Post-transplant lymphoproliferative disorders and Epstein-Barr virus DNAemia in a cohort of lung transplant recipients

<p>Abstract</p> <p>Background</p> <p>Post-transplant lymphoproliferative disorders (PTLD) are serious complications in lung transplant recipients. No consensus on EBV DNAemia levels predictive of PTLD has been reached. In addition, in many instances EBV DNAemia is determined in patients with suggestive symptoms only.</p> <p>Methods</p> <p>The characteristics of five patients with PTLD as well as the prevalence of EBV DNAmia in a cohort of 137 consecutive patients receiving lung transplantation are described.</p> <p>Results</p> <p>Twenty-six out of 137 patients (18.9%) were excluded from the analysis because lost at follow-up or dead from PTLD-independent reasons within three months of transplantation. EBV DNA in peripheral blood mononuclear cells (PBMC) was determined in 83/111 patients (74.8%) because of potential PTLD-related symptoms, while 28 patients (25.2%) showed no symptoms and were not examined. EBV DNAemia was positive in 53/83 patients (63.8%), and negative in 30/83 patients (36.2%). PTLD was diagnosed in five (4.5%) patients at a median time of 270 (range 120-870) days following transplantation. All five PTLD (three large B-cell lymphomas, one Hodgkin lymphoma and one possible pre-neoplastic lesion) were potentially associated with EBV infection. However, only 3/5 patients with PTLD had detectable EBV DNAemia: < 1,000 copies EBV DNA/1 × 10⁵PBMC in one patient and > 1,000 copies EBV DNA/1 × 10⁵PBMC in two patients.</p> <p>Conclusion</p> <p>A systematic multidisciplinary (clinical, radiologic, virologic and histologic) approach is mandatory for the diagnosis and management of PTLD in lung transplant recipients, while monitoring of symptomatic patients only may provide an incomplete or late picture of the clinical problem. In addition, staining for EBV antigens and quantification of EBV DNA in biopsy specimens should always be performed to understand the role of EBV infection in the pathogenesis of PTLD.</p

A phase II trial of the oral mTOR inhibitor everolimus in relapsed aggressive lymphoma

The phosphatidylinositol 3-kinase signal transduction pathway members are often activated in tumor samples from patients with non-Hodgkin's lymphoma (NHL). Everolimus is an oral agent that targets the raptor mammalian target of rapamycin (mTORC1). The goal of this trial was to learn the antitumor activity and toxicity of single-agent everolimus in patients with relapsed/refractory aggressive NHL. Patients received everolimus 10 mg PO daily. Response was assessed after two and six cycles, and then every three cycles until progression. A total of 77 patients with a median age of 70 years were enrolled. Patients had received a median of three previous therapies and 32% had undergone previous transplant. The overall response rate (ORR) was 30% (95% confidence interval: 20–41%), with 20 patients achieving a partial remission and 3 a complete remission unconfirmed. The ORR in diffuse large B cell was 30% (14/47), 32% (6/19) in mantle cell and 38% (3/8) in follicular grade 3. The median duration of response was 5.7 months. Grade 3 or 4 anemia, neutropenia and thrombocytopenia occurred in 14, 18 and 38% of patients, respectively. Everolimus has single-agent activity in relapsed/refractory aggressive NHL and provides proof-of-concept that targeting the mTOR pathway is clinically relevant

Modulation of apoptosis in human hepatocellular carcinoma (HepG2 cells) by a standardized herbal decoction of Nigella sativa seeds, Hemidesmus indicus roots and Smilax glabra rhizomes with anti- hepatocarcinogenic effects

<p>Abstract</p> <p>Background</p> <p>A standardized poly-herbal decoction of <it>Nigella sativa </it>seeds, <it>Hemidesmus indicus </it>roots and <it>Smilax glabra </it>rhizomes used traditionally in Sri Lanka for cancer therapy has been demonstrated previously, to have anti-hepatocarcinogenic potential. Cytotoxicity, antioxidant activity, anti-inflammatory activity, and up regulation of p53 and p21 activities are considered to be some of the possible mechanisms through which the above decoction may mediate its anti-hepatocarcinogenic action. The main aim of the present study was to determine whether apoptosis is also a major mechanism by which the decoction mediates its anti-hepatocarcinogenic action.</p> <p>Methods</p> <p>Evaluation of apoptosis in HepG2 cells was carried out by (a) microscopic observations of cell morphology, (b) DNA fragmentation analysis, (c) activities of caspase 3 and 9, as well as by (d) analysis of the expression of pro-apoptotic (Bax) and anti-apoptotic (Bcl-2) proteins associated with cell death.</p> <p>Results</p> <p>The results demonstrated that in HepG2 cells, the decoction can induce (a) DNA fragmentation and (b) characteristic morphological changes associated with apoptosis (nuclear condensation, membrane blebbing, nuclear fragmentation and apoptotic bodies). The decoction could also, in a time and dose dependent manner, up regulate the expression of the pro-apoptotic gene <it>Bax </it>and down regulate expression of anti-apoptotic <it>Bcl-2 </it>gene (as evident from RT-PCR analysis, immunohistochemistry and western blotting). Further, the decoction significantly (<it>p </it>< .001) enhanced the activities of caspase-3 and caspase-9 in a time and dose dependent manner.</p> <p>Conclusions</p> <p>Overall findings provide confirmatory evidence to demonstrate that the decoction may mediate its reported anti-hepatocarcinogenic effect, at least in part, through modulation of apoptosis.</p

A mouse model for oral squamous cell carcinoma

Despite recent advances, the prognosis of oral squamous cell carcinoma is still poor. Therapeutic options such as radiotherapy, chemotherapy, surgery and the novel treatment option gene therapy are being investigated in animal models. Diverse models have been studied to induce oral squamous cell carcinomas. The carcinogenic 4-nitroquinoline-1-oxide (4NQO) model has proven to be successful although until now it is unknown at what time point the established tumor is a representative squamous cell carcinoma and has a suitable volume for scientific treatment. For this end we applied 4NQO 3 times a week during 16 weeks and measured the volume of tumor tissue each week until the end of the experiment at 40 weeks. Concurrent histopathology at different time points up to the end of the experiment revealed that all mice bearing oral tumors were diagnosed with squamous cell carcinoma. Immunohistochemistry with markers cyclin D1 and E-cadherin revealed that the generated mouse oral tumors showed strong similarities with the described immunopathology in human oral tumors. The 4NQO model is a suitable alternative for preclinical gene therapy experiments with primary oral tumors. Future survey of therapeutic options in the carcinogenic 4NQO model should be conducted around 40 weeks after the start of the treatment

In vivo and in vitro assessment of pathways involved in contrast media-induced renal cells apoptosis

Contrast-induced nephropathy accounts for >10% of all causes of hospital-acquired renal failure, causes a prolonged in-hospital stay and represents a powerful predictor of poor early and late outcome. Mechanisms of contrast-induced nephropathy are not completely understood. In vitro data suggests that contrast media (CM) induces a direct toxic effect on renal tubular cells through the activation of the intrinsic apoptotic pathway. It is unclear whether this effect has a role in the clinical setting. In this work, we evaluated the effects of CM both in vivo and in vitro. By analyzing urine samples obtained from patients who experienced contrast-induced acute kidney injury (CI-AKI), we verified, by western blot and immunohistochemistry, that CM induces tubular renal cells apoptosis. Furthermore, in cultured cells, CM caused a dose–response increase in reactive oxygen species (ROS) production, which triggered Jun N-terminal kinases (JNK1/2) and p38 stress kinases marked activation and thus apoptosis. Inhibition of JNK1/2 and p38 by different approaches (i.e. pharmacological antagonists and transfection of kinase-death mutants of the upstream p38 and JNK kinases) prevented CM-induced apoptosis. Interestingly, N-acetylcysteine inhibited ROS production, and thus stress kinases and apoptosis activation. Therefore, we conclude that CM-induced tubular renal cells apoptosis represents a key mechanism of CI-AKI

Efficacy of adenovirally expressed soluble TRAIL in human glioma organotypic slice culture and glioma xenografts

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in malignant cells, including gliomas, and is currently in anticancer clinical trials. However, the full-length and tagged forms of TRAIL, unlike the untagged ligand (soluble TRAIL (sTRAIL)), exhibits toxicity against normal cells. Here, we report the generation and testing of an adenovirus (AdsTRAIL) that expresses untagged sTRAIL in an intracranial xenograft model and a human glioma organotypic slice culture model. AdsTRAIL efficiently induced apoptosis in glioma cell lines, including those resistant to sTRAIL, but not in normal human astrocytes (NHAs). It inhibited anchorage-independent glioma growth and exerted a bystander effect in transwell assays. Intratumoral injections of AdsTRAIL in a rodent intracranial glioma model resulted in reduced tumor growth and improved survival compared with Ad-enhanced green fluorescent protein (EGFP)- or vehicle-treated controls without toxicity. Human glioma organotypic slices treated with AdsTRAIL demonstrated apoptosis induction and caspase activation

Efficient Induction of Extrinsic Cell Death by Dandelion Root Extract in Human Chronic Myelomonocytic Leukemia (CMML) Cells

BACKGROUND: Chronic Myelomonocytic Leukemia (CMML) is a heterogeneous disease that is not only hard to diagnose and classify, but is also highly resistant to treatment. Available forms of therapy for this disease have not shown significant effects and patients rapidly develop resistance early on in therapy. These factors lead to the very poor prognosis observed with CMML patients, with median survival duration between 12 and 24 months after diagnosis. This study is therefore centered around evaluating the selective efficacy of a natural extract from dandelion roots, in inducing programmed cell death in aggressive and resistant CMML cell lines. METHODOLOGY/PRINCIPAL FINDINGS: To confirm the induction of programmed cell death in three human CMML cell lines, nuclear condensation and externalization of the phosphatidylserine, two main characteristics of apoptosis, were detected using Hoechst staining and annexin-V binding assay. The induction of another mode of cell death, autophagy, was determined using a monodansylcadaverine (MDC) stain, to detect the formation of autophagy vacuoles. The results from this study indicate that Dandelion Root Extract (DRE) is able to efficiently and selectively induce apoptosis and autophagy in these cell lines in a dose and time dependent manner, with no significant toxicity on non-cancerous peripheral blood mononuclear cells. More importantly, we observed early activation of initiator caspase-8, which led to mitochondrial destabilization and the induction of autophagy, suggesting that DRE acts through the extrinsic pathway of apoptosis. The inability of DRE to induce apoptosis in dominant-negative FADD cells, confirms the mechanism of action of DRE in in vitro models of CMML. CONCLUSION: The results from this study indicate that natural products, in particular Dandelion Root Extract, have great potential, as non-toxic and effective alternatives to conventional modes of chemotherapy available today

Genomic Profiling of Smoldering Multiple Myeloma Identifies Patients at a High Risk of Disease Progression

PURPOSE: Smoldering multiple myeloma (SMM) is a precursor condition of multiple myeloma (MM) with a 10% annual risk of progression. Various prognostic models exist for risk stratification; however, those are based on solely clinical metrics. The discovery of genomic alterations that underlie disease progression to MM could improve current risk models. METHODS: We used next-generation sequencing to study 214 patients with SMM. We performed whole-exome sequencing on 166 tumors, including 5 with serial samples, and deep targeted sequencing on 48 tumors. RESULTS: We observed that most of the genetic alterations necessary for progression have already been acquired by the diagnosis of SMM. Particularly, we found that alterations of the mitogen-activated protein kinase pathway (KRAS and NRAS single nucleotide variants [SNVs]), the DNA repair pathway (deletion 17p, TP53, and ATM SNVs), and MYC (translocations or copy number variations) were all independent risk factors of progression after accounting for clinical risk staging. We validated these findings in an external SMM cohort by showing that patients who have any of these three features have a higher risk of progressing to MM. Moreover, APOBEC associated mutations were enriched in patients who progressed and were associated with a shorter time to progression in our cohort. CONCLUSION: SMM is a genetically mature entity whereby most driver genetic alterations have already occurred, which suggests the existence of a right-skewed model of genetic evolution from monoclonal gammopathy of undetermined significance to MM. We identified and externally validated genomic predictors of progression that could distinguish patients at high risk of progression to MM and, thus, improve on the precision of current clinical models

Transcriptional profiling of Epstein–Barr virus (EBV) genes and host cellular genes in nasal NK/T-cell lymphoma and chronic active EBV infection

Nasal NK/T-cell lymphoma is an aggressive subtype of non-Hodgkin lymphoma (NHL) that is closely associated with Epstein–Barr virus (EBV). The clonal expansion of EBV-infected NK or T cells is also seen in patients with chronic active EBV (CAEBV) infection, suggesting that two diseases might share a partially similar mechanism by which EBV affects host cellular gene expression. To understand the pathogenesis of EBV-associated NK/T-cell lymphoproliferative disorders (LPD) and design new therapies, we employed a novel EBV DNA microarray to compare patterns of EBV expression in six cell lines established from EBV-associated NK/T-cell LPD. We found that expression of BZLF1, which encodes the immediate-early gene product Zta, was expressed in SNK/T cells and the expression levels were preferentially high in cell lines from CAEBV infection. We also analyzsd the gene expression patterns of host cellular genes using a human oligonucleotide DNA microarray. We identified a subset of pathogenically and clinically relevant host cellular genes, including TNFRSF10D, CDK2, HSPCA, IL12A as a common molecular biological properties of EBV-associated NK/T-cell LPD and a subset of genes, such as PDCD4 as a putative contributor for disease progression. This study describes a novel approach from the aspects of viral and host gene expression, which could identify novel therapeutic targets in EBV-associated NK/T-cell LPD