CURIE LUNG CANCER Mutation analysis

This document summarizes procedures for generating Curie Lung cancer mutation data output. Data analysis has been performed by Dr Ivan Bièche in Hospital platform for cancer molecular genetics at Institut Curie.


PDX was analysed by targeted next generation sequencing (NGS) of 95 genes, chosen among the most frequently mutated genes in lung cancer (>1%) and including potential therapeutic targets (SAFIR Lung panel) (see table below).

NGS was performed on an Illumina HiSeq2500 sequencer and the genomic variants were annotated with COSMIC and 1000 genome databases.

Reads were aligned using BWA allowing up to 4% of mismatches with the reference.Only those reads with a mapping quality higher than 20 were used for variant calling, performed with GATK unified genotyper.

NGS primers were selected based on their specificity on the human genome. Genomic alterations included single nucleotide variations of significantly mutated genes (ie, base substitutions and short insertions/deletions). Deleterious genomic alterations were defined as follows:

  1. For oncogenes, only mutations driving to gain of function were considered (ie, hotspots missense mutations, in-frame insertions/deletions/splicing described as oncogenic),
  2. For tumour suppressor genes, only mutations driving to loss of function were considered (ie. biallelic truncating alterations (nonsense mutations, frameshift insertions/deletions/splicing) or monoallelic truncating alterations associated with heterozygous deletion detected by copy number analysis). Variants with low allelic frequency (<5%) or low coverage (<100x) were excluded from the analysis.

Genomic variants were biologically validated by comparison with COSMIC, TumorPortal and cBioportal databases.

Gene type Mutation covered number
AKT1 HotSpot p.E17K 1 amplicon
AKT2 HotSpot p.E17Klike 1 amplicon
AKT3 HotSpot p.E17Klike 1 amplicon
ALK HotSpot ex 20,22,23,24,25 8 amplicons
ATM FullCDS 96.40% 147 amplicons
BRAF HotSpot Ex 11&15 2 amplicons
BRCA1 FullCDS 92.80% 123 amplicons
BRCA2 FullCDS 99.10% 73 amplicons
DDR2 HotSpot Ex6,9,14-16,18 7 amplicons
EGFR HotSpot Ex3,6,7,15,18,19,20,21 11 amplicons
ESR1 HotSpot Ex4,5,7,8 4 amplicons
HER2 /Erbb2 HotSpot Ex8, 17-22, 24 9 amplicons
HER3/ Erbb3 HotSpot Ex2,3,6-8 6 amplicons
HER4 / Erbb4 HotSpot Ex1 1 amplicon
FBXW7 HotSpot Ex5,8-11 6 amplicons
FGFR1 HotSpot ex4,7,12,14,15 5 amplicons
FGFR2 HotSpot Ex3,7,9,12,14 17 amplicons
FGFR3 HotSpot Ex7,9 (couv 52%),14,16,18 6 amplicons
FGFR4 HotSpot Ex11,12,14,16 4 amplicons
HRAS HotSpot Ex2,3,4 4 amplicons
IDH1 HotSpot Ex4 1 amplicon
IDH2 HotSpot Ex4 2 amplicons
KDR HotSpot Ex7,8,23 3 amplicons
KEAP1 HotSpot Ex2,3,4,6 8 amplicons
KIT HotSpot Ex8 à 14, 17, 18 15 amplicons
KRAS HotSpot Ex 2,3,4 3 amplicons
MAP2K1 HotSpot Ex3,4,6,7,11 5 amplicons
MAP2K2 HotSpot Ex2,4 2 amplicons
MAP2K4 FullCDS 99.30% 20 amplicons
MAP3K1 FullCDS 95.40% 58 amplicons
MET HotSpot Ex14 (+introns), 16,19-21 9 amplicons
MTOR FullCDS 99.10% 113 amplicons
NF1 FullCDS 98.50% 137 amplicons
NFE2L2 HotSpot Ex2 3 amplicons
NRAS HotSpot Ex 2,3,4 4 amplicons
PALB2 FullCDS 96.60% 44 amplicons
PDGFRA HotSpot Ex12,14,15,18 6 amplicons
PIK3CA HotSpot Ex2,5,8,10,14,21 11 amplicons
PIK3R1 HotSpot Ex9-15 8 amplicons
POLE HotSpot Ex6,9,11,13,14,16,19,31,36 11 amplicons
PPP2R1A HotSpot Ex5,6 2 amplicons
PTEN FullCDS 88.60% 19 amplicons
PTPN11 HotSpot Ex3,13 2 amplicons
RAD51B FullCDS 94.00% 22 amplicons
RAD51C FullCDS 100% 22 amplicons
RET HotSpot Ex10,11,13,15,16 8 amplicons
ROS1 HotSpot Ex 38 1 amplicon
STK11 FullCDS 94.30% 23 amplicons
TP53 FullCDS 97.70% 22 amplicons
TSC1 FullCDS 97.90% 49 amplicons
TSC2 FullCDS 97.20% 93 amplicons
VHL FullCDS 97.80% 8 amplicons

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