Methods for determining the cause of somatic mutagenesis

Abstract

A method can determine the likelihood that targeted somatic mutagenesis of a nucleic acid molecule by a mutagenic agent has occurred. The method includes analyzing the sequence of the nucleic acid molecule to determine, for a number of mutations of a mutation type at one or more motifs recognized or targeted by the mutagenic agent, the codon context of those mutations to thereby identify the location of a mutation and mutation type for each of the mutated codons in the nucleic acid molecule. The codon context of an individual mutation can be determined by determining at which of the three positions of a corresponding mutated codon the individual mutation occurs. The mutagenic agent can be one of aflatoxin, activation-induced cytidine deaminase (AID), or an apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) cytidine deaminase.

What is claimed is: 
     
         1 . A method for determining the likelihood that targeted somatic mutagenesis of a nucleic acid molecule by a mutagenic agent has occurred, the method comprising:
 analyzing the sequence of the nucleic acid molecule to determine, for a plurality of mutations of a mutation type at one or more motifs recognized or targeted by the mutagenic agent, the codon context of those mutations to thereby identify the location of a mutation and mutation type for each of a plurality of mutated codons in the nucleic acid molecule, wherein the codon context of an individual mutation is determined by determining at which of the three positions of a corresponding mutated codon the individual mutation occurs; and   determining that targeted somatic mutagenesis is likely to have occurred when there is a higher than expected percentage or number of mutations of a mutation type at one of the three positions in the plurality of mutated codons; wherein   the mutagenic agent is selected from among aflatoxin, activation-induced cytidine deaminase (AID), and an apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) cytidine deaminase.   
     
     
         2 . The method of  claim 1 , wherein the expected percentage or number of mutations is calculated by assuming that mutations occur independently of codon context. 
     
     
         3 . The method of  claim 2 , wherein the expected percentage of mutations is approximately 11% or 17%. 
     
     
         4 . The method of  claim 2 , wherein the expected number of mutations is approximately 1 of every 9 mutations or 1 of every 6 mutations. 
     
     
         5 . The method of  claim 1 , wherein the percentage of mutations is observed to be at least 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 80%, 85%, 90%, 95% or more. 
     
     
         6 . The method of  claim 1 , wherein the APOBEC cytidine deaminase is selected from among APOBEC1, APOBEC3A, APOBEC3B, APOBEC3C, APOBEC3D, APOBEC3F, APOBEC3G and APOBEC3H. 
     
     
         7 . The method of  claim 1 , wherein targeted somatic mutagenesis is determined to be likely to have occurred when:
 the number or percentage of observed G to A mutations in GYW motifs at MC-2 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed C>T mutations in WRC motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed G>A mutations in CG motifs at MC-2 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed C>T mutations in CG motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed C>T mutations in CA motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed G>A mutations in GA motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed G>A mutations in TG motifs at MC-2 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed G>T mutations in GG motifs at MC-3 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected;   the number or percentage of observed C>T mutations in CC motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected; or   the number or percentage of observed A>G mutations in WA motifs at MC-2 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected.   
     
     
         8 . The method of  claim 7 , wherein the mutagenic agent is AID if the number or percentage of observed G>A mutations in GYW motifs at MC-2 sites, A>G mutations in WA motifs at MC-2 sites, and/or C>T mutations in WRC motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected. 
     
     
         9 . The method of  claim 7 , wherein the mutagenic agent is APOBEC3G if the number or percentage of observed G>A mutations in CG motifs at MC-2 sites, C>T mutations in CC motifs at MC-1 sites or C>T mutations in CG motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected. 
     
     
         10 . The method of  claim 7 , wherein the mutagenic agent is APOBEC1 if the number or percentage of observed C>T mutations in CA motifs at MC-1 sites or G>A mutations in TG motifs at MC-2 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected. 
     
     
         11 . The method of  claim 7 , wherein the mutagenic agent is APOBEC3H if the number or percentage of observed G>A mutations in GA motifs at MC-1 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected. 
     
     
         12 . The method of  claim 7 , wherein the mutagenic agent is aflatoxin if the number or percentage of observed G>T mutations in GG motifs at MC-3 sites in the non-transcribed strand of the nucleic acid molecule is higher than expected. 
     
     
         13 . The method of  claim 1 , comprising first isolating the nucleic acid molecule. 
     
     
         14 . The method of  claim 1 , comprising sequencing all or a part of the nucleic acid molecule. 
     
     
         15 . The method of  claim 1 , wherein the nucleic acid molecule comprises all or part of a single gene or the cDNA of a single gene, or all or part of two or more genes or the cDNA of two or more genes. 
     
     
         16 . The method of  claim 15 , wherein the gene is a gene associated with cancer. 
     
     
         17 . The method of  claim 15 , wherein the gene is selected from among TP53, PIK3CA, ERBB2, DIRAS3, TET2 and nitric oxide synthase (NOS) genes. 
     
     
         18 . The method of  claim 1 , wherein nucleic acid molecules that constitute the whole exome or the whole genome of a cell are analyzed. 
     
     
         19 . The method of  claim 1 , wherein all or a part of the method is performed by a processing system.