A unique spectrum of somatic PIK3CA (p110alpha) mutations within primary endometrial carcinomas

Abstract

Purpose: The goal of this study was to comprehensively define the incidence of mutations in all exons of PIK3CA in both endometrioid endometrial cancer (EEC) and nonendometrioid endometrial cancer (NEEC).

Experimental design: We resequenced all coding exons of PIK3CA and PTEN, and exons 1 and 2 of KRAS, from 108 primary endometrial tumors. Somatic mutations were confirmed by sequencing matched normal DNAs. The biochemical properties of a subset of novel PIK3CA mutations were determined by exogenously expressing wild type and mutant constructs in U2OS cells and measuring levels of AKT(Ser473) phosphorylation.

Results: Somatic PIK3CA mutations were detected in 52.4% of 42 EECs and 33.3% of 66 NEECs. Half (29 of 58) of all nonsynonymous PIK3CA mutations were in exons 1-7 and half were in exons 9 and 20. The exons 1-7 mutations localized to the ABD, ABD-RBD linker and C2 domains of p110α. Within these regions, Arg88, Arg93, Gly106, Lys111, Glu365, and Glu453, were recurrently mutated; Arg88, Arg93, and Lys111 formed mutation hotspots. The p110α-R93W, -G106R, -G106V, -K111E, -delP449-L455, and -E453K mutants led to increased levels of phospho-AKT(Ser473) compared to wild-type p110α. Overall, 62% of exons 1-7 PIK3CA mutants and 64% of exons 9-20 PIK3CA mutants were activating; 72% of exon 1-7 mutations have not previously been reported in endometrial cancer.

Conclusions: Our study identified a new subgroup of endometrial cancer patients with activating mutations in the amino-terminal domains of p110α; these patients might be appropriate for consideration in clinical trials of targeted therapies directed against the PI3K pathway.

Figure 1. Distribution of somatic PIK3CA mutations in endometrial cancers and coexistence with PTEN and KRAS mutations

(A) Localization of PIK3CA mutations in EECs (orange triangles) and NEECs (blue triangles), relative to functional domains of p110α. Amino acid positions are indicated. (B) Patterns of PIK3CA, PTEN, and KRAS utations among 42 EECs (top) and 66 NEECs (bottom). Each column represents an individual tumor. The percentage of mutated tumors is indicated. Cases with a mutation in exons 1–7 of PIK3CA are indicted by a central dot.

Figure 2. A subset of amino terminal mutants of p110a is associated with increased AKT phosphorylation on Ser473

(A). Total protein from stably transfected, serum starved U2OS cells expressing vector, wild-type FLAG-p110α, or mutant Flag-p110α were analyzed by western blotting. Constructs expressing the known activating mutants, p110α-H1047R and p110α-E365K, served as positive controls for AKT phosphorylation (B) FLAG- p110α and p-AKT^Ser473 bands were normalized to Actin. The ratio of normalized p-AKT^Ser473 to normalized FLAG- p110α relative to wild-type p110α is shown.