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Melanocytic neoplasms with spitzoid histomorphology are often difficult to classify without identifying genetic drivers such as kinase fusions. Traditional diagnostic methods, such as immunohistochemistry, can yield inconclusive results, and advanced techniques such as the Archer fusion assay are often inaccessible and costly. The Idylla GeneFusion Assay might offer a rapid and cost-effective alternative. This study compared Idylla and Archer in identifying ALK, pan-NTRK, RET, and ROS1 gene fusions. Of the 147 samples where next-generation sequencing did not detect genetic drivers, 89 (60.5%) meeting the tissue requirements were further analyzed using Idylla (Cohort A). Idylla demonstrated a sensitivity of 75% and a specificity of 100% in detecting these fusions. Additionally, among 27 randomly selected cases (Cohort B) that failed to meet the inclusion criteria, Idylla maintained the same levels of sensitivity and specificity. Our findings also show that Idylla can be effectively conducted with isolated RNA, broadening its applicability beyond tissue samples. Although the Idylla assay may not replace more comprehensive molecular assays such as Archer, it could serve as a valuable initial screening tool in diagnosing spitzoid melanocytic tumors.
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CONTEXT: DNA mismatch repair (MMR) deficiency (dMMR) testing is now recommended in endometrial cancer. Defect identification in the molecules participating in this pathway, or the presence of microsatellite instability, are commonly employed for this purpose. Novel methods are continuously evolving to report dMMR/microsatellite instability and to easily perform routine diagnoses. OBJECTIVE: The main aim of this study was to compare the concordance of the Idylla microsatellite instability test for the identification of dMMR endometrial cancer samples defined by immunohistochemistry and MMR genomic status. DESIGN: We applied the Idylla MSI test to 126 early-stage endometrial cancer cases with MMR testing by immunohistochemistry and genomic characterization (methylation in MLH1 and sequence alterations in MLH1, PMS2, MSH2 and MSH6). Individual markers and overall specific performance indicators were explored. RESULTS: The Idylla platform achieved a higher global concordance rate with MMR genomic status than with immunohistochemistry (75 % and 66 %, respectively). Sensitivity and specificity are also higher (75 % vs 66 % and 96 % vs 90 %, respectively). Clustering analysis split the patients into 2 well-differentiated clusters, the pMMR and the dMMR group, represented by MLH1/PMS2 loss and the MLH1 methylated promoter. Overall, immunohistochemistry and MMR genomic status identified more dMMR cases than did the Idylla test, although correlations were improved with a modified Idylla test cut-off. CONCLUSIONS: Performance of the Idylla test was better correlated with MMR genomic status than MMR immunohistochemistry status, which improved with a modified test cut-off. Further studies are needed to confirm the cut-off accuracy.
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Malignant pleural effusion (MPE) from patients with advanced non-small-cell lung cancer (NSCLC) has been proven valuable for molecular analysis; however, simultaneous detection of driver fusions in MPE is still challenging. In this study, we investigated the Idylla™ GeneFusion Panel, a stand-alone test in tissue samples, in the evaluation of ALK, ROS1, RET and MET ex14 skipping mutations in MPE and compared its performance with routine reference methods (Real-time-based and Next-generation Sequencing-NGS). The inclusion criteria for sample selection were as follows: advanced NSCLC harboring ALK, ROS1, RET fusions or MET exon-skipping alterations and the availability of MPE collected at diagnosis or disease progression. Molecular alterations have been investigated on tissue by fluorescence in situ hybridization (FISH) or Real-time PCR or NGS. For molecular profiling with the Idylla™ GeneFusion, 200 µL of MPE supernatants combined with 50 µL of RNA Later solution were loaded into the Idylla™ cartridge without cfRNA extraction. The Idylla™ GeneFusion Assay performed on MPEs was able to confirm molecular profile, previously diagnosed with conventional methods, in all cases. Our data confirm that MPE are suitable material for investigating fusion alterations. The Idylla™ GeneFusion, although indicated for investigation of tissue samples, offers the possibility of performing a molecular characterization of supernatants without undertaking the entire cfRNA extraction procedure providing a rapid and reliable strategy for the detection of actionable genetic alterations.
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Carcinoma Pulmonar de Células não Pequenas , Sequenciamento de Nucleotídeos em Larga Escala , Neoplasias Pulmonares , Reação em Cadeia da Polimerase em Tempo Real , Humanos , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Projetos Piloto , Masculino , Feminino , Idoso , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real/métodos , Derrame Pleural Maligno/genética , Derrame Pleural Maligno/patologia , Derrame Pleural Maligno/diagnóstico , Proteínas de Fusão Oncogênica/genética , Fusão Gênica , Adulto , Mutação , Quinase do Linfoma Anaplásico/genética , Idoso de 80 Anos ou mais , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Tirosina Quinases , Proteínas Proto-OncogênicasRESUMO
INTRODUCTION: Microsatellite instability (MSI) is an important prognostic molecular biomarker for gastric cancer (GC). MSI status may be detected by immunohistochemistry (IHC) for mismatch repair (MMR) proteins and polymerase chain reaction (PCR). Idylla™ MSI assay has not been validated for GC but may prove to be a valid alternative. METHODS: In a series of 140 GC cases, MSI status was evaluated by IHC for MLH1, PMS2, MSH2, and MSH6; gold-standard pentaplex PCR panel (PPP) (BAT-25, BAT-26, NR-21, NR-24, and NR-27); and Idylla. Statistical analysis was performed using SPSS 27.0. RESULTS: PPP identified 102 microsatellite stable (MSS) cases and 38 MSI-high cases. Only 3 cases showed discordant results. Compared with PPP, the sensitivity was 100% for IHC and 94.7% for Idylla. Specificity was 99% for IHC and 100% for Idylla. MLH1 IHC alone showed sensitivity and specificity of 97.4% and 98.0%, respectively. IHC identified three indeterminate cases; all were MSS according to PPP and Idylla. CONCLUSION: IHC for MMR proteins represents an optimal screening tool for MSI status in GC. If resources are limited, isolated MLH1 evaluation may constitute a valuable option for preliminary screening. Idylla may help detect rare MSS cases with MMR-loss and define MSI status in indeterminate cases.
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Neoplasias Colorretais , Neoplasias Gástricas , Humanos , Instabilidade de Microssatélites , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/genética , Biomarcadores Tumorais/análise , Imuno-Histoquímica , Neoplasias Colorretais/genética , Repetições de MicrossatélitesRESUMO
Targeting EGFR alterations, particularly the L858R (Exon 21) mutation and Exon 19 deletion (del19), has significantly improved the survival of lung cancer patients. From now on, the issue is to shorten the time to treatment. Here, we challenge two well-known rapid strategies for EGFR testing: the cartridge-based platform Idylla™ (Biocartis) and a digital droplet PCR (ddPCR) approach (ID_Solution). To thoroughly investigate each testing performance, we selected a highly comprehensive cohort of 39 unique del19 (in comparison, the cbioportal contains 40 unique del19), and 9 samples bearing unique polymorphisms in exon 19. Additional L858R (N = 24), L861Q (N = 1), del19 (N = 63), and WT samples (N = 34) were used to determine clear technical and biological cutoffs. A total of 122 DNA samples extracted from formaldehyde-fixed samples was used as input. No false positive results were reported for either of the technologies, as long as careful droplet selection (ddPCR) was ensured for two polymorphisms. ddPCR demonstrated higher sensitivity in detecting unique del19 (92.3%, 36/39) compared to Idylla (67.7%, 21/31). However, considering the prevalence of del19 and L858R in the lung cancer population, the adjusted theranostic values were similar (96.51% and 95.26%, respectively). ddPCR performs better for small specimens and low tumoral content, but in other situations, Idylla is an alternative (especially if a molecular platform is absent).
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/genética , Receptores ErbB/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Técnicas de Diagnóstico Molecular , Mutação , Reação em Cadeia da Polimerase/métodos , Medicina de PrecisãoRESUMO
Recent diagnostic and therapeutic progresses have increased the need of searching for microsatellite instability (MSI) in cancer samples beyond colorectal cancer (CRC) ones. The availability of the fully-automated Idylla MSI test (Biocartis), implementable easily in pathology laboratories, offers the opportunity to reconsider MSI diagnostic strategies towards rapid and in-house diagnosis. In this study, we evaluate the performances and cost-effectiveness of an in-house Idylla MSI testing in comparison with an externalized testing of about 54 non-CRC tumor samples. The Idylla MSI test concluded in valid analyses in 53/54 (98.1%) tumor samples with MSI statuses concordant with external molecular and immunohistochemical testing in 50/53 (94.3%) samples. Wrong Idylla MSI test results were obtained in 3/53 (5.7%) samples. Manual checking of microsatellite analyses results and confrontation between the results of Idylla and immunohistochemical analyses have permitted detection and correction of the discrepancies. The implementation of an in-house Idylla MSI testing for non-CRC tumors, necessarily combined with immunohistochemistry searching for MSI tumors, appeared not only valuable in terms of performances, but also in terms of cost-effectiveness without increasing the analyses-related costs but decreasing dramatically their turnaround times to one single working day.
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Neoplasias Colorretais , Neoplasias , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Análise Custo-Benefício , Humanos , Imuno-Histoquímica , Instabilidade de Microssatélites , Repetições de MicrossatélitesRESUMO
OBJECTIVE: KRAS is a frequently mutated gene in cancers, and with recent FDA-approved targeted therapy for the G12C mutation, testing for KRAS variants is essential. We evaluated the performance of the Idylla KRAS assay on extracted DNA and cytology smears in order to expand the utility of the assay. METHODS: In total, fifty-seven human samples were analyzed. Idylla results from sixteen DNA extracted from formalin-fixed, paraffin-embedded tissues (FFPE DNA) and thirty cytology smears were compared to the reference method. We evaluated the performance of the Idylla assay using corresponding cytology smears to rescue cellblocks or surgical blocks that were quantity not sufficient (QNS) for next generation sequencing (NGS). RESULT: In the FFPE DNA cohort, 10 ng DNA input yielded valid results in all 16 samples, with 15 of 16 (93 %) concordant with NGS findings. In the cytology smear cohort, the Idylla KRAS assay demonstrated 100 % concordance with previous NGS results in 30 cases. In the QNS cohort, the assay was valid in all cases and KRAS mutations were identified in 3 of 11 cytology smears, including one G12C mutation. CONCLUSION: The Idylla KRAS assay is a high-performing, feasible, and convenient option for testing extracted DNA and cytology smears. It rescues QNS samples allowing it to be integrated into the molecular workflow as an initial screening test with remarkably quick turnaround times.
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Citodiagnóstico , Proteínas Proto-Oncogênicas p21(ras) , DNA , Análise Mutacional de DNA/métodos , Formaldeído , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genéticaRESUMO
For cancer treatment, diagnostics concerning tumor type and determination of molecular markers in short TAT is critical. The fully automated, real-time PCR-based molecular diagnostic Idylla assays are well established in many laboratories for qualitative detection, short TAT and routine screening of clinically relevant oncogenic mutations. According to the manufacturer, all IVD assays are recommended for use only with FFPE tissue samples of 5-10 µM dissections with at least 10% tumor content. In this study, we tested the performance and accuracy of the IVD assays along with the gene fusion assay (RUO) with different tissue/source materials like isolated DNA/RNA, cryomaterial, etc. The study also included testing archival FFPE tissue sections dating back from 20 years and a performance check for different pan-cancer samples individually. All the assays tested with FFPE sections and gDNA/RNA input showed above 96% accuracy and sensitivity, individually with 100% specificity. The Idylla assays also performed exceptionally well on the archival FFPE tissues, and the use of assays for other solid tumors was also remarkable. The performance test and accuracy of Idylla assays showed high efficiency with certain limitations. For the use of Idylla assays, both qualitative and quantitative applicability of different tumor source materials could produce efficient results in different diagnostic settings within a short TAT.
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Neoplasias , Humanos , Análise Mutacional de DNA/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Neoplasias/diagnóstico , Neoplasias/genética , Patologia Molecular , RNA , MutaçãoRESUMO
Molecular analyses have become mandatory for treatment choices in patients with various advanced cancers. Beside next generation sequencing (NGS) analyzing genes panels, non-NGS targeted analyses about the main biomarkers remain of interest. In this article, we review the data about the fast and fully automated real-time PCR platform Idylla™ (Biocartis, Mechelen, Belgium) permitting the mutational analyses of BRAF, KRAS, NRAS, EGFR and microsatellite instability notably in melanoma, non-small-cell lung cancer and colorectal cancer samples. Future applications as well as the implementation of Idylla™ in the workflow of pathology and/or molecular biology laboratories are also discussed.
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Colorretais , Neoplasias Pulmonares , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Análise Mutacional de DNA , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Mutação , Medicina de Precisão , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genéticaRESUMO
BACKGROUND: Pancreatic carcinoma carries a devastating prognosis and is the 4th leading cause for cancer related death in the US and most European countries. Apart from imaging and CA 19-9, pancreatic carcinoma is still lacking reliable markers to assess tumor dynamics and to monitor treatment response over time. The aim of this study was to evaluate the feasibility of cell free tumor-DNA (cft-DNA), respectively KRAS mutation in peripheral blood, detection as a prognostic and predictive value for chemotherapy monitoring. METHODS: Serial plasma samples from 42 patients with KRAS mutated pancreatic cancer were prospectively collected and the ctKRAS Mutation Assay (Idylla™, Biocartis, Mechelen, Belgium) of cft-DNA was performed on 29 patients that did not receive curative surgery and went on to palliative chemotherapy. To monitor cft-DNA KRAS mutation levels during treatment quantitative assessment of cft-DNA was performed at baseline and during follow up at predetermined times. RESULTS: All 29 patients included in our analyses had a detected KRAS mutation in the tumor biopsy. In almost half (48.2%) of patients a KRAS mutation could also be detected in peripheral plasma. Patients with detectable KRAS mutations before treatment start in plasma had a significantly worse survival (16.8 months vs not reached, p < 0.031 and HR 3.303). Looking for a dynamic assessment of tumor response, we found a statistically significant association between the KRAS mutant ratio from first staging CT scan to basal levels with tumor response or progress (p = 0.014). CONCLUSION: Performing KRAS testing from peripheral blood for patients, who have no elevated tumor markers, might be a novel option for treatment monitoring complementing routine imaging techniques.
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DNA Tumoral Circulante , Neoplasias Pancreáticas , Proteínas Proto-Oncogênicas p21(ras)/genética , Biomarcadores Tumorais/genética , Análise Mutacional de DNA , Humanos , Mutação , Neoplasias Pancreáticas/genética , Prognóstico , Neoplasias PancreáticasRESUMO
The current standard of care for advanced non-small-cell lung cancer is based on detecting actionable mutations that can benefit from targeted therapy. Comprehensive genetic tests can have long turn-around times, and because EGFR mutations are the most prevalent actionable mutation, a quick detection would enable a prompt initiation of targeted therapy. Furthermore, the scarcity of diagnostic material means that sometimes only cytologic material is available. The Idylla™ EGFR assay is a real-time PCR-based method able to detect 51 EGFR mutations in 2.5 h. Idylla is validated for use only on FFPE sections, but some researchers described their experiences with cytological material. We reviewed the relevant literature, finding four articles describing 471 cases and many types of cytological input material: smears, cell-block sections, suspensions, and extracted DNA. The sensitivity, specificity, and limit of detection appear comparable to those obtained with histological input material, with one exception: the usage of scraped stained smears as input may reduce the accuracy of the test. In conclusion, usage of cytological material as input to the Idylla EGFR test is possible. A workflow where common mutations are tested first and fast, leaving rarer mutations for subsequent comprehensive profiling, seems the most effective approach.
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Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Análise Mutacional de DNA , Testes Genéticos , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Receptores ErbB/genética , Humanos , Mutação/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
INTRODUCTION: The currently approved techniques for RAS mutations testing in colorectal cancer (CRC) tissue are labor-intensive and time consuming. The Idylla technology (IT) is a rapid and fully automated diagnostics system. The primary aim of this study is to compare the Idylla performance against that of conventional techniques (CT). METHODOLOGY: Archival CRC tumor samples from 2 hospitals were tested for KRAS and NRAS mutations using the IT. Results were compared to those obtained earlier by CT performed in accredited laboratories. Unexplained discordant results were verified locally by next generation sequencing (NGS) to ascertain the accuracy of IT. RESULTS: Forty five samples were processed. All samples underwent dual testing (CT & IT) for KRAS mutations. IT identified mutations in 2 samples that were not detected by CT. Primary concordance rate for KRAS was 93.3% and the accuracy rate improved to 100% after verification and explanation of discordant results. Only 18 samples underwent dual testing for NRAS. Primary concordance and accuracy rates for NRAS were 94.4%. The mean time from dispatching the specimen for RAS testing by CT until receipt of results was 12 (7-28) days compared to few hours when IT was used. CONCLUSION: IT provides a quick and reliable mean for RAS testing. In addition, it identifies mutations that are not detected by CT and thus may provide better guidance to treatment choices.
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Neoplasias Colorretais/genética , Análise Mutacional de DNA/métodos , GTP Fosfo-Hidrolases/genética , Proteínas de Membrana/genética , Técnicas de Diagnóstico Molecular/métodos , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias Colorretais/diagnóstico , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BRAF mutation detection is worthwhile for the management of patients with some advanced cancers. The tumor samples are sometimes difficult to analyze using DNA-based molecular methods because of poor tumor DNA quality or quantity. Anti-BRAFV600E VE1 immunohistochemistry (IHC) has been proposed as a valuable ancillary tool to analyze some "molecularly challenging" tumor samples. In this technical study, we focused on its application in the field of decalcified tumor samples. We selected four patients with known BRAFV600E-mutated cancer (3 metastatic melanomas and 1 hairy cell leukemia) and paired non-decalcified/decalcified tumor samples. Molecular analyses failed in the four decalcified samples (3 bone metastases and 1 osteo-medullar biopsy) with non-contributive mutation status. Whereas non-decalcified tumor samples were all positive using anti-BRAFV600E VE1 IHC, the four decalcified samples were concluded negative. Because decalcified tumor samples are difficult to analyze from a molecular point of view, it is tempting to use IHC instead of DNA-based methods searching for BRAFV600E mutations in these samples. Nevertheless, the decalcification process may also cause false-negative results using VE1 IHC. Decalcified samples require specific and optimized IHC and molecular protocols and quality controls.
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Leucemia de Células Pilosas/diagnóstico , Melanoma/diagnóstico , Proteínas Proto-Oncogênicas B-raf/genética , Adulto , Substituição de Aminoácidos , Biópsia , Técnica de Descalcificação , Reações Falso-Negativas , Feminino , Humanos , Imuno-Histoquímica , Leucemia de Células Pilosas/genética , Leucemia de Células Pilosas/patologia , Masculino , Melanoma/genética , Melanoma/patologia , Pessoa de Meia-Idade , Mutação , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: Testing for KRAS mutations in metastatic colorectal cancer (mCRC) on formalin-fixed, paraffin embedded (FFPE) tumor tissue has become standard of care. Different molecular methods exist to determine hotspot KRAS mutations in exon 2, 3 and 4, but testing is often limited by the sensitivity and the speed of analysis. The aim of this retrospective study was to establish the clinical performance of the Idylla™ KRAS Mutation Test on FFPE tumor samples of patients with mCRC. METHODS: KRAS mutation analysis was performed using the therascreen KRAS on the RotorGene Q platform (CE-IVD; Qiagen) and results were subsequently compared to the Idylla™ KRAS Mutation Test. Discordant result testing was performed with massive parallel sequencing or alternative routine approaches. RESULTS: Data from 182 samples were used to show that the overall agreement between the two methods for mutation characterization was 96.7% [95%CI: 93.0%-98.5%]. Six out of 182 samples (3.3%) showed true discordant results. CONCLUSION: The Idylla™ KRAS Mutation Test allows for a fast and reliable analysis of FFPE samples with a turnaround-time of two hours without the need of molecular infrastructure or expertise in order to guide the personalized treatment of colorectal cancer patients.
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Neoplasias Colorretais/genética , Análise Mutacional de DNA/métodos , Proteínas Proto-Oncogênicas p21(ras)/genética , Neoplasias Colorretais/diagnóstico , Formaldeído , Humanos , Limite de Detecção , Mutação/genética , Inclusão em ParafinaRESUMO
IDH1 and IDH2 mutational status is a critical biomarker with diagnostic, prognostic, and treatment implications in glioma. Although IDH1 p.R132H-specific immunohistochemistry is available, it is unable to identify other mutations in IDH1/2. Next-generation sequencing can accurately determine IDH1/2 mutational status but suffers from long turnaround time when urgent treatment planning and initiation is medically necessary. The Idylla assay can detect IDH1/2 mutational status from unstained formalin-fixed paraffin-embedded (FFPE) slides in as little as a few hours. In a clinical validation, we demonstrate clinical accuracy of 97% compared to next-generation sequencing. Sensitivity studies demonstrated a limit of detection of 2.5-5% variant allele frequency, even at DNA inputs below the manufacturer's recommended threshold. Overall, the assay is an effective and accurate method for rapid determination of IDH1/2 mutational status.
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Neoplasias Encefálicas , Glioma , Isocitrato Desidrogenase , Mutação , Humanos , Isocitrato Desidrogenase/genética , Glioma/genética , Glioma/patologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/enzimologia , Análise Mutacional de DNA/métodos , Inclusão em Parafina , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/análise , Sequenciamento de Nucleotídeos em Larga Escala , Formaldeído , Fixação de Tecidos/métodos , Reprodutibilidade dos TestesRESUMO
Fifteen percent of all colorectal cancers have detectable defects in the mismatch repair system (dMMR). MMR status is used to identify possible Lynch Syndrome (LS) and to determine prognosis and choice of treatment. Two standard techniques for determining MMR status are immunohistochemistry (IHC) and analysis for microsatellite instability (MSI) by PCR. Recently, our department introduced Idylla™ MSI assay as an alternative option to IHC, and as part of this, we introduced a decision algorithm. The purpose of this study was to review the use of the new method and our algorithm and to assess possible false-positive results. Retrospectively, we identified 629 cases of colorectal cancer in which either IHC (336 cases) or Idylla™ MSI (293 cases) was performed. Similar results were obtained by the two methods. IHC detected dMMR in 55 cases (16%) and Idylla™ MSI in 52 cases (18%). In all 52 cases of MSI, subsequent IHC was performed. One case was not confirmed by IHC, but was confirmed by another PCR-based method. Overall, we found that the Idylla™ MSI works well as a screening method for dMMR with no false-positive cases detected. The proposed algorithm was useful and easily applicable.
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Algoritmos , Neoplasias Colorretais , Imuno-Histoquímica , Instabilidade de Microssatélites , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/patologia , Estudos Retrospectivos , Masculino , Feminino , Imuno-Histoquímica/métodos , Pessoa de Meia-Idade , Idoso , Reparo de Erro de Pareamento de DNA/genética , Adulto , Idoso de 80 Anos ou mais , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Reação em Cadeia da Polimerase/métodosRESUMO
Background: Epidermal growth factor receptor (EGFR) mutation detection is essential for the therapy of lung cancer. A sensitive, specific, and cost-effective standardized method to quickly and accurately detect EGFR mutations is urgently needed. Methods: We evaluated the Idylla™ EGFR Mutation Assay for EGFR mutations in formalin-fixed, paraffin-embedded (FFPE) tumor samples from 232 lung cancer patients, and compared the results with amplification refractory mutation system (ARMS) (n=146) and next-generation sequencing (NGS) (n=86). The surgical tumor sections and cell blocks derived from the same FFPE section were compared. Overall concordance, specificity, sensitivity, cost-effectiveness and turnaround time were compared among the three methods. Results: The overall concordance between Idylla and ARMS was 89.51% [95% confidence interval (CI): 83.31% to 93.64%] and the specificity of Idylla was 88.68% (95% CI: 80.69% to 93.76%). A concordance of 97.67% (95% CI: 91.41% to 99.86%) was obtained between Idylla and NGS, the specificity of Idylla was 96.30% (95% CI: 86.16% to 99.36%). Compared to the ARMS and NGS, the Idylla™ system significantly reduces the turnaround time. Combining labor, equipment, reagents and time costs, Idylla is more affordable. Conclusions: Clinically urgent cases with adequate cellularity, can first perform Idylla to detect critical markers, then perform NGS for a comprehensive mutation analysis. Besides, with limited molecular expertise or infrastructure, the Idylla has the potential to extend EGFR testing to more pathology laboratories in primary hospitals.
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The current study assessed the performance of the fully automated RT-PCR-based Idylla™ GeneFusion Assay, which simultaneously covers the advanced non-small cell lung carcinoma (aNSCLC) actionable ALK, ROS1, RET, and MET exon 14 rearrangements, in a routine clinical setting involving 12 European clinical centers. The Idylla™ GeneFusion Assay detects fusions using fusion-specific as well as expression imbalance detection, the latter enabling detection of uncommon fusions not covered by fusion-specific assays. In total, 326 archival aNSCLC formalin-fixed paraffin-embedded (FFPE) samples were included of which 44% were resected specimen, 46% tissue biopsies, and 9% cytological specimen. With a total of 179 biomarker-positive cases (i.e., 85 ALK, 33 ROS1, 20 RET fusions and 41 MET exon 14 skipping), this is one of the largest fusion-positive datasets ever tested. The results of the Idylla™ GeneFusion Assay were compared with earlier results of routine reference technologies including fluorescence in situ hybridization, immunohistochemistry, reverse-transcription polymerase chain reaction, and next-generation sequencing, establishing a high sensitivity/specificity of 96.1%/99.6% for ALK, 96.7%/99.0% for ROS1, 100%/99.3% for RET fusion, and 92.5%/99.6% for MET exon 14 skipping, and a low failure rate (0.9%). The Idylla™ GeneFusion Assay was found to be a reliable, sensitive, and specific tool for routine detection of ALK, ROS1, RET fusions and MET exon 14 skipping. Given its short turnaround time of about 3 h, it is a time-efficient upfront screening tool in FFPE samples, supporting rapid clinical decision making. Moreover, expression-imbalance-based detection of potentially novel fusions may be easily verified with other routine technologies without delaying treatment initiation.
Assuntos
Quinase do Linfoma Anaplásico , Carcinoma Pulmonar de Células não Pequenas , Éxons , Neoplasias Pulmonares , Proteínas de Fusão Oncogênica , Proteínas Tirosina Quinases , Proteínas Proto-Oncogênicas c-met , Proteínas Proto-Oncogênicas c-ret , Proteínas Proto-Oncogênicas , Humanos , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas Proto-Oncogênicas c-ret/genética , Quinase do Linfoma Anaplásico/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas c-met/genética , Éxons/genética , Proteínas Proto-Oncogênicas/genética , Proteínas de Fusão Oncogênica/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/análise , Rearranjo Gênico , Hibridização in Situ Fluorescente/métodos , Reação em Cadeia da Polimerase MultiplexRESUMO
Background: The impact of KRAS mutation testing on pancreatic ductal adenocarcinoma (PDAC) samples by endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) for reducing the need to repeat EUS-FNA has been demonstrated. Such testing however is not part of standard practice for endoscopic ultrasound-guided fine-needle biopsy (EUS-FNB). Objectives: We aim to analyse the proportion of non-contributive samples by EUS-FNB and to evaluate the impact of KRAS mutation testing on the diagnosis, theranostics and survival. Design: In this retrospective study, the impact on diagnosis and survival of KRAS testing for contributive and non-contributive samples by EUS-FNB was analysed. Methods: The EUS-FNB samples, combined with KRAS testing using the Idylla® technique on liquid-based cytology from patients with PDAC between February 2019 and May 2023, were retrospectively reviewed. The cytology results were classified according to the guidelines of the World Health Organization System for Reporting Pancreaticobiliary Cytopathology (WHOSRPC). Results: A total of 85 EUS-FNB specimens were reviewed. In all, 25 EUS-FNB samples did not lead to a formal diagnosis of PDAC according to the WHOSRPC (30.2%). Out of these 25, 11 (44%) could have been considered positive for a PDAC diagnosis thanks to the KRAS mutation test without carrying out further diagnosis procedures. The sensitivity of KRAS mutation testing using the Idylla technique was 98.6%. According to the available data, survival rates were not statistically different depending on the type of mutation. Conclusion: KRAS mutation testing on liquid-based cytology using the Idylla or equivalent technique, combined with the PDAC EUS-FNB sample, should become a standard for diagnosis to avoid delaying treatment by doing another biopsy. Furthermore, knowledge of the KRAS status from treatment initiation could be used to isolate mutations requiring targeted treatments or inclusion in clinical research trials, especially for wild-type KRAS PDAC.
Diagnostic and theranostic interest of searching for a KRAS mutation in echoendoscopic ultrasound biopsies of pancreatic adenocarcinomas The echoendoscopic ultrasound diagnostic of pancreatic adenocarcinomas sometimes remains difficult due to the nature of these tumors with a particular microenvironment. For more than 30 years, several authors have underlined the importance of searching for a KRAS mutation on samples taken by echoendoscopic ultrasound to improve diagnostic performance. However, this research is not common practice. Our retrospective study made it possible to review the files of 85 patients with pancreatic adenocarcinoma in whom an echoendoscopic ultrasound biopsy was performed with a search for the KRAS mutation (with second-generation fine needle biopsy). Forty-four percent could have been considered positive for the diagnosis of PDAC thanks to the search for the KRAS mutation without repeating new samples. Furthermore, knowledge of the KRAS mutation type from diagnosis would make it possible to isolate mutations justifying possible targeted treatments.
RESUMO
Molecular diagnostics for lung cancer is a well-established standard of care, but how to use the available diagnostic tools for optimal and cost-effective patient care remains unresolved. Here, we show that DNA-only, small gene next-generation sequencing (sNGS) panels (<50 genes) combined with ultra-rapid reflex testing for common fusion transcripts using the Idylla Genefusion assay provide a cost-effective and sufficiently comprehensive testing modality for the majority of lung cancer cases. We also demonstrate the need for additional reflex testing capability on larger DNA and fusion panels for a small subset of lung cancers bearing rare single-nucleotide variants, indels and fusion transcripts and secondary, post-treatment resistance mutations. A similar testing workflow could be adopted for other solid tumor types for which extensive gene/fusion variant profiles are available both in the treatment-naïve and post-therapy settings.