MCAM and its Isoforms as Novel Targets in Angiogenesis Research and Therapy

Melanoma cell adhesion molecule (MCAM) (CD146) is a membrane glycoprotein of the mucin family. It is one of the numerous proteins composing the junction of the vascular endothelium, and it is expressed in other cell types such as cancer cells, smooth muscle cells, and pericytes. Some recent works were designed to highlight its structural features, its location in the endothelium, and its role in angiogenesis, vascular permeability, and monocyte transmigration, but also in the maintenance of endothelial junctions and tumor development. MCAM exists in different splice variants and is shedded from the vascular membrane by metalloproteases. Studies about MCAM spliced and cleaved variant on human angiogenic physiological and pathological models permit a better understanding on the roles initially described for this protein. Furthermore, this knowledge will help in the future to develop therapeutic and diagnostic tools targeting specifically the different MCAM variant. Recent advances in research on angiogenesis and in the implication of MCAM in this process are discussed in this chapter

Expression of the Neuroblastoma-Associated ALK-F1174L Activating Mutation During Embryogenesis Impairs the Differentiation of Neural Crest Progenitors in Sympathetic Ganglia

Neuroblastoma (NB) is an embryonal malignancy derived from the abnormal differentiation of the sympathetic nervous system. The Anaplastic Lymphoma Kinase (ALK) gene is frequently altered in NB, through copy number alterations and activating mutations, and represents a predisposition in NB-genesis when mutated. Our previously published data suggested that ALK activating mutations may impair the differentiation potential of neural crest (NC) progenitor cells. Here, we demonstrated that the expression of the endogenous ALK gene starts at E10.5 in the developing sympathetic ganglia (SG). To decipher the impact of deregulated ALK signaling during embryogenesis on the formation and differentiation of sympathetic neuroblasts, Sox10-Cre;LSL-ALK-F1174L embryos were produced to restrict the expression of the human ALK-F1174L transgene to migrating NC cells (NCCs). First, ALK-F1174L mediated an embryonic lethality at mid-gestation and an enlargement of SG with a disorganized architecture in Sox10-Cre;LSL-ALK-F1174L embryos at E10.5 and E11.5. Second, early sympathetic differentiation was severely impaired in Sox10-Cre;LSL-ALK-F1174L embryos. Indeed, their SG displayed a marked increase in the proportion of NCCs and a decrease of sympathetic neuroblasts at both embryonic stages. Third, neuronal and noradrenergic differentiations were blocked in Sox10-Cre;LSL-ALK-F1174L SG, as a reduced proportion of Phox2b+ sympathoblasts expressed βIII-tubulin and almost none were Tyrosine Hydroxylase (TH) positive. Finally, at E10.5, ALK-F1174L mediated an important increase in the proliferation of Phox2b+ progenitors, affecting the transient cell cycle exit observed in normal SG at this embryonic stage. Altogether, we report for the first time that the expression of the human ALK-F1174L mutation in NCCs during embryonic development profoundly disturbs early sympathetic progenitor differentiation, in addition to increasing their proliferation, both mechanisms being potential crucial events in NB oncogenesis

Identification of genetic variants associated with Huntington's disease progression: a genome-wide association study

Background Huntington's disease is caused by a CAG repeat expansion in the huntingtin gene, HTT. Age at onset has been used as a quantitative phenotype in genetic analysis looking for Huntington's disease modifiers, but is hard to define and not always available. Therefore, we aimed to generate a novel measure of disease progression and to identify genetic markers associated with this progression measure. Methods We generated a progression score on the basis of principal component analysis of prospectively acquired longitudinal changes in motor, cognitive, and imaging measures in the 218 indivduals in the TRACK-HD cohort of Huntington's disease gene mutation carriers (data collected 2008–11). We generated a parallel progression score using data from 1773 previously genotyped participants from the European Huntington's Disease Network REGISTRY study of Huntington's disease mutation carriers (data collected 2003–13). We did a genome-wide association analyses in terms of progression for 216 TRACK-HD participants and 1773 REGISTRY participants, then a meta-analysis of these results was undertaken. Findings Longitudinal motor, cognitive, and imaging scores were correlated with each other in TRACK-HD participants, justifying use of a single, cross-domain measure of disease progression in both studies. The TRACK-HD and REGISTRY progression measures were correlated with each other (r=0·674), and with age at onset (TRACK-HD, r=0·315; REGISTRY, r=0·234). The meta-analysis of progression in TRACK-HD and REGISTRY gave a genome-wide significant signal (p=1·12 × 10−10) on chromosome 5 spanning three genes: MSH3, DHFR, and MTRNR2L2. The genes in this locus were associated with progression in TRACK-HD (MSH3 p=2·94 × 10−8 DHFR p=8·37 × 10−7 MTRNR2L2 p=2·15 × 10−9) and to a lesser extent in REGISTRY (MSH3 p=9·36 × 10−4 DHFR p=8·45 × 10−4 MTRNR2L2 p=1·20 × 10−3). The lead single nucleotide polymorphism (SNP) in TRACK-HD (rs557874766) was genome-wide significant in the meta-analysis (p=1·58 × 10−8), and encodes an aminoacid change (Pro67Ala) in MSH3. In TRACK-HD, each copy of the minor allele at this SNP was associated with a 0·4 units per year (95% CI 0·16–0·66) reduction in the rate of change of the Unified Huntington's Disease Rating Scale (UHDRS) Total Motor Score, and a reduction of 0·12 units per year (95% CI 0·06–0·18) in the rate of change of UHDRS Total Functional Capacity score. These associations remained significant after adjusting for age of onset. Interpretation The multidomain progression measure in TRACK-HD was associated with a functional variant that was genome-wide significant in our meta-analysis. The association in only 216 participants implies that the progression measure is a sensitive reflection of disease burden, that the effect size at this locus is large, or both. Knockout of Msh3 reduces somatic expansion in Huntington's disease mouse models, suggesting this mechanism as an area for future therapeutic investigation

ALK IN THE CONTROL OF THE SYMPATHOADRENAL LINEAGE DIFFERENTIATION AND NEUROBLASTOMA TUMOR PROGRESSION

Neuroblastoma (NB) is an embryonal malignancy derived from the abnormal differentiation of the sympathetic nervous system. NB is a highly heterogeneous disease in terms of clinical, biological and genetic features. NB display an intratumoral heterogeneity and are composed of both neuroblastic and mesenchymal-like cells, displaying a cellular plasticity between both phenotypes. The Anaplastic Lymphoma Kinase (ALK) gene is frequently altered in NB through copy number changes and mutations, with ALK-F1174L and ALK-R1275Q being the most potent activating mutations. ALK amplification and mutations are associated to decrease in the overall free-survival of NB patients. Increased frequencies of ALK mutations have been found at relapse in NB. ALK thus represents a bona fide target for NB therapy. We studied the impact of deregulated ALK signaling during embryonic development on the formation and differentiation of sympathetic neuroblasts. We report that the expression of the human ALK-F1174L mutation in migrating neural crest cells disturbs early sympathetic progenitor differentiation, in addition to increasing their proliferation, both mechanisms being potentially crucial events in the development of embryonal tumors such as NB. Our study suggests that dysregulated signaling of ALK during embryogenesis promotes a precancerous state and may represent an initial and developmental path for NB oncogenesis. In the second part of this work, we investigated the impact of ALK-wt, ALK-R1275Q and ALK-F1174L -mediated signaling on NB tumorigenesis and progression. ALK isoforms were overexpressed in the human NB cell lines SK-N-Be2c (adrenergic and tumorigenic) and SH-EP (mesenchymal and weakly tumorigenic) through lentiviral vectors. Transduced cells were further injected orthotopically in the adrenal gland of athymic Swiss nude mice or subcutaneously in NOD-SCID- mice, respectively. The overexpression of the ALK variants had a low impact on the aggressive NB cell line SK-N-Be2c. In contrast, the expression of ALK-wt, ALK-R1275Q and ALK-F1174L in the SH- EP cell line promotes an aggressive phenotype of the tumors. Indeed, histological and transcriptomic analyses revealed that the overexpression of ALK mediates a change in the cellular phenotype and increases the metastatic potential of the SH-EP tumor cells. Furthermore, we identified the downregulation of genes related to axon guidance, extracellular matrix (ECM) organization in ALK-wt and ALK-F1174L tumors, while these pathways were found upregulated in ALK-R1275Q tumors. Further investigations are needed to validate these data and to understand the consequences of their deregulation in NB pathogenesis. To conclude, this work contributed to increase the understanding of ALK mutation as a path to initiate NB-genesis during embryonic development. Moreover, our observations suggest that ALK overexpression and mutations may promote an aggressive phenotype in NB tumors, further opening new research perspectives for the role of ALK deregulated signaling in NB. -- Le neuroblastome (NB) est une tumeur se développant à partir du système nerveux sympathique (SNS). Le NB est une maladie très hétérogène en termes d’évolution clinique, caractéristiques biologiques et altérations génétiques. Les tumeurs de NB présentent également une hétérogénéité intratumorale. Elles sont composées de cellules adrénergiques et mésenchymateuses qui présentent une plasticité cellulaire. Le gène Anaplastic lymphoma kinase (ALK) est fréquemment amplifié ou muté dans le NB, et ces altérations sont associées à un mauvais pronostic chez les patients. Les mutations ALK-F1174L et ALK-R1275Q sont les mutations les plus fréquentes. De plus, une augmentation du nombre de mutations ponctuelles du gène ALK a été décrite lors des rechutes. ALK constitue ainsi une cible thérapeutique importante dans le NB. Nous avons étudié l'impact de la mutation ALK-F1174L sur la formation et la différentiation du SNS lors du développement embryonnaire. L'expression de la mutation ALK-F1174L dans les cellules de crête neurale perturbe la formation du SNS, en bloquant la différenciation des progéniteurs sympathiques et en augmentant leur prolifération. Ces deux mécanismes sont des événements cruciaux pour le développement du NB. Notre étude suggère ainsi que l’expression de la mutation ALK- F1174L pendant l'embryogenèse favorise un état précancéreux et serait ainsi impliqué dans l’initiation de la maladie. Dans un second temps, nous avons étudié l'implication de ALK-wt et des mutations ALK-F1174L et ALK-R1275Q dans la pathogenèse du NB. Les variants ont été surexprimés dans les lignées de NB SK-N-Be2c (adrénergique et tumorigène) et SH- EP (mésenchymateuse et peu tumorigène) par infection lentivirale. Les cellules transduites ont ensuite été injectées respectivement de façon orthotopique dans la glande surrénale de souris athymic Swiss nude mice, ou en sous-cutanée dans des souris NOD-SCID- mice. Alors que la surexpression des variants ALK n’a que peu d’impact dans la lignée de NB agressive SK-N-Be2c, la surexpression de ALK-wt, ALK- F1174L et ALK-R1275Q dans la lignée peu tumorigène SH-EP induit un phénotype agressif des tumeurs de NB. En effet, les analyses histologiques et transcriptomiques suggèrent que la surexpression du gène ALK favorise un changement du phénotype cellulaire et montrent une augmentation du potentiel métastatique des tumeurs. De plus, des gènes impliqués dans la formation axonale et l’organisation de la matrice extracellulaire (ECM) sont négativement régulés dans les tumeurs ALK-wt et ALK- F1174L, alors que ceux-ci sont positivement régulés dans les tumeurs ALK-R1275Q. Des études approfondies seront nécessaires pour valider ces données et comprendre les conséquences de la modulation de ces gènes dans la pathogenèse du NB. Pour conclure, ce travail a permis d’apporter une meilleure compréhension de l’impact des mutations du gène ALK au niveau du développement embryonnaire. De plus, nos observations suggèrent que l’amplification et les mutations induisent un phénotype agressif des tumeurs de NB. Ces données ouvrent de nouvelles perspectives de recherche quant au rôle de ALK dans l’initiation et le développement du NB

Therapeutic targeting of soluble CD146/MCAM with the M2J‐1 monoclonal antibody prevents metastasis development and procoagulant activity in CD146‐positive invasive tumors

International audienceInitially discovered in human melanoma, CD146/MCAM is expressed on many tumors and is correlated with cancer progression and metastasis. However, targeting CD146 remains challenging since it is also expressed on other cell types, as vessel cells, where it displays important physiological functions. We previously demonstrated that CD146 is shed as a soluble form (sCD146) that vectorizes the effects of membrane CD146 on tumor angiogenesis, growth and survival. We thus generated a novel monoclonal antibody, the M2J-1 mAb, which specifically targets sCD146, but not membrane CD146, and counteracts these effects. In our study, we analyzed the effects of sCD146 on the dissemination and the associated procoagulant phenotype in two highly invasive human CD146-positive cancer cell lines (ovarian and melanoma). Results show that sCD146 induced epithelial to mesenchymal transition, favored the generation of cancer stem cells and increased the membrane expression of tissue factor. Treatment of cancer cells with sCD146 in two experimental models (subcutaneous xenografting and intracardiac injection of cancer cells in nude mice) led to increased tumor dissemination and procoagulant activity. The M2J-1 mAb drastically reduced metastasis but also procoagulant activity, in particular by decreasing the number of circulating tumor microparticles, and blocked the relevant signaling pathways as demonstrated by RNA expression profiling experiments. Thus, our findings demonstrate that sCD146 mediates important pro-metastatic and procoagulant effects in two CD146-positive tumors. Targeting sCD146 with the newly generated M2J-1 mAb could constitute an innovative strategy for preventing dissemination and thromboembolism in many CD146-positive tumors

Therapeutic targeting of soluble CD146/MCAM with the M2J‐1 monoclonal antibody prevents metastasis development and procoagulant activity in CD146‐positive invasive tumors

International audienceInitially discovered in human melanoma, CD146/MCAM is expressed on many tumors and is correlated with cancer progression and metastasis. However, targeting CD146 remains challenging since it is also expressed on other cell types, as vessel cells, where it displays important physiological functions. We previously demonstrated that CD146 is shed as a soluble form (sCD146) that vectorizes the effects of membrane CD146 on tumor angiogenesis, growth and survival. We thus generated a novel monoclonal antibody, the M2J-1 mAb, which specifically targets sCD146, but not membrane CD146, and counteracts these effects. In our study, we analyzed the effects of sCD146 on the dissemination and the associated procoagulant phenotype in two highly invasive human CD146-positive cancer cell lines (ovarian and melanoma). Results show that sCD146 induced epithelial to mesenchymal transition, favored the generation of cancer stem cells and increased the membrane expression of tissue factor. Treatment of cancer cells with sCD146 in two experimental models (subcutaneous xenografting and intracardiac injection of cancer cells in nude mice) led to increased tumor dissemination and procoagulant activity. The M2J-1 mAb drastically reduced metastasis but also procoagulant activity, in particular by decreasing the number of circulating tumor microparticles, and blocked the relevant signaling pathways as demonstrated by RNA expression profiling experiments. Thus, our findings demonstrate that sCD146 mediates important pro-metastatic and procoagulant effects in two CD146-positive tumors. Targeting sCD146 with the newly generated M2J-1 mAb could constitute an innovative strategy for preventing dissemination and thromboembolism in many CD146-positive tumors

Reduced Cancer Incidence in Huntington's Disease: Analysis in the Registry Study

Background: People with Huntington's disease (HD) have been observed to have lower rates of cancers. Objective: To investigate the relationship between age of onset of HD, CAG repeat length, and cancer diagnosis. Methods: Data were obtained from the European Huntington's disease network REGISTRY study for 6540 subjects. Population cancer incidence was ascertained from the GLOBOCAN database to obtain standardised incidence ratios of cancers in the REGISTRY subjects. Results: 173/6528 HD REGISTRY subjects had had a cancer diagnosis. The age-standardised incidence rate of all cancers in the REGISTRY HD population was 0.26 (CI 0.22-0.30). Individual cancers showed a lower age-standardised incidence rate compared with the control population with prostate and colorectal cancers showing the lowest rates. There was no effect of CAG length on the likelihood of cancer, but a cancer diagnosis within the last year was associated with a greatly increased rate of HD onset (Hazard Ratio 18.94, p < 0.001). Conclusions: Cancer is less common than expected in the HD population, confirming previous reports. However, this does not appear to be related to CAG length in HTT. A recent diagnosis of cancer increases the risk of HD onset at any age, likely due to increased investigation following a cancer diagnosis

A global metagenomic map of urban microbiomes and antimicrobial resistance

We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms. Profiles of AMR genes varied widely in type and density across cities. Cities showed distinct microbial taxonomic signatures that were driven by climate and geographic differences. These results constitute a high-resolution global metagenomic atlas that enables discovery of organisms and genes, highlights potential public health and forensic applications, and provides a culture-independent view of AMR burden in cities.Funding: the Tri-I Program in Computational Biology and Medicine (CBM) funded by NIH grant 1T32GM083937; GitHub; Philip Blood and the Extreme Science and Engineering Discovery Environment (XSEDE), supported by NSF grant number ACI-1548562 and NSF award number ACI-1445606; NASA (NNX14AH50G, NNX17AB26G), the NIH (R01AI151059, R25EB020393, R21AI129851, R35GM138152, U01DA053941); STARR Foundation (I13- 0052); LLS (MCL7001-18, LLS 9238-16, LLS-MCL7001-18); the NSF (1840275); the Bill and Melinda Gates Foundation (OPP1151054); the Alfred P. Sloan Foundation (G-2015-13964); Swiss National Science Foundation grant number 407540_167331; NIH award number UL1TR000457; the US Department of Energy Joint Genome Institute under contract number DE-AC02-05CH11231; the National Energy Research Scientific Computing Center, supported by the Office of Science of the US Department of Energy; Stockholm Health Authority grant SLL 20160933; the Institut Pasteur Korea; an NRF Korea grant (NRF-2014K1A4A7A01074645, 2017M3A9G6068246); the CONICYT Fondecyt Iniciación grants 11140666 and 11160905; Keio University Funds for Individual Research; funds from the Yamagata prefectural government and the city of Tsuruoka; JSPS KAKENHI grant number 20K10436; the bilateral AT-UA collaboration fund (WTZ:UA 02/2019; Ministry of Education and Science of Ukraine, UA:M/84-2019, M/126-2020); Kyiv Academic Univeristy; Ministry of Education and Science of Ukraine project numbers 0118U100290 and 0120U101734; Centro de Excelencia Severo Ochoa 2013–2017; the CERCA Programme / Generalitat de Catalunya; the CRG-Novartis-Africa mobility program 2016; research funds from National Cheng Kung University and the Ministry of Science and Technology; Taiwan (MOST grant number 106-2321-B-006-016); we thank all the volunteers who made sampling NYC possible, Minciencias (project no. 639677758300), CNPq (EDN - 309973/2015-5), the Open Research Fund of Key Laboratory of Advanced Theory and Application in Statistics and Data Science – MOE, ECNU, the Research Grants Council of Hong Kong through project 11215017, National Key RD Project of China (2018YFE0201603), and Shanghai Municipal Science and Technology Major Project (2017SHZDZX01) (L.S.

Clinical and genetic characteristics of late-onset Huntington's disease

Background: The frequency of late-onset Huntington's disease (&gt;59 years) is assumed to be low and the clinical course milder. However, previous literature on late-onset disease is scarce and inconclusive. Objective: Our aim is to study clinical characteristics of late-onset compared to common-onset HD patients in a large cohort of HD patients from the Registry database. Methods: Participants with late- and common-onset (30–50 years)were compared for first clinical symptoms, disease progression, CAG repeat size and family history. Participants with a missing CAG repeat size, a repeat size of ≤35 or a UHDRS motor score of ≤5 were excluded. Results: Of 6007 eligible participants, 687 had late-onset (11.4%) and 3216 (53.5%) common-onset HD. Late-onset (n = 577) had significantly more gait and balance problems as first symptom compared to common-onset (n = 2408) (P &lt;.001). Overall motor and cognitive performance (P &lt;.001) were worse, however only disease motor progression was slower (coefficient, −0.58; SE 0.16; P &lt;.001) compared to the common-onset group. Repeat size was significantly lower in the late-onset (n = 40.8; SD 1.6) compared to common-onset (n = 44.4; SD 2.8) (P &lt;.001). Fewer late-onset patients (n = 451) had a positive family history compared to common-onset (n = 2940) (P &lt;.001). Conclusions: Late-onset patients present more frequently with gait and balance problems as first symptom, and disease progression is not milder compared to common-onset HD patients apart from motor progression. The family history is likely to be negative, which might make diagnosing HD more difficult in this population. However, the balance and gait problems might be helpful in diagnosing HD in elderly patients