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Post-transplant lymphoproliferative disorder is a life-threatening complication of immunosuppression following transplantation mediated by failure of T cells to control Epstein–Barr virus (EBV)–infected and transformed B cells. Typically, a modification or reduction of immunosuppression is recommended, but insufficiently defined thus far. In order to help delineate this, we characterized EBV-antigen–specific T cells and lymphoblastoid cell lines from healthy donors and in patients with a kidney transplant in the absence or presence of the standard immunosuppressants tacrolimus, cyclosporin A, prednisolone, rapamycin, and mycophenolic acid. Phenotypes of lymphoblastoid cell-lines and T cells, T cell–receptor–repertoire diversity, and T-cell reactivity upon co-culture with autologous lymphoblastoid cell lines were analyzed. Rapamycin and mycophenolic acid inhibited lymphoblastoid cell-line proliferation. T cells treated with prednisolone and rapamycin showed nearly normal cytokine production. Proliferation and the viability of T cells were decreased by mycophenolic acid, while tacrolimus and cyclosporin A were strong suppressors of T-cell function including their killing activity. Overall, our study provides a basis for the clinical decision for the modification and reduction of immunosuppression and adds information to the complex balance of maintaining anti-viral immunity while preventing acute rejection. Thus, an immunosuppressive regime based on mTOR inhibition and reduced or withdrawn calcineurin inhibitors could be a promising strategy for patients with increased risk of or manifested EBV-associated post-transplant lymphoproliferative disorder.

Chimeric antigen receptor (CAR) T cells have emerged as a potent therapeutic approach for patients with certain haematological cancers, with multiple CAR T cell products currently approved by the FDA for those with relapsed and/or refractory B cell malignancies. However, in order to derive the desired level of effectiveness, patients need to successfully receive the CAR T cell infusion in a timely fashion. This process entails apheresis of the patient’s T cells, followed by CAR T cell manufacture. While awaiting infusion at an authorized treatment centre, patients may receive interim disease-directed therapy. Most patients will also receive a course of pre-CAR T cell lymphodepletion, which has emerged as an important factor in enabling durable responses. The time between apheresis and CAR T cell infusion is often not a simple journey, with each milestone being a critical step that can have important downstream consequences for the ability to receive the infusion and the strength of clinical responses. In this Review, we provide a summary of the many considerations for preparing patients with B cell non-Hodgkin lymphoma or acute lymphoblastic leukaemia for CAR T cell therapy, and outline current limitations and areas for future research.

Viral infections, such as with cytomegalovirus (CMV), remain a major risk factor for mortality and morbidity of transplant recipients because of their requirement for lifelong immunosuppression (IS). Antiviral drugs often cause toxicity and sometimes fail to control disease. Thus, regeneration of the antiviral immune response by adoptive antiviral T cell therapy is an attractive alternative. Our recent data, however, show only short-term efficacy in some solid organ recipients, possibly because of malfunction in transferred T cells caused by ongoing IS. We developed a vector-free clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-based good manufacturing practice (GMP)-compliant protocol that efficiently targets and knocks out the gene for the adaptor protein FK506-binding protein 12 (FKBP12), required for the immunosuppressive function of tacrolimus. This was achieved by transient delivery of ribonucleoprotein complexes into CMV-specific T cells by electroporation. We confirmed the tacrolimus resistance of our gene-edited T cell products in vitroand demonstrated performance comparable with non-tacrolimus-treated unmodified T cells. The alternative calcineurin inhibitor cyclosporine A can be administered as a safety switch to shut down tacrolimus-resistant T cell activity in case of adverse effects. Furthermore, we performed safety assessments as a prerequisite for translation to first-in-human applications.

The discovery of the highly efficient site-specific nuclease system CRISPR–Cas9 from Streptococcus pyogeneshas galvanized the field of gene therapy1,2. The immunogenicity of Cas9 nuclease has been demonstrated in mice3,4. Preexisting immunity against therapeutic gene vectors or their cargo can decrease the efficacy of a potentially curative treatment and may pose significant safety issues3–6. S. pyogenesis a common cause for infectious diseases in humans, but it remains unclear whether it induces a T cell memory against the Cas9 nuclease7,8. Here, we show the presence of a preexisting ubiquitous effector T cell response directed toward the most widely used Cas9 homolog from S. pyogenes(SpCas9) within healthy humans. We characterize SpCas9-reactive T cells within the CD4/CD8 compartments for multi-effector potency, cytotoxicity, and lineage determination. In-depth analysis of SpCas9-reactive T cells reveals a high frequency of SpCas9-reactive regulatory T cells that can mitigate SpCas9-reactive effector T cell proliferation and function in vitro. Our results shed light on T cell–mediated immunity toward CRISPR-associated nucleases and offer a possible solution to overcome the problem of preexisting immunity.

We have demonstrated how to create a parent surfactant that is open to manipulation, while maintaining its surface activity, robustness, and compatibility, to expand the landscape of surfactant-dependent microfluidic-droplet-based assays.To create a library of functional surfactants from the parent surfactant, an oxidation-responsive small building block with thioethers and multiple 1,2-diols was used as a head group, while a non-polar perfluoropolyether chain was used as a tail.

Activity of the CXCR3alt-CXCL11 chemokine system in CD8+stem-like T cells predicts chemotherapy response in patients with bladder cancer.

Background. Endometriosis is a chronic and estrogen-dependent pelvic inflammatory disease, which may have various causes, such as oxidative stress. Dysmenorrhea, dyspareunia, and pelvic pain are well-known symptoms of endometriosis. The present clinical trial assessed the role of supplementation with antioxidant vitamins on the indices of oxidative stress as well as the severity of pain in women with endometriosis. Materials and Methods. We enrolled 60 reproductive-aged (15–45 years) women with pelvic pain in this triple-blind clinical trial. They had 1–3 stages of laparoscopic-proven endometriosis. The participants were randomized to group A (n = 30), given vitamin C (1000 mg/day, 2 tablets of 500 mg each) and vitamin E (800 IU/day, 2 tablets of 400 IU each) combination, or group B (n = 30), given placebo pills daily for 8 weeks. Results. Following treatment with vitamin C and vitamin E, we found a significant reduction in MDA and ROS compared with the placebo group. There was no significant decline in total antioxidant capacity after treatment. However, the severity of pelvic pain (p value <0.001), dysmenorrhea (p value <0.001), and dyspareunia (p value <0.001) significantly decreased in the treatment group after 8 weeks of supplementation. Conclusions. The present findings support the potential role of antioxidants in the management of endometriosis. The intake of vitamin C and vitamin E supplements effectively reduced dysmenorrhea severity and improved dyspareunia and severity of pelvic pain.

Background Health-promoting lifestyle is an effective strategy for maintaining and controlling health, especially in immigrant women as a vulnerable group. Regarding this, the present study was conducted to determine the health promoting lifestyle and its associated factors in Afghan migrant women in Iran.Method This was a population based cross-sectional study in which 255 Afghan women of reproductive age. The study population was selected using the continuous sampling method from all Afghan women who referred to health centers of southwest of Tehran in 2018. The data were collected through the socio-demographic and Health Promoting Lifestyle Profile-II (HPLP-II) questionnaire.Results The total HPLP-II mean score of women was (124.05 ± 17.28). The lowest score was related to physical activity dimension (14.70 ± 3.78) and the highest score was related to spiritual growth (24.56 ± 5.06). Although some factors such as age, duration of education, income level, ability to speak Persian, education level of husband, and number of children significantly related factors to participant’s lifestyle (P<.05), but multiple regression model showed that income level and ability to speak Persian are final statistically related factors to Afghan women health promoting lifestyle in Iran.Conclusion Familiarization of the individual with the Persian language and the promotion of employment status should be considered due to the low socio-economic level and the fact that most women participating in the study are housewives.

Objective:To assess perceptions about vaginal examinations (VEs) during labor among women referred to the Akbarabadi Hospital in Tehran, Iran. Methods:This was a cross-sectional study conducted from December 2015 to May 2016. It included 200 primiparous women. Convenience sampling was used to select participants. The data collection tool was a questionnaire that collected both demographic and obstetric data from participants, as well as data in relation to women’s experiences according to a designated and validated scale, 24 hours after childbirth. Higher scores were indicative of a more positive perception in relation to a participant’s experience of VE. Results:The mean ± SD score for the perception of the VE and number of examinations in the active phase of labor was 62.9 ± 26.2 and 7.4 ± 2.44, respectively. Multiple linear regression analysis showed a significant relationship between perceived duration of examination (short, average [B = −1.03], long [B = 3.84]), feeling of comfort with the examiner (B = 2.73), and number of examiners (B = −0.81) with the mean scores of their perceptions (P< .05). These 3 variables accounted for 8% of the changes in the women’s perceptions score of VE. Conclusion:The majority of participants in this study underwent excessive VEs during labor. If obstetrically safe, a reduction in the number of examinations during labor along with decreased duration based on women’s perceptions could improve women’s perceptions of VE overall. Additionally, should each VE be performed by the same clinician, women’s perceptions in relation to VEs may also improve.

ObjectiveTo assess whether reshaping of the immune balance by infusion of autologous natural regulatory T cells (nTregs) in patients after kidney transplantation is safe, feasible, and enables the tapering of lifelong high dose immunosuppression, with its limited efficacy, adverse effects, and high direct and indirect costs, along with addressing several key challenges of nTreg treatment, such as easy and robust manufacturing, danger of over immunosuppression, interaction with standard care drugs, and functional stability in an inflammatory environment in a useful proof-of-concept disease model.DesignInvestigator initiated, monocentre, nTreg dose escalation, phase I/IIa clinical trial (ONEnTreg13).SettingCharité-University Hospital, Berlin, Germany, within the ONE study consortium (funded by the European Union).ParticipantsRecipients of living donor kidney transplant (ONEnTreg13, n=11) and corresponding reference group trial (ONErgt11-CHA, n=9).InterventionsCD4+ CD25+ FoxP3+ nTreg products were given seven days after kidney transplantation as one intravenous dose of 0.5, 1.0, or 2.5-3.0×106cells/kg body weight, with subsequent stepwise tapering of triple immunosuppression to low dose tacrolimus monotherapy until week 48.Main outcome measuresThe primary clinical and safety endpoints were assessed by a composite endpoint at week 60 with further three year follow-up. The assessment included incidence of biopsy confirmed acute rejection, assessment of nTreg infusion related adverse effects, and signs of over immunosuppression. Secondary endpoints addressed allograft functions. Accompanying research included a comprehensive exploratory biomarker portfolio.ResultsFor all patients, nTreg products with sufficient yield, purity, and functionality could be generated from 40-50 mL of peripheral blood taken two weeks before kidney transplantation. None of the three nTreg dose escalation groups had dose limiting toxicity. The nTreg and reference groups had 100% three year allograft survival and similar clinical and safety profiles. Stable monotherapy immunosuppression was achieved in eight of 11 (73%) patients receiving nTregs, while the reference group remained on standard dual or triple drug immunosuppression (P=0.002). Mechanistically, the activation of conventional T cells was reduced and nTregs shifted in vivo from a polyclonal to an oligoclonal T cell receptor repertoire.ConclusionsThe application of autologous nTregs was safe and feasible even in patients who had a kidney transplant and were immunosuppressed. These results warrant further evaluation of Treg efficacy and serve as the basis for the development of next generation nTreg approaches in transplantation and any immunopathologies.Trial registrationNCT02371434(ONEnTreg13) and EudraCT:2011-004301-24 (ONErgt11).

le Coutre: Novartis: Honoraria.

le Coutre: Novartis: Honoraria.

To date no single molecular biomarker, with proven clinical utility, that is predictive and identifies patients with more aggressive chronic myeloid leukaemia (CML) at diagnosis, has been reported. Such a molecular biomarker would enable stratification of patients according to aggressiveness of the malignancy and therefore at greater risk of progressing to blast crisis (BC). If these patients could be identified a more potent TKI could be prescribed at presentation or be considered for stem cell transplant up front. Recently a common polymorphic variant of growth factor independent 1 (GFI1), a zinc finger DNA binding transcriptional repressor, whereby serine is substituted by asparagine at residue 36 (S36N), was reported (Khandanpour et al.Blood. 2010; 115:22462) to be associated with a 1.6 fold increased risk of developing acute myeloid leukemia (AML). In addition, investigators reported a frequency of 11% of the S36N GFI1allele among AML patients, compared with 3-7% among the European populations. Reports suggest S36N GFI1variant fails to repress the target genes, including HOXA9, and as a consequence their expression levels are elevated. These observations are consistent with a critical role for GFI1in myeloid differentiation. In view of these findings we hypothesized that the GFI1S36N allele might influence CML progression, viaup-regulation of HOXA9, from a relatively indolent chronic phase (CP) to invariably fatal BC, characterized by presence of leukaemic stem cells, resistant to tyrosine kinase inhibitors in peripheral blood. Therefore, we assessed if S36N allele frequency was increased in CML and quantified GFI1expression.

Musashi 2(also known as MSI2), a mRNA binding protein, is reported to control critical stem cell fate decisions by binding to the 3’untranslated region of target mRNAs, thereby inhibiting translation. MSI2is preferentially expressed in hematopoietic tissue and in particular in early myeloid progenitors. Furthermore, investigators have shown MSI2expression is upregulated by HOXA9resulting in dysfunction of the regulatory pathway, leading to hematopoietic stem cell proliferation, impaired myeloid differentiation and is associated with worse prognosis in chronic myeloid leukemia (CML) and acute myeloid leukemia (Kharas, et al.Nat Med 2010; 16:903; Ito, et al.Nature 2010; 466:765). We verified MSI2levels are significantly increased in CML patients in blast crisis compared with those in chronic phase, irrespective of lymphoid or myeloid transformation (Kaeda, et al.Blood. 2011;118;supplement). Given these observations we sought to assess MSI2and HOXA9expression in BCR-ABL1negative myeloproliferative neoplasm (MPN) patients.

Kaeda: Novartis: Research Funding. le Coutre:Novartis: Research Funding.

Increased expression of Musashi 2 (also known as MSI2),a mRNA binding protein,is reported to trigger progression of chronic myeloid leukemia (CML) from chronic phase (CP) to blast crisis (BC), which is frequently fatal. The data imply that MSI2is upregulated by HOXA9leading to disruption of the critical hematopoietic stem cell (HSC) fate decisions viathe NUMB-NOTCH signaling pathway. These events lead to HSC proliferation, impaired myeloid differentiation and are associated with worse prognosis in CML and acute myeloid leukemia (Kharas, et al.Nat Med 2010; 16:903; Ito, et al.Nature 2010; 466:765). Previously, we confirmed increased MSI2levels in CML patients in blast crisis (BC) compared with those in CP, irrespective of lymphoid or myeloid transformation (Kaeda, et al.Blood. 2011;118;supplement). To further elucidate this regulatory pathway we assessed whether HOXA9expression correlated with increased MSI2mRNA levels and upregulation of NOTCH.