González, Á. (Álvaro)

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    Espectrometría de masas en los laboratorios clínicos de proteínas
    (2024) Mugueta, C. (Carmen); González, Á. (Álvaro); Deza, S. (Sara); Sin Autoridad; Puig, N. (Noemí); Varo, N. (Nerea)
    Joseph John Thomson fue un ingeniero y matemático inglés descubridor del electrón, que recibió el Premio Nobel de Física en 1906, el mismo año en que Santiago Ramón y Cajal recibía el de Medicina. Thomson ya describió en 1899 un instrumento parecido a un espectrómetro de masas. Fueron sus discípulos, Aston y Dempster, de la Universidad de Chicago, quienes construyeron en la década siguiente los primeros espectrómetros de masas tal y como se conocen en la actualidad. Desde entonces, la tecnología ha avanzado de manera extraordinaria, primero con la introducción de instrumentos de tiempo de vuelo o cuadrupolo. El electrospray resolvió después el problema de la ionización de proteínas de gran tamaño y amplió el rango de análisis, previamente restringido a compuestos pequeños. En su conferencia por el Premio Nobel de Química en 2002, Fenn, se refirió a esto como dotar de “alas de electrospray a elefantes moleculares”. Estas mejoras y otras posteriores como el Matrix Assisted Laser Desportion/Ionization (MALDI) y la trampa iónica, han convertido a la espectrometría de masas (EM) en una herramienta analítica potente, versátil, precisa y sensible cuyo uso se ha extendido a ámbitos muy diferentes, hasta finalmente llamar también a las puertas del Laboratorio Clínico. Hasta ahora, su uso en rutina en los laboratorios clínicos se ha restringido al análisis de fármacos, hormonas esteroideas y otros metabolitos. Sin embargo, por sus características, el espectro de potenciales aplicaciones de la EM es muy amplio. De hecho, en los últimos años, su uso se ha extendido al análisis de moléculas más grandes como las proteínas, incluyendo la inmunoglobulina monoclonal empleada como biomarcador para el diagnóstico y seguimiento de las gammapatías monoclonales (GM).
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    Study of circulating MicroRNA-125b levels in serum exosomes in advanced melanoma
    (College of american pathologists, 2014) Alegre, E. (Estibaliz); Martin-Algarra, S. (Salvador); Carranza, O. (Omar); Rodríguez, C. (Carmen); González, Á. (Álvaro); Fernandez-Sanmamed, M. (Miguel)
    Context: Malignant melanoma is an aggressive tumor that produces exosomes, which contain microRNAs (miRNAs) that could be of utility in following tumoral cell dysregulation. MicroR-125b is a miRNA whose down-regulation seems to be implicated in melanoma progression. Objective: To analyze miR-125b levels in serum, and in exosomes obtained from serum, from patients with advanced melanoma. Design: Serum samples were obtained from 21 patients with advanced melanoma, from 16 disease-free patients with melanoma, and from 19 healthy volunteers. Exosomes were isolated from serum by precipitation, and miR-16 and miR-125b levels were quantified by real-time polymerase chain reaction. Results: MicroR-16, but not miR-125b, was detected in all samples, and miR-16 levels were significantly higher in serum than they were in exosomes. MicroR-16 expression levels did not differ significantly between the 2 groups (patients with melanoma and healthy donors). There was a significant relationship between miR-125b and miR-16 levels in exosomes. Additionally, miR-125b levels in exosomes were significantly lower in patients with melanoma compared with disease-free patients with melanoma and healthy controls. Conclusions: Exosomes can provide a suitable material to measure circulating miRNA in melanoma, and miR-16 can be used as an endogenous normalizer. Lower levels of miR-125b in exosomes obtained from serum are associated with advanced melanoma disease, probably reflecting the tumoral cell dysregulation.
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    Serum interleukin-8 reflects tumor burden and treatment response across malignancies of multiple tissue origins
    (AACR, 2014) Mazzolini, G. (Guillermo); Alfaro, C. (Carlos); Rizzo, M. (Manglio); Perez-Gracia, J.L. (Jose Luis); Munoz-Calleja, C. (Cecilia); Martin-Algarra, S. (Salvador); Rodriguez, I. (Inmaculada); Rodriguez-Ruiz, M.E. (María Esperanza); Carranza, O. (Omar); Fernández-Landázuri, S. (Sara); Lopez-Picazo, J.M. (José M.); Sangro, B. (Bruno); Oñate, C. (Carmen); Andueza, M.P. (Maria P.); Melero, I. (Ignacio); Gross, S. (Stefanie); Perez, G. (Guiomar); González, Á. (Álvaro); Pascual, J.I. (Juan Ignacio); Fernandez-Sanmamed, M. (Miguel)
    Purpose: Interleukin-8 (IL8) is a chemokine produced by malignant cells of multiple cancer types. It exerts various functions in shaping protumoral vascularization and inflammation/immunity. We evaluated sequential levels of serum IL8 in preclinical tumor models and in patients to assess its ability to estimate tumor burden. Experimental Design:IL8levels were monitored by sandwich ELISAsin cultured tumor cells supernatants, tumor-xenografted mice serum, and in samples from 126 patients with cancer. We correlated IL8 serum levels with baseline tumor burden and with treatment-induced changes in tumor burden, as well as with prognosis. Results: IL8 concentrations correlated with the number of IL8-producing tumor cells in culture. In xenografted neoplasms, IL8 serum levels rapidly dropped after surgical excision, indicating an accurate correlation with tumor burden. In patients with melanoma (n ¼ 16), renal cell carcinoma (RCC; n ¼ 23), non–small cell lung cancer (NSCLC; n ¼ 21), or hepatocellular carcinoma (HCC; n ¼ 30), serum IL8 concentrations correlated with tumor burden and stage, survival (melanoma, n ¼ 16; RCC, n ¼ 23; HCC, n ¼ 33), and objective responses to therapy, including those to BRAF inhibitors (melanoma, n ¼ 16) and immunomodulatory monoclonal antibodies (melanoma, n ¼ 8). IL8 concentrations in urine (n ¼ 18) were mainly elevated in tumors with direct contact with the urinary tract. Conclusions: IL8 levels correlate with tumor burden in preclinical models and in patients with cancer. IL8 is a potentially useful biomarker to monitor changes in tumor burden following anticancer therapy, and has prognostic significance.
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    Genomic characterization of individuals presenting extreme phenotypes of high and low risk to develop tobacco-induced lung cancer
    (2018) Pajares, M.J. (María José); Gil-Bazo, I. (Ignacio); Patiño-García, A. (Ana); Pio, R. (Rubén); Lozano, M.D. (María Dolores); Alonso, R. (Rosario); Casanova, C. (Ciro); Perez-Gracia, J.L. (Jose Luis); Benitez, J. (Javier); Rodriguez-Ruiz, M.E. (María Esperanza); Agudo, A. (Antonio); Baz-Dávila, R. (Rebeca); Bou-i-Sala, N. (Núria); Lopez-Picazo, J.M. (José M.); Fusco, J.P. (Juan Pablo); Torres, J.P. (Juan P.) de; Gurpide, A. (Alfonso); Andueza, M.P. (Maria P.); Montuenga-Badia, L.M. (Luis M.); Melero, I. (Ignacio); Ardanaz, E. (Eva); González, Á. (Álvaro); Gonzalez-Neira, A. (Anna); Alvarez, N. (Nuria); Fernandez-Sanmamed, M. (Miguel); Zulueta, J. (Javier); Pita, G. (Guillermo)
    Single nucleotide polymorphisms (SNPs) may modulate individual susceptibility to carcinogens. We designed a genome-wide association study to characterize individuals presenting extreme phenotypes of high and low risk to develop tobacco-induced non-small cell lung cancer (NSCLC), and we validated our results. We hypothesized that this strategy would enrich the frequencies of the alleles that contribute to the observed traits. We genotyped 2.37 million SNPs in 95 extreme phenotype individuals, that is: heavy smokers that either developed NSCLC at an early age (extreme cases); or did not present NSCLC at an advanced age (extreme controls), selected from a discovery set (n=3631). We validated significant SNPs in 133 additional subjects with extreme phenotypes selected from databases including >39,000 individuals. Two SNPs were validated: rs12660420 (p(combined)=5.66x10(-5); ORcombined=2.80), mapping to a noncoding transcript exon of PDE10A; and rs6835978 (p(combined)=1.02x10(-4); ORcombined=2.57), an intronic variant in ATP10D. We assessed the relevance of both proteins in early-stage NSCLC. PDE10A and ATP10D mRNA expressions correlated with survival in 821 stage I-II NSCLC patients (p=0.01 and p<0.0001). PDE10A protein expression correlated with survival in 149 patients with stage I-II NSCLC (p=0.002). In conclusion, we validated two variants associated with extreme phenotypes of high and low risk of developing tobacco-induced NSCLC. Our findings may allow to identify individuals presenting high and low risk to develop tobacco-induced NSCLC and to characterize molecular mechanisms of carcinogenesis and resistance to develop NSCLC.
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    Relevance of MIA and S100 serum tumor markers to monitor BRAF inhibitor therapy in metastatic melanoma patients
    (Elsevier BV, 2014) Alegre, E. (Estibaliz); Lozano, M.D. (María Dolores); Perez-Gracia, J.L. (Jose Luis); Martin-Algarra, S. (Salvador); Carranza, O. (Omar); Fernández-Landázuri, S. (Sara); Rodríguez, C. (Carmen); Echeveste, J.I. (José I.); Zubiri, L. (Leire); González, Á. (Álvaro); Fernandez-Sanmamed, M. (Miguel)
    BRAF V600 mutation has been reported in more than 50% of melanoma cases and its presence predicts clinical activity of BRAF inhibitors (iBRAF). We evaluated the role of MIA, S100 and LDH to monitor iBRAF efficiency in advanced melanoma patients presenting BRAF V600 mutations. This was a prospective study of melanoma patients harboring the BRAF V600 mutation and treated with iBRAF within a clinical trial (dabrafenib) or as part of an expanded access program (vemurafenib). MIA, S100 and LDH were analyzed in serum at baseline, and every 4–6 weeks during treatment. Eighteen patients with melanoma stages IIIc–IV were enrolled with 88.8% of response rate to iBRAF. Baseline concentrations of all the tumor markers correlated with tumor burden. MIA and S100 concentrations decreased significantly one month after the beginning of treatment and, upon progression, their concentrations increased significantly above the minimum levels previously achieved. MIA levels lower than 9 μg/L one month after the beginning of treatment and S100 concentrations lower than 0.1 μg/L at the moment of best response were associated with improved progression-free survival. In conclusion, MIA and S100 are useful to monitor response in melanoma patients treated with iBRAF
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    Utility of recombinant human TSH stimulation test in the follow-up of patients with differentiated thyroid cancer depending on basal thyroglobulin results
    (De Gruyter, 2020) Alegre-Martinez, E. (Estibaliz); Perdomo-Zelaya, C.M. (Carolina M.); Sandúa-Condado, A. (Amaia); Ferrer, R. (Roser); Macías, M. (Mónica); González, Á. (Álvaro); Galofre, J.C. (Juan Carlos)
    Background: Thyroglobulin (Tg) is fundamental for differentiated thyroid cancer (DTC) monitoring. Tg detection can be enhanced using recombinant human thyroidstimulating hormone (TSH) (rhTSH). This study is aimed to evaluate the use of the rhTSH stimulation test when using a high-sensitivity Tg assay. Methods: We retrospectively studied 181 rhTSH tests from 114 patients with DTC and negative for antithyroglobulin antibodies (anti-TgAb). Image studies were performed in all cases. Serum Tg and anti-TgAb were measured using specific immunoassays. Results: rhTSH stimulation in patients with basal serum Tg (b-Tg) concentrations lower than 0.2 ng/mL always resulted in rhTSH-stimulated serum Tg (s-Tg) concentrations lower than 1.0 ng/mL and negative structural disease. In patients with bTg concentration between 0.2 and 1.0 ng/mL, s-Tg detected one patient (1/30) who showed biochemical incomplete response. Patients with negative images had lower s-Tg than thosewith nonspecific or abnormal findings (p<0.05).Receiver operating characteristic curve analysis of the s-Tg to detect altered images showed an area under the curve of 0.763 (p<0.05).With an s-Tg cutoff of 0.85 ng/mL, the sensitivity was 100%, decreasing to 96.15% with an s-Tg cutoff of 2 ng/mL. Conclusions: Patients with DTC with b-Tg concentrations equal or higher than 0.2 ng/mL can benefit from the rhTSH stimulation test.
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    A model based on the quantification of complement C4c, CYFRA 21-1 and CRP exhibits high specificity for the early diagnosis of lung cancer
    (Elsevier, 2021) Pio, R. (Rubén); Sainz, C. (Cristina); Lozano, M.D. (María Dolores); Remirez, A. (Ana); Perez-Gracia, J.L. (Jose Luis); Perez-Palacios, R. (Rosa); Massion, P.P. (Pierre P.); Montuenga-Badia, L.M. (Luis M.); Bertolo, C. (Cristina); Martin, A.C. (Ana C.); Mesa-Guzmán, M.A. (Miguel Alejandro); González, Á. (Álvaro); Ajona, D. (Daniel); Zulueta, J. (Javier); Varo-Cenarruzabeitia, M.N. (Miren Nerea)
    Lung cancer screening detects early-stage cancers, but also a large number of benign nodules. Molecular markers can help in the lung cancer screening process by refining inclusion criteria or guiding the management of indeterminate pulmonary nodules. In this study, we developed a diagnostic model based on the quantification in plasma of complement-derived fragment C4c, cytokeratin fragment 21-1 (CYFRA 21-1) and C-reactive protein (CRP). The model was first validated in two independent cohorts, and showed a good diagnostic performance across a range of lung tumor types, emphasizing its high specificity and positive predictive value. We next tested its utility in two clinically relevant contexts: assessment of lung cancer risk and nodule malignancy. The scores derived from the model were associated with a significantly higher risk of having lung cancer in asymptomatic individuals enrolled in a computed tomography (CT)-screening program (OR = 1.89; 95% CI = 1.20–2.97). Our model also served to discriminate between benign and malignant pulmonary nodules (AUC: 0.86; 95% CI = 0.80–0.92) with very good specificity (92%). Moreover, the model performed better in combination with clinical factors, and may be used to reclassify patients with intermediate-risk indeterminate pulmonary nodules into patients who require a more aggressive work-up. In conclusion, we propose a new diagnostic biomarker panel that may dictate which incidental or screening-detected pulmonary nodules require a more active work-up.
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    The dynamic use of EGFR mutation analysis in cell-free DNA as a follow-up biomarker during different treatment lines in non-small-cell lung cancer patients
    (Hindawi Limited, 2019) Gil-Bazo, I. (Ignacio); Alegre-Martinez, E. (Estibaliz); Patiño-García, A. (Ana); Perez-Gracia, J.L. (Jose Luis); Alkorta-Aranburu, G. (Gorka); Lopez-Picazo, J.M. (José M.); Gurpide, A. (Alfonso); Andueza, M.P. (Maria P.); Mateos, B. (Beatriz); Macías, M. (Mónica); González, Á. (Álvaro)
    Epidermal growth factor receptor (EGFR) mutational testing in advanced non-small-cell lung cancer (NSCLC) is usually performed in tumor tissue, although cfDNA (cell-free DNA) could be an alternative. We evaluated EGFR mutations in cfDNA as a complementary tool in patients, who had already known EGFR mutations in tumor tissue and were treated with either EGFR-tyrosine kinase inhibitors (TKIs) or chemotherapy. We obtained plasma samples from 21 advanced NSCLC patients with known EGFR tumor mutations, before and during therapy with EGFR-TKIs and/or chemotherapy. cfDNA was isolated and EGFR mutations were analyzed with the multiple targeted cobas EGFR Mutation Test v2. EGFR mutations were detected at baseline in cfDNA from 57% of patients. The semiquantitative index (SQI) significantly decreased from the baseline (median = 11, IQR = 9 5-13) to the best response (median = 0, IQR = 0-0, p < 0 01), followed by a significant increase at progression (median = 11, IQR = 11-15, p < 0 01) in patients treated with either EGFR-TKIs or chemotherapy. The SQI obtained with the cobas EGFR Mutation Test v2 did not correlate with the concentration in copies/mL determined by droplet digital PCR. Resistance mutation p.T790M was observed at progression in patients with either type of treatment. In conclusion, cfDNA multiple targeted EGFR mutation analysis is useful for treatment monitoring in tissue of EGFR-positive NSCLC patients independently of the drug received.
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    Evaluation of multiple serum markers in advanced melanoma
    (Springer, 2011) Alegre-Martinez, E. (Estibaliz); Gonzalez-Cao, M. (María); Martin-Algarra, S. (Salvador); Arroyo, A. (Ainhoa); Zudaire, M.E. (Maria E.); González, Á. (Álvaro); Viteri, S. (S.); Diaz-Lagares, A. (Ángel)
    The aim of this retrospective study was to analyse in advanced melanoma the potential tumor markers S-100B, melanoma inhibiting activity protein (MIA) and YKL-40 compared to LDH. Serum levels of S-100B, MIA, LDH and YKL-40 were measured in 110 patients with advanced melanoma (36 in stage IIIB/C and 74 in stage IV), in 66 disease-free patients and in 65 healthy controls. Results show that S-100B, MIA and LDH levels were significantly higher in patients with advanced melanoma than in disease-free patients or healthy controls. The combination of S-100B plus MIA had the best diagnostic sensitivity, and the addition of LDH did not further increase this sensitivity. MIA was an independent prognostic factor of overall survival. Patients with both S-100B and MIA elevated had a significant shorter survival than those with both S-100B and MIA under the cut-off. YKL-40 levels did not differentiate patients with advanced melanoma from controls. We concluded that the combination of MIA plus S-100B showed a better prognostic value in advanced melanoma compared to LDH.