What is the best surgical option after failure of graciloplasty in patients with recurrent rectovaginal fistula? A study of 19 consecutive patients.

BACKGROUND: Management of recurrent rectovaginal fistula (rRVF) remains challenging despite the good results of graciloplasty reported in the literature. However, little is known about how to avoid a permanent stoma if graciloplasty fails. The aim of our study was to report the management of rRVF after failure of graciloplasty. METHODS: A retrospective study was performed on consecutive patients with rRVF after failure of graciloplasty treated at our institution in January 2005-December 2021. RESULTS: There were 19 patients, with a median age at graciloplasty of 39 years (range 25-64 years). Etiologies of RVF were Crohn's disease (CD) (n = 10), postoperative (n = 5), post-obstetrical (n = 3), and unknown (n = 1). After failure of graciloplasty, 45 new procedures were performed, all of them with a covering stoma: trans-anal repairs (n = 31), delayed colo-anal anastomosis (DCAA) (n = 4), biological mesh interposition (n = 3), second graciloplasty (n = 3), stoma only (n = 2) and redo ileal pouch-anal anastomosis (IPAA) (n = 2). One patient was not re-operated on and instead treated medically for CD. After a mean follow-up of 63 ± 49 months, success (i.e., absence of stoma or RVF) was obtained in 11 patients (58%): 4/4 DCAA (100%), 5/31 after local repair (16%), 1 after stoma creation alone (50%) and 1 after redo IPAA (50%). Second graciloplasty and biologic mesh interposition all failed. All 8 patients with failed intervention had CD. CONCLUSIONS: In cases of rRVF after failed graciloplasty, reoperation is possible, although the chance of success is relatively low. The best results were obtained with DCAA. CD is a predictor of poor outcome.

Short-term rapamycin treatment increases life span and attenuates aortic aneurysm in a murine model of Marfan-Syndrome.

BACKGROUND: Marfan syndrome (MFS) is a genetic disorder leading to medial aortic degeneration and life-limiting dissections. To date, there is no causal prevention or therapy. Rapamycin is a potent and selective inhibitor of the mechanistic target of rapamycin (mTOR) protein kinase, regulating cell growth and metabolism. The mgR/mgR mice represent an accepted MFS model for studying aortic pathologies to understand the underlying molecular pathomechanisms. This study investigated whether rapamycin inhibits the development of thoracic aortic aneurysms and dissections in mgR/mgR mice. METHODS: Isolated primary aortic smooth muscle cells (mAoSMCs) from mgR/mgR mice were used for in vitro studies. Two mg kg/BW rapamycin was injected intraperitoneally daily for two weeks, beginning at 7-8 weeks of age. Mice were sacrificed 30 days post-treatment. Histopathological and immunofluorescence analyses were performed using adequate tissue specimens and techniques. Animal survival was evaluated accompanied by periodic echocardiographic examinations of the aorta. RESULTS: The protein level of the phosphorylated ribosomal protein S6 (p-RPS6), a downstream target of mTOR, was significantly increased in the aortic tissue of mgR/mgR mice. In mAoSMCs isolated from these animals, expression of mTOR, p-RPS6, tumour necrosis factor α, matrix metalloproteinase-2 and -9 was significantly suppressed by rapamycin, demonstrating its anti-inflammatory capacity. Short-term rapamycin treatment of Marfan mice was associated with delayed aneurysm formation, medial aortic elastolysis and improved survival. CONCLUSIONS: Short-term rapamycin-mediated mTOR inhibition significantly reduces aortic aneurysm formation and thus increases survival in mgR/mgR mice. Our results may offer the first causal treatment option to prevent aortic complications in MFS patients.

[Treatment recommendations in cardio-oncology: where are we?] / Therapieempfehlungen in der Kardioonkologie ­ wo stehen wir?

This article provides an overview of current prevention and treatment options for typical cardiovascular side effects of oncological therapies as well as cardiovascular complications of malignant disease. Focus is put on the prevention and treatment of heart failure under potentially cardiotoxic cancer therapies. In addition, current options for the treatment of common venous thromboembolism in cancer patients will be discussed.

AAV-mediated cardiac gene transfer of wild-type desmin in mouse models for recessive desminopathies.

Mutations in the human desmin gene cause autosomal-dominant and recessive cardiomyopathies and myopathies with marked phenotypic variability. Here, we investigated the effects of adeno-associated virus (AAV)-mediated cardiac wild-type desmin expression in homozygous desmin knockout (DKO) and homozygous R349P desmin knockin (DKI) mice. These mice serve as disease models for two subforms of autosomal-recessive desminopathies, the former for the one with a complete lack of desmin protein and the latter for the one with solely mutant desmin protein expression in conjunction with protein aggregation pathology in striated muscle. Two-month-old mice were injected with either a single dose of 5 × 1012 AAV9-hTNT2-mDes (AAV-Des) vector genomes or NaCl as control. One week after injection, mice were subjected to a forced swimming exercise protocol for 4 weeks. Cardiac function was monitored over a period of 15 month after injection and before the mice were sacrificed for biochemical and morphological analysis. AAV-mediated cardiac expression of wild-type desmin in both the homozygous DKO and DKI backgrounds reached levels seen in wild-type mice. Notably, AAV-Des treated DKO mice showed a regular subcellular distribution of desmin as well as a normalization of functional and morphological cardiac parameters. Treated DKI mice, however, showed an aberrant subcellular localization of desmin, unchanged functional cardiac parameters, and a trend toward an increased cardiac fibrosis. In conclusion, the effect of a high-dose AAV9-based desmin gene therapy is highly beneficial for the heart in DKO animals, but not in DKI mice.

AAV-9 mediated phosphatase-1 inhibitor-1 overexpression improves cardiac contractility in unchallenged mice but is deleterious in pressure-overload.

The downregulation of ß-adrenergic receptors (ß-AR) and decreased cAMP-dependent protein kinase activity in failing hearts results in decreased phosphorylation and inactivation of phosphatase-inhibitor-1 (I-1), a distal amplifier element of ß-adrenergic signaling, leading to increased protein phosphatase 1 activity and dephosphorylation of key phosphoproteins, including phospholamban. Downregulated and hypophosphorylated I-1 likely contributes to ß-AR desensitization; therefore its modulation is a promising approach in heart failure treatment. Aim of our study was to assess the effects of adeno-associated virus serotype 9 (AAV9) - mediated cardiac-specific expression of constitutively active inhibitor-1 (I-1c) and to investigate whether I-1c is able to attenuate the development of heart failure in mice subjected to transverse aortic constriction (TAC). 6-8 week old C57BL/6 N wild-type mice were subjected to banding of the transverse aorta (TAC). Two days later 2.8 × 1012 AAV-9 vector particles harbouring I-1c cDNA under transcriptional control of a human troponin T-promoter (AAV9/I-1c) were intravenously injected into the tail vein of these mice (n=12). AAV9 containing a Renilla luciferase reporter (AAV9/hRluc) was used as a control vector (n=12). Echocardiographic analyses were performed weekly to evaluate cardiac morphology and function. 4 weeks after TAC pressure- volume measurements were performed and animals were sacrificed for histological and molecular analyses. Both groups exhibited progressive contractile dysfunction and myocardial remodeling. Surprisingly, echocardiographic assessment and histological analyses showed significantly increased left ventricular hypertrophy in AAV9/I-1c treated mice compared to AAV9/hRluc treated controls as well as reduced contractility. Pressure-volume loops revealed significantly impaired contractility after AAV9/I-1c treatment. At the molecular level, hearts of AAV9/I-1c treated TAC mice showed a hyperphosphorylation of the SR Ca2+-ATPase inhibitor phospholamban. In contrast, expression of AAV9/I-1c in unchallenged animals resulted in selective enhancement of phospholamban phosphorylation and augmented cardiac contractility. Our data suggest that AAV9-mediated cardiac-specific overexpression of I-1c, previously associated with enhanced calcium cycling, improves cardiac contractile function in unchallenged animals but failed to protect against cardiac remodeling induced by hemodynamic stress questioning the use of I-1c as a potential strategy to treat heart failure in conditions with increased afterload.

Missed opportunities for earlier diagnosis of rifampicin-resistant tuberculosis despite access to Xpert® MTB/RIF.

OBJECTIVE: To assess the proportion of rifampicin-resistant tuberculosis (RR-TB) patients with potential earlier RR-TB diagnoses in Khayelitsha, South Africa. DESIGN: We conducted a retrospective analysis among RR-TB patients diagnosed from 2012 to 2014. Patients were considered to have missed opportunities for earlier diagnosis if 1) they were incorrectly screened according to the Western Cape diagnostic algorithm; 2) the first specimen was not tested using Xpert® MTB/RIF; 3) no specimen was ever tested; or 4) the initial Xpert test showed a negative result, but no subsequent specimen was sent for follow-up testing in human immunodeficiency virus-positive patients. RESULTS: Among 543 patients, 386 (71%) were diagnosed with Xpert and 112 (21%) had had at least one presentation at a health care facility within the 6 months before the presentation at which RR-TB was diagnosed. Overall, 95/543 (18%) patients were screened incorrectly at some point: 48 at diagnostic presentation only, 38 at previous presentation only, and 9 at both previous and diagnostic presentations. CONCLUSIONS: These data show that a significant proportion of RR-TB patients might have been diagnosed earlier, and suggest that case detection could be improved if diagnostic algorithms were followed more closely. Further training and monitoring is required to ensure the greatest benefit from universal Xpert implementation.

Identification of circular RNAs with host gene-independent expression in human model systems for cardiac differentiation and disease.

AIMS: Cardiovascular disease, one of the most common causes of death in western populations, is characterized by changes in RNA splicing and expression. Circular RNAs (circRNA) originate from back-splicing events, which link a downstream 5' splice site to an upstream 3' splice site. Several back-splicing junctions (BSJ) have been described in heart biopsies from human, rat and mouse hearts (Werfel et al., 2016; Jakobi et al., 2016 ). Here, we use human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) to identify circRNA and host gene dynamics in cardiac development and disease. In parallel, we explore candidate interactions of selected homologs in mouse and rat via RIP-seq experiments. METHODS AND RESULTS: Deep RNA sequencing of cardiomyocyte development and ß-adrenergic stimulation uncovered 4518 circRNAs. The set of circular RNA host genes is enriched for chromatin modifiers and GTPase activity regulators. RNA-seq and qRT-PCR data showed that circular RNA expression is highly dynamic in the hiPSC-CM model with 320 circRNAs showing significant expression changes. Intriguingly, 82 circRNAs are independently regulated to their host genes. We validated the same circRNA dynamics for circRNAs from ATXN10, CHD7, DNAJC6 and SLC8A1 in biopsy material from human dilated cardiomyopathy (DCM) and control patients. Finally, we could show that rodent homologs of circMYOD, circSLC8A1, circATXN7 and circPHF21A interact with either the ribosome or Argonaute2 protein complexes. CONCLUSION: CircRNAs are dynamically expressed in a hiPSC-CM model of cardiac development and stress response. Some circRNAs show similar, host-gene independent expression dynamics in patient samples and may interact with the ribosome and RISC complex. In summary, the hiPSC-CM model uncovered a new signature of potentially disease relevant circRNAs which may serve as novel therapeutic targets.

AAV9-mediated gene transfer of desmin ameliorates cardiomyopathy in desmin-deficient mice.

Mutations of the human desmin (DES) gene cause autosomal dominant and recessive myopathies affecting skeletal and cardiac muscle tissue. Desmin knockout mice (DES-KO), which develop progressive myopathy and cardiomyopathy, mirror rare human recessive desminopathies in which mutations on both DES alleles lead to a complete ablation of desmin protein expression. Here, we investigated whether an adeno-associated virus-mediated gene transfer of wild-type desmin cDNA (AAV-DES) attenuates cardiomyopathy in these mice. Our approach leads to a partial reconstitution of desmin protein expression and the de novo formation of the extrasarcomeric desmin-syncoilin network in cardiomyocytes of treated animals. This finding was accompanied by reduced fibrosis and heart weights and improved systolic left-ventricular function when compared with control vector-treated DES-KO mice. Since the re-expression of desmin protein in cardiomyocytes of DES-KO mice restores the extrasarcomeric desmin-syncoilin cytoskeleton, attenuates the degree of cardiac hypertrophy and fibrosis, and improves contractile function, AAV-mediated desmin gene transfer may be a novel and promising therapeutic approach for patients with cardiomyopathy due to the complete lack of desmin protein expression.

Treatment of multifocal breast cancer by systemic delivery of dual-targeted adeno-associated viral vectors.

Adeno-associated viral (AAV) vectors yield high potential for clinical gene therapy but, like for other vectors systems, they frequently do not sufficiently transduce the target tissue and their unspecific tropism prevents their application for multifocal diseases such as disseminated cancer. Targeted AAV vectors have been obtained from random AAV display peptide libraries but so far, all vector variants selected from AAV libraries upon systemic administration in vivo retained some collateral tropism, frequently the heart. Here we explored, if this impediment can be overcome by microRNA-regulated transgene cassettes as the combination of library-derived capsid targeting and micro-RNA control has not been evaluated so far. We used a tumor-targeted AAV capsid variant (ESGLSQS) selected from random AAV-display peptide libraries in vivo with remaining off-target tropism toward the heart and regulated targeted transgene expression in vivo by complementary target elements for heart-specific microRNA (miRT-1d). Although this vector still maintained its strong transduction capacity for tumor target tissue after intravenous injection, transgene expression in the heart was almost completely abrogated. This strong and completely tumor-specific transgene expression was used for therapeutic gene transfer in an aggressive multifocal, transgenic, polyoma middle T-induced, murine breast cancer model. A therapeutic suicide gene, delivered systemically by this dual-targeted AAV vector to multifocal breast cancer, significantly inhibited tumor growth after one single vector administration while avoiding side effects compared with untargeted vectors.

Therapeutic safety of high myocardial expression levels of the molecular inotrope S100A1 in a preclinical heart failure model.

Low levels of the molecular inotrope S100A1 are sufficient to rescue post-ischemic heart failure (HF). As a prerequisite to clinical application and to determine the safety of myocardial S100A1 DNA-based therapy, we investigated the effects of high myocardial S100A1 expression levels on the cardiac contractile function and occurrence of arrhythmia in a preclinical large animal HF model. At 2 weeks after myocardial infarction domestic pigs presented significant left ventricular (LV) contractile dysfunction. Retrograde application of AAV6-S100A1 (1.5 × 10(13) tvp) via the anterior cardiac vein (ACV) resulted in high-level myocardial S100A1 protein peak expression of up to 95-fold above control. At 14 weeks, pigs with high-level myocardial S100A1 protein overexpression did not show abnormalities in the electrocardiogram. Electrophysiological right ventricular stimulation ruled out an increased susceptibility to monomorphic ventricular arrhythmia. High-level S100A1 protein overexpression in the LV myocardium resulted in a significant increase in LV ejection fraction (LVEF), albeit to a lesser extent than previously reported with low S100A1 protein overexpression. Cardiac remodeling was, however, equally reversed. High myocardial S100A1 protein overexpression neither increases the occurrence of cardiac arrhythmia nor causes detrimental effects on myocardial contractile function in vivo. In contrast, this study demonstrates a broad therapeutic range of S100A1 gene therapy in post-ischemic HF using a preclinical large animal model.

Direct visualization of cerebellar nuclei in patients with focal cerebellar lesions and its application for lesion-symptom mapping.

As yet, human cerebellar lesion studies have not taken advantage of direct magnetic resonance imaging (MRI) of the cerebellar nuclei in individual patients. In the present study, susceptibility weighted imaging (SWI) was used to visualize lesions of the dentate nuclei in patients with chronic focal lesions. Fifteen patients with cerebellar lesions either due to stroke or tumor surgery underwent SWI imaging using a 1.5T MRI scanner. Dentate nuclei were seen as hypointensities in all patients. Three of the patients underwent additional SWI imaging at 3T and 7T. Compared to 1.5T, corrugation of the dentate wall was seen with greater precision and the dorsal, iron-poorer part was seen more fully. Lesion-symptom mapping was performed based on the 1.5T MR images. Patients were divided into two groups with and without upper limb ataxia. A region-of-interest-(ROI)-driven normalization technique was used which had initially been developed by Diedrichsen et al. (2011) for functional MRI (fMRI) of the dentate nuclei. Compared to conventional normalization of the cerebellum, overlap of dentate lesions improved and lead to increased sensitivity of lesion-symptom maps. Subtraction analysis revealed that the more dorsal and rostral parts of the dentate nuclei were related to upper limb ataxia. Findings were in good accordance with the dentate hand area shown in recent fMRI studies. These data provide evidence that direct identification of dentate lesions together with the ROI-driven normalization technique allows for improved lesion-symptom mapping at the level of the cerebellar nuclei already at conventional 1.5T MRI field strength.

Novel random peptide libraries displayed on AAV serotype 9 for selection of endothelial cell-directed gene transfer vectors.

We have demonstrated the potential of random peptide libraries displayed on adeno-associated virus (AAV)2 to select for AAV2 vectors with improved efficiency for cell type-directed gene transfer. AAV9, however, may have advantages over AAV2 because of a lower prevalence of neutralizing antibodies in humans and more efficient gene transfer in vivo. Here we provide evidence that random peptide libraries can be displayed on AAV9 and can be utilized to select for AAV9 capsids redirected to the cell type of interest. We generated an AAV9 peptide display library, which ensures that the displayed peptides correspond to the packaged genomes and performed four consecutive selection rounds on human coronary artery endothelial cells in vitro. This screening yielded AAV9 library capsids with distinct peptide motifs enabling up to 40-fold improved transduction efficiencies compared with wild-type (wt) AAV9 vectors. Incorporating sequences selected from AAV9 libraries into AAV2 capsids could not increase transduction as efficiently as in the AAV9 context. To analyze the potential on endothelial cells in the intact natural vascular context, human umbilical veins were incubated with the selected AAV in situ and endothelial cells were isolated. Fluorescence-activated cell sorting analysis revealed a 200-fold improved transduction efficiency compared with wt AAV9 vectors. Furthermore, AAV9 vectors with targeting sequences selected from AAV9 libraries revealed an increased transduction efficiency in the presence of human intravenous immunoglobulins, suggesting a reduced immunogenicity. We conclude that our novel AAV9 peptide library is functional and can be used to select for vectors for future preclinical and clinical gene transfer applications.

The assembly-activating protein promotes capsid assembly of different adeno-associated virus serotypes.

Adeno-associated virus type 2 (AAV2) capsid assembly requires the expression of a virally encoded assembly-activating protein (AAP). By providing AAP together with the capsid protein VP3, capsids are formed that are composed of VP3 only. Electron cryomicroscopy analysis of assembled VP3-only capsids revealed all characteristics of the wild-type AAV2 capsids. However, in contrast to capsids assembled from VP1, VP2, and VP3, the pores of VP3-only capsids were more restricted at the inside of the 5-fold symmetry axes, and globules could not be detected below the 2-fold symmetry axes. By comparing the capsid assembly of several AAV serotypes with AAP protein from AAV2 (AAP-2), we show that AAP-2 is able to efficiently stimulate capsid formation of VP3 derived from several serotypes, as demonstrated for AAV1, AAV2, AAV8, and AAV9. Capsid formation, by coexpressing AAV1-, AAV2-, or AAV5-VP3 with AAP-1, AAP-2, or AAP-5 revealed the ability of AAP-1 and AAP-2 to complement each other in AAV1 and AAV2 assembly, whereas for AAV5 assembly more specific conditions are required. Sequence alignment of predicted AAP proteins from the known AAV serotypes indicates a high degree of homology of all serotypes to AAP-2 with some divergence for AAP-4, AAP-5, AAP-11, and AAP-12. Immunolocalization of assembled capsids from different serotypes confirmed the preferred nucleolar localization of capsids, as observed for AAV2; however, AAV8 and AAV9 capsids could also be detected throughout the nucleus. Taken together, the data show that AAV capsid assembly of different AAV serotypes also requires the assistance of AAP proteins.

Prevalencia de enfermedad renal crónica en centros urbanos de atención primaria / Prevalence of chronic kidney disease in subjects consulting in urban primary care clinics

Background: Chronic kidney disease (CKD) is a major worldwide public health problem and is associated with increased risk of cardiovascular disease and death. Aim: To assess CKD prevalence in urban Primary Care Services (PCS) of Concepcion, Chile. Material and Methods: The clinical records of 27.894 adults aged 55 ± 18 years (66 percent females), consulting in outpatient clinics and in whom serum creatinine was measured, with or without assessment of urine albumin levels, were reviewed. The glomerular filtration rate (eGFR) was estimated using the Modification of Diet in Renal Disease (MDRD)-4 equation. CKD was defined as an eGFR < 60 ml/min/1.73 m2 and classified according to the National Kidney Foundation Kidney Disease Outcomes Quality Initiative (NFK-KDOQI) guidelines. Results: Mean eGFR was 77.1 ± 16.3 ml/min/1.73 m2. Twelve percent of subjects had CKD (women, 14.5 percent and men 7,4 percent, p < 0,05). The prevalence of stages 3, 4 and 5 of CKD were 11.6, 0.3 and 0.2 percent respectively. eGFR was negatively correlated with age ( r = -0,54, p < 0,05). Among patients with an eGFR < 60 ml/min/1.73 m2, 96.3 percent had eGFR 30-59, 2.3 percent 15-29 and 1.4 percent < of 15. Seventy nine percent were women. 75.1 percent were aged 65 years or more, 26.8 percent had a serum creatinine equal or less than 1.0 mg/dL and 40.5 percent had microalbuminuria. Only 1 percent of outpatients ascribed to Cardiovascular or Diabetes Programs had the diagnosis of CKD registered. Independent risk predictors of CKD were age > 60 years, female sex and microalbuminuria. Conclusions: This study showed a high prevalence of CKD in ambulatory patients, mainly among women and older people. The low level of diagnosis of CKD in cardiovascular and diabetes programs is of concern.

[Coronary artery disease diagnostic with fractional flow reserve]. / Diagnostic de la maladie coronarienne stable en 2011: importance de la méthode dite "fractional flow reserve".

Fractional flow reserve (FFR) is an invasive pressure-derived index of epicardial stenosis severity used in the catheterization laboratory to assess the hemodynamic significance of coronary lesions when non-invasive functional assessment has either not been performed or is inconclusive. The use of invasive coronary physiology has demonstrated favorable outcomes for decision making in patients with intermediate single-vessel disease, with left main stenosis, complex bifurcations lesions and multivessel coronary artery disease. Specifically, in patients with stable angina revascularization with either percutaneous coronary intervention (PCI) or coronary artery bypass graft (CABG) surgery should be deferred for epicardial coronary stenosis with an FFR > 0.8.

[Validation of pre-cystectomy nomograms for the prediction of locally advanced urothelial bladder cancer in a multicentre study: are we able to adequately predict locally advanced tumour stages before surgery?]. / Validierung von Präzystektomienomogrammen zur Vorhersage lokal fortgeschrittener Harnblasenkarzinome in einer multizentrischen Studie : Können wir lokal fortgeschrittene Tumorstadien präoperativ ausreichend sicher vorhersagen?

OBJECTIVE: Pre-cystectomy nomograms with a high predictive ability for locally advanced urothelial carcinomas of the bladder would enhance individual treatment tailoring and patient counselling. To date, there are two currently not externally validated nomograms for prediction of the tumour stages pT3-4 or lymph node involvement. MATERIALS AND METHODS: Data from a German multicentre cystectomy series comprising 2,477 patients with urothelial carcinoma of the bladder were applied for the validation of two US nomograms, which were originally based on the data of 726 patients (nomogram 1: prediction of pT3-4 tumours, nomogram 2: prediction of lymph node involvement). Multivariate regression models assessed the value of clinical parameters integrated in both nomograms, i.e. age, gender, cT stage, TURB grade and associated Tis. Discriminative abilities of both nomograms were assessed by ROC analyses; calibration facilitated a comparison of the predicted probability and the actual incidence of locally advanced tumour stages. RESULTS: Of the patients, 44.5 and 25.8% demonstrated tumour stages pT3-4 and pN+, respectively. If only one case of a previously not known locally advanced carcinoma (pT3-4 and/or pN+) is considered as a staging error, the rate of understaging was 48.9% (n=1211). The predictive accuracies of the validated nomograms were 67.5 and 54.5%, respectively. The mean probabilities of pT3-4 tumours and lymph node involvement predicted by application of these nomograms were 36.7% (actual frequency 44.5%) and 20.2% (actual frequency 25.8%), respectively. Both nomograms underestimated the real incidence of locally advanced tumours. CONCLUSIONS: The present study demonstrates that prediction of locally advanced urothelial carcinomas of the bladder by both validated nomograms is not conferrable to patients of the present German cystectomy series. Hence, there is still a need for statistical models with enhanced predictive accuracy.

[Influence of older age on survival after radical cystectomy due to urothelial carcinoma of the bladder: survival analysis of a German multi-centre study after curative treatment of urothelial carcinoma of the bladder]. / Einfluss des Alters auf das karzinomspezifische Überleben nach radikaler Zystektomie : Überlebensanalyse aus einer deutschen Multicenterstudie nach kurativer Therapie eines Urothelkarzinoms der Harnblase.

BACKGROUND: The therapeutic gold standard of muscle-invasive tumour stages is radical cystectomy (RC), but there are still conflicting reports about associated morbidity and mortality and the oncologic benefit of RC in elderly patients. The aim of the present study was the comparison of overall (OS) and cancer-specific survival (CSS) in patients <75 and >75 years of age (median follow-up was 42 months). PATIENTS AND METHODS: Clinical and histopathological data of 2,483 patients with urothelial carcinoma and consecutive RC were collated. The study group was dichotomized by the age of 75 years at RC. Statistical analyses comprising an assessment of postoperative mortality within 90 days, OS and CSS were assessed. Multivariate logistic regression and survival analyses were performed. RESULTS: The 402 patients (16.2%) with an age of ≥75 years at RC showed a significantly higher local tumour stage (pT3/4 and/or pN+) (58 vs 51%; p=0.01), higher tumour grade (73 vs 65%; p=0.003) and higher rates of upstaging in the RC specimen (55 vs 48%; p=0.032). Elderly patients received significantly less often adjuvant chemotherapy (8 vs 15%; p<0.001). The 90-day mortality was significantly higher in patients ≥75 years (6.2 vs 3.7%; p=0.026). When adjusted for different variables (gender, tumour stage, adjuvant chemotherapy, time period of RC), only in male patients and locally advanced tumour stages was an association with 90-day mortality noticed. The multivariate analysis showed that patients ≥75 years of age have a significantly worse OS (HR=1.42; p<0.001) and CSS (HR=1.27; p=0.018). CONCLUSIONS: An age of ≥75 years at RC is associated with a worse outcome. Prospective analyses including an assessment of the role of comorbidity and possibly age-dependent tumour biology are warranted.

microRNA122-regulated transgene expression increases specificity of cardiac gene transfer upon intravenous delivery of AAV9 vectors.

Adeno-associated virus (AAV) vectors with capsids of AAV serotype 9 enable an efficient transduction of the heart upon intravenous injection of adult mice but also transduce the liver. The aim of this study was to improve specificity of AAV9 vector-mediated cardiac gene transfer by microRNA (miR)-dependent control of transgene expression. We constructed plasmids and AAV vectors containing target sites (TSs) of liver-specific miR122, miR192 and miR148a in the 3' untranslated region (3'UTR) of a luciferase expression cassette. Luciferase expression was efficiently suppressed in liver cell lines expressing high levels of the corresponding miRs, whereas luciferase expression was unaffected in cardiac myocytes. Intravenous injections of AAV9 vectors bearing three repeats of miR122 TS in the 3'UTR of an enhanced green fluorescent expression (EGFP) expression cassette resulted in the absence of EGFP expression in the liver of adult mice, whereas the control vectors without miR TS displayed significant hepatic EGFP expression. EGFP expression levels in the heart, however, were comparable between miR122-regulated and control vectors. The liver-specific de-targeting in vivo using miR122 was even more efficient than transcriptional targeting with a cardiac cytomegalovirus (CMV)-enhanced myosin light chain (MLC) promoter. These data indicate that miR-regulated targeting is a powerful new tool to further improve cardiospecificity of AAV9 vectors.