Serum-based measurements of stromal activation through ADAM12 associate with poor prognosis in colorectal cancer.

BACKGROUND Recently it has been recognized that stromal markers could be used as a clinically relevant biomarker for therapy response and prognosis. Here, we report on a serum marker for stromal activation, A Disintegrin and Metalloprotease 12 (ADAM12) in colorectal cancer (CRC). METHODS Using gene expression databases we investigated ADAM12 expression in CRC and delineated the source of ADAM12 expression. The clinical value of ADAM12 was retrospectively assessed in the CAIRO2 trial in metastatic CRC with 235 patients (31% of total cohort), and an independent rectal cancer cohort (n = 20). RESULTS ADAM12 is expressed by activated CRC associated fibroblasts. In the CAIRO2 trial cohort, ADAM12 serum levels were prognostic (ADAM12 low versus ADAM12 high; median OS 25.3 vs. 17.1 months, HR 1.48 [95% CI 1.11-1.96], P = 0.007). The prognostic potential was specifically high for metastatic rectal cancer (HR 1.78 [95% CI 1.06-3.00], P = 0.030) and mesenchymal subtype tumors (HR 2.12 [95% CI 1.25-3.60], P = 0.004). ADAM12 also showed potential for predicting recurrence in an exploratory analysis of non-metastatic rectal cancers. CONCLUSIONS Here we describe a non-invasive marker for activated stroma in CRC which associates with poor outcome, especially for primary cancers located in the rectum

A genome-wide association study of myasthenia gravis

IMPORTANCE: Myasthenia gravis is a chronic, autoimmune, neuromuscular disease characterized by fluctuating weakness of voluntary muscle groups. Although genetic factors are known to play a role in this neuroimmunological condition, the genetic etiology underlying myasthenia gravis is not well understood. OBJECTIVE: To identify genetic variants that alter susceptibility to myasthenia gravis, we performed a genome-wide association study. DESIGN, SETTING, AND PARTICIPANTS: DNA was obtained from 1032 white individuals from North America diagnosed as having acetylcholine receptor antibody–positive myasthenia gravis and 1998 race/ethnicity-matched control individuals from January 2010 to January 2011. These samples were genotyped on Illumina OmniExpress single-nucleotide polymorphism arrays. An independent cohort of 423 Italian cases and 467 Italian control individuals were used for replication. MAIN OUTCOMES AND MEASURES: We calculated P values for association between 8114394 genotyped and imputed variants across the genome and risk for developing myasthenia gravis using logistic regression modeling. A threshold P value of 5.0 × 10(−8) was set for genome-wide significance after Bonferroni correction for multiple testing. RESULTS: In the over all case-control cohort, we identified association signals at CTLA4 (rs231770; P = 3.98 × 10(−8); odds ratio, 1.37; 95% CI, 1.25–1.49), HLA-DQA1 (rs9271871; P = 1.08 × 10(−8); odds ratio, 2.31; 95% CI, 2.02 – 2.60), and TNFRSF11A (rs4263037; P = 1.60 × 10(−9); odds ratio, 1.41; 95% CI, 1.29–1.53). These findings replicated for CTLA4 and HLA-DQA1 in an independent cohort of Italian cases and control individuals. Further analysis revealed distinct, but overlapping, disease-associated loci for early- and late-onset forms of myasthenia gravis. In the late-onset cases, we identified 2 association peaks: one was located in TNFRSF11A (rs4263037; P = 1.32 × 10(−12); odds ratio, 1.56; 95% CI, 1.44–1.68) and the other was detected in the major histocompatibility complex on chromosome 6p21 (HLA-DQA1; rs9271871; P = 7.02 × 10(−18); odds ratio, 4.27; 95% CI, 3.92–4.62). Association within the major histocompatibility complex region was also observed in early-onset cases (HLA-DQA1; rs601006; P = 2.52 × 10(−11); odds ratio, 4.0; 95% CI, 3.57–4.43), although the set of single-nucleotide polymorphisms was different from that implicated among late-onset cases. CONCLUSIONS AND RELEVANCE: Our genetic data provide insights into aberrant cellular mechanisms responsible for this prototypical autoimmune disorder. They also suggest that clinical trials of immunomodulatory drugs related to CTLA4 and that are already Food and Drug Administration approved as therapies for other autoimmune diseases could be considered for patients with refractory disease

Consensus-based care recommendations for adults with myotonic dystrophy type 1

Purpose of review Myotonic dystrophy type 1 (DM1) is a severe, progressive genetic disease that affects between 1 in 3,000 and 8,000 individuals globally. No evidence-based guideline exists to inform the care of these patients, and most do not have access to multidisciplinary care centers staffed by experienced professionals, creating a clinical care deficit. Recent findings The Myotonic Dystrophy Foundation (MDF) recruited 66 international clinicians experienced in DM1 patient care to develop consensus-based care recommendations. MDF created a 2-step methodology for the project using elements of the Single Text Procedure and the Nominal Group Technique. The process generated a 4-page Quick Reference Guide and a comprehensive, 55-page document that provides clinical care recommendations for 19 discrete body systems and/or care considerations. Summary The resulting recommendations are intended to help standardize and elevate care for this patient population and reduce variability in clinical trial and study environments. Described as “one of the more variable diseases found in medicine,” myotonic dystrophy type 1 (DM1) is an autosomal dominant, triplet-repeat expansion disorder that affects somewhere between 1:3,000 and 1:8,000 individuals worldwide.1 There is a modest association between increased repeat expansion and disease severity, as evidenced by the average age of onset and overall morbidity of the condition. An expansion of over 35 repeats typically indicates an unstable and expanding mutation. An expansion of 50 repeats or higher is consistent with a diagnosis of DM1. DM1 is a multisystem and heterogeneous disease characterized by distal weakness, atrophy, and myotonia, as well as symptoms in the heart, brain, gastrointestinal tract, endocrine, and respiratory systems. Symptoms may occur at any age. The severity of the condition varies widely among affected individuals, even among members of the same family. Comprehensive evidence-based guidelines do not currently exist to guide the treatment of DM1 patients. As a result, the international patient community reports varied levels of care and care quality, and difficulty accessing care adequate to manage their symptoms, unless they have access to multidisciplinary neuromuscular clinics. Consensus-based care recommendations can help standardize and improve the quality of care received by DM1 patients and assist clinicians who may not be familiar with the significant variability, range of symptoms, and severity of the disease. Care recommendations can also improve the landscape for clinical trial success by eliminating some of the inconsistencies in patient care to allow more accurate understanding of the benefit of potential therapies

Navigating, Negotiating, and Narrating: Re-Envisioning Patient-Centered Chronic Illness Care

Patient-centered care (PCC) is ubiquitous in how we think about patient-practitioner encounters. But such a taken-for granted stance may unknowingly obscure how conversations actually unfold in real life. The purpose of this work is to unravel the disconnect between how patient-centered care is talked about and how it is implemented in the real world. The overarching research question that framed this study was: What are the influences that shape the unfolding of the conversations that occur at chronic illness health encounters and how does this unfolding influence the learning and execution of PCC? The aim of this research was to offer a conceptualization of how patients’ and practitioners’ approaches to interactions at health encounters influence how stories unfold. Two major research studies framed this work addressing these research questions: (1) how do patients prepare and shape their stories of illness in order to interact productively with health practitioners and (2) how do interactions between patients and health practitioners shape the stories told at encounters? Using a constructivist grounded theory approach (CGT) a total of 32 participants -patients and practitioners- were interviewed using a semi-structured interview guide. Data collection and analysis was iterative using the constant comparative method. We found that patients did a lot of work to engage in health interactions and that this work was invisible to practitioners. Despite this work, however, patients were often left feeling that the stories that unfolded at health encounters often remained incomplete. Combining the perspectives of both patients and practitioners, we described different types of incomplete stories, namely the hidden story, the interpreted story, and the tailored story. The shared dimensions of making choices, balancing time, and targeting priorities informed the conceptualization of ‘Getting Airtime’ as a framework to understand how chronic illness interactions unfold at encounters. Using the framework of Patient-Centered Clinical Method (PCCM) to address educational considerations and the framework of Minimally Disruptive Medicine (MDM) to address practice considerations, we propose a re-envisioning of patient-centered encounters that reduces patients’ health interaction work, builds information-sharing capacity, and prevents harmful gaps in storytelling

Detection of mutant protein in complex biological samples: glucocerebrosidase mutations in Gaucher's disease

We report a sensitive method to detect point mutations in proteins from complex samples. The method is based on surface-enhanced laser desorption/ionization time-of-flight (SELDI-ToF) MS but can be extended to other MS platforms. The target protein in this study is the lysosomal enzyme glucocerebrosidase (GC), the key enzyme in Gaucher's disease. Deficiency of GC activity results in accumulation of glucosylceramide in macrophages. The relationship between GC genotypes and Gaucher's patient phenotypes is not strict. The possibility to measure protein levels of GC in clinical samples may provide deeper insight into the phenomenology of Gaucher's disease. For this purpose, GC was isolated in a single enrichment step through interaction with an immobilized monoclonal antibody, 8E4. After on-chip digestion of the antibody-antigen complex with trypsin, a total of 25 GC peptides were identified (sequence coverage approximately 60%), including several peptides containing mutated amino acid residues. The described methodology allows mutational analysis on the protein level, directly measured on complex biological samples without the necessity of elaborate purification procedure

Nanomolar affinity, iminosugar-based chemical probes for specific labeling of lysosomal glucocerebrosidase

Three different photoprobes were synthesized to label beta-glucosidases; one probe was based on glucose, two probes on the iminosugar deoxynojirimycin. The affinity of the probes for three different beta-glucosidases was determined. Furthermore, their labeling efficiencies, binding specificities through competition with deoxynojirimycin, and binding specificities in the presence of cell lysate, were evaluated. Especially one showed very high affinity towards non-lysosomal glucoceramidase (IC(50)=20nM

CCL18: a urinary marker of Gaucher cell burden in Gaucher patients

Glucosylceramide-laden tissue macrophages in Gaucher patients secrete large quantities of chitotriosidase and CC chemokine ligand 18 (CCL18), resulting in markedly increased plasma levels. We have comparatively investigated the occurrence of both parameters in plasma and urine samples of Gaucher patients. Chitotriosidase was high in urine samples of some symptomatic patients, but elevations did not correlate with increased plasma concentrations. Urinary chitotriosidase was particularly high in a patient with severe kidney involvement and local storage cell infiltration. Urinary levels of CCL18 were also highly elevated in samples from Gaucher patients as compared to controls. The median value of the CCL18/creatinine ratio in urine samples of 31 Gaucher patients was 143.3 pg/micromol (range 32-551) and in those of 12 normal subjects was 4.1 pg/micromol (range 1.3-6.8). In sharp contrast to chitotriosidase, increases in the low-molecular-mass chemokine CCL18 in urine and plasma specimens of Gaucher patients correlated well. A correlation was also observed for reductions in urinary and plasma CCL18 following therapy. It is concluded that assessment of urinary CCL18 of Gaucher patients gives insight into the total body burden on Gaucher cells, whereas that of chitotriosidase does not. Urinary chitotriosidase appears rather to be a reflection of renal patholog

HEPES activates a MiT/TFE-dependent lysosomal-autophagic gene network in cultured cells: A call for caution

In recent years, the lysosome has emerged as a highly dynamic, transcriptionally regulated organelle that is integral to nutrient-sensing and metabolic rewiring. This is coordinated by a lysosome-to-nucleus signaling nexus in which MTORC1 controls the subcellular distribution of the microphthalmia-transcription factor E (MiT/TFE) family of "master lysosomal regulators". Yet, despite the importance of the lysosome in cellular metabolism, the impact of traditional in vitro culture media on lysosomal dynamics and/or MiT/TFE localization has not been fully appreciated. Here, we identify HEPES, a chemical buffering agent that is broadly applied in cell culture, as a potent inducer of lysosome biogenesis. Supplementation of HEPES to cell growth media is sufficient to decouple the MiT/TFE family members-TFEB, TFE3 and MITF-from regulatory mechanisms that control their cytosolic retention. Increased MiT/TFE nuclear import in turn drives the expression of a global network of lysosomal-autophagic and innate host-immune response genes, altering lysosomal dynamics, proteolytic capacity, autophagic flux, and inflammatory signaling. In addition, siRNA-mediated MiT/TFE knockdown effectively blunted HEPES-induced lysosome biogenesis and gene expression profiles. Mechanistically, we show that MiT/TFE activation in response to HEPES requires its macropinocytic ingestion and aberrant lysosomal storage/pH, but is independent of MTORC1 signaling. Altogether, our data underscore the cautionary use of chemical buffering agents in cell culture media due to their potentially confounding effects on experimental result

Synthesis and evaluation of dimeric lipophilic iminosugars as inhibitors of glucosylceramide metabolism

Four dimeric and four monomeric lipophilic iminosugars were synthesized and subsequently evaluated on their inhibitory potential towards mammalian glucosylceramide synthase, glucocerebrosidase, beta-glucosidase 2, sucrase and lysosomal alpha-glucosidase. Compared to their monomeric counterparts the dimeric inhibitors showed decreased inhibition of glucosylceramide synthase and generally a comparable inhibitory potency for the glycosidases. (C) 2009 Elsevier Ltd. All rights reserve