VE-cadherin RGD motifs promote metastasis and constitute a potential therapeutic target in melanoma and breast cancers

13 p.-6 fig.We have investigated the role of vascular-endothelial (VE)-cadherin in melanoma and breast cancer metastasis. We found that VE-cadherin is expressed in highly aggressive melanoma and breast cancer cell lines. Remarkably, inactivation of VEcadherin triggered a significant loss of malignant traits (proliferation, adhesion, invasion and transendothelial migration) in melanoma and breast cancer cells. These effects, except transendothelial migration, were induced by the VE-cadherin RGD motifs. Co-immunoprecipitation experiments demonstrated an interaction between VE-cadherin and α2β1 integrin, with the RGD motifs found to directly affect β1 integrin activation. VE-cadherin-mediated integrin signaling occurred through specific activation of SRC, ERK and JNK, including AKT in melanoma. Knocking down VEcadherin suppressed lung colonization capacity of melanoma or breast cancer cells inoculated in mice, while pre-incubation with VE-cadherin RGD peptides promoted lung metastasis for both cancer types. Finally, an in silico study revealed the association of high VE-cadherin expression with poor survival in a subset of melanoma patients and breast cancer patients showing low CD34 expression. These findings support a general role for VE-cadherin and other RGD cadherins as critical regulators of lung and liver metastasis in multiple solid tumours. These results pave the way for cadherin-specific RGD targeted therapies to control disseminated metastasis in multiple cancers.BEP was an FPI fellow from Ministry of Economy and Competitiveness (MINECO). This research was supported by grants BIO2012-31023 and BIO2015-66849 from MINECO and PRB2 (IPT13/0001-ISCIII-SGEFI/FEDER) to JIC.Peer reviewe

A prognostic six-gene expression risk-score derived from proteomic profiling of the metastatic colorectal cancer secretome

14 p.-6 fig.-1 tab.The necessity to accurately predict recurrence and clinical outcome in early stage colorectal cancer (CRC) is critical to identify those patients who may benefit from adjuvant chemotherapy. Here, we developed and validated a gene-based risk-score algorithm for patient stratification and personalised treatment in early stage disease based on alterations in the secretion of metastasis-related proteins. A quantitative label-free proteomic analysis of the secretome of highly and poorly metastatic CRC cell lines with different genetic backgrounds revealed 153 differentially secreted proteins (fold-change >5). These changes in the secretome were validated at the transcriptomic level. Starting from 119 up-regulated proteins, a six-gene/protein-based prognostic signature composed of IGFBP3, CD109, LTBP1, PSAP, BMP1, and NPC2 was identified after sequential discovery, training,and validation in four different cohorts. This signature was used to develop a risk-score algorithm, named SEC6,for patient stratification. SEC6 risk-score components showed higher expression in the poor prognosis CRC sub types: consensus molecular subtype 4 (CMS4), CRIS-B, and stem-like. High expression of the signature was also associated with patients showing dMMR, CIMP+ status, and BRAF mutations. In addition, the SEC6 signature was associated with lower overall survival, progression-free interval, and disease-specific survival in stage II and III patients. SEC6-based risk stratification indicated that 5-FU treatment was beneficial for low-risk patients,whereas only aggressive treatments (FOLFOX and FOLFIRI) provided benefits to high-risk patients in stages II and III. In summary, this novel risk-score demonstrates the value of the secretome compartment as a reliable source for the retrieval of biomarkers with high prognostic and chemotherapy-predictive capacity, providing a potential new tool for tailoring decision-making in patient care.This project was supported by grants RTI2018-095055-B-100 and PID2021-122227OB-I00 from the MICYT, IND2019/BMD-17153 from the Comunidad de Madrid and PRB3 (ISCIII-SGEFI/FEDER- PT17/0019/0008) from the ISCIII.Peer reviewe

Elective Cancer Surgery in COVID-19-Free Surgical Pathways During the SARS-CoV-2 Pandemic: An International, Multicenter, Comparative Cohort Study.

PURPOSE: As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19-free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS: This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19-free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS: Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19-free surgical pathways. Patients who underwent surgery within COVID-19-free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19-free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score-matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19-free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION: Within available resources, dedicated COVID-19-free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks

Elective cancer surgery in COVID-19-free surgical pathways during the SARS-CoV-2 pandemic: An international, multicenter, comparative cohort study

PURPOSE As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19–free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19–free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19–free surgical pathways. Patients who underwent surgery within COVID-19–free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19–free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score–matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19–free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION Within available resources, dedicated COVID-19–free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks

An RGD motif present in cadherin 17 induces integrin activation and tumor growth

15 p.-9 fig.Little is known about the mechanism of integrin activation by cadherin 17 (CDH17). Here we observed the presence of a tri-peptide motif, RGD, in domain 6 of the human CDH17 sequence and other cadherins such as cadherin 5 and cadherin 6. The use of CDH17 RAD mutants demonstrated a considerable decrease of proliferation and adhesion in RKO and KM12SM colon cancer cells. Furthermore, RGD peptides inhibited the adhesion of both cell lines to recombinant CDH17 domain 6. The RGD motif added exogenously to the cells provoked a change in β1 integrin to an active, high-affinity conformation and an increase in focal adhesion kinase and ERK1/2 activation. In vivo experiments with Swiss nude mice demonstrated that cancer cells expressing the CDH17 RAD mutant showed a considerable delay in tumor growth and liver homing. CDH17 RGD effects were also active in pancreatic cancer cells. Our results suggest that α2β1 integrin interacts with two different ligands, collagen IV and CDH17, using two different binding sites. In summary, the RGD binding motif constitutes a switch for integrin pathway activation and shows a novel capacity of CDH17 as an integrin ligand. This motif could be targeted to avoid metastatic dissemination in tumors overexpressing CDH17 and other RGD-containing cadherins.This research was supported by grant BIO2012-31023 from the Spanish Ministry of Economy and Competitiveness, by a grant to established research groups (AECC), and by grant S2011/BMD-2344/ (Colomics2) from Comunidad de Madrid and ProteoRed-ISCIII.Peer reviewe

El factor de intercambio de nucleótidos de guanina C3G adopta una conformación cerrada estabilizada por una interacción cabeza-cola

Resumen del póster presentado al XXXVIII Congreso de la Sociedad Española de Bioquímica y Biología Molecular, celebrado en Valencia (España) del 7 al 10 de septiembre de 2015.C3G es un factor de intercambio de nucleótidos de guanina (GEF) que activa las GTPasas Rap1 y R-Ras. C3G participa en adhesión y migración celular, en remodelación de actina, transformación celular, apoptosis, proliferación celular, maduración vascular, maduración y expansión de timocitos, transporte de glucosa y diferenciación. C3G (120 kDa) tiene una estructura tripartita. La región amino (N-C3G), cuya estructura se desconoce y que carece de similitud con otras proteínas, alberga un sitio de unión a E-cadherina. La región central contiene 5 motivos ricos en Pro que median la unión a proteínas con dominios SH3 como Crk, p130Cas, Grb2, Hck y c-Abl; a esta región también se unen la fosfatasa TC-PTP y ß-catenina, que carecen de dominios SH3. La región catalítica C-terminal contiene un dominio Cdc25H y un dominio REM. La deleción de la mitad amino de C3G aumenta la actividad GEF, sugiriendo que esa región actúa como un elemento regulador negativo, si bien se desconocen los mecanismos de autoinhibición. Aquí hemos detectado y caracterizado una interacción intramolecular entre N-C3G y la región catalítica, y hemos identificado residuos importantes para el mantenimiento de la conformación cerrada. Esta interacción cabeza-cola en C3G es reminiscente de conformaciones autoinhibidas de otros GEFs Cdc25H como Sos1, Epac2 y RasGRP1. Por último, empleando dicroísmo circular en el UV-lejano hemos estimado que N-C3G tiene un alto contenido en hélice-¿. La naturaleza anfipática de las potenciales hélices de N-C3G sugiere que podrían formar un haz de hélices.Peer Reviewe

Structural organization of the guanine nucleotide exchange factor C3G

Resumen del póster presentado al 5th International Iberian Biophysics Congress, celebrado en Porto (Portugal) del 15 al 17 de junio de 2016.C3G is a guanine nucleotide exchange factor (GEF) that activates the small GTPases Rap1 and R-Ras. C3G is involved in multiple cellular functions including adhesion, migration, cytoskeletal remodeling, cell proliferation, differentiation, transformation, and apoptosis. C3G (120 kDa) has a tripartite structure. The N-terminal region (N-C3G) mediates binding to E-cadherin at nascent adherent junctions. The structure of N-C3G is unknown and bears no similarity to other proteins. The central region contains five Pro-rich sequence motifs that mediate the interaction with protein that contain SH3 domains, such as Crk, p130Cas, Grb2, Hck and c-Abl. The central region also mediates binding to the TC-PTP phosphatase and to ß-catenin, which do not contain SH3 domains. The C-terminal catalytic region consists of a REM (Ras exchange motif) and a Cdc25H domain. Deletion of the N-terminal half of C3G increases the GEF activity, suggesting that this N-terminal half acts as a regulatory element. Yet, the mechanisms of autoinhibition of C3G remain unknown. Here we have identified and characterized an intramolecular interaction between the N-C3G and the catalytic region. We have identified residues important for maintaining the close conformation. This head-tail interaction in C3G resembles autoinhibitory conformations of other GEFs of the Cdc25H family, such as Sos1, Epac2, and RasGRP1. Finally, using recombinant N-C3G fragments we show that this region has a high content of α-helical structure. Sequence analysis suggests the presence of amphipathic helices that could fold in a helical bundle.Peer Reviewe

PAUF/ZG16B promotes colorectal cancer progression through alterations of the mitotic functions and the Wnt/β-catenin pathway

37 p.-6 fig.Pancreatic adenocarcinoma–upregulated factor (PAUF), also known as ZG16B, was previously found in the secretome of metastatic colorectal cancer cells. Here, we demonstrated the presence of PAUF at the intracellular level and its multiple effects on cancer progression. An initial decline of PAUF expression was observed at early stages of colorectal cancer followed by an increase at the metastastic site. PAUF was located at different cellular compartments: membrane-associated vesicles, endosomes, microtubule-associated vesicles, cell growth cones and the cell nucleus. PAUF loss in two colorectal cancer cell lines caused severe alterations in the cell phenotype and cell cycle, including tetraploidy, extensive genomic alterations, micronuclei, and increased apoptosis. An exhaustive analysis of the PAUF interactome using different proteomic approaches revealed the presence of multiple components of the cell cycle, mitotic checkpoint, Wnt pathway and intracellular transport. Among the interacting proteins we found ZW10, a moonlighting protein with a dual function in membrane trafficking and mitosis. In addition, PAUF silencing was associated to APC loss and increased β-catenin nuclear expression. Altogether, our results suggest that PAUF depletion increases aneuploidy, promotes apoptosis and activates the Wnt/β-catenin pathway in colorectal cancer cells facilitating cancer progression. In summary, PAUF behaves as a multifunctional protein, with different roles in cancer progression according to the extra- or intracellular expression, suggesting a therapeutic value for colorectal cancer.This research was supported by grants from the MINECO (BIO2015-66489-R), Fundación Areces and PRB3 (IPT17/0019 - ISCIII-SGEFI /ERDF).Peer reviewe

Twist1-induced activation of human fibroblasts promotes matrix stiffness by upregulating palladin and collagen α1(VI)

The transcription factor Twist1 is involved in the epithelial–mesenchymal transition and contributes to cancer metastasis through mostly unknown mechanisms. In colorectal cancer, Twist1 expression is mainly restricted to the tumor stroma. We found that human fibroblast cell lines stably transfected with Twist1 acquired characteristics of activated cancer-associated fibroblasts (CAFs), such as hyperproliferation, an increased ability to migrate and an alignment of the actin cytoskeleton. Further, Twist1-activated fibroblasts promoted increased matrix stiffness. Using quantitative proteomics, we identified palladin and collagen α1(VI) as two major mediators of the Twist1 effects in fibroblast cell lines. Co-immunoprecipitation studies indicated that palladin and Twist1 interact within the nucleus, suggesting that palladin could act as a transcription regulator. Palladin was found to be more relevant for the cellular biomechanical properties, orientation and polarity, and collagen α1(VI) for the migration and invasion capacity, of Twist1-activated fibroblasts. Both palladin and collagen α1(VI) were observed to be overexpressed in colorectal CAFs and to be associated with poor colorectal cancer patient survival and relapse prediction. Our results demonstrate that Twist1-expressing fibroblasts mimic the properties of CAFs present at the tumor invasive front, which likely explains the prometastatic activities of Twist1. Twist1 appears to require both palladin and collagen α1(VI) as downstream effectors for its prometastatic effects, which could be future therapeutic targets in cancer metastasis.IG-P was supported by a contract S2010/BMD-2344/ Colomics2 from the Comunidad de Madrid. ST was a recipient of a Juan de la Cierva programme. RAB was supported by a grant to established research groups of the Asociación Española Contra el Cáncer (AECC). ML-L was a recipient of a ProteoRed contract. AP-G and BE-P were FPI fellows from the Ministry of Economy and Competitiveness (MINECO). This research was supported by grants to established research groups of the S2010/BMD-2344/Colomics2 from the Comunidad de Madrid, ‘Asociación Española Contra el Cancer (AECC)’, BIO2012-31023 from the MINECO, PRB2 (IPT13/0001-ISCIII-SGEFI/FEDER), RD12/0036/0041 and RD12/0036/0021 from the Instituto de Salud Carlos III-FEDER.Peer reviewe

Cerebellar Hemangioblastoma with Leptomeningeal Spread and a Fatal Outcome: A Rare Case Report with MDM2 and EGFR Alterations

Luis Miguel Chinchilla-Tábora,1 Javier Ortiz Rodríguez-Parets,1 Álvaro Otero-Rodríguez,2 Laura Ruiz Martín,2 Juan Carlos Paniagua Escudero,3 Luis Miguel Navarro Martín,4 Belén Cigarral García,4 Adelaida Nieto Palacios,5 Idalia González Morais,1 José María Sayagués,1 María Dolores Ludeña de la Cruz1 1Department of Pathology, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Salamanca, Spain; 2Department of Neurosurgery, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Salamanca, Spain; 3Department of Radiology, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Salamanca, Spain; 4Department of Clinical Oncology, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Salamanca, Spain; 5Department of Radiation Oncology, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Salamanca, SpainCorrespondence: Luis Miguel Chinchilla-Tábora, Department of Pathology, University Hospital of Salamanca and Institute for Biomedical Research of Salamanca, Paseo de San Vicente, 58-182, Salamanca, PC. 37007, Spain, Tel +34 923291100. Extension: 56738, Email [email protected]: Hemangioblastoma (HB) is a Central Nervous System (CNS) tumor with a generally favorable behavior and prognosis, classified as WHO grade 1. Sporadic HB is not related to any inherited disease, and it usually appears in a single location. Sporadic or VHL-related HBs show variable patterns of growth velocity. Cases of growing HB can cause mild symptoms such as headache, but some cases develop serious complications such as accumulation of cerebrospinal fluid in the brain with secondary neurological damage sometimes being irreversible when early treatment is not started. Our case showed some clinical characteristics more frequently observed in VHL-related HB rather than sporadic HB, and the presence of alterations in MDM2 and EGFR that could be related to the oncogenesis of these tumors. Even when the treatment of choice for HB is surgery, the presence of these genetic alterations could open a new window for research aimed at assessing the possibility of new therapies with TKIs-EGFR and anti-MDM2 inhibitors in those HB cases with multifocal recurrences or cases with an adverse clinical behavior.Keywords: haemangioblastoma, mouse double minute 2, epidermal growth factor receptor, hydrocephalus, immunohistochemistry, fluorescence in situ hybridizatio