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TP53 intron 1 hotspot rearrangements are specific to sporadic osteosarcoma and can cause Li-Fraumeni syndrome.

Ribi S, Baumhoer D, Lee K - Oncotarget (2015)

Bottom Line: Using whole-genome sequencing of OS, we found features of TP53 intron 1 rearrangements suggesting a unique mechanism correlated with transcription.We revisited a four-generation LFS family where no TP53 mutation had been identified and found a 445 kb inversion spanning from the TP53 intron 1 towards the centromere.Cancers in this family had loss of heterozygosity, retaining the rearranged allele and resulting in TP53 expression loss.

View Article: PubMed Central - PubMed

Affiliation: Cancer Therapeutics & Stratified Oncology, Genome Institute of Singapore, Singapore 138672, Singapore.

ABSTRACT
Somatic mutations of TP53 are among the most common in cancer and germline mutations of TP53 (usually missense) can cause Li-Fraumeni syndrome (LFS). Recently, recurrent genomic rearrangements in intron 1 of TP53 have been described in osteosarcoma (OS), a highly malignant neoplasm of bone belonging to the spectrum of LFS tumors. Using whole-genome sequencing of OS, we found features of TP53 intron 1 rearrangements suggesting a unique mechanism correlated with transcription. Screening of 288 OS and 1,090 tumors of other types revealed evidence for TP53 rearrangements in 46 (16%) OS, while none were detected in other tumor types, indicating this rearrangement to be highly specific to OS. We revisited a four-generation LFS family where no TP53 mutation had been identified and found a 445 kb inversion spanning from the TP53 intron 1 towards the centromere. The inversion segregated with tumors in the LFS family. Cancers in this family had loss of heterozygosity, retaining the rearranged allele and resulting in TP53 expression loss. In conclusion, intron 1 rearrangements cause p53-driven malignancies by both germline and somatic mechanisms and provide an important mechanism of TP53 inactivation in LFS, which might in part explain the diagnostic gap of formerly classified "TP53 wild-type" LFS.

No MeSH data available.


Related in: MedlinePlus

TP53 intron 1 rearrangement in a family with LFS(A) Pedigree of a family with LFS. Squares and circles represent males and females, respectively. Filled symbols indicate individuals with early onset cancer. Symbols with a diagonal line indicate that the individual was deceased at the time of recruitment. Diamonds represent several additional family members whose gender should not be disclosed. Individuals of whom DNA samples have been obtained are numbered with individuals tested positive for TP53 exon 1 deletion by MLPA in red. TP53 rearrangement carrier without cancer at the age of 10 years is indicated by a white symbol with a black dot. (B) PCR analysis of positive control and TP53 intron 1 rearrangement points (LFS-BP1 and LFS-BP2) of family members of whom high quality DNA was available. DNA quality of P12 did not allow PCR amplification, including positive control (not shown). a, LFS breakpoint 1 (BP1); b, LFS breakpoint 2 (BP2); c, positive control amplicon at the RNA polymerase II locus (POLR2A); sample IDs correspond to A with affected individuals in red. L, ladder. (C) Sanger sequencing of rearrangement points LFS-BP1 and LFS-BP2 at the TP53 locus of P2. Micro homologies between the two participating break point regions are illustrated by green vertical lines. Genomic coordinates are based on NCBI build 37.
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Figure 3: TP53 intron 1 rearrangement in a family with LFS(A) Pedigree of a family with LFS. Squares and circles represent males and females, respectively. Filled symbols indicate individuals with early onset cancer. Symbols with a diagonal line indicate that the individual was deceased at the time of recruitment. Diamonds represent several additional family members whose gender should not be disclosed. Individuals of whom DNA samples have been obtained are numbered with individuals tested positive for TP53 exon 1 deletion by MLPA in red. TP53 rearrangement carrier without cancer at the age of 10 years is indicated by a white symbol with a black dot. (B) PCR analysis of positive control and TP53 intron 1 rearrangement points (LFS-BP1 and LFS-BP2) of family members of whom high quality DNA was available. DNA quality of P12 did not allow PCR amplification, including positive control (not shown). a, LFS breakpoint 1 (BP1); b, LFS breakpoint 2 (BP2); c, positive control amplicon at the RNA polymerase II locus (POLR2A); sample IDs correspond to A with affected individuals in red. L, ladder. (C) Sanger sequencing of rearrangement points LFS-BP1 and LFS-BP2 at the TP53 locus of P2. Micro homologies between the two participating break point regions are illustrated by green vertical lines. Genomic coordinates are based on NCBI build 37.

Mentions: Since mutations of TP53 are associated with LFS it seemed possible that TP53 intron 1 rearrangements could constitute a previously underappreciated category of alterations that can cause LFS. We revisited an LFS family with 12 affected members with cancer across four generations in which we previously had been unable to identify a coding TP53 mutation or a co-segregating, potentially damaging and disease-causing alteration based on exome sequencing of patients P1, P2 and P13 (Figure 3A and data not shown). Copy number analysis of DNA from blood of patients P1 and P13 by single nucleotide polymorphism (SNP) array revealed evidence for an approximately 2.5 kb deletion in intron 1 of TP53 including exon 1 (Supplementary Figure S13 and Table S12). However, we were not able to amplify the fusion point of a deletion by PCR. We then used a custom sequence capture assay for targeted paired-end sequencing of the TP53 locus to search for a rearrangement point. We identified a 445 kb inversion spanning from the breakpoint cluster region in intron 1 of TP53 towards the centromere (upstream of TP53) with a loss of 2,275 bp of intron 1/exon 1 of TP53 (Figure 4A and 4B). The lost sequence was in agreement with the deletion identified by SNP arrays. In the adult family members tested, we found this specific rearrangement co-segregating with the disease, implicating the rearrangement as the causative alteration (Figure 3B and 3C).


TP53 intron 1 hotspot rearrangements are specific to sporadic osteosarcoma and can cause Li-Fraumeni syndrome.

Ribi S, Baumhoer D, Lee K - Oncotarget (2015)

TP53 intron 1 rearrangement in a family with LFS(A) Pedigree of a family with LFS. Squares and circles represent males and females, respectively. Filled symbols indicate individuals with early onset cancer. Symbols with a diagonal line indicate that the individual was deceased at the time of recruitment. Diamonds represent several additional family members whose gender should not be disclosed. Individuals of whom DNA samples have been obtained are numbered with individuals tested positive for TP53 exon 1 deletion by MLPA in red. TP53 rearrangement carrier without cancer at the age of 10 years is indicated by a white symbol with a black dot. (B) PCR analysis of positive control and TP53 intron 1 rearrangement points (LFS-BP1 and LFS-BP2) of family members of whom high quality DNA was available. DNA quality of P12 did not allow PCR amplification, including positive control (not shown). a, LFS breakpoint 1 (BP1); b, LFS breakpoint 2 (BP2); c, positive control amplicon at the RNA polymerase II locus (POLR2A); sample IDs correspond to A with affected individuals in red. L, ladder. (C) Sanger sequencing of rearrangement points LFS-BP1 and LFS-BP2 at the TP53 locus of P2. Micro homologies between the two participating break point regions are illustrated by green vertical lines. Genomic coordinates are based on NCBI build 37.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480712&req=5

Figure 3: TP53 intron 1 rearrangement in a family with LFS(A) Pedigree of a family with LFS. Squares and circles represent males and females, respectively. Filled symbols indicate individuals with early onset cancer. Symbols with a diagonal line indicate that the individual was deceased at the time of recruitment. Diamonds represent several additional family members whose gender should not be disclosed. Individuals of whom DNA samples have been obtained are numbered with individuals tested positive for TP53 exon 1 deletion by MLPA in red. TP53 rearrangement carrier without cancer at the age of 10 years is indicated by a white symbol with a black dot. (B) PCR analysis of positive control and TP53 intron 1 rearrangement points (LFS-BP1 and LFS-BP2) of family members of whom high quality DNA was available. DNA quality of P12 did not allow PCR amplification, including positive control (not shown). a, LFS breakpoint 1 (BP1); b, LFS breakpoint 2 (BP2); c, positive control amplicon at the RNA polymerase II locus (POLR2A); sample IDs correspond to A with affected individuals in red. L, ladder. (C) Sanger sequencing of rearrangement points LFS-BP1 and LFS-BP2 at the TP53 locus of P2. Micro homologies between the two participating break point regions are illustrated by green vertical lines. Genomic coordinates are based on NCBI build 37.
Mentions: Since mutations of TP53 are associated with LFS it seemed possible that TP53 intron 1 rearrangements could constitute a previously underappreciated category of alterations that can cause LFS. We revisited an LFS family with 12 affected members with cancer across four generations in which we previously had been unable to identify a coding TP53 mutation or a co-segregating, potentially damaging and disease-causing alteration based on exome sequencing of patients P1, P2 and P13 (Figure 3A and data not shown). Copy number analysis of DNA from blood of patients P1 and P13 by single nucleotide polymorphism (SNP) array revealed evidence for an approximately 2.5 kb deletion in intron 1 of TP53 including exon 1 (Supplementary Figure S13 and Table S12). However, we were not able to amplify the fusion point of a deletion by PCR. We then used a custom sequence capture assay for targeted paired-end sequencing of the TP53 locus to search for a rearrangement point. We identified a 445 kb inversion spanning from the breakpoint cluster region in intron 1 of TP53 towards the centromere (upstream of TP53) with a loss of 2,275 bp of intron 1/exon 1 of TP53 (Figure 4A and 4B). The lost sequence was in agreement with the deletion identified by SNP arrays. In the adult family members tested, we found this specific rearrangement co-segregating with the disease, implicating the rearrangement as the causative alteration (Figure 3B and 3C).

Bottom Line: Using whole-genome sequencing of OS, we found features of TP53 intron 1 rearrangements suggesting a unique mechanism correlated with transcription.We revisited a four-generation LFS family where no TP53 mutation had been identified and found a 445 kb inversion spanning from the TP53 intron 1 towards the centromere.Cancers in this family had loss of heterozygosity, retaining the rearranged allele and resulting in TP53 expression loss.

View Article: PubMed Central - PubMed

Affiliation: Cancer Therapeutics & Stratified Oncology, Genome Institute of Singapore, Singapore 138672, Singapore.

ABSTRACT
Somatic mutations of TP53 are among the most common in cancer and germline mutations of TP53 (usually missense) can cause Li-Fraumeni syndrome (LFS). Recently, recurrent genomic rearrangements in intron 1 of TP53 have been described in osteosarcoma (OS), a highly malignant neoplasm of bone belonging to the spectrum of LFS tumors. Using whole-genome sequencing of OS, we found features of TP53 intron 1 rearrangements suggesting a unique mechanism correlated with transcription. Screening of 288 OS and 1,090 tumors of other types revealed evidence for TP53 rearrangements in 46 (16%) OS, while none were detected in other tumor types, indicating this rearrangement to be highly specific to OS. We revisited a four-generation LFS family where no TP53 mutation had been identified and found a 445 kb inversion spanning from the TP53 intron 1 towards the centromere. The inversion segregated with tumors in the LFS family. Cancers in this family had loss of heterozygosity, retaining the rearranged allele and resulting in TP53 expression loss. In conclusion, intron 1 rearrangements cause p53-driven malignancies by both germline and somatic mechanisms and provide an important mechanism of TP53 inactivation in LFS, which might in part explain the diagnostic gap of formerly classified "TP53 wild-type" LFS.

No MeSH data available.


Related in: MedlinePlus