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Review
. 2019 Apr 23;9(4):158.
doi: 10.3390/biom9040158.

Mouse Models for Exploring the Biological Consequences and Clinical Significance of PIK3CA Mutations

Camilla B Mitchell  1 , Wayne A Phillips  2   3
Affiliations
Review

Mouse Models for Exploring the Biological Consequences and Clinical Significance of PIK3CA Mutations

Camilla B Mitchell et al. Biomolecules. .

Abstract

The phosphatidylinositol 3-kinase (PI3K) pathway is involved in a myriad of cellular signalling pathways that regulate cell growth, metabolism, proliferation and survival. As a result, alterations in the PI3K pathway are frequently associated with human cancers. Indeed, PIK3CA-the gene encoding the p110α catalytic subunit of PI3K-is one of the most commonly mutated human oncogenes. PIK3CA mutations have also been implicated in non-malignant conditions including congenital overgrowth syndromes and vascular malformations. In order to study the role of PIK3CA mutations in driving tumorigenesis and tissue overgrowth and to test potential therapeutic interventions for these conditions, model systems are essential. In this review we discuss the various mouse models currently available for preclinical studies into the biological consequences and clinical significance of PIK3CA mutations.

Keywords: PI 3-kinase; PI3K; PIK3CA; PROS; cancer; knock-in; mouse model; overgrowth; transgenic.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Schematic overviews of the transgenic p110α fusion models. (a) A simple transgene with a murine mammary tumour virus (MMTV) promoter upstream of a murine Pik3ca cDNA with a Src myristoylation sequence (MYR) added to the 5’-terminal end resulting in the expression of a myristoylated-p110α protein [30]. (b) A splice acceptor sequence (SA) and a loxP-flanked transcriptional stop cassette (STOP) upstream of a Pik3ca cDNA with a 5’-terminal myristoylation sequence (MYR), followed by an IRES-EGFP reporter element (an internal ribosome entry site (IRES) upstream of an enhanced green fluorescent protein (EGFP) cDNA), inserted into the ROSA26 gene locus. Upon expression of Cre recombinase (Cre) the stop cassette is excised allowing expression of the myristoylated-p110α and the EGFP reporter, under the control of the endogenous ROSA26 promoter [38]. (c) An α myosin heavy chain (MyHC) promoter-driven transgene containing a bovine PIK3CA construct with a 5’ sequence coding for the inter-SH2 domain (iSH2) of p85 separated by a glycine linker sequence, which generates a chimeric protein with p85 iSH2 fused to the N-terminus of p110α by a flexible linker (iSH2p110α*) [32]. (d) A splice acceptor sequence (SA) and a loxP-flanked transcriptional stop cassette (STOP) upstream of a p85 iSH2-linker-PIK3CA cDNA construct, followed by an IRES-GFP reporter element, inserted into the ROSA26 gene locus. Upon expression of Cre recombinase (Cre) the stop cassette is excised allowing expression of the iSH2p110α* protein and the GFP reporter, under the control of the endogenous ROSA26 promoter [33]. Triangles represent loxP sites. Blue colour indicates endogenous gene sequence, orange indicates modified PIK3CA constructs and purple indicates vector DNA sequence.
Figure 2
Figure 2
Schematic overviews of Pik3ca mutation specific transgenic models. (a) A mouse Pik3ca cDNA (modified to express the H1047R mutation), preceded by a splice acceptor sequence (SA) and a loxP-flanked transcriptional stop cassette (STOP), inserted into the ROSA26 gene locus. Upon expression of Cre recombinase (Cre) the stop cassette is excised allowing expression of mutant Pik3ca under the control of the endogenous ROSA26 promoter [40]. (b) A construct with a chicken β-actin (CAGS) promoter and a loxP-STOP-loxP sequence upstream of a 5’-terminally hemagglutinin (HA)-tagged human PIK3CA cDNA (wild type or modified to express H1047R or E545K mutations) and an IRES2-EGFP reporter element (an internal ribosome entry site (IRES) upstream of an enhanced green fluorescent protein (EGFP) cDNA), inserted into the ROSA26 locus. Expression of Cre recombinase results in the excision of the stop cassette allowing CAGS promoter-driven expression of the Pik3ca cDNAs and the green fluorescent protein (GFP) reporter [44,64]. (c) A DNA transgene consisting of seven direct repeats of the tetracycline operator (TetO) sequence followed by human PIK3CA cDNA containing a H1047R mutation. Reversible expression of the transgene requires the expression of a reverse tetracycline transactivator protein (rtTA) and is controlled by the administration of doxycycline (dox) [39]. (d) Similar to (C) but includes a downstream IRES and luciferase reporter gene [41]. Triangles represent loxP sites. Blue colour indicates endogenous gene sequence, orange indicates Pik3ca cDNA and purple indicates vector DNA sequence.
Figure 3
Figure 3
Schematic overviews of conditional knock-in PIK3CA mutation models. Triangles represent loxP sites. (A) Homologous recombination was used to insert a loxP-flanked transcriptional stop cassette (STOP) immediately upstream of the exon containing the initiation codon and replace exon 9 with an exon containing a E545K mutation, in one allele of the endogenous Pik3ca gene. Expression of the mutant allele is prevented by the stop cassette until removed by the expression of Cre recombinase (Cre) [63]. (B) A neomycin selection cassette (pgk neo) flanked by frt sites was inserted in the intron between exons 19 and 20 and point mutations resulting in a H1047R mutation introduced into exon 20, of one allele of the endogenous Pik3ca gene. The presence of the neo cassette suppresses the expression of the targeted allele until removed by Flp recombinase [65]. (C) LoxP sites were inserted on either side of the endogenous wild type exon 20 and a copy of exon 20 coding for a H1047R mutation inserted downstream of the endogenous stop codon (and after the downstream LoxP site) in one allele of the endogenous Pik3ca gene. The modified allele continues to express wild type protein (with expression of the inserted mutant exon prevented by the stop codon in the endogenous exon 20) until the expression of Cre recombinase which removes the wild type exon 20, replacing it with the mutated exon 20 [46]. (D) Similar to (C) but includes a transcriptional stop cassette downstream of the endogenous gene and before the inserted mutant exon [65]. (E) Point mutations coding for a E545K mutation were introduced into exon 9 of one allele of the endogenous Pik3ca gene and a loxP-flanked splice acceptor site (SA) and transcriptional stop cassette inserted into the intron immediately upstream of the mutated exon. Expression of the mutated exon is prevented by the stop cassette until removed by Cre recombinase [67]. (F) Similar to (E) but targeting exon 20 (H1047R) [68]. Triangles represent loxP sites and hexagons represent frt sites. Blue colour indicates endogenous gene sequence, orange indicates modified Pik3ca exon and purple indicates vector DNA sequence.
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References

    1. Vanhaesebroeck B., Guillermet-Guibert J., Graupera M., Bilanges B. The emerging mechanisms of isoform-specific PI3K signalling. Nat. Rev. Mol. Cell. Biol. 2010;11:329–341. doi: 10.1038/nrm2882. - DOI - PubMed
    1. Zhao L., Vogt P.K. Class I PI3K in oncogenic cellular transformation. Oncogene. 2008;27:5486–5496. doi: 10.1038/onc.2008.244. - DOI - PMC - PubMed
    1. Fruman D.A., Chiu H., Hopkins B.D., Bagrodia S., Cantley L.C., Abraham R.T. The PI3K Pathway in Human Disease. Cell. 2017;170:605–635. doi: 10.1016/j.cell.2017.07.029. - DOI - PMC - PubMed
    1. Thorpe L.M., Yuzugullu H., Zhao J.J. PI3K in cancer: divergent roles of isoforms, modes of activation and therapeutic targeting. Nat. Rev. Cancer. 2015;15:7–24. doi: 10.1038/nrc3860. - DOI - PMC - PubMed
    1. Marone R., Cmiljanovic V., Giese B., Wymann M.P. Targeting phosphoinositide 3-kinase—moving towards therapy. Biochim. Biophys. Acta Proteins Proteom. 2008;1784:159–185. doi: 10.1016/j.bbapap.2007.10.003. - DOI - PubMed

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