Background
Forkhead box (FOX) proteins are evolutionarily conserved transcription factor family of proteins, which are characterized by their forkhead winged helix-turn-helix DNA binding domain composed of three α–helices and two loop or “wing” domains. Currently, more than 2000 members have been found in this family of transcription factors based on sequence homology, which are ubiquitously expressed across a range of species from yeast to human [
1,
2]. FOX proteins regulate a wide spectrum of biological processes involved in normal homeostasis and development [
3,
4]. Although the forkhead DNA binding domain with ~ 100 amino acid residues is highly conserved, the other domains are very divergent in FOX proteins. So they have very different binding specificities and cellular effects. According to additional domains and sequence conservation, FOX family is further grouped into various subfamilies, namely FOXM, FOXK, FOXA and FOXO families [
5‐
7].
The forkhead box class O (FOXO) family is a ubiquitously expressed transcription factor that plays important role in higher organisms. The first member of this family with
fork head was described in
Drosophila, which plays key roles in the terminal development of Drosophila embryo [
8]. The mammalian system consists of four members, namely FOXO1, FOXO3a, FOXO4, and FOXO6, which are known to be regulated by the phosphoinositol-3-kinase (PI3K)-PKB signaling pathway [
9‐
11]. FOXO family has been shown to regulate developmental processes and energy metabolism as well as tumorigenesis in many tissues. All these functions are mediated by the specific activation of a coordinated transcriptional program [
12]. The deregulation of FOXO functions will cause uncontrolled cell proliferation and accumulation of DNA damage, which results in carcinogenesis.
The member of FOXO subfamily, FOXO3a, also known as FOXO3 or forehead in rhabdomyosarcoma-like 1 (FKHRL1), was first identified in human placental cosmid. The
FOXO3a gene is located on chromosome 6q21 [
13] and it plays vital role in regulating a variety of cellular processes through targeting the expression and activity of effector genes. The subcellular localization of FOXO3a is important for its activities and functions [
14]. The phosphorylation of FOXO3a leads to its translocation from nucleus to cytoplasm, where it associates with 14–3-3 protein and this binding prevents its reentry into the nucleus [
15,
16]. In this review, we focus on the recent findings and important progress made in identification of FOXO3a functions and its target molecules and we have also presented an overview of the current understanding of the influence of FOXO3a activity on cancer.
FOXO3a as biomarker and therapeutic target in cancer
Currently, due to the physiological and anatomical features of tumor, it is difficult to observe obvious early symptoms of patients, leading to a large number of patients diagnosed at an advanced stage. Therefore, valuable biomarkers for early diagnosis and prognosis of cancer are required in clinical practice. FOXO3a has recently emerged as a potential biomarker for the diagnosis, prognosis and treatment of multiple malignant tumors. For example, FOXO3a expression is identified as a cancer-initiating cells biomarker in Hodgkin’s lymphoma [
119]. Many studies showed that FOXO3a expression acts as a prognostic biomarker in multiple cancers [
94,
98,
120‐
127]. Interestingly, overexpression of FOXO3a is associated with poor prognosis in triple-negative breast cancer [
120], hepatocellular carcinoma [
121], glioblastoma [
94] and gastric cancer [
122] patients, whereas low expression of FOXO3a is associated with poor prognosis in glioma [
126] and ovarian cancer [
127] patients. The expression of phosphorylated FOXO3a is also identified as a prognostic biomarker in ovarian cancer [
128] and acute myeloid leukemia [
129]. The nuclear localization of FOXO3a is demonstrated as a prognostic biomarker in luminal-like breast cancer [
130]. In addition, the subcellular localization of FOXO3a is identified as a biomarker for predicting response to the chemotherapy and radiotherapy in cervical carcinoma, breast cancer and esophageal cancer [
131,
132]. Although the potential value of FOXO3a as a biomarker has been established in small-scale studies, it is difficult to validate it in large cohorts of patients with cancer. Therefore, further large-scale studies on patient populations are required to confirm the utility of FOXO3a as a biomarker in cancer.
FOXO3a has become a potential target of chemotherapeutic drugs due to its central role in in carcinogenesis. Many chemical and pharmacological agents targeting FOXO3a have been tested in clinical as well as experimental settings. FOXO3 is an indirect target of BMS-345541 (a highly selective IKK inhibitor) in T-cell acute lymphoblastic leukemia (T-ALL) in which the expression of
p21Cip1 is up-regulation by increased nuclear translocation of FOXO3a after treatment with BMS-345541. This process is independent of PKB and ERK 1/2 signaling, which indicates that the loss of FOXO3a tumor suppressor function could be mainly due to overactivation of IKK [
133]. In BCR-ABL-positive chronic myeloid leukaemia cell lines, STI571 (also called imatinib or Glivec), an inhibitor of BCR-ABL oncoprotein, increases FOXO3a mediated apoptosis by triggering FOXO3a dependent cell cycle arrest and Bim expression [
134]. Epigallocatechin-3-gallate (EGCG), the major constituent of green tea, can induce apoptosis by targeting FOXO3a in pancreatic carcinoma [
135] and breast carcinoma cells [
136]. FOXO3a is also an indirect target of many anticancer agents including paclitaxel [
137], cisplatin [
138], imatinib [
139] and lidamycin [
140] in breast cancer cells. All these compounds activate FOXO3a by decreasing PKB activity. However, Paclitaxel also enhances JNK activity, which targets both FOXO3a and 14–3-3 proteins. JNK regulates the activity or stability of FOXO3a by phosphorylation, and this phosphorylation event additionally reduces its interaction with 14–3-3 proteins, which results in the nuclear export of FOXO3a.
The PI3K-PKB pathway is a major downstream signaling pathway of epidermal growth factor receptor (EGFR), which is a crucial cell surface receptor involved in cancer cell proliferation. Thus, the inhibition of EGFR by chemotherapeutic drugs (trastuzumab, lapatinib, afatinib, cetuximab, gefitinib and neratinib) provide a novel and valuable therapeutic strategy for treating breast, colon, prostate, ovarian, lung and head and neck cancers [
141,
142] by replenishing the activity of FOXO3a through inhibition of PI3K-PKB.
BNIP3L is a pro-apoptotic gene, which is required for chemosensitization of cancer cells. This gene is one of the targets of FOXO3a. In breast cancer cell lines, the blockade of EGFR by antibodies or small-molecule inhibitors induces nuclear translocation of FOXO3a and promotes the expression of
BNIP3L gene, which consequently results in apoptotic death of breast cancer cells [
143]. Knockdown of FOXO3a also promotes the response to cetuximab treatment in colorectal cancer [
144]. These findings indicate that FOXO3a could be a crucial target of small-molecule EGFR inhibitors, and its activity also increases chemosensitivity of cancer cells to agents such as lapatinib. In agreement with this, the activation of FOXO3a by other anticancer agents also sensitize cancer cells with resistance to apoptosis. For instance, FOXO3a transcriptional activity and its target gene
Bim expression level is increased in Saos2 (a p53-null osteosarcoma cell line) upon ionizing radiation, which indicates that FOXO3a is a crucial effector of radiation-inducing apoptosis [
100]. However, there is a drawback in therapeutically targeting FOXO3a for some type of cancers.
IGFR1 and
PI3KCA have been identified as target genes of FOXO3a in a colon carcinoma cell line [
115], which indicates that FOXO3a may activate PI3K–PKB signaling pathway by multiple mechanisms and it could contribute to drug resistance in colon cancer. However, the majority of studies have revealed that the activation of FOXO3a is highly associated with apoptotic pathway in tumor cells.
FOXO3a activity is directly regulated by a large number of miRNAs. This indicates that the screening or synthesis of novel chemotherapeutic drugs targeting these miRNAs may also be a valuable strategy to treat cancer. Although valuable progress has been made in FOXO3a-based therapeutics for cancer, the most important challenges such as the detailed mechanism of FOXO3a in sensitivity and resistance of chemotherapeutic drugs remain to be solved before its translation in to clinic.
Conclusions
FOXO3a is a core regulator of multiple physiological and pathological processes by directly inducing or mediating the expression of genes associated with cell proliferation, growth and survival. The deregulation of FOXO3a signaling significantly contributes to the development and progression of many disorders, including cancer. There is a complicated cross-talk between FOXO3a and other key signaling pathways (such as p53 and ER) involved in carcinogenesis. Therefore, FOXO3a is a valuable therapeutic target for a wide range of cancers. The unique role of FOXO3a in the carcinogenesis is that certain tissues offers exciting possibility for cancer-tissue-specific therapeutic strategies. Current studies have shown that FOXO3a targeted chemotherapy has lower toxicity in normal tissues compared with tumor tissues. In chemotherapy-resistant breast cancer cell lines, FOXO3a activation is vital for sensitizing cells to chemotherapeutic agents. ERα is a critical regulator in breast cancer development and it is an efficient target for endocrine therapy [
145]. The expression of ERα is considered as a marker for favorable prognosis and the level of functional ERα plays a key role in a successful endocrine treatment for breast cancer [
146]. It is well documented that FOXO3a and FOXM1 regulate the expression of ERα [
117]. Thus, FOXO3a could be a critical factor in determining the sensitivity and resistance of endocrine treatment. The PI3K-PKB signaling pathway is a relatively stable signaling pathway, which is not commonly mutated in cancers. Therefore, it is a promising strategy to identify novel inhibitors of FOXO3a for future anti-cancer drug design by targeting a downstream node of the PI3K-PKB pathway. As FOXO3a requires the recruitment of co-activators or suppressor for its activity or its inactivation, the therapeutic targeting of the coactivators or corepressors of FOXO3a could also be another way to manipulate FOXO3a functions in cancer cells. This strategy, along with therapeutic manipulation of PTM of FOXO3a would help to avoid the potential side effects in long term due to total inhibition of FOXO3a, which is required for normal cell functions. Given the fact that FOXO3a network is complex and considering its crosstalk with other transcription factors, the influence of FOXO3a in carcinogenesis need to be further investigated in order to develop an efficient FOXO3a based therapeutic strategies. The clinical applications of FOXO3a are potentially promising to limit the progression of human cancers in the future.