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Browsing by Author "Vicha, Ales"

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  • Frequency and Prognostic Impact of ALK Amplifications and Mutations in the European Neuroblastoma Study Group (SIOPEN) High-Risk Neuroblastoma Trial (HR-NBL1)
    Publication . Bellini, Angela; Pötschger, Ulrike; Bernard, Virginie; Lapouble, Eve; Baulande, Sylvain; Ambros, Peter F.; Auger, Nathalie; Beiske, Klaus; Bernkopf, Marie; Betts, David R.; Bhalshankar, Jaydutt; Bown, Nick; de Preter, Katleen; Clément, Nathalie; Combaret, Valérie; Font de Mora, Jaime; George, Sally L.; Jiménez, Irene; Jeison, Marta; Marques, Barbara; Martinsson, Tommy; Mazzocco, Katia; Morini, Martina; Mühlethaler-Mottet, Annick; Noguera, Rosa; Pierron, Gaelle; Rossing, Maria; Taschner-Mandl, Sabine; Van Roy, Nadine; Vicha, Ales; Chesler, Louis; Balwierz, Walentyna; Castel, Victoria; Elliott, Martin; Kogner, Per; Laureys, Geneviève; Luksch, Roberto; Malis, Josef; Popovic-Beck, Maja; Ash, Shifra; Delattre, Olivier; Valteau-Couanet, Dominique; Tweddle, Deborah A.; Ladenstein, Ruth; Schleiermacher, Gudrun
    Purpose: In neuroblastoma (NB), the ALK receptor tyrosine kinase can be constitutively activated through activating point mutations or genomic amplification. We studied ALK genetic alterations in high-risk (HR) patients on the HR-NBL1/SIOPEN trial to determine their frequency, correlation with clinical parameters, and prognostic impact. Materials and methods: Diagnostic tumor samples were available from 1,092 HR-NBL1/SIOPEN patients to determine ALK amplification status (n = 330), ALK mutational profile (n = 191), or both (n = 571). Results: Genomic ALK amplification (ALKa) was detected in 4.5% of cases (41 out of 901), all except one with MYCN amplification (MNA). ALKa was associated with a significantly poorer overall survival (OS) (5-year OS: ALKa [n = 41] 28% [95% CI, 15 to 42]; no-ALKa [n = 860] 51% [95% CI, 47 to 54], [P < .001]), particularly in cases with metastatic disease. ALK mutations (ALKm) were detected at a clonal level (> 20% mutated allele fraction) in 10% of cases (76 out of 762) and at a subclonal level (mutated allele fraction 0.1%-20%) in 3.9% of patients (30 out of 762), with a strong correlation between the presence of ALKm and MNA (P < .001). Among 571 cases with known ALKa and ALKm status, a statistically significant difference in OS was observed between cases with ALKa or clonal ALKm versus subclonal ALKm or no ALK alterations (5-year OS: ALKa [n = 19], 26% [95% CI, 10 to 47], clonal ALKm [n = 65] 33% [95% CI, 21 to 44], subclonal ALKm (n = 22) 48% [95% CI, 26 to 67], and no alteration [n = 465], 51% [95% CI, 46 to 55], respectively; P = .001). Importantly, in a multivariate model, involvement of more than one metastatic compartment (hazard ratio [HR], 2.87; P < .001), ALKa (HR, 2.38; P = .004), and clonal ALKm (HR, 1.77; P = .001) were independent predictors of poor outcome. Conclusion: Genetic alterations of ALK (clonal mutations and amplifications) in HR-NB are independent predictors of poorer survival. These data provide a rationale for integration of ALK inhibitors in upfront treatment of HR-NB with ALK alterations.
    2021-06-11Journal article Open access Show more
  • Genetic alterations of ALK in high-risk neuroblastoma patients. A SIOPEN study
    Publication . Bellini, Angela; Bernard, Virginie; Ambros, Inge; F. Ambros, Peter; de Preter, Katleen; Combaret, Valérie; Beiske, Klaus; Jeison, Marta; Marques, Barbara; Morini, Martina; Mazzocco, Katia; Defferrari, Raffaella; Betts, David; Martinsson, Tommy; Mühlethaler-Mottet, Annick; Noguera, Rosa; Font de Mora, Jaime; Vicha, Ales; Ladenstein, Ruth; Valteau-Couanet, Dominique; Rossing, Caroline Maria; Bown, Nick; Tweddle, Deborah; Avigad, Smadar; Lapouble, Eve; Chicard, Mathieu; Leprovost, Nada; Clement, Nathalie; Baulande, Sylvain; Pierron, Gaelle; Irene, Jimenez; Jaydutt, Bhalshankar; Delattre, Olivier; Michon, Jean; Schleiermacher, Gudrun
    Background: In neuroblastoma (NB), the ALK receptor tyrosine kinase can be constitutively activated through genomic amplification or activating point mutations. We studied ALK genetic alterations in high-risk NB patients to determine their frequency and prognostic impact. Methods: Diagnostic NB samples from 1039 patients enrolled in the SIOPEN-HR-NBL1 trial were studied to determine the ALK amplification status (copy number analysis; n=337), the ALK mutational profile (Sanger and/or NGS including deep sequencing, n=203) or both (n=499). The sensitivity of ALK mutated/ALK amplified or ALK wildtype NB cell lines ((CLB-GA (R1275Q), CLB-GE (F1174V; ALK-A), SKNBE-2C (ALK wt)) to simultaneous or consecutive combinations of ALK TKIs (crizotinib/lorlatinib) and/or chemotherapy (Etoposide and Doxorubicin) was then tested. Results: Genomic ALK amplifications were detected in 4.4% of cases (37/836); all but 2 showed MYCN amplification. ALK mutations were detected at a clonal level (>20% mutated allele fraction, MAF) in 9.8% of cases (69/702) (F1174 n=25, R1275 n=32, both F1174 and R1275 n=1, F1245 n=6, others n=5) and at a subclonal level (MAF 0.5-20%) in 3.7% of patients (22/586) (F1174 n=11, R1275 n=6, both F1174 and R1275 or F1174 and F1245 n=3, other n=2). A significantly poorer OS and EFS was observed in cases with clonal ALK mutations, versus all others (3-years OS 47% +/-6.4% versus 65% +/-2%, logrank, p< 0.0001) and in those with ALK amplifications, versus all others (3-years OS 31% +/-8.5% versus 66% +/- 1.9%; logrank, p<0.0001). A Cox proportional hazards procedure (450 patients with complete clinical/biological datasets) retained stage 4 disease (as opposed to non-stage 4) and ALK amplification as factors with a higher hazard of relapse/progression (hazard 2.3 and 2.2, respectively), whereas ALK mutation, MYCN amplification and age>18 months were not retained. The consecutive treatment of Doxorubicin followed by Lorlatinib had a synergistic effect in ALK mutated/amplified NB cell lines. Conclusion: Genetic alterations of ALK (clonal mutations, amplifications) in high-risk NB patients are associated with poorer survival. Further preclinical data are required to determine optimal treatment modalities for integration of TKI in upfront treatment strategies of HR NB patients with ALK alterations.
    2019-10-26Conference object Open access Show more
  • Genetic alterations of ALK in high-risk neuroblastoma patients: a SIOPEN study
    Publication . Bellini, Angela; Bernard, Virginie; Ambros, Inge; Ambros, Peter F.; de Preter, Katleen; Combaret, Valérie; Beiske, Klaus; Jeison, Marta; Marques, Barbara; Morini, Martina; Mazzocco, Katia; Defferrari, Raffaella; Betts, David; Martinsson, Tommy; Mühlethaler-Mottet, Annick; Noguera, Rosa; Font de Mora, Jaime; Vicha, Ales; Ladenstein, Ruth; Valteau-Couanet, Dominique; Rossing, Caroline Maria; Bown, Nick; Tweddle, Deborah; Avigad, Smadar; Lapouble, Eve; Chicard, Mathieu; Leprovost, Nada; Clement, Nathalie; Baulande, Sylvain; Pierron, Gaelle; Irene, Jimenez; Jaydutt, Bhalshankar; Delattre, Olivier; Michon, Jean; Schleiermacher, Gudrun
    Background: In neuroblastoma (NB), the ALK receptor tyrosine kinase can be constitutively activated either through genomic amplification or activating point mutations. We studied ALK genetic alterations in high-risk NB patients to determine their frequency and prognostic impact.
    2018-10-10Conference object Open access Show more
  • High frequency of subclonal ALK mutations in high risk neuroblastoma patients. A SIOPEN study
    Publication . Bellini, Angela; Bernard, Virginie; Lapouble, Eve; Clement, Nathalie; Pierron, Gaelle; Ambros, Inge M.; Preter, Katleen de; Van Roy, Nadine; Vicha, Ales; Combaret, Valérie; Betts, David; Jeison, Marta; Avigad, Smadar; Morini, Martina; Varesio, Luigi; Marques9, Barbara; Muhlethaler, Annick; Noguera, Rosa; Berbegall, Ana; Mora, Jaime Font de; Ambros, Peter F.; Ladenstein, Ruth; Valteau-Couanet, Dominique; Michon, Jean; Delattre, Olivier; Bown, Nick; Tweddle, Deborah; Schleiermache, Gudrun
    Introduction: In neuroblastoma (NB), activating ALK receptor tyrosine kinase point mutations are detected in 8–10% at diagnosis using conventional sequencing. To determine the potential occurrence and the prognostic impact of ALK mutations in a series of high risk NB patients we studied ALK variation frequencies using targeted deep sequencing in samples of patients enrolled in the SIOPEN HR-NBL01 study
    2015-11Conference object Open access Show more
  • A multilocus technique for risk evaluation of patients with neuroblastoma
    Publication . Ambros, Inge M.; Brunner, Bettina; Aigner, Gerhard; Bedwell, Clare; Beiske, Klaus; Bénard, Jean; Bown, Nick; Combaret, Valerie; Couturier, Jerome; Defferrari, Raffaella; Gross, Nicole; Jeison, Marta; Lunec, John; Marques, Barbara; Martinsson, Tommy; Mazzocco, Katia; Noguera, Rosa; Schleiermacher, Gudrun; Speleman, Frank; Stallings, Ray; Tonini, Gian Paolo; Tweddle, Deborah A.; Valent, Alexander; Vicha, Ales; Van Roy, Nadine; Villamon, Eva; Ziegler, Andrea; Preuner, Sandra; Drobics, Mario; Ladenstein, Ruth; Amann, Gabriele; Schuit, Robert J.L.; Pötschger, Ulrike; Ambros, Peter F.
    Purpose: Precise and comprehensive analysis of neuroblastoma genetics is essential for accurate risk evaluation and only pangenomic/multilocus approaches fulfill the present-day requirements. We present the establishment and validation of the PCR-based multiplex ligation-dependent probe amplification (MLPA) technique for neuroblastoma. Experimental Design: A neuroblastoma-specific MLPA kit was designed by the SIOP Europe Neuroblastoma Biology Committee in cooperation with MRC-Holland. The contained target sequences cover 19 chromosomal arms and reference loci. Validation was performed by single locus and pangenomic techniques (n ¼ 174). Dilution experiments for determination of minimal tumor cell percentage were performed and testing of reproducibility was checked by interlaboratory testing (n ¼ 15). Further 156 neuroblastomas were used for establishing the amplification cutoff level. Results: The MLPA technique was tested in 310 neuroblastomas and 8 neuroblastoma cell lines (including validation and amplification cutoff level testing). Intertechnique validation showed a high concordance rate (99.5%). Interlaboratory MLPA testing (k ¼ 0.95, P < 0.01) revealed 7 discrepant of 1,490 results (0.5%). Validation by pangenomic techniques showed a single discordance of 190 consensus results (0.5%). The test results led to formulation of interpretation standards and to a kit revision. The minimal tumor cell percentage was fixed at 60%. Conclusions: The recently designed neuroblastoma-specific MLPA kit covers all chromosomal regions demanded by the International Neuroblastoma Risk Group for therapy stratification and includes all hitherto described genetic loci of prognostic interest for future studies and can be modified or extended at any time. Moreover, the technique is cost effective, reliable, and robust with a high interlaboratory and intertechnique concordance.
    2011-02-15Journal article Restricted access Show more