed us to analyze how those relate to the issues of scientific misconduct, prioritization, and noncompliance with the biobank's mission. We discuss those issues in light of the actions and motivations used by TNGB in the access decision-making process. Based on this analysis, we suggest that a cross-implementation of a checklist for access assessment would improve the whole access process, ensuring a more transparent and smoother governance. Finally, we conclude that the TNGB's Charter and approach toward access governance could contribute as an important reference point to deal with the issues that have emerged in the international discussion on the topic.

"Integrating China in the International Consortium for Personalized Medicine": The Coordination and Support Action to Foster Collaboration in Personalized Medicine Development between Europe and China.

"Integrating China in the International Consortium for Personalized Medicine" (IC2PerMed) is a coordination and support action funded within the Horizon 2020 work program. Following the guidance of the International Consortium for Personalized Medicine (ICPerMed), the project's overarching aim is to align the European Union and China's research agendas in the field of personalized medicine (PM) to enable a swift development of PM approaches in the EU with strong leverage upon EU-Chinese collaboration. Living in the CO-VID-19 era, we are witnessing how the challenges imposed by the pandemic all around the globe have been acting as a catalyst for collaborations and knowledge sharing among national health systems worldwide. Given the strong interest on behalf of both Europe and China in the advancement of PM approaches, now more than ever, a cross-border collaboration between the 2 powers can accelerate the effective translation of such innovation to healthcare systems, advance research, and ensure that such change follows the directions toward the path of sustainability. IC2PerMed developments will be led by European and Chinese experts equally assembled into 3 Working Groups: (1) people and organization, (2) innovation and market, and (3) research and clinical studies in PM. This complex and dynamic network of actions thrives on dialog, cooperation, and alignment of research at national and global levels; work in the direction taken by IC2PerMed shall pave the way toward the realization of PM's full potential, prevent it from becoming a burden for healthcare systems, and, rather, prove that it provides an essential and irreplaceable contribution to their effectiveness, efficiency, and sustainability.

The RD-Connect Registry & Biobank Finder: a tool for sharing aggregated data and metadata among rare disease researchers.

In rare disease (RD) research, there is a huge need to systematically collect biomaterials, phenotypic, and genomic data in a standardized way and to make them findable, accessible, interoperable and reusable (FAIR). RD-Connect is a 6 years global infrastructure project initiated in November 2012 that links genomic data with patient registries, biobanks, and clinical bioinformatics tools to create a central research resource for RDs. Here, we present RD-Connect Registry & Biobank Finder, a tool that helps RD researchers to find RD biobanks and registries and provide information on the availability and accessibility of content in each database. The finder concentrates information that is currently sparse on different repositories (inventories, websites, scientific journals, technical reports, etc.), including aggregated data and metadata from participating databases. Aggregated data provided by the finder, if appropriately checked, can be used by researchers who are trying to estimate the prevalence of a RD, to organize a clinical trial on a RD, or to estimate the volume of patients seen by different clinical centers. The finder is also a portal to other RD-Connect tools, providing a link to the RD-Connect Sample Catalogue, a large inventory of RD biological samples available in participating biobanks for RD research. There are several kinds of users and potential uses for the RD-Connect Registry & Biobank Finder, including researchers collaborating with academia and the industry, dealing with the questions of basic, translational, and/or clinical research. As of November 2017, the finder is populated with aggregated data for 222 registries and 21 biobanks.

The alliance between genetic biobanks and patient organisations: the experience of the telethon network of genetic biobanks.

BACKGROUND: Rare diseases (RDs) are often neglected because they affect a small percentage of the population (6-8 %), which makes research and development of new therapies challenging processes. Easy access to high-quality samples and associated clinical data is therefore a key prerequisite for biomedical research. In this context, Genetic Biobanks are critical to developing basic, translational and clinical research on RDs. The Telethon Network of Genetic Biobanks (TNGB) is aware of the importance of biobanking as a service for patients and has started a dialogue with RD-Patient Organisations via promotion of dedicated meetings and round-tables, as well as by including their representatives on the TNGB Advisory Board. This has enabled the active involvement of POs in drafting biobank policies and procedures, including those concerning ethical issues. Here, we report on our experience with RD-Patient Organisations who have requested the services of existing biobanks belonging to TNGB and describe how these relationships were established, formalised and maintained. RESULTS: The process of patient engagement has proven to be successful both for lay members, who increased their understanding of the complex processes of biobanking, and for professionals, who gained awareness of the needs and expectations of the people involved. This collaboration has resulted in a real interest on the part of Patient Organisations in the biobanking service, which has led to 13 written agreements designed to formalise this process. These agreements enabled the centralisation of rare genetic disease biospecimens and their related data, thus making them available to the scientific community. CONCLUSIONS: The TNGB experience has proven to be an example of good practice with regard to patient engagement in biobanking and may serve as a model of collaboration between disease-oriented Biobanks and Patient Organisations. Such collaboration serves to enhance awareness and trust and to encourage the scientific community to address research on RDs.

Cbl-b mediates TGFß sensitivity by downregulating inhibitory SMAD7 in primary T cells.

T cell-intrinsic transforming growth factor ß (TGFß) receptor signaling plays an essential role in controlling immune responses. The RING-type E3 ligase Cbl-b has been shown to mediate the sensitivity of T cells to TGFß; however, the mechanism underlying this process is unknown. This study shows that SMAD7, an established negative regulator of TGFß receptor (TGFßR) signaling, is a key downstream effector target of Cbl-b. SMAD7 protein levels, but not SMAD7 mRNA levels, are upregulated in cblb(-/-) T cells. Cbl-b directly interacts with and ubiquitinates SMAD7, suggesting that Cbl-b posttranscriptionally regulates SMAD7. In support of this notion, concomitant genetic loss of SMAD7 in cblb(-/-) mice restored TGFß sensitivity on T cell cytokine responses and abrogated the tumor rejection phenotype of cblb(-/-) mice. These results demonstrate an essential and non-redundant role for Cbl-b in controlling TGFßR signaling by directly targeting SMAD7 for degradation during T cell responses in vitro and in vivo.

Targeting Cx43 and N-cadherin, which are abnormally upregulated in venous leg ulcers, influences migration, adhesion and activation of Rho GTPases.

BACKGROUND: Venous leg ulcers can be very hard to heal and represent a significant medical need with no effective therapeutic treatment currently available. PRINCIPAL FINDINGS: In wound edge biopsies from human venous leg ulcers we found a striking upregulation of dermal N-cadherin, Zonula Occludens-1 and the gap junction protein Connexin43 (Cx43) compared to intact skin, and in stark contrast to the down-regulation of Cx43 expression seen in acute, healing wounds. We targeted the expression of these proteins in 3T3 fibroblasts to evaluate their role in venous leg ulcers healing. Knockdown of Cx43 and N-cadherin, but not Zonula Occludens-1, accelerated cell migration in a scratch wound-healing assay. Reducing Cx43 increased Golgi reorientation, whilst decreasing cell adhesion and proliferation. Furthermore, Connexin43 and N-cadherin knockdown led to profound effects on fibroblast cytoskeletal dynamics after scratch-wounding. The cells exhibited longer lamelipodial protrusions lacking the F-actin belt seen at the leading edge in wounded control cells. This phenotype was accompanied by augmented activation of Rac-1 and RhoA GTPases, as revealed by Förster Resonance Energy Transfer and pull down experiments. CONCLUSIONS: Cx43 and N-cadherin are potential therapeutic targets in the promotion of healing of venous leg ulcers, by acting at least in part through distinct contributions of cell adhesion, migration, proliferation and cytoskeletal dynamics.

Abnormal connexin expression underlies delayed wound healing in diabetic skin.

OBJECTIVE: Dynamically regulated expression of the gap junction protein connexin (Cx)43 plays pivotal roles in wound healing. Cx43 is normally downregulated and Cx26 upregulated in keratinocytes at the edge of the wound as they adopt a migratory phenotype. We have examined the dynamics of Cx expression during wound healing in diabetic rats, which is known to be slow. RESEARCH DESIGN AND METHODS: We induced diabetes with streptozotocin and examined Cx expression and communication in intact and healing skin. RESULTS: We found that diabetes decreased Cx43 and Cx26 protein and communication in the intact epidermis and increased Cx43 protein and communication in the intact dermis. Diabetes also altered the dynamic changes of Cxs associated with wound healing. Within 24 h, Cx43 was upregulated in a thickened bulb of keratinocytes at the wound edge (rather than downregulated as in controls, which formed a thin process of migratory cells). Cx43 decline was delayed until 48 h, when reepithelialization began. Although Cx26 was upregulated as normal after wounding in diabetic skin, its distribution at the wound edge was abnormal, being more widespread. Application of Cx43-specific antisense gel to diabetic wounds prevented the abnormal upregulation of Cx43 and doubled the rate of reepithelialization, which exceeded control levels. CONCLUSIONS: Cx expression in diabetic skin is abnormal, as is the dynamic response of Cx43 to injury, which may underlie the delayed healing of diabetic wounds. Preventing the upregulation of Cx43 in diabetic wounds significantly improves the rate of healing and clearly has potential therapeutic value.

Acute downregulation of connexin43 at wound sites leads to a reduced inflammatory response, enhanced keratinocyte proliferation and wound fibroblast migration.

Experimental downregulation of connexin43 (Cx43) expression at skin wound sites appears to markedly improve the rate and quality of healing, but the underlying mechanisms are currently unknown. Here, we have compared physiological and cell biological aspects of the repair process with and without Cx43 antisense oligodeoxynucleotide treatment. Treated wounds exhibited accelerated skin healing with significantly increased keratinocyte and fibroblast proliferation and migration. In vitro knockdown of Cx43 in a fibroblast wound-healing model also resulted in significantly faster healing, associated with increased mRNA for TGF-beta1, and collagen alpha1 and general collagen content at the wound site. Treated wounds showed enhanced formation of granulation tissue and maturation with more rapid angiogenesis, myofibroblast differentiation and wound contraction appeared to be advanced by 2-3 days. Recruitment of both neutrophils and macrophages was markedly reduced within treated wounds, concomitant with reduced leukocyte infiltration. In turn, mRNA levels of CC chemokine ligand 2 and TNF-alpha were reduced in the treated wound. These data suggest that, by reducing Cx43 protein with Cx43-specific antisense oligodeoxynucleotides at wound sites early in the skin healing process repair is enhanced, at least in part, by accelerating cell migration and proliferation, and by attenuating inflammation and the additional damage it can cause.