Plant-growth promoting activity of three fungal endophytes isolated from plants living in dehesas and their effect on Lolium multiflorum.

Endophytic fungi have been demonstrated to produce bioactive secondary metabolites, some of which promote plant growth. Three endophytic fungi isolated from healthy plants living in dehesas of Extremadura (Spain) were identified and evaluated for their ability to produce phytohormone-like substances, antioxidant activity, total polyphenol content, phosphate solubilization ability and siderophore and ammonia production. The filtrates and extracts produced by the three endophytes were applied to Lolium multiflorum seeds and seedlings under both in vitro and greenhouse conditions, to analyse their influence on plant growth traits such as germination, vigour index, chlorophyll data, number and length of leaves and roots, and dry weight. All three endophytes, which were identified as Fusarium avenaceum, Sarocladium terricola and Xylariaceae sp., increased the germination of L. multiflorum seeds by more than 70%. Shoot and root length, plant dry weight and the number of roots were positively affected by the application of fungal filtrates and/or extracts, compared with controls. The tentative HPLC-MS identification of phytohormone-like substances, such as gibberellin A2 and zeatin, or the antioxidant acetyl eugenol, may partially explain the mechanisms of L. multiflorum plant growth promotion after the application of fungal filtrates and/or extracts.

A closer look at 30 day hospital readmissions after head and neck cancer surgery.

PURPOSE: 30-day hospital readmissions after head and neck cancer surgery continue to be a significant source of patient harm and healthcare expenditure. While there is substantial data in the literature assessing predictive factors for readmissions after head and neck cancer surgery, there are a paucity of studies which attempt to understand if such readmissions are preventable. The goal of this paper is to determine factors associated with 30-day hospital readmissions after head and neck cancer surgery and to understand if these readmissions were preventable. MATERIALS AND METHODS: Retrospective review from a single academic tertiary care center. Patients readmitted within 30 days after undergoing surgery for cancers of the head and neck between 2015 and 2018 were identified. RESULTS: Over a 3-year period, 26 patients undergoing resection with or without reconstruction of head and neck cancers were readmitted to the hospital within 30 days of discharge. There were 15 (58%) men and 11 (42%) women with a mean age of 68 years (SD 14 years). Twenty-one (81%) patients had squamous cell carcinoma and 13 (50%) had a primary site in the oral cavity. Thirteen (50%) had undergone free or regional flap reconstruction. The indication for readmission was related to the surgical wound in 19 (73%) and to medical complications in 7 (27%). Each case was categorized as "possibly preventable" versus "uncertain if preventable" based on whether a reasonable and feasible change in management may have prevented readmission. Six (23%) readmissions were deemed possibly preventable. Four were related to the surgical wound where initial free or regional flaps may have prevented complication. Two were medical complications that may have benefited from longer inpatient observation. CONCLUSIONS: For a subset of patients readmitted within 30 days of head and neck cancer surgery, a reasonable and feasible change in management may have prevented their hospital readmission. The significance of better understanding this patient population is underscored by the high mortality rate.

Mitochondrial DNA copy number is associated with all-cause mortality and cardiovascular events in patients with peripheral arterial disease.

BACKGROUND: Dysfunctional mitochondria have an influence on inflammation and increased oxidative stress due to an excessive production of reactive oxygen species. The mitochondrial DNA copy number (mtDNA-CN) is a potential biomarker for mitochondrial dysfunction and has been associated with various diseases. However, results were partially contrasting which might have been caused by methodological difficulties to quantify mtDNA-CN. OBJECTIVE: We aimed to investigate whether mtDNA-CN is associated with peripheral arterial disease (PAD) as well as all-cause mortality and cardiovascular events during seven years of follow-up. METHODS: A total of 236 male patients with PAD from the Cardiovascular Disease in Intermittent Claudication (CAVASIC) study were compared with 249 age- and diabetes-matched controls. MtDNA-CN was measured with a well-standardized plasmid-normalized quantitative PCR-based assay determining the ratio between mtDNA-CN and nuclear DNA. RESULTS: Individuals in the lowest quartile of mtDNA-CN had a twofold increased risk for PAD which, however, was no longer significant after adjusting for leukocytes and platelets. About 67 of the 236 patients had already experienced a cardiovascular event at baseline and those in the lowest mtDNA-CN quartile had a 2.34-fold increased risk for these events (95% CI 1.08-5.13). During follow-up, 37 PAD patients died and 66 patients experienced a cardiovascular event. Patients in the lowest mtDNA-CN quartile had hazard ratios of 2.66 (95% CI 1.27-5.58) for all-cause-mortality and 1.82 (95% CI 1.02-3.27) for cardiovascular events compared with the combined quartile 2-4 (adjusted for age, smoking, CRP, diabetes, prevalent cardiovascular disease, leukocytes and platelets). CONCLUSION: This investigation supports the hypothesis of mitochondrial dysfunction in peripheral arterial disease and shows an association of low mtDNA-CNs with all-cause-mortality and prevalent and incident cardiovascular disease in PAD patients with intermittent claudication.

Stromal-derived IGF2 promotes colon cancer progression via paracrine and autocrine mechanisms.

The insulin-like growth factor (IGF)2/IGF1 receptor (IGF1R) signaling axis has an important role in intestinal carcinogenesis and overexpression of IGF2 is an accepted risk factor for colorectal cancer (CRC) development. Genetic amplifications and loss of imprinting contribute to the upregulation of IGF2, but insufficiently explain the extent of IGF2 expression in a subset of patients. Here, we show that IGF2 was specifically induced in the tumor stroma of CRC and identified cancer-associated fibroblasts (CAFs) as the major source. Further, we provide functional evidence that stromal IGF2, via the paracrine IGF1R/insulin receptor axis, activated pro-survival AKT signaling in CRC cell lines. In addition to its effects on malignant cells, autocrine IGF2/IGF1R signaling in CAFs induced myofibroblast differentiation in terms of alpha-smooth muscle actin expression and contractility in floating collagen gels. This was further augmented in concert with transforming growth factor-ß (TGFß) signaling suggesting a cooperative mechanism. However, we demonstrated that IGF2 neither induced TGFß/smooth muscle actin/mothers against decapentaplegic (SMAD) signaling nor synergized with TGFß to hyperactivate this pathway in two dimensional and three dimensional cultures. IGF2-mediated physical matrix remodeling by CAFs, but not changes in extracellular matrix-modifying proteases or other secreted factors acting in a paracrine manner on/in cancer cells, facilitated subsequent tumor cell invasion in organotypic co-cultures. Consistently, colon cancer cells co-inoculated with CAFs expressing endogenous IGF2 in mouse xenograft models exhibited elevated invasiveness and dissemination capacity, as well as increased local tumor regrowth after primary tumor resection compared with conditions with IGF2-deficient CAFs. In line, expression of IGF2 correlated with elevated relapse rates and poor survival in CRC patients. In agreement with our results, high-level coexpression of IGF2 and TGFß was predicting adverse outcome with higher accuracy than increased expression of the individual genes alone. Taken together, we demonstrate that stroma-induced IGF2 promotes colon cancer progression in a paracrine and autocrine manner and propose IGF2 as potential target for tumor stroma cotargeting strategies.

Risk of sinusoidal obstruction syndrome in allogeneic stem cell transplantation after prior gemtuzumab ozogamicin treatment: a retrospective study from the Acute Leukemia Working Party of the EBMT.

Gemtuzumab ozogamicin (GO) may increase the risk of sinusoidal obstruction syndrome (SOS) when used prior to allogeneic stem cell transplantation (HSCT). We assessed SOS incidence and outcomes after HSCT of 146 adults, with a median age of 50 years, previously receiving GO. SOS prophylaxis was used in 69 patients (heparin n=57, ursodeoxycholic acid n=8, defibrotide n=4). Cumulative incidence (CI) of SOS was 8% (n=11), with death in 3 patients. Median interval between last GO dose and HSCT was 130 days. Overall survival (OS) and SOS incidence did not differ for patients receiving GO ⩽3.5 months before HSCT and the others. CI of acute and chronic GVHD was 31% and 25%, respectively. Probability of OS and leukemia-free survival (LFS) at 5 years was 40% and 37%, respectively. Relapse incidence and non-relapse mortality were 42% and 21%, respectively. In multivariate analysis, active disease at HSCT was associated with relapse and worse LFS and OS (P<0.03). Liver abnormalities before HSCT correlated with worse OS (P<0.03). Use of low-dose GO prior to HSCT is associated with an acceptable SOS incidence. Prospective studies investigating the role and the utility of SOS prophylaxis are warranted.

The proteome pattern cGvHD_MS14 allows early and accurate prediction of chronic GvHD after allogeneic stem cell transplantation.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) may be curative, but is associated with significant morbidity and mortality. Chronic graft-versus-host disease (cGvHD), characterized by inflammation and fibrosis of multiple target organs, considerably contributes to the morbidity and mortality even years after allo-HSCT. Diagnosis of cGvHD is based on clinical features and histology of biopsies. Here, we report the generation of a urinary cGvHD-specific proteome-pattern (cGvHD_MS14) established by capillary electrophoresis-mass spectrometry to predict onset and severity of cGvHD as an unbiased laboratory test. cGvHD_MS14 was evaluated on samples from 412 patients collected prospectively in four transplant centers. Sensitivity and specificity was 84 and 76% by cGvHD_MS14 classification. Sensitivity further increased to 93% by combination of cGvHD_MS14 with relevant clinical variables to a logistic regression model. cGvHD was predicted up to 55 days prior to clinical diagnosis. Acute GvHD is not recognized by cGvHD_MS14. cGvHD_MS14 consists of 14 differentially excreted peptides, six of those have been sequenced to date and are fragments from thymosin ß-4, eukaryotic translation initiation factor 4γ2, fibrinogen ß-chain or collagens. In conclusion, the cGvHD_MS14-pattern allows early, highly sensitive and specific prediction of cGvHD as an independent diagnostic criterion of clinical diagnosis potentially allowing early therapeutic intervention.

Efficacy and safety of deferasirox in non-thalassemic patients with elevated ferritin levels after allogeneic hematopoietic stem cell transplantation.

Elevated serum ferritin contributes to treatment-related morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). The multicenter DE02 trial assessed the safety, efficacy and impact of deferasirox on iron homeostasis after allogeneic HSCT. Deferasirox was administered at a starting dose of 10 mg/kg per day to 76 recipients of allogeneic HSCT, with subsequent dose adjustments based on efficacy and safety. Deferasirox was initiated at a median of 168 days after HSCT, with 84% of patients still on immunosuppression. Baseline serum ferritin declined from 2045 to 957 ng/mL. Deferasirox induced a negative iron balance in 84% of patients. Hemoglobin increased in the first 3 months, and trough serum cyclosporine levels were stable. Median exposure was 330 days, with a median compliance rate of >80%. The most common investigator-reported drug-related adverse events (AEs) were increased blood creatinine (26.5%), nausea (9.0%) and abdominal discomfort (8.3%). Fifty-four (71.1%) patients experienced drug-related AEs, which occasionally resulted in discontinuation (gastrointestinal (n=6), skin (n=3), elevated transaminases (n=1) and creatinine (n=1)). The incidence of AEs appeared to be dose related, with 7.5 mg/kg per day being the best-tolerated dose. Low-dose deferasirox is an effective chelation therapy after allogeneic HSCT, with a manageable safety profile, even in patients receiving cyclosporine.

Tyrosine phosphatase SHP2 increases cell motility in triple-negative breast cancer through the activation of SRC-family kinases.

Tumor cell migration has a fundamental role in early steps of metastasis, the fatal hallmark of cancer. In the present study, we investigated the effects of the tyrosine phosphatase, SRC-homology 2 domain-containing phosphatase 2 (SHP2), on cell migration in metastatic triple-negative breast cancer (TNBC), an aggressive disease associated with a poor prognosis for which a targeted therapy is not yet available. Using mouse models and multiphoton intravital imaging, we have identified a crucial effect of SHP2 on TNBC cell motility in vivo. Further, analysis of TNBC cells revealed that SHP2 also influences cell migration, chemotaxis and invasion in vitro. Unbiased phosphoproteomics and biochemical analysis showed that SHP2 activates several SRC-family kinases and downstream targets, most of which are inducers of migration and invasion. In particular, direct interaction between SHP2 and c-SRC was revealed by a fluorescence resonance energy transfer assay. These results suggest that SHP2 is a crucial factor during early steps of TNBC migration to distant organs.

Treatment of rare co-occurrence of Epstein-Barr virus-driven post-transplant lymphoproliferative disorder and hemophagocytic lymphohistiocytosis after allogeneic stem cell transplantation.

In both conditions, post-transplant lymphoproliferative disorder (PTLD) and hemophagocytic lymphohistiocytosis (HLH), infection with Epstein-Barr virus (EBV) is a key mechanism: almost all PTLD in allogeneic stem cell transplantation (alloSCT) is caused by EBV-related neoplastic lymphoproliferation, and secondary HLH is most frequently triggered by EBV infection. Therefore, concomitant EBV-driven PTLD and HLH early after alloSCT require an approach to eliminate EBV and balance immune activation simultaneously. We report on a patient who developed simultaneous PTLD and signs of HLH on day 64 after alloSCT. Treatment was comprised of stopping cyclosporine, short-course dexamethasone, and 3 courses of rituximab. The patient showed full recovery and complete remission of lymphadenopathy. This result indicates that immediate reduction in EBV-carrying B cells by rituximab, suppression of general inflammation, and parallel support of reconstitution of long-term T-cell function, might be an appropriate therapeutic approach in this rare situation.

Risk-adapted, treosulfan-based therapy with auto- and allo-SCT for relapsed/refractory aggressive NHL: a prospective phase-II trial.

Since the outcome of relapsed/refractory aggressive non-Hodgkin's lymphoma (NHL) is highly variable, a risk-adapted treatment approach was evaluated. After two cycles of DHAP, patients received high-dose treosulfan/etoposide/carboplatinum (TEC) and autologous stem cell rescue. After TEC, low-risk patients with late relapse (>1 year after first CR who achieved CR after DHAP received no further treatment. Patients with late relapse who achieved CR or PR only after TEC underwent a second cycle of TEC. High-risk patients with early relapse/refractory disease received treosulfan/fludarabine followed by allogeneic transplantation. Rituximab was added in patients with B-cell lymphoma (86%). At entry, 36% of all 57 patients had refractory disease, 32% early and 32% late relapse. During DHAP treatment, progression occurred in 32% of patients. Of 33 patients who received TEC, 5 received second TEC and 15 allogeneic transplantation. Main toxicity after TEC was oral mucositis (CTC grades 3 and 4 in 50% and 13%, respectively). In total, 42% patients achieved CR. Median OS was 21.4 months for all patients and 32.6 for those who underwent allogeneic transplantation. International prognostic index (IPI) at study entry was highly discriminative at predicting OS (P<0.0001). Risk-adapted, treosulfan-based therapy with auto- and allo-SCT is feasible. Long-term survival is possible with allogeneic transplantation.

Proteomic peptide profiling for preemptive diagnosis of acute graft-versus-host disease after allogeneic stem cell transplantation.

Allogeneic hematopoietic stem cell transplantation is one curative treatment for hematological malignancies, but is compromised by life-threatening complications, such as severe acute graft-versus-host disease (aGvHD). Prediction of severe aGvHD as early as possible is crucial to allow timely initiation of treatment. Here we report on a multicentre validation of an aGvHD-specific urinary proteomic classifier (aGvHD_MS17) in 423 patients. Samples (n=1106) were collected prospectively between day +7 and day +130 and analyzed using capillary electrophoresis coupled on-line to mass spectrometry. Integration of aGvHD_MS17 analysis with demographic and clinical variables using a logistic regression model led to correct classification of patients developing severe aGvHD 14 days before any clinical signs with 82.4% sensitivity and 77.3% specificity. Multivariate regression analysis showed that aGvHD_MS17 positivity was the only strong predictor for aGvHD grade III or IV (P<0.0001). The classifier consists of 17 peptides derived from albumin, ß2-microglobulin, CD99, fibronectin and various collagen α-chains, indicating inflammation, activation of T cells and changes in the extracellular matrix as early signs of GvHD-induced organ damage. This study is currently the largest demonstration of accurate and investigator-independent prediction of patients at risk for severe aGvHD, thus allowing preemptive therapy based on proteomic profiling.

Expansion of recipient-derived antiviral T cells may influence donor chimerism after allogeneic stem cell transplantation.

Donor chimerism (DC) analysis is an important marker in the hematopoietic stem cell transplant follow-up. Here, we present evidence for a possible relationship of infectious complications and declines in DC. We analyzed the DC in patients experiencing cytomegalovirus (CMV) reactivation. In addition, in some patients chimerism analyses of T-cell subsets were performed. CMV-specific cytotoxic T-lymphocytes (CMV-CTL) were monitored using human leukocyte antigen-restricted multimer staining. Interestingly, CMV reactivation was accompanied by changes in DC in 11 of 67 patients transplanted. For example, DC declined in a cord blood recipient, in both total leukocytes and CD4 and CD8 T-cell subsets upon CMV reactivation. The latter was controlled after only 5 days through expanding CMV-CTL of 96% recipient origin, according to chimerism analysis of CMV-CTL (enriched beyond 50%). In another patient, transplanted after reduced-intensity conditioning from a DQB1 mismatched, CMV seronegative donor, incipient CMV reactivation was completely aborted by CMV-CTL of recipient origin. However, at the same time, mixed chimerism dropped from 51% to 0% donor type, resulting in late graft rejection. Our data indicate that chimerism analyses in subset populations lead to a better understanding of declining total leukocyte chimerism. Furthermore, recipient-derived CMV-CTL may be able to control CMV reactivation after reduced-intensity conditioning. We speculate that autologous CMV-CTL may be instrumental to overcome recurrent CMV reactivations, especially in patients transplanted from CMV-seronegative donors. In addition, the expansion of recipient-derived CMV-CTL may contribute to both, graft failure or to conversion to full DC.

Randomized comparison of prophylactic and minimal residual disease-triggered imatinib after allogeneic stem cell transplantation for BCR-ABL1-positive acute lymphoblastic leukemia.

Minimal residual disease (MRD) after allogeneic stem cell transplantation (SCT) for Ph+ acute lymphoblastic leukemia (ALL) is predictive of relapse. Imatinib administration subsequent to SCT may prevent relapse, but the role of scheduling and its impact on outcome are not known. In a prospective, randomized multicenter trial, we compared the tolerability and efficacy of post-transplant imatinib administered either prophylactically (arm A; n=26) or following detection of MRD (arm B; n=29). Prophylactic imatinib significantly reduced the incidence of molecular recurrence after SCT compared with MRD-triggered imatinib (40% vs 69%; P=0.046). Median duration of PCR negativity was 26.5 and 6.8 months, respectively (P=0.065). Five-year survival in both interventional groups was high (80 and 74.5%), despite premature discontinuation of imatinib in the majority of patients because of poor tolerability. Relapse probability was significantly higher in patients who became MRD positive (P=0.017). In conclusion, post-transplant imatinib results in a low relapse rate, durable remissions and excellent long-term outcome in patients with BCR-ABL1-positive ALL irrespective of whether it is given prophylactically or MRD-triggered. Reappearance of BCR-ABL1 transcripts early after SCT or at higher levels identifies a small subset of patients who do not benefit sufficiently from imatinib, and in whom alternative approaches should be explored.

Renal comorbidity after solid organ and stem cell transplantation.

After transplantation of solid organs or hematopoietic stem cells, a significant acute decrease in renal function occurs in the majority of patients. Depending on the degree of kidney injury, a large number of patients develop chronic kidney disease (CKD) and some develop end-stage renal disease requiring renal replacement therapy. The incidence varies depending on the transplanted organ, but important risk factors for the development of CKD are preexisting renal disease, hepatitis C, diabetes, hypertension, age, sex, posttransplant acute kidney injury and thrombotic microangiopathy. This review article focuses on the risk factors of posttransplant chronic kidney disease after organ transplantation, considering the current literature and integrates the incidence and the associated mortality rates of acute and chronic kidney disease. Furthermore, we introduce the RECAST (REnal Comorbidity After Solid organ and hematopoietic stem cell Transplantation) registry.

Outcomes and prognostic factors of adults with acute lymphoblastic leukemia who relapse after allogeneic hematopoietic cell transplantation. An analysis on behalf of the Acute Leukemia Working Party of EBMT.

To describe outcomes, treatment and prognostic factors that influence survival of adult patients with acute lymphoblastic leukemia (ALL), who relapsed after allogeneic hematopoietic cell transplantation (HCT), we retrospectively analyzed 465 ALL adult patients from European Group for Blood and Marrow Transplantation (EBMT) centers who relapsed after a first HCT performed in complete remission (CR1 65%, CR2/3 35%). Salvage treatments were: supportive care (13%), cytoreductive therapy (43%), donor lymphocyte infusion without or with prior chemotherapy (23%) and second HCT (20%). Median time from HCT to relapse was 6.9 months, median follow-up was 46 months and median survival after relapse was 5.5 months. Estimated 1-, 2- and 5-year post-relapse survival was 30 ± 2%, 16 ± 2% and 8 ± 1%, respectively. In a multivariate analysis, adverse factors for survival were: late CR (CR2/3) at transplant (P<0.012), early relapse after transplant (<6.9 months, P <0.0001) and peripheral blast percent at relapse (P <0.0001). On the basis of multivariate model for survival, three groups of patients were identified with estimated 2 year survival of 6 ± 2, 17 ± 3 and 30 ± 7%. Outcome of ALL patients relapsing after HCT is dismal and there is a need for new therapies. Our study provides the standard expectations in ALL relapse and may help in the decision of post-relapse therapy.

Tetramer monitoring to assess risk factors for recurrent cytomegalovirus reactivation and reconstitution of antiviral immunity post allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Reactivation of cytomegalovirus (CMV) is a major cause of morbidity after allogeneic hematopoietic stem cell transplantation (HSCT). In healthy individuals, virus-specific T cells (CMV-CTL) control the reactivation of latent CMV. The monitoring of virus-epitope-binding CD8(+) T cells using major histocompatibility complex-I-peptide complexes (tetramers) has recently been established, allowing assessment of the reconstitution of CMV-CTL post HSCT. PATIENTS AND METHODS: In order to study immune reconstitution and reactivation control through CMV-CTL, we regularly monitored all patients undergoing allogeneic HSCT in our department for 2 years, who matched at least 1 of 6 commercially available tetramers for common human leukocyte antigen (HLA) types. To verify risk factors for CMV reactivations in our cohorts, clinical characteristics of all patients transplanted within the last 10 years were included in statistical analyses determining the relative risk for single and recurrent CMV reactivations. RESULTS: As expected, CMV serostatus, HLA match, and donor source significantly influenced the risk of recurrent CMV reactivation. Applying CMV-CTL tetramer monitoring for 2 years allowed the monitoring of 114 (85%) of 134 patients, by testing a set of tetramers representing 6 epitopes from 3 different CMV proteins. The presence of CMV-CTL before day + 50 and their expansion post reactivation seem to protect against recurrent CMV reactivations. The mean number of CMV-CTL by day +100 was >5-fold higher in the recipient CMV-positive/donor-positive (R +/D +) group (91/µL) compared with the R +/ D- (13/µL) and the R -/D +(2/µL) group. Seventy-nine percent of patients from the R +/D + setting recovered >10 CMV-CTL per µL by day + 100, while almost 50% of the other groups failed to mount a CMV-specific response by that time (R +/D -: 58%; R -/D +: 43%). CONCLUSION: Tetramer monitoring can help to predict (recurrent) CMV reactivation and is a useful approach to monitor individual patients with increased risk for recurrent reactivation post HSCT; thus, it could help to identify patients in need of adoptive transfer of CMV-CTL or to optimize the use of antiviral drugs.