Recommendations for in vitro evaluation of blood components collected, prepared and stored in non-DEHP medical devices

© 2022 International Society of Blood Transfusion. Funding Information: T.R.L.K., S.B. and D.K. led the working party and contributed to the writing of the manuscript. A.L., O.E.S., M.D.W., C.G., P.J.M.B., R.E., L.L., S.T., T.N., W.B., J.E. and B.M. contributed to the writing of the manuscript. Publisher Copyright: © 2022 International Society of Blood Transfusion.BACKGROUND AND OBJECTIVES: DEHP, di(2-ethylhexyl) phthalate, is the most common member of the class of ortho-phthalates, which are used as plasticizers. The Medical Device Regulation has restricted the use of phthalates in medical devices. Also DEHP has been added to the Annex XIV of REACH, "Registration, Evaluation, Authorisation and Restriction of Chemicals" due to its endocrine disrupting properties to the environment. As such, the sunset date for commercialisation of DEHP-containing blood bags is May 27th 2025. There are major concerns in meeting this deadline as these systems have not yet been fully validated and/or CE-marked. Also, since DEHP is known to affect red cell quality during storage, it is imperative to transit to non-DEHP without affecting blood product quality. Here, EBA members aim to establish common grounds on the evaluation and assessment of blood components collected, prepared and stored in non-DEHP devices. MATERIALS AND METHODS: Based on data as well as the input of relevant stakeholders a rationale for the validation of each component was composed. RESULTS: The red cell components will require the most extensive validation as their quality is directly affected by the absence of DEHP, as opposed to platelet and plasma components. CONCLUSION: Studies in the scope of evaluating the quality of blood products obtained with non-DEHP devices, under the condition that they are carried out according to these recommendations, could be used by all members of the EBA to serve as scientific support in the authorization process specific to their jurisdiction or for their internal validation use.Peer reviewe

Diffusion timescale conundrum in contact metamorphism: Oxygen isotope and trace element profiles in dolomite

Successful application of diffusion chronometry is based on detailed knowledge of diffusion parameters obtained from laboratory experiments, which also need to be upscaled to be applied to natural samples. While generally applicable, several examples of disagreement with other dating techniques and discrepancies with geological observations highlight the importance of validating diffusion parameters and their use to geochronologic application by testing them on samples from well-characterized geologic environments. The incomplete dissolution and reprecipitation reaction of dolomite associated with infiltration along grain boundaries of a fluid of magmatic origin in a contact metamorphic environment is the starting point of this study. The reaction produces dolomite rims, which are characterized by lower & delta;18O values (about 16%o) and slightly higher Fe (-1000 ppm) and Mn (-200 ppm) concentrations. The cores, in comparison, have no Fe and Mn and a & delta;18O value of about 30%o. This compositional and isotopic gradient led to chemical re-equilibration through diffusion at the reaction interface. We acquired oxygen isotope profiles using a secondary ion mass spectrometer (SIMS), with spot sizes as small as 3 & mu;m, and iron and manganese profiles with a NanoSIMS and a beam diameter of about 240 nm. These profiles were then compared against cumulative theoretical diffusion distances calculated for a modeled time - temperature path of the contact aureole using the experimentally determined diffusion parameters for O, Fe and Mn in dolomite. The oxygen isotope profiles yielded diffusion distances of ca. 10 & mu;m, corresponding to a formation temperature of about 550 degrees C, which is concordant with phase petrology and a timing for fluid infiltration at the onset of cooling of the contact aureole (i.e., non-isothermal evolution of the dolomite marbles). The Fe and Mn profiles were very sharp, with a diffusion length of about 150 nm. Such short diffusion length would be achieved at nearsurface temperature and corresponding cooling timescale are several orders of magnitude smaller than those derived from the oxygen diffusion profiles. From the mineralogy and textures, a decoupling of the oxygen isotopes and Fe-Mn reaction interface can be ruled out, suggesting that the published diffusion parameters are not fully representative, either due to the experimental conditions or by the assumed diffusion mechanism of natural processes. This case study is not the first example in the literature where the diffusion rate of Fe and Mn in natural dolomite seems to be substantially slower than predicted by experiments. Thus, there is a need to reassess the experimental diffusivities before Fe and Mn diffusion in dolomite can be reliably used to determine duration, cooling or heating paths of metamorphic processes.LG

Therapeutic assessment of targeting ASNS combined with L-Asparaginase treatment in solid tumors and investigation of resistance mechanisms

AbstractAsparagine deprivation by L-Asparaginase (L-ASNase) is an effective therapeutic strategy in Acute Lymphoblastic Leukemia, with resistance occurring due to upregulation of ASNS, the only human enzyme synthetizing Asparagine1. L-Asparaginase efficacy in solid tumors is limited by dose-related toxicities 2. Large-scale loss of function genetic in vitro screens identified ASNS as a cancer dependency in several solid malignancies 3,4. Here we evaluate the therapeutic potential of targeting ASNS in melanoma cells. While we confirm in-vitro dependency on ASNS silencing, this is largely dispensable for in vivo tumor growth, even in face of asparagine deprivation, prompting us characterize such resistance mechanism to devise novel therapeutic strategies. Using ex vivo quantitative proteome and transcriptome profiling, we characterize the compensatory mechanism elicited by ASNS knockout melanoma cells allowing their survival. Mechanistically, a genome-wide CRISPR screen revealed that such resistance mechanism is elicited by a dual axis: GCN2-ATF4 aimed at restoring amino acids levels and MAPK-BCLXL to promote survival. Importantly, pharmacological inhibition of such nodes synergizes with L-Asparaginase-mediated Asparagine deprivation in ASNS deficient cells suggesting novel potential therapeutic combinations in melanoma.</jats:p

Therapeutic Assessment of Targeting ASNS Combined with l-Asparaginase Treatment in Solid Tumors and Investigation of Resistance Mechanisms

Asparagine deprivation by L-Asparaginase (L-ASNase) is an effective therapeutic strategy in Acute Lymphoblastic Leukemia, with resistance occurring due to upregulation of ASNS,the only human enzyme synthetizing Asparagine1. L-Asparaginase efficacy in solid tumors is limited by dose-related toxicities 2. Large-scale loss of function genetic in vitro screens identified ASNSas a cancer dependency in several solid malignancies 3,4. Here we evaluate the therapeutic potential of targeting ASNS in melanoma cells. While we confirm in-vitro dependency on ASNS silencing, this is largely dispensable for in vivo tumor growth, even in face of asparagine deprivation, suggesting the need to further characterize such pathway to devise novel therapeutic strategies. Using ex vivo quantitative proteome and transcriptome profiling, we characterize the compensatory mechanism elicited by ASNS knockout melanoma cells allowing their survival. Additionally, genome-wide CRISPR screens upon manipulation of aminoacids levels identify BCLXL, MAPK and GCN2 as critical nodes mediating the observed resistance mechanism. Importantly, pharmacological inhibition of such hits synergizes with L-Asparaginase-mediated Asparagine deprivation in ASNS deficient cells suggesting novel potential therapeutic combinations in melanoma

Assessment of the soluble proteins HMGB1 , CD40L and CD62P during various platelet preparation processes and the storage of platelet concentrates: The BEST collaborative study

International audienceAbstract Background Structural and biochemical changes in stored platelets are influenced by collection and processing methods. This international study investigates the effects of platelet (PLT) processing and storage conditions on HMGB1, sCD40L, and sCD62P protein levels in platelet concentrate supernatants (PCs). Study Design/Methods PC supernatants ( n = 3748) were collected by each international centre using identical centrifugation methods ( n = 9) and tested centrally using the ELISA/Luminex platform. Apheresis versus the buffy coat (BC‐PC) method, plasma storage versus PAS and RT storage versus cold (4°C) were investigated. We focused on PC preparation collecting samples during early (RT: day 1–3; cold: day 1–5) and late (RT: day 4–7; cold: day 7–10) storage time points. Results HMGB1, sCD40L, and sCD62P concentrations were similar during early storage periods, regardless of storage solution (BC‐PC plasma and BC‐PC PAS‐E) or temperature. During storage and without PAS, sCD40L and CD62P in BC‐PC supernatants increased significantly (+33% and +41%, respectively) depending on storage temperature (22 vs. 4°C). However, without PAS‐E, levels decreased significantly (−31% and −20%, respectively), depending on storage temperature (22 vs. 4°C). Contrastingly, the processing method appeared to have greater impact on HMGB1 release versus storage duration. These data highlight increases in these parameters during storage and differences between preparation methods and storage temperatures. Conclusions The HMGB1 release mechanism/intracellular pathways appear to differ from sCD62P and sCD40L. The extent to which these differences affect patient outcomes, particularly post‐transfusion platelet increment and adverse events, warrants further investigation in clinical trials with various therapeutic indications