Ultrasensitive detection of TKI resistant mutations using Illumina NGS in chronic myeloid leukemia patients

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F65269705%3A_____%2F18%3A00068763" target="_blank" >RIV/65269705:_____/18:00068763 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216224:14740/18:00103392

Výsledek na webu

<a href="http://phdretreat.ceitec.cz/ceitec-phd-and-postdoc-retreat-2018/" target="_blank" >http://phdretreat.ceitec.cz/ceitec-phd-and-postdoc-retreat-2018/</a>

DOI - Digital Object Identifier

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Jazyk výsledku

angličtina

Název v původním jazyce

Ultrasensitive detection of TKI resistant mutations using Illumina NGS in chronic myeloid leukemia patients

Popis výsledku v původním jazyce

The discovery of tyrosine kinase inhibitors (TKI) brought a revolution in the manage-ment of chronic myeloid leukemia (CML), increasing survival and life-quality of pa-tients. However, mutations in kinase domain (KD) of BCR-ABL1 may occur and result in therapy failure. Although KD mutation screening is routinely done by Sanger sequenc-ing (detection limit &gt;20%VAF), numerous studies showed enhanced sensitivity (&lt;15%) using next generation sequencing (NGS). Majority of NGS studies used two rounds of PCR to amplify KD sequence. Such approach may introduce PCR-based errors, restrict-ing the detection limit to &gt;1%. We aimed to develop new Illumina-based one-round PCR amplification protocol and bioinformatics pipeline (BI) with sensitivity LESS-THAN OR EQUAL TO1%. Our protocol was tested on 6 healthy controls (HC) and 34 retrospective samples from 13 CML patients with TKI resistant mutations (15 FUP samples at the time of SS muta-tion detection, 11 FUP samples collected 1-18 month before SS mutation detection, 8 diagnostic samples). The performance was assessed in comparison to routinely used SS. The BI was designed with respect to biological input (cDNA), allowing detection of SNVs with 0.1%VAF. Overall, no mutations were detected in ABL1 KD of HC, showing decreased background noise. NGS detected mutations in 100% of FUP samples previously positive by SS, showing high correlation of VAF (r2=0.96) and confirming its specificity. Due to high sensitivity, Illumina-based NGS analysis could detect mutations in 24% more samples than SS and proved to be suitable for earlier detection of TKI resistant mutations at very low frequencies (GREATER-THAN OR EQUAL TO0.1 %).

Název v anglickém jazyce

Ultrasensitive detection of TKI resistant mutations using Illumina NGS in chronic myeloid leukemia patients

Popis výsledku anglicky

The discovery of tyrosine kinase inhibitors (TKI) brought a revolution in the manage-ment of chronic myeloid leukemia (CML), increasing survival and life-quality of pa-tients. However, mutations in kinase domain (KD) of BCR-ABL1 may occur and result in therapy failure. Although KD mutation screening is routinely done by Sanger sequenc-ing (detection limit &gt;20%VAF), numerous studies showed enhanced sensitivity (&lt;15%) using next generation sequencing (NGS). Majority of NGS studies used two rounds of PCR to amplify KD sequence. Such approach may introduce PCR-based errors, restrict-ing the detection limit to &gt;1%. We aimed to develop new Illumina-based one-round PCR amplification protocol and bioinformatics pipeline (BI) with sensitivity LESS-THAN OR EQUAL TO1%. Our protocol was tested on 6 healthy controls (HC) and 34 retrospective samples from 13 CML patients with TKI resistant mutations (15 FUP samples at the time of SS muta-tion detection, 11 FUP samples collected 1-18 month before SS mutation detection, 8 diagnostic samples). The performance was assessed in comparison to routinely used SS. The BI was designed with respect to biological input (cDNA), allowing detection of SNVs with 0.1%VAF. Overall, no mutations were detected in ABL1 KD of HC, showing decreased background noise. NGS detected mutations in 100% of FUP samples previously positive by SS, showing high correlation of VAF (r2=0.96) and confirming its specificity. Due to high sensitivity, Illumina-based NGS analysis could detect mutations in 24% more samples than SS and proved to be suitable for earlier detection of TKI resistant mutations at very low frequencies (GREATER-THAN OR EQUAL TO0.1 %).

Druh

O - Ostatní výsledky

CEP obor

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OECD FORD obor

10606 - Microbiology

Projekt

<a href="/cs/project/NV17-30397A" target="_blank" >NV17-30397A: Mutační analýza primitivních buněčných populací u chronické myeloidní leukémie: za hranicí BCR-ABL1</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2018

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů