Technical Evaluation of the COBAS EGFR Semiquantitative Index (SQI) for Plasma cfDNA Testing in NSCLC Patients with EGFR Exon 19 Deletions.

The cobas® EGFR Test provides a semiquantitative index (SQI) that reflects the proportion of mutated versus wild-type copies of the EGFR gene in plasma. The significance of SQI as an indirect measure of the variant allele frequency (VAF) or mutated copies/mL remains unclear. The aim of this study was to evaluate the correlation of SQI with the VAF and the number of mutated copies/mL obtained by a digital droplet PCR (ddPCR) test in NSCLC samples. The study included 118 plasma samples from a retrospective cohort of 25 stage IV adenocarcinoma patients with EGFR exon 19 deletions (Ex19Del), obtained before and during tyrosine kinase inhibitor (TKI) treatment. Both SQI and VAF and SQI and mutated copies/mL showed the same significant correlation (r2 = 0.79, p < 0.00001) across the whole study cohort. We found better correlation in samples collected at the baseline between SQI and VAF (r2 = 0.94, p < 0.00001) and SQI and mutated copies/mL (r2 = 0.97, p < 0.00001) compared to samples collected during TKI treatment: r2 = 0.76; p < 0.00001 for SQI and VAF and r2 = 0.75; p < 0.00001 for SQI and mutated copies/mL. The study indicates that SQI is a robust quantitative indirect measure of VAF and the number of mutated copies/mL in plasma from patients with an EGFR Ex19Del mutation. Further studies are desirable to assess the SQI cut-off values related to the clinical status of the patient.

Newborn Screening for SCID. Experience in Spain (Catalonia).

Newborn screening (NBS) for severe combined immunodeficiency (SCID) started in Catalonia in January-2017, being the first Spanish and European region to universally include this testing. In Spain, a pilot study with 5000 samples was carried out in Seville in 2014; also, a research project with about 35,000 newborns will be carried out in 2021-2022 in the NBS laboratory of Eastern Andalusia. At present, the inclusion of SCID is being evaluated in Spain. The results obtained in the first three and a half years of experience in Catalonia are presented here. All babies born between January-2017 and June-2020 were screened through TREC-quantification in DBS with the Enlite Neonatal TREC-kit from PerkinElmer. A total of 222,857 newborns were screened, of which 48 tested positive. During the study period, three patients were diagnosed with SCID: an incidence of 1 in 74,187 newborns; 17 patients had clinically significant T-cell lymphopenia (non-SCID) with an incidence of 1 in 13,109 newborns who also benefited from the NBS program. The results obtained provide further evidence of the benefits of early diagnosis and curative treatment to justify the inclusion of this disease in NBS programs. A national NBS program is needed, also to define the exact SCID incidence in Spain.

Molecular characterization of advanced non-small cell lung cancer patients by cfDNA analysis: experience from routine laboratory practice.

BACKGROUND: Analysis of circulating free DNA (cfDNA) by the real-time PCR cobas® EGFR Mutation Test v2 (cobas® EGFR Test) is a diagnostic approach used in clinical practice for the characterization of advanced non-small cell lung cancer (NSCLC) patients. The test additionally outputs a semiquantitative index (SQI) which reflects the proportion of mutated versus wild-type copies of the EGFR gene in cfDNA with potential use as a biomarker. CfDNA concentration and cfDNA fragmentation pattern have also shown potential utility as biomarkers for cancer patients. We evaluated the implementation of EGFR testing and cfDNA related parameters in NSCLC patients in routine clinical setting as biomarkers for disease stage and diagnosis. METHODS: A prospective cohort of 173 locally advanced or metastatic NSCLC TKI-naïve patients analyzed by the cobas® EGFR Test were included in the study. Reproducibility of the test was assessed in 56 patients. The concentration of cfDNA and fragment size pattern was measured using fluorometry and microchip electrophoresis respectively. RESULTS: The test showed high diagnostic accuracy when compared to the gold standard of biopsy tumor tissue testing. The SQI value showed a moderate reproducibility (r2=0.70) and did not correlate with cfDNA concentration (r2=0.17, P=0.28) or disease stage (stage III patients SQI =9.1±3.1 and stage IV patients SQI =11.5±4.8, P=0.41). We found differences in SQI values according to the type of EGFR mutation (Ex19Del mutations, SQI =13.6; p.L858R, SQI =8.88; P=0.001). Stage IV patients had higher concentrations of cfDNA (P<0.0001) and higher fractions of cfDNA 100-250 base pairs (bp) fragments (P=0.01) compared to stage III patients. From the ROC curve analysis, cfDNA concentration showed higher AUC compared to cfDNA 100-250 bp fragments (0.86 vs. 0.71). We obtained a cut-off value for cfDNA concentration of 20.3 ng/mL with 72.3% sensitivity and 95% specificity for predicting disease stage in TKI-naïve advanced NSCLC patients. CONCLUSIONS: The study indicates that cfDNA analysis in plasma for EGFR testing by RT-PCR is an accurate and fast method to initially stratify NSCLC patients in a real-world clinical setting. However, the SQI has limited clinical value. The cfDNA concentration and fragmentation pattern have clear potential clinical utility for tumor staging in NSCLC patients.

[Response of the Catalonia neonatal screening laboratory to the pandemic of SARS-CoV-2.] / Respuesta del Laboratorio de Cribado Neonatal de Cataluña ante la pandemia por SARS-CoV-2.

Faced with the prospect of a collapsed health system due to the COVID-19 pandemic, the professionals involved in the Neonatal Screening Programme (NSP) of Catalonia had to adapt to this situation in a flexible, forceful and efficient manner. The most important goals were to prevent the risk of infection in the professionals, in families and their newborns, as well as to ensure the same effectiveness for the early detection of the diseases included in our programme. To this end, the laboratory was reorganised by dividing the staff into groups and the spaces were redistributed. It was also necessary to modify several protocols and circuits, especially for the management of early discharges from maternity centres, and for the collection of the necessary second samples (from newborns with inconclusive results or for low quality samples). In general, a 36% reduction in the time of arrival of these second samples at the laboratory was achieved with respect to the previous circuit. In the specific case of cystic fibrosis detection, the implementation of a new strategy meant a 100% reduction in the request for second samples and a 70% reduction in the age of diagnosis of the newborn. After evaluating these changes, it can be concluded that in the face of the pandemic, the NSP of Catalonia showed determined leadership, aligning all its professionals, ensuring the continuity of the activity in the programme and generating new opportunities. The new processes and circuits implemented have been definitively consolidated, improving the efficiency of the programme.

Ante la crisis de un sistema sanitario colapsado debido a la pandemia por la COVID-19, los profesionales implicados en el Programa de Cribado Neonatal (PCN) de Cataluña nos tuvimos que adaptar a dicha situación de forma ágil, contundente y eficiente. Los objetivos prioritarios fueron prevenir el riesgo de contagio tanto en los profesionales sanitarios como en las familias y sus recién nacidos, así como asegurar la misma eficacia para la detección precoz de las enfermedades incluidas en el PCN. Para ello, se reorganizó el laboratorio dividiendo en grupos al personal y se redistribuyeron los espacios. También fue necesario modificar varios protocolos y circuitos, en especial para la gestión de las altas precoces de los centros maternales y para la toma de las segundas muestras necesarias (de recién nacidos que presentaron resultados dudosos o por muestra inválida). En general, se consiguió una reducción del 36% del tiempo de llegada de estas segundas muestras al laboratorio respecto al circuito anterior. Para la detección de la fibrosis quística, la implementación de una nueva estrategia supuso una reducción del 100% en la solicitud de segundas muestras y del 70% en la edad de diagnóstico del recién nacido. Tras la evaluación de estos cambios, se puede concluir que ante la pandemia el PCN de Cataluña mostró un liderazgo decidido, alineando a todos sus profesionales, asegurando la continuidad de la actividad en el programa y generando nuevas oportunidades. Los nuevos procesos y circuitos de trabajo implantados han quedado definitivamente consolidados, mejorando la eficiencia del programa.

[Impact of the inclusion of second-tier tests in the newborn screening program of Catalonia and in other international programs.] / Impacto de la inclusión de pruebas de segundo nivel en el programa de cribado neonatal de Cataluña y en otros programas internacionales.

OBJECTIVE: Newborn screening programmes (NBSP) have experienced a qualitative breakthrough due to the implementation of tandem mass spectrometry. However, the tests used give rise to false positives (FP) generating an excessive request for second samples with the consequent anxiety of the families. In order to avoid this problem several programmes have developed second-tier tests (2TT). METHODS: This article presents our experience in the implementation of 2TT in the NBSP of Catalonia, as well as in other international programmes. RESULTS: From 2004 to the present, 2TT tests have been developed for more than 30 diseases. The use of 2TT helps to decrease the FP rate and increase the positive predictive value (PPV). In the NBSP of Catalonia, the implementation of 2TT for the detection of methylmalonic and propionic acidemias, homocystinurias, maple syrup disease and citrulinaemia, has managed to increase the PPV to 95% and decrease the PF rate to less than 0.01%. In cystic fibrosis, the application of 2TT slightly increases PPV but with a significant decrease in the request for second samples and in the number of cases referred to clinical units. CONCLUSIONS: The introduction of 2TT in the NBSP allows to reduce considerably the FP, decreases the number of requested samples, as well as both anxiety and stress of the families, at the same time that the hospital costs are reduced and the PPV is increased, improving notably the efficiency of the NBSP.

OBJETIVO: Los programas de cribado neonatal (PCN) han experimentado un gran avance cualitativo debido a la implementación de la espectrometría de masas en tándem. Sin embargo, las pruebas utilizadas dan lugar a falsos positivos (FP) generando una excesiva solicitud de segundas muestras con la consiguiente ansiedad de las familias. Con el fin de evitar este problema diversos programas han desarrollado pruebas de segundo nivel (2TT). METODOS: En este artículo se presenta nuestra experiencia en la implementación de 2TT en el PCN de Cataluña, así como en otros programas internacionales. RESULTADOS: Desde el año 2004 hasta la actualidad se han desarrollado pruebas de 2TT para más de 30 enfermedades. La utilización de 2TT ayuda a disminuir la tasa de FP y aumentar el valor predictivo positivo (VPP). En el PCN de Cataluña, la implementación de 2TT para la detección de acidemias metilmalónicas y propiónica, homocistinurias, jarabe de arce y citrulinemia, ha conseguido aumentar el VPP a un 95% y disminuir la tasa de FP a menos del 0,01%. En la fibrosis quística la aplicación de 2TT aumenta ligeramente el VPP pero con disminución significativa de la solicitud de segundas muestras y de los casos referidos a las unidades clínicas. CONCLUSIONES: La introducción de los 2TT en los PCN permite reducir considerablemente los FP, disminuye el número de muestras solicitadas, así como la ansiedad y el estrés de las familias, a la vez que se reducen los costes hospitalarios y se aumenta el VPP, mejorando notablemente la eficiencia de los PCN.