Revistas
Autores:
Lopez-Borrego, S.; Campos-Silva, C.; Sandua, Amaia; et al.
Revista:
FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY
ISSN:
2296-634X
Año:
2023
Vol.:
10
Págs.:
1055288
Metastatic melanoma presents, in many cases, oncogenic mutations in BRAF, a MAPK involved in proliferation of tumour cells. BRAF inhibitors, used as therapy in patients with these mutations, often lead to tumour resistance and, thus, the use of MEK inhibitors was introduced in clinics. BRAFi/MEKi, a combination that has modestly increased overall survival in patients, has been proven to differentially affect immune ligands, such as NKG2D-ligands, in drug-sensitive vs. drug-resistant cells. However, the fact that NKG2D-ligands can be released as soluble molecules or in extracellular vesicles represents an additional level of complexity that has not been explored. Here we demonstrate that inhibition of MAPK using MEKi, and the combination of BRAFi with MEKi in vitro, modulates NKG2D-ligands in BRAF-mutant and WT melanoma cells, together with other NK activating ligands. These observations reinforce a role of the immune system in the generation of resistance to directed therapies and support the potential benefit of MAPK inhibition in combination with immunotherapies. Both soluble and EV-associated NKG2D-ligands, generally decreased in BRAF-mutant melanoma cell supernatants after MAPKi in vitro, replicating cell surface expression. Because potential NKG2D-ligand fluctuation during MAPKi treatment could have different consequences for the immune response, a pilot study to measure NKG2D-ligand variation in plasma or serum from metastatic melanoma patients, at different time points during MAPKi treatment, was performed. Not all NKG2D-ligands were equally detected. Further, EV detection did not parallel soluble protein. Altogether, our data confirm the heterogeneity between melanoma lesions, and suggest testing several NKG2D-ligands and other melanoma antigens in serum, both as soluble or vesicle-released proteins, to help classifying immune competence of patients.
Revista:
CLINICA CHIMICA ACTA
ISSN:
0009-8981
Año:
2023
Vol.:
543
Págs.:
117303
Aims: Characterization of PSA in extracellular microvesicles (EVs) and its reactivity to commercial methods.
Materials and methods: EVs derived from serum of 47 prostate cancer (PCa) patients, 27 benign prostatic hyperplasia (BPH) patients and 42 healthy controls were analyzed. EVs isolation and quantification of PSA immunoreactive to total (ev-T-PSA) or free (ev-F-PSA) PSA immunoassays, were performed using commercial assays. PSA in CD81+ or CD63+ EVs was determined directly in serum by an immunocapture-ELISA (IC-ELISA).
Results: Ev-T-PSA immunoreactive to Elecsys assay was detected in all samples. Median T-PSA ev/srm ratio was 2.20 % (Q1-Q3: 0.80-4.00 %), although in some samples this ratio reached 59 %. T-PSA ev/srm ratio was higher in those samples with serum T-PSA below 4 µg/L than in those exceeding that cut-off (p < 0.001). T-PSA ev/srm ratio was lower in PCa patients compared to healthy controls and BPH patients (p < 0.001). Elecsys immunoassays detected higher concentrations of ev-T-PSA and ev-F-PSA than Immulite (p < 0.001). PSA was detected by IC-ELISA more intensely in CD81+ EVs than in CD63+ EVs, and ev-T-PSA correlated with PSA+ CD63+ (p < 0.001) but not with PSA+ CD81+.
Conclusion: EVs-bound PSA is another form of circulating PSA whose measurement could be easily performed in clinical laboratories by automated immunoassays.
Autores:
Campos-Silva, C.; Cáceres-Martell, Y.; Sánchez-Herrero, E.; et al.
Revista:
JOURNAL OF NANOBIOTECHNOLOGY
ISSN:
1477-3155
Año:
2022
Vol.:
20
N°:
1
Págs.:
72
Background: Extracellular vesicles (EVs), released by most cell types, provide an excellent source of biomarkers in biological fluids. However, in order to perform validation studies and screenings of patient samples, it is still necessary to develop general techniques permitting rapid handling of small amounts of biological samples from large numbers of donors. Results: Here we describe a method that, using just a few microliters of patient's plasma, identifies tumour markers exposed on EVs. Studying physico-chemical properties of EVs in solution, we demonstrate that they behave as stable colloidal suspensions and therefore, in immunocapture assays, many of them are unable to interact with a stationary functionalised surface. Using flocculation methods, like those used to destabilize colloids, we demonstrate that cationic polymers increase EV zeta-potential, diameter, and sedimentation coefficient and thus, allow a more efficient capture on antibody-coated surfaces by both ELISA and bead-assisted flow cytometry. These findings led to optimization of a protocol in microtiter plates allowing effective immunocapture of EVs, directly in plasma without previous ultracentrifugation or other EV enrichment. The method, easily adaptable to any laboratory, has been validated using plasma from lung cancer patients in which the epithelial cell marker EpCAM has been detected on EVs. Conclusions: This optimized high throughput, easy to automate, technology allows screening of large numbers of patients to phenotype tumour markers in circulating EVs, breaking barriers for the validation of proposed EV biomarkers and the discovery of new ones.
Revista:
SCIENTIFIC REPORTS
ISSN:
2045-2322
Año:
2022
Vol.:
12
N°:
1
Págs.:
9752
Breath tests used to evaluate carbohydrates malabsorption require baseline H2 and CH4 levels as low as possible. Test cancellation is recommended when exceeding certain cut-offs (H2 ¿ 20 ppm and CH4 ¿ 10 ppm). Although following preparation protocols, many patients have baseline levels above those cut-offs. We investigated if light walking can reduce baseline H2 and CH4 levels. We retrospectively analyzed baseline H2 and CH4 levels from 1552 breath tests. Baseline levels (B1), especially in H2, were lower when obtained at later hours of the day. In those with baseline levels above cut-off, re-sampling (B2) after light walking for one hour, decreased H2 levels 8 ppm (Q1-Q3: 1-18 ppm), and 2 ppm (Q1-Q3: 0-3 ppm) for CH4. Consequently, 40% of tests with elevated B1 levels, presented B2 levels below mentioned cut-offs. Ten percent of tests considered negative when using B1 for calculations, turned positive when using B2 instead. All positive tests when using B1 values, remained elevated when using B2. Re-sampling after light walking for one hour could allow test performance in those with previous elevated baseline levels, avoiding diagnosis delays. Using the second sample for delta calculations identifies positive patients for malabsorption that would have been considered negative.
Revista:
ADVANCES IN LABORATORY MEDICINE / AVANCES EN MEDICINA DE LABORATORIO
ISSN:
2628-491X
Año:
2020
Vol.:
1
N°:
1
Págs.:
20190017
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
Nacionales y Regionales
Título:
GRANATE - GRUPO DE RADIOTERAPIA AVANZADA DE NAVARRA ¿ TERAPIA Y EFICACIA
Código de expediente:
0011-1411-2022-000066
Investigador principal:
Ana Patiño García
Financiador:
GOBIERNO DE NAVARRA
Convocatoria:
2022 GN PROYECTOS ESTRATEGICOS DE I+D 2022-2025
Fecha de inicio:
01/06/2022
Fecha fin:
31/12/2024
Importe concedido:
536.739,00€
Otros fondos:
-