Novel somatic mutations to PI3K pathway genes in metastatic melanoma

Abstract

Background: BRAF(V600) inhibitors have offered a new gateway for better treatment of metastatic melanoma. However, the overall efficacy of BRAF(V600) inhibitors has been lower than expected in clinical trials, and many patients have shown resistance to the drug's effect. We hypothesized that somatic mutations in the Phosphoinositide 3-Kinase (PI3K) pathway, which promotes proliferation and survival, may coincide with BRAF(V600) mutations and contribute to chemotherapeutic resistance.

Methods: We performed a somatic mutation profiling study using the 454 FLX pyrosequencing platform in order to identify candidate cancer genes within the MAPK and PI3K pathways of melanoma patients. Somatic mutations of theses candidate cancer genes were then confirmed using Sanger sequencing.

Results: As expected, BRAF(V600) mutations were seen in 51% of the melanomas, whereas NRAS mutations were seen in 19% of the melanomas. However, PI3K pathway mutations, though more heterogeneous, were present in 41% of the melanoma, with PTEN being the highest mutated PI3K gene in melanomas (22%). Interestingly, several novel PI3K pathway mutations were discovered in MTOR, IRS4, PIK3R1, PIK3R4, PIK3R5, and NFKB1. PI3K pathway mutations co-occurred with BRAF(V600) mutations in 17% of the tumors and co-occurred with 9% of NRAS mutant tumors, implying cooperativity between these pathways in terms of melanoma progression.

Conclusions: These novel PI3K pathway somatic mutations could provide alternative survival and proliferative pathways for metastatic melanoma cells. They therefore may be potential chemotherapeutic targets for melanoma patients who exhibit resistance to BRAF(V600) inhibitors.

Figure 1. Distribution of allele frequencies of non-synonymous and synonymous germline and somatic variants in melanoma patients.

(a) Allele frequencies of germline and somatic variants. The mean allele frequencies were: higher for germline variants (ns = 6.125, s = 7.412) than for the somatic variants, (ns = 2.685, s = 1.857). (b) Comparison of the ratios of non-synonymous to synonymous germline and somatic variants. Both were significantly different from the 2∶1 ratio expected by chance alone, but in opposite directions (somatic, binomial p = 1.15354×10⁻⁵; germline binomial p = 6.3953×10⁻¹²⁶).

Figure 2. Counts and ratios of variants discovered in each sequenced gene.

The absolute number of non-synonymous somatic variants for each gene is shown in (a) and the comparison of NS/S ratios for the germline and somatic variants of each gene is shown in (b). Blue shaded bars in (a) are non-synonymous mutations. The orange shaded bars are the NS/S ratios for the somatics variants, and the green shaded bars are the NS/S ratios for the germline variants.

Figure 3. Distribution of melanoma patients according to pathway somatic mutations.

(a) Percentages of melanomas that have: a BRAF^V600 mutation without a PI3K pathway mutation, a BRAF^V600 mutation with a PI3K pathway mutation, a NRAS mutation without a PI3K pathway mutation, a NRAS mutations with a PI3K pathway mutation, and a PI3K pathway mutation with wild-type BRAF^V600 and NRAS. (b) Number of patients who carried the somatic mutations identified by Sanger sequencing.