Evaluation Of Novel Treatment Strategies Research Articles

Prognostic impact of clinical stage on non-surgical patients with pleural mesothelioma.

e20096 Background: Most patients with pleural mesothelioma (PM) are not surgical candidates and receive systemic therapy. Clinical stage is usually not reported in mesothelioma clinical trials. We evaluated the prognostic impact of clinical stage in mesothelioma using a large database with the hypothesis that M1 patients will have worse OS compared to M0 patients. Methods: The Surveillance, Epidemiology and End Results (SEER) Program was used to identify adult patients with PM diagnosed between 2004 and 2020, who received chemotherapy and did not undergo surgery. Patients with unknown stage were excluded. Staging was subdivided into I, II-III and IV for better accuracy. Age, sex, and stage were included in multivariable analyses (MVAs) using a Cox proportional hazard regression model. Unadjusted OS was estimated by the Kaplan-Meier method and compared using log-rank tests. Results: Among the 10,422 patients, 2,644 (25.4%) met the inclusion criteria. Histology classification was not available for 1,120 patients (42.4%), which were reported as malignant mesothelioma. Most patients were male (78.3%) and older than 70 years (54.5%). There were 970 patients (36.7%) with stage I, 893 (33.8%) with stage II-III and 781 (29.5%) with stage IV. The median OS for all patients was 11 months, with 1-year, 3-year and 5-year OS of 43.9%, 10.3% and 4.3% respectively. The median OS for stages I, II-III, and IV was 13 months, 11 months, and 9 months respectively, with 1-year OS of 50.5%, 42.8% and 36.9% respectively and 5-year OS for stages I, II-III, and IV of 5.3%, 3.0% and 4.9% respectively. The median and 1-year OS were 12 months and 46.8% respectively for M0 and 9 months and 36.9% respectively for M1 (p < 0.001). For patient with M0, the median OS for N0, N1 and N2 was 12 months, 11 months, and 11 months, respectively. In MVA, worse OS was observed in patients with age 71 or older compared to < 60 (HR 1.24, 1.09-1.4), and in male patients compared to females (HR 1.27, 1.15.-1.4). Stage IV was associated with worse OS when compared to stage II-III (HR 1.11, 1.001-1.23, p = 0.04) and stage I (HR 1.34, 1.21-1.49, p < 0.001) while stage II-III was associated with worse OS compared to stage I (HR 1.21, 1.1-1.33, p = 0.0001). Conclusions: Clinical stage is a significant predictor for outcomes in patients with unresected pleural mesothelioma treated with chemotherapy, has been underreported, and should be included in clinical trials for a better evaluation of novel treatment strategies.

Clinical and economic burden of systemic lupus erythematosus in Colombia

Aims Our cost-of-illness (COI) model adopted payer and societal perspectives over five years to measure the economic burden of Systemic Lupus Erythematosus (SLE) in Colombia. Materials and methods A prevalence-based model was constructed to estimate costs and economic consequences for SLE patients in Colombia. The model included four health states: three phenotypes of SLE representing mild, moderate, and severe states and death. The clinical inputs were captured from the published literature and validated by the Delphi panel. Our model measured direct medical and indirect costs, including disease management, transient events, and indirect costs. One-way sensitivity analysis was also performed. Results The number of Colombian SLE patients was 37,498. The number of SLE patients with mild, moderate, and severe phenotypes was 5343, 28757 and 3,397, respectively. SLE-patients with moderate (Colombian pesos; COP 146 billion) and severe phenotypes (COP276 billion) incurred higher costs than those with mild phenotypes (COP2 billion), over 5 years. The total SLE cost in Colombia over five years from the payer and societal perspectives was estimated to be COP 915 billion and 8 trillion, respectively. The costs per patient per year from the payer and societal perspectives were COP 4,881,902 ($3,510) and COP 46,637,054 ($33,528), respectively. Conclusion The burden of SLE in Colombia over five years is substantially high, mainly due to the consequences of economic loss because it affects women and men of working age, in addition to the costs of SLE management and its consequences, such as flares, infection, and organ damage. Our COI indicated that disease management costs among patients with moderate and severe SLE were substantially higher than those among patients with a mild phenotype. Therefore, more attention should be paid to limiting the progression of SLE and the occurrence of flares, with the need for further economic evaluation of novel treatment strategies that help in disease control.

Window-of-opportunity clinical trial platform for evaluation of novel treatment strategies in renal cell cancer (WIRE).

TPS406 Background: Renal cell cancer (RCC) is the 7th commonest cancer in the UK. The WIRE trial ‘exploits’ a window-of-opportunity between clear cell RCC diagnosis and nephrectomy to use vascular endothelial growth factor tyrosine kinase inhibitors, PARP inhibitors, and immune checkpoint inhibitor therapies in isolation and in combination. WIRE aims to provide biological proof-of-mechanism of these therapies, and build on encouraging pre-clinical evidence of DNA damage response inhibition to be an effective future therapeutic strategy in RCC. Methods: WIRE is a phase II, multi-IMP, multi-arm, non-randomised clinical trial featuring a Bayesian adaptive design with five treatment arms, and three recruitment stages per arm. The treatment groups: cediranib, cediranib and olaparib, olaparib, durvalumab, and durvalumab and olaparib form the initial set of therapy options with the opportunity for additional treatments to be deployed on this platform-trial in future. The primary endpoint for the first three arms is tumour capillary permeability, measured using multiparametric MRI, defined as a 30% reduction in Ktrans compared to pre-treatment. Whereas in the remaining two arms the primary endpoint is changed to intra-tumoural CD8 positive T cells, defined as an increase of 30% cell count compared to pre-treatment. Up to 76 evaluable participants will be recruited. Monotherapy arms will recruit a maximum of 12 participants each, whereas a maximum of 20 participants will be recruited into each dual-therapy arm. Participants are assigned to the current recruiting arm for treatment lasting between 14 and 28 days before proceeding with standard-of-care nephrectomy. Participants will undergo a biopsy and imaging before and after treatment for primary endpoint analysis, alongside blood and urine collections for translational endpoints. A final two follow-up visits are scheduled before concluding the trial pathway. Rapid interim analysis will follow at the end of a recruitment stage to determine whether to cease recruitment to the current treatment arm due to biological response, or due to futility – defined as lacking in promising evidence for proof of mechanism. Alternatively, if more data is required, recruitment will continue within the same treatment arm until the next interim analysis, where endpoints are re-examined. This highly-efficient design accelerates the exploration of multiple therapies with a strong emphasis on determining early proof-of-mechanism. Promising therapies can then be taken forward to later phase trials in RCC with greater confidence. Clinical trial information: NCT03741426.

A dental implant-on-a-chip for 3D modeling of host-material-pathogen interactions and therapeutic testing platforms.

The precise spatiotemporal control and manipulation of fluid dynamics on a small scale granted by lab-on-a-chip devices provide a new biomedical research realm as a substitute for in vivo studies of host-pathogen interactions. While there has been a rise in the use of various medical devices/implants for human use, the applicability of microfluidic models that integrate such functional biomaterials is currently limited. Here, we introduced a novel dental implant-on-a-chip model to better understand host-material-pathogen interactions in the context of peri-implant diseases. The implant-on-a-chip integrates gingival cells with relevant biomaterials - keratinocytes with dental resin and fibroblasts with titanium while maintaining a spatially separated co-culture. To enable this co-culture, the implant-on-a-chip's core structure necessitates closely spaced, tall microtrenches. Thus, an SU-8 master mold with a high aspect-ratio pillar array was created by employing a unique backside UV exposure with a selective optical filter. With this model, we successfully replicated the morphology of keratinocytes and fibroblasts in the vicinity of dental implant biomaterials. Furthermore, we demonstrated how photobiomodulation therapy might be used to protect the epithelial layer from recurrent bacterial challenges (∼3.5-fold reduction in cellular damage vs. control). Overall, our dental implant-on-a-chip approach proposes a new microfluidic model for multiplexed host-material-pathogen investigations and the evaluation of novel treatment strategies for infectious diseases.

The WIRE study a phase II, multi-arm, multi-centre, non-randomised window-of-opportunity clinical trial platform using a Bayesian adaptive design for proof-of-mechanism of novel treatment strategies in operable renal cell cancer \u2013 a study protocol

BackgroundWindow-of-opportunity trials, evaluating the engagement of drugs with their biological target in the time period between diagnosis and standard-of-care treatment, can help prioritise promising new systemic treatments for later-phase clinical trials. Renal cell carcinoma (RCC), the 7th commonest solid cancer in the UK, exhibits targets for multiple new systemic anti-cancer agents including DNA damage response inhibitors, agents targeting vascular pathways and immune checkpoint inhibitors. Here we present the trial protocol for the WIndow-of-opportunity clinical trial platform for evaluation of novel treatment strategies in REnal cell cancer (WIRE).MethodsWIRE is a Phase II, multi-arm, multi-centre, non-randomised, proof-of-mechanism (single and combination investigational medicinal product [IMP]), platform trial using a Bayesian adaptive design. The Bayesian adaptive design leverages outcome information from initial participants during pre-specified interim analyses to determine and minimise the number of participants required to demonstrate efficacy or futility. Patients with biopsy-proven, surgically resectable, cT1b+, cN0–1, cM0–1 clear cell RCC and no contraindications to the IMPs are eligible to participate. Participants undergo diagnostic staging CT and renal mass biopsy followed by treatment in one of the treatment arms for at least 14 days. Initially, the trial includes five treatment arms with cediranib, cediranib + olaparib, olaparib, durvalumab and durvalumab + olaparib. Participants undergo a multiparametric MRI before and after treatment. Vascularised and de-vascularised tissue is collected at surgery. A ≥ 30% increase in CD8+ T-cells on immunohistochemistry between the screening and nephrectomy is the primary endpoint for durvalumab-containing arms. Meanwhile, a reduction in tumour vascular permeability measured by Ktrans on dynamic contrast-enhanced MRI by ≥30% is the primary endpoint for other arms. Secondary outcomes include adverse events and tumour size change. Exploratory outcomes include biomarkers of drug mechanism and treatment effects in blood, urine, tissue and imaging.DiscussionWIRE is the first trial using a window-of-opportunity design to demonstrate pharmacological activity of novel single and combination treatments in RCC in the pre-surgical space. It will provide rationale for prioritising promising treatments for later phase trials and support the development of new biomarkers of treatment effect with its extensive translational agenda.Trial registrationClinicalTrials.gov: NCT03741426 / EudraCT: 2018–003056-21.

Slice Culture Modeling of CNS Viral Infection.

The complexity of the central nervous system (CNS) is not recapitulated in cell culture models. Thin slicing and subsequent culture of CNS tissue has become a valued means to study neuronal and glial biology within the context of the physiologically relevant tissue milieu. Modern membrane-interface slice culturing methodology allows for straightforward access to both CNS tissue and feeding medium, enabling experimental manipulations and analyses that would otherwise be impossible in vivo. CNS slices can be successfully maintained in culture for up to several weeks for investigation of evolving pathology and long-term intervention in models of chronic neurologic disease.Herein, membrane-interface slice culture models for studying viral encephalitis and myelitis are detailed, with emphasis on the use of these models for investigation of pathogenesis and evaluation of novel treatment strategies. We describe techniques to (1) generate brain and spinal cord slices from rodent donors, (2) virally infect slices, (3) monitor viral replication, (4) assess virally induced injury/apoptosis, (5) characterize "CNS-specific" cytokine production, and, (6) treat slices with cytokines/pharmaceuticals. Although our focus is on CNS viral infection, we anticipate that the described methods can be adapted to address a wide range of investigations within the fields of neuropathology, neuroimmunology, and neuropharmacology.

The cause of death in bacterial meningitis

BackgroundDeath from bacterial meningitis is rarely attributed to the actual event causing death.The present study therefore categorized and characterized the cause and time of death due to bacterial meningitis.MethodsIn a cohort of patients > 15 years of age with community acquired bacterial meningitis the medical records were reviewed, and a clinical cause of death categorized into six main categories: 1) CNS complications, 2) Systemic complications, 3) Combination of systemic and CNS complications, 4) Sudden death, 5) Withdrawal of care, or 6) Unknown.ResultsWe identified 358 patients of which 84 (23%) died in-hospital. Causes of death were ascribed to CNS complications in 43%, Systemic complications in 39%, Combined CNS and systemic complications in 4%, Sudden death in 7% and withdrawal of care in 5%. Brain herniation, circulatory failure, intractable seizures and other brain injury were the most common specific causes of death within 14 days from admission (55%).ConclusionFatal complications due to the primary infection – meningitis - is most common within 14 days of admission. The diversity of complications causing death in meningitis suggest that determining the clinical cause of death is essential to the evaluation of novel treatment strategies.

Pancreatic grade 3 neuroendocrine tumors behave similarly to neuroendocrine carcinomas following resection: a multi-center, international appraisal of the WHO 2010 and WHO 2017 staging schema for pancreatic neuroendocrine lesions

BackgroundIn 2017, the WHO updated their 2010 classification of pancreatic neuroendocrine tumors, introducing a well-differentiated, highly proliferative grade 3 tumor, distinct from neuroendocrine carcinomas. The aim of this study was to investigate the clinical significance of this update in a large cohort of resected tumors. MethodsUsing a multicenter, international dataset of patients with pancreatic neuroendocrine lesions, patients were classified both according to the WHO 2010 and 2017 schema. Multivariable survival analyses were performed, and the models were evaluated for discrimination ability and goodness of fit. ResultsExcluding patients with a known germline MEN1 mutation and incomplete data, 544 patients were analyzed. The performance of the WHO 2010 and 2017 models was similar, however surgically resected grade 3 tumors behaved very similarly to neuroendocrine carcinomas. ConclusionThe addition of a grade 3 NET classification may be of limited utility in surgically resected patients, as these lesions have similar postoperative survival compared to carcinomas. While the addition may allow for a more granular evaluation of novel treatment strategies, surgical intervention for high grade tumors should be considered judiciously.

496P - Predictors for early trial discontinuation of patients with cancer participating in phase I clinical trials

BackgroundPatients with cancer who are refractory to standard treatments may participate in phase I clinical trials. Despite stringent eligibility criteria for trial participation, early discontinuation often occurs. These patients do not benefit from study treatment and may experience additional burden from participation, while adequate evaluation of novel treatment strategies is hampered. The aim of this study was to identify predictors for early discontinuation of phase I clinical trials. MethodsData from patients with solid tumors who participated in phase I trials in our center were pooled for the current analysis. Early trial discontinuation was defined as (i) trial discontinuation within 28 days after administration of the first dose of the investigated drug or (ii) discontinuation before administration of the first dose in patients who were found to be eligible. Based on the literature, the following potential predictors were examined: opioid use, number of metastatic sites, body mass index, ECOG/WHO performance status, comorbidity history of thromboembolism, hemoglobin level, platelet count, leukocytes, lymphocytes, absolute neutrophil count, serum sodium level, creatinine clearance, serum albumin level, serum alkaline phosphatase level, serum aspartate aminotransferase level (AST), serum lactate dehydrogenase level and. Multilevel logistic regression analyses were conducted and Odds ratio (OR) and 95% confidence interval (95%CI) were reported. ResultsData from 154 patients recruited in 8 phase I clinical trials were analyzed. Thirty-six (23%) participants discontinued the trial early. Baseline hyponatremia (OR=3.69, 95%CI=1.09-12.47) and an elevated AST level (OR=2.57, 95%CI=1.10-6.01) were independent predictors for early trial discontinuation. ConclusionsHyponatremia and an elevated serum AST level were identified as significant independent predictors for early trial discontinuation in patients with cancer participating in phase I clinical trials. These predictors will be further investigated in a prospective study, to determine their added value in minimizing early trial discontinuation of patients participating in phase I oncology trials. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

Abstract 4093: Preclinical evaluation of novel treatment strategies in patient-derived xenograft (PDX) models of pancreatic cancer

Abstract Pancreatic cancer (PC) remains a lethal disease with only 3 - 8% of patients surviving 5 years after diagnosis of the tumor (WHO, 2012). Within the EU project “CAM-PaC” a comprehensive panel of thirty patient-derived PC xenografts (PDX) was established and used for the efficacy screening of new therapeutic options. Within this study, responders to the MPS-1 inhibitor BAY1161909, the Super Enhancer disrupting agent Minnelide and the MEK inhibitor Trametinib were identified and analyzed for potential biomarkers. Patient tumors were collected during surgery and circulating tumorigenic cancer stem cells were isolated from the peripheral blood using VAR2CSA-coated magnetic beads. Both were transplanted subcutaneously into NOD/SCID/IL2y- mice and propagated in NMRI:nu/nu mice after engraftment. These were morphologically and molecularly characterized by histopathological revision and with NGS panels, designed based on pathway aggregated genes identified with the International Cancer consortium (described by Bailey et al., Nature 531, 2016). Standard drugs were applied using clinically relevant dosages and schedule. MPS-1 inhibitor BAY1161909 was given in monotherapy and in combination with Abraxane. Minnelide (MTD) was applied second line after three cycles of chemotherapy (Cisplatin, Abraxane, Gemcitabine) and Trametinib was tested as monotherapy. All PDX correlated with histopathological and molecular characteristics of patient tumours. BAY1161909 monotherapy showed moderate anti-tumor efficacy with an average tumor growth inhibition of 30% (p > 0.05). However, tumor relapse after the end of chemotherapy was delayed in mice treated with the combination of BAY1161909 and Abraxane compared to Abraxane alone. 13 out of 22 PDX models tested to date were identified as responders (tumor growth inhibition > 50%) to Minnelide and 5 out of 11 to Trametinib. While Minnelide induced tumor growth inhibition above 80% in 32% of the models, Trametinib achieved the same efficacy in only 9% of the tested PDX models. The described PDX panel clearly reflects clinical situation of pancreatic cancer due to their histologic growth and detection of inherent and acquired treatment resistance as well as recurrent disease. In a few cases, the tested drugs induced complete remissions. We are currently analyzing the molecular data to determine response markers. Our approach may offer personalized treatment options for PC patients. Citation Format: Diana Behrens, Britta Büttner, Rita L. Lawlor, Christopher Heeschen, Malte Buchholz, Jens Siveke, Antje M. Wengner, Ashok Saluja, Jens Hoffmann. Preclinical evaluation of novel treatment strategies in patient-derived xenograft (PDX) models of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4093.

Modelling invasive group A streptococcal disease using bioluminescence

BackgroundThe development of vaccines and evaluation of novel treatment strategies for invasive group A streptococcal (iGAS) disease requires suitable models of human infection that can be monitored longitudinally and are preferably non-invasive. Bio-photonic imaging provides an opportunity to reduce use of animals in infection modelling and refine the information that can be obtained, however the range of bioluminescent GAS strains available is limited. In this study we set out to develop bioluminescent iGAS strains for use in in vivo pneumonia and soft tissue disease models.ResultsUsing clinical emm1, emm3, and emm89 GAS strains that were transformed with constructs carrying the luxABCDE operon, growth and bioluminescence of transformed strains were characterised in vitro and in vivo.Emm3 and emm89 strains expressed detectable bioluminescence when transformed with a replicating plasmid and light production correlated with viable bacterial counts in vitro, however plasmid instability precluded use in the absence of antimicrobial pressure. Emm89 GAS transformed with an integrating construct demonstrated stable bioluminescence that was maintained in the absence of antibiotics. Bioluminescence of the emm89 strain correlated with viable bacterial counts both in vitro and immediately following infection in vivo. Although bioluminescence conferred a detectable fitness burden to the emm89 strain during soft tissue infection in vivo, it did not prevent dissemination to distant tissues.ConclusionDevelopment of stably bioluminescent GAS for use in vitro and in vivo models of infection should facilitate development of novel therapeutics and vaccines while also increasing our understanding of infection progression and transmission routes.

Systemic antibody responses induced by a two-component Clostridium difficile toxoid vaccine protect against C. difficile-associated disease in hamsters

Clostridium difficile infection (CDI) has been identified as the leading cause of nosocomial diarrhoea and pseudomembranous colitis associated with antibiotic therapy. Recent epidemiological changes as well as increases in the number of outbreaks of strains associated with increased virulence and higher mortality rates underscore the importance of identifying alternatives to antibiotics to manage this important disease. Animal studies have clearly demonstrated the roles that toxins A and B play in gut inflammation as well as diarrhoea; therefore it is not surprising that serum anti-toxin A and B IgG are associated with protection against recurrent CDI. In humans, strong humoral toxin-specific immune responses elicited by natural C. difficile infection is associated with recovery and lack of disease recurrence, whereas insufficient humoral responses are associated with recurrent CDI. The first generation of C. difficile vaccine that contained inactivated toxin A and B was found to be completely protective against death and diarrhoea in the hamster C. difficile challenge model. When tested in young healthy volunteers in Phase I clinical trials, this investigational vaccine was shown to be safe and immunogenic. Moreover, in a separate study this vaccine was able to prevent further relapses in three out of three patients who had previously suffered from chronic relapsing C. difficile-associated diarrhoea. Herein we examined the immunogenicity and protective activity of a next-generation Sanofi Pasteur two-component highly purified toxoid vaccine in a C. difficile hamster model. This model is widely recognized as a stringent and relevant choice for the evaluation of novel treatment strategies against C. difficile and was used in preclinical testing of the first-generation vaccine candidate. Intramuscular (i.m.) immunizations with increasing doses of this adjuvanted toxoid vaccine protected hamsters from mortality and disease symptoms in a dose-dependent manner. ELISA measurements of pre-challenge sera showed that the median anti-toxin A and anti-toxin B IgG titres in the group of surviving animals were significantly higher than the median values in the group of animals that did not survive challenge. Assessment of the neutralizing activity of these sera revealed a statistically significant difference between the levels of both toxin A and toxin B neutralizing titres in protected versus unprotected animals as the median anti-toxin A and anti-toxin B neutralizing titres from surviving animals were higher than the median values from animals that succumbed to challenge. Statistically significant correlations between the toxin-specific binding titres and toxin neutralizing titres were seen for both toxin A and toxin B responses. The role of circulating anti-toxin antibodies in immunity against disease was evaluated by passive transfer of immune sera against C. difficile toxoids to naïve hamsters. Passively immunized animals were protected against morbidity and mortality associated with C. difficile challenge. Taken together, these results indicate the ability of i.m. immunization with inactivated toxins A and B to induce robust dose-dependent anti-toxin A and anti-toxin B IgG responses, the principal role of circulating anti-toxin antibody in immunity against disease and that antibody toxin binding and neutralization titres can serve as correlates of protection in the hamster challenge model of C. difficile.

Lung tumour growth kinetics in SPC-c-Raf-1-BB transgenic mice assessed by longitudinal in-vivo micro-CT quantification

BackgroundSPC-c-Raf-1-BxB transgenic mice develop genetically induced disseminated lung adenocarcinoma allowing examination of carcinogenesis and evaluation of novel treatment strategies. We report on assessment of lung tumour growth kinetics using a semiautomated region growing segmentation algorithm.Methods156 non contrast-enhanced respiratory gated micro-CT of the lungs were obtained in 12 SPC-raf transgenic (n = 9) and normal (n = 3) mice at different time points. Region-growing segmentation of the aerated lung areas was obtained as an inverse surrogate for tumour burden. Time course of segmentation volumes was assessed to demonstrate the potential of the method for follow-up studies.ResultsMicro-CT allowed assessment of tumour growth kinetics and semiautomated region growing enabled quantitative analysis. Significant changes of the segmented lung volumes over time could be shown (p = 0.009). Significant group differences could be detected between transgenic and normal animals for time points 8 to 13 months (p = 0.043), when marked tumour progression occurred.ConclusionThe presented region-growing segmentation algorithm allows in-vivo quantification of multifocal lung adenocarcinoma in SPC-raf transgenic mice. This enables the assessment of tumour load and progress for the study of carcinogenesis and the evaluation of novel treatment strategies.

Reactivation of Silenced WT1 Transgene by Hypomethylating Agents - Implications forin vitroModeling of Chemoimmunotherapy

BackgroundA cell line with transfected Wilms' tumor protein 1 (WT1) is has been used for the preclinical evaluation of novel treatment strategies of WT1 immunotherapy for leukemia due to the lack of appropriate murine leukemia cell line with endogenous WT1. However, silencing of the transgene occurs. Regarding the effects of hypomethylating agents (HMAs) on reactivation of silenced genes, HMAs are considered to be immune enhancers.MethodsWe treated murine WT1- transfected C1498 (mWT1-C1498) with increasing doses of decitabine (DAC) and azacitidine (AZA) to analyze their effects on transgene reactivation.ResultsDAC and AZA decreased the number of viable cells in a dose- or time-dependent manner. Quantification of WT1 mRNA level was analyzed by real-time polymerase chain reaction after mWT1-C1498 treated with increasing dose of HMA. DAC treatment for 48 h induced 1.4-, 14.6-, and 15.5-fold increment of WT1 mRNA level, compared to untreated sample, at 0.1, 1, and 10µM, respectively. Further increment of WT1 expression in the presence of 1 and 10µM DAC was evident at 72 h. AZA treatment also induced up-regulation of mRNA, but not to the same degree as with DAC treatment. The correlation between the incremental increases in WT1 mRNA by DAC was confirmed by Western blot and concomitant down-regulation of WT1 promoter methylation was revealed.ConclusionThe in vitro data show that HMA can induce reactivation of WT1 transgene and that DAC is more effective, at least in mWT1-C1498 cells, which suggests that the combination of DAC and mWT1-C1498 can be used for the development of the experimental model of HMA-combined WT1 immunotherapy targeting leukemia.

In vivo monitoring of cystic fibrosis-like lung disease in mice by volumetric computed tomography

The onset and spontaneous development of cystic fibrosis (CF) lung disease remain poorly understood. In the present study, we used volumetric computed tomography (VCT) as a new method for longitudinal in vivo monitoring of early lesions and disease progression in CF-like lung disease in β-epithelial Na(+) channel (ENaC)-transgenic (TG) mice. Using a VCT scanner prototype (80 kV, 50 mA·s, scan time 19 s and spatial resolution 200 μm), βENaC-TG mice and wild-type (WT) littermates were examined longitudinally at 10 time-points from neonatal to adult ages, and VCT images were assessed by qualitative and quantitative morphological parameters. We demonstrate that VCT detected early-onset airway mucus obstruction, diffuse infiltrates, atelectasis and air trapping as characteristic abnormalities in βENaC-TG mice. Furthermore, we show that early tracheal mucus obstruction predicted mortality in βENaC-TG mice and that the density of lung parenchyma was significantly reduced at all time-points in βENaC-TG compared with WT mice (median ± sem -558 ± 8 HU in WT versus -686 ± 16 HU in βENaC-TG at 6 weeks of age; p < 0.005). Our study demonstrates that VCT is a sensitive, noninvasive technique for early detection and longitudinal monitoring of morphological abnormalities of CF-like lung disease in mice, and may thus provide a useful tool for pre-clinical in vivo evaluation of novel treatment strategies for CF.

Depression in adolescents: current treatments, suicidality and evaluation of novel treatment strategies.

Multiple studies have examined the age of onset of major depression, indicating it is most frequent in adolescence and young adulthood. In this context, the offspring of depressed parents have a 2 to 4 time increased risk for depression compared with children of non-depressed parents.Treatment for depression in adolescents can be divided into psychosocial, psychopharmacologic, somatic and combined psychosocial-psychopharmacologic, psychosocial-psychosomatic and psychopharmacologic-psychosomatic.Depression in the children and adolescent population has been an area of research for over 20 years. Among novel therapeutic strategies, transcranial magnetic stimulation (TMS) has demonstrated the most favorable side effect profile. Until this time there are no published suicide attempts associated with this treatment and it may offer an option that is not associated with stigma of electroconvulsive therapy (ECT) or medications. Further research may provide more access to this therapy and hope to children, adolescents with depression and their families.

Gene therapy for thyroid cancer: current status and future prospects.

Despite multimodality treatment for thyroid cancer, including surgical resection, radioiodine therapy, thyrotropin (TSH)-suppressive thyroxine treatment, and chemotherapy/radiotherapy, survival rates have not improved over the last decades. Therefore, development and evaluation of novel treatment strategies, including gene therapy, are urgently needed. A variety of gene therapy approaches have been evaluated for the treatment of follicular cell-derived and medullary thyroid cancer, including corrective gene therapy (p53 restoration, expression of a dominant negative RET mutant), cytoreductive gene therapy (suicide gene/prodrug strategy herpes simplex virus-thymidine kinase [HSV-tk]/ganciclovir, antiangiogenic therapy with endostatin) and immunomodulatory gene therapy (expression of interleukin (IL)-2 and IL-12). Furthermore, cloning of the sodium iodide symporter (NIS) gene has paved the way for the development of a novel cytoreductive gene therapy strategy based on NIS gene transfer followed by the application of radioiodine therapy ((131)I). NIS gene delivery into medullary and follicular cell-derived thyroid cancer cells has been shown to be capable of establishing or restoring radioiodine accumulation and might therefore represent an effective therapy for medullary and dedifferentiated thyroid tumors that lack iodide accumulating activity. The data summarized in this review article clearly demonstrate that the currently available strategies represent potentially curative novel therapeutic approaches for future gene therapy of thyroid cancer. The combination of different therapeutic genes has been demonstrated to be very useful to enhance therapeutic efficacy and seems to have a promising role at least as part of a multimodality approach for advanced thyroid cancer.