p53-PHLDA3-Akt Network: The Key Regulators of Neuroendocrine Tumorigenesis

Abstract

p53 is a well-known tumor suppressor gene and one of the most extensively studied genes in cancer research. p53 functions largely as a transcription factor and can trigger a variety of antiproliferative programs via induction of its target genes. We identified PHLDA3 as a p53 target gene and found that its protein product is a suppressor of pancreatic neuroendocrine tumors (PanNETs) and a repressor of Akt function. PHLDA3 is frequently inactivated by loss of heterozygosity (LOH) and methylation in human PanNETs, and LOH at the PHLDA3 gene locus correlates with PanNET progression and poor prognosis. In addition, in PHLDA3-deficient mice, pancreatic islet cells proliferate abnormally and acquire resistance to apoptosis. In this article, we briefly review the roles of p53 and Akt in human neuroendocrine tumors (NETs) and describe the relationship between the p53-PHLDA3 and Akt pathways. We also discuss the role of PHLDA3 as a tumor suppressor in various NETs and speculate on the possibility that loss of PHLDA3 function may be a useful prognostic marker for NET patients indicating particular drug therapies. These results suggest that targeting the downstream PHLDA3-Akt pathway might provide new therapies to treat NETs.

Keywords: Akt; PHLDA3; neuroendocrine tumor; p53.

Figure 1

PHLDA3 is a target gene of p53 and encodes a protein that functions as an endogenous inhibitor of Akt. (A) A schematic diagram of PHLDA3. Pleckstrin Homology (PH) domain is the only known protein domain in PHLDA3. (B) A model of Akt repression by PHLDA3. PHLDA3 functions as a dominant negative repressor of Akt. PHLDA3 localizes to the plasma membrane via binding to phosphatidylinositol phosphates (PIPs), thereby competing with Akt binding to PIPs and suppressing Akt-mediated survival signaling.

Figure 2

PHLDA3 gene defects are observed in lung and pancreatic neuroendocrine tumors (NETs). (A) Among lung NETs having wild-type p53, 63% exhibited loss of PHLDA3, whereas among lung NETs having a nonfunctional p53, only 13% exhibited loss of PHLDA3. Nonfunctional p53 can be caused by deletion as well as mutation. Abnormalities in p53 can result in upregulated expression of the protein, which may be detected by immunohistochemistry. Genomic sequencing is also preformed to analyze p53 mutations. Chromosome copy number alterations in PHLDA3 are analyzed by comparative genomic hybridization (CGH). (B) High frequency of PHLDA3 loss of heterozygosity (LOH) is found in pancreatic neuroendocrine tumors (PanNETs), which commonly have wild-type p53. (C) Two-hit inactivation of PHLDA3 in PanNETs. One of the PHLDA3 loci undergoes LOH and the other undergoes methylation. (D) PanNET tumorigenesis requires the functional loss of both PHLDA3 and MEN1. PHLDA3 and MEN1 suppress cell proliferation in normal islet cells. Loss of both PHLDA3 and MEN1 function is necessary for PanNET tumorigenesis. (E) Functional loss of PHLDA3 contributes to PanNET progression. Loss of MEN1 function leads to islet hyperplasia and/or atypia over time, and additional loss of PHLDA3 function is required for tumor formation and progression.

Figure 3

PHLDA3-deficient mice exhibit islet hyperplasia and resistance of β cells to apoptosis. (A) HE staining of islets from wild type and PHLDA3-deficient 10 month-old mice. (B) Apoptosis of β cells is observed in wild type mice treated with streptozotocin (STZ) but is suppressed in PHLDA3-deficient mice.

Figure 4

A model of NET tumorigenesis resulting from functional loss of both PHLDA3 and MEN1. PHLDA3 is a repressor of Akt. Loss of PHLDA3 leads to Akt activation, increased cell proliferation and decreased apoptosis. PHLDA3 functions independent of MEN1, and PanNET tumorigenesis therefore requires the functional loss of both PHLDA3 and MEN1.