Next Article in Journal
Treatment with Soluble Activin Receptor Type IIB Alters Metabolic Response in Chemotherapy-Induced Cachexia
Next Article in Special Issue
Epigenetic Regulation of p21cip1/waf1 in Human Cancer
Previous Article in Journal
Hyper-Activation of STAT3 Sustains Progression of Non-Papillary Basal-Type Bladder Cancer via FOSL1 Regulome
Previous Article in Special Issue
p21 in Cancer Research
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Cancers
Volume 11
Issue 9
10.3390/cancers11091220
cancers-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Review

The Multifaceted p21 (Cip1/Waf1/CDKN1A) in Cell Differentiation, Migration and Cancer Therapy

by
Nina-Naomi Kreis
*,
Frank Louwen
and
Juping Yuan
Department of Gynecology and Obstetrics, University Hospital, J. W. Goethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt, Germany
*
Author to whom correspondence should be addressed.
Cancers 2019, 11(9), 1220; https://doi.org/10.3390/cancers11091220
Submission received: 19 July 2019 / Revised: 15 August 2019 / Accepted: 17 August 2019 / Published: 21 August 2019
(This article belongs to the Special Issue p21 – An Underestimated Driver for Cancers)
Browse Figure
Versions Notes

Abstract

:
Loss of cell cycle control is characteristic of tumorigenesis. The protein p21 is the founding member of cyclin-dependent kinase inhibitors and an important versatile cell cycle protein. p21 is transcriptionally controlled by p53 and p53-independent pathways. Its expression is increased in response to various intra- and extracellular stimuli to arrest the cell cycle ensuring genomic stability. Apart from its roles in cell cycle regulation including mitosis, p21 is involved in differentiation, cell migration, cytoskeletal dynamics, apoptosis, transcription, DNA repair, reprogramming of induced pluripotent stem cells, autophagy and the onset of senescence. p21 acts either as a tumor suppressor or as an oncogene depending largely on the cellular context, its subcellular localization and posttranslational modifications. In the present review, we briefly mention the general functions of p21 and summarize its roles in differentiation, migration and invasion in detail. Finally, regarding its dual role as tumor suppressor and oncogene, we highlight the potential, difficulties and risks of using p21 as a biomarker as well as a therapeutic target.

1. Introduction

p21, encoded by the gene CDKN1A, is the founding member of cyclin-dependent kinase (Cdk) inhibitors (CKI) [1], a pivotal cell cycle regulator ensuring genomic stability and often deregulated in human cancer [2,3,4]. Due to the lack of a defined tertiary structure, p21 is able to interact with a number of proteins involved in many key biological processes [5]. It is widely accepted that p21 acts through a “folding-on-binding mechanism” [6] with a prominent “binding promiscuity” [7]. As a critical effector of a variety of intra- and extracellular stress signals [8], p21 is exceedingly regulated by a myriad of transcriptional factors [9,10], distinct posttranscriptional regulators like microRNAs (miRNAs) and RNA binding proteins [11] as well as various posttranslational modifications. The short-lived p21 protein is degraded via different ubiquitin-dependent and -independent pathways [3,12]. Reversible protein phosphorylation by diverse kinases serves as an additional posttranslational mechanism controlling p21′s function, localization, binding partners, stability and degradation [3,13]. In particular, phosphorylation at distinct sites by various kinases can cause the cytoplasmic translocation of p21 greatly affecting its functionality and the therapeutic response of many chemotherapeutic interventions [2,3,4,14,15,16]. Importantly, p21 is not solely acting as a tumor suppressor but also as an oncogene depending on the cellular context leading to a negative or rather positive impact on tumor development and progression.

2. Functions of p21

2.1. p21 in General

The independent discovery of p21 by different groups leads to its numerous names [1,17,18,19,20,21], and reflects its multifaceted and heterogeneous cellular functionality. Rather than simply being a cell cycle inhibitor, senescence inducer and tumor suppressor, it is now appreciated as a much more complex and broader regulator [22]. It has additional key roles in apoptosis, differentiation, reprogramming of induced pluripotent stem cells, DNA repair, transcription and cell migration [2,3]. p21 is important for G2/M transition and mitotic progression, its loss prolongs mitosis resulting in mitotic defects and possibly in genomic instability [23,24]. p21 maintains chromosomal integrity in vivo [25]. As “linchpin” [26] or “molecular switch” [27] of cell fate, p21 is tightly involved in balancing and coordinating proliferation with all other cellular processes.
In general, p21 is an important regulator of cell cycle checkpoints ensuring proper cell division. p21 inhibits cell proliferation directly through binding to Cdks [1,18] and proliferating cell nuclear antigen (PCNA) [28], or indirectly at the transcriptional level [29,30,31]. Besides p53 [17], plenty of oncogenes, tumor suppressors, inflammatory cytokines and nutrients can initiate p21 transcription [9]. Interestingly, myelocytomatosis oncogene cellular homolog (c-Myc) has been shown to inhibit p21 gene expression [32] and, beyond that, it is able to interact with p21 disrupting its interaction with PCNA and causing a decrease in DNA synthesis inhibition [33]. The protective function of p21 is therefore disabled in tumor cells with high c-Myc levels [8]. In response to noxious environmental stimuli, p21 participates in the checkpoint control and initiates a temporary cell cycle arrest [34,35]. Using live single-cell measurements it has recently been shown that the amount of p21 and the time period of its increased expression contribute to the decision between cell cycle arrest and the repair of incomplete replicated DNA maintaining genomic stability [36].
Intriguingly, contradictory to its function as an inhibitor, low concentrations of p21 promote proliferation through the assembly and activation of cyclin D/Cdk4 or Cdk6 complexes in cancer cell lines [37]. This is supported by another study showing that primary mouse embryonic fibroblasts (MEFs) lacking p21 and its family member p27 failed to assemble detectable amounts of cyclin D/Cdk4 complexes [38]. Furthermore, hyper-phosphorylated p21 activates Cdk1 at G2/M transition [39]. Hence, the p21-describing term “cell cycle inhibitor” is a “confusing misnomer” [14]. The data highly suggest that p21 is not only an inhibitor but also an activator of the cell cycle depending on the cellular context and its expression levels.
Additionally, several studies revealed that p21 is involved in the major DNA repair pathways, which is attributed to its high PCNA binding capacity [28]. However, p21’s function in DNA repair is contradictory: p21 is reported to be inhibitory [40,41,42], unnecessary [43,44] or even required [45,46,47], mostly depending on the type of lesion/repair mechanism, the experimental setup, the cell system or/and the extent of DNA damage. A model was proposed in which the balance between p21 expression and DNA damage is crucial for the decision between DNA repair or rather apoptosis induction [48]. Low DNA damage tends to provoke a p21-dependent inhibition of apoptosis enabling DNA repair, whereas severe DNA damage causes the induction of apoptosis [48], probably through cleavage of p21 by caspase-3 [49,50].
The “antagonistic duality” of p21 [51] is particularly reflected in cell death mechanisms: p21 is regarded as a modulator of apoptosis [51,52] and is involved in macro-autophagy [53]. Cytoplasmic p21 leads to an inhibition of multiple caspases and apoptotic effectors including pro-caspase-3, caspase-8, caspase-10, apoptosis signal-regulating kinase 1 and stress-activated protein kinase [2,3]. Paradoxically, p21 can also promote apoptosis, as summarized in [52]. Notably, p21 activates autophagy to accelerate cell death in cancer cells [54,55], while it inhibits autophagy and induces apoptosis under death stimuli in normal cells [56]. Beyond that, p21 induces a senescent arrest in normal and tumor cells independent of p53 [57,58,59], identified through its overexpression in senescent human diploid fibroblasts [19].
In essence, p21 does not work solely as a tumor suppressor preserving genomic stability, permitting cell cycle arrest or senescence and enabling DNA repair. Meanwhile, there is increasing awareness of its oncogenic potential, often attributed to its cytoplasmic localization [3], promoting cell cycle progression, favoring migration [60,61] and inhibiting apoptosis [51], besides p21’s sustained (over)expression leads to bypassing/escaping from senescent cell arrest [62]. Its dual behavior in various processes often leads to opposing cellular responses (Table 1).
Of note, the individual roles of p21 should not be viewed in isolation. Instead, its multifaceted functions are interconnected and reciprocally influenced by each other.

2.2. p21 in Differentiation and Stem Cell Renewal

From its cDNA isolation, p21 is associated with differentiation [20]. Using subtractive hybridization, p21 was isolated based on its increased expression in human melanoma cells that were induced to differentiate, therefore, p21 was named melanoma differentiation-associated gene 6 (mda6) [20]. Given that cell proliferation and differentiation show a notable inverse relationship, their temporal coupling is crucial for growth and development, and is critical for tissue homeostasis [68]. Cell cycle arrest or rather withdrawal is a prerequisite and the first step towards terminal differentiation, and into quiescence or senescence, often accompanied by an increased p21 expression. Activation of p53, the major activator of p21 transcription [17], leads to cell cycle arrest via the suppression of cell cycle genes by the dimerization partner, RB-like, E2F and MuvB (DREAM) complex [29]. p21′s transcription is also activated during differentiation induced by p53-independent mechanisms through various agents including phorbol ester, okadaic acid, vitamin D3, tumor necrosis factor-α, myoblast determination protein, nerve growth factor together with aphidicolin, calcium, interleukin-6 or interferon-α [9].
Several lines of evidence indicate that p21 plays either a positive or negative role in differentiation depending on the cell type and the stage of differentiation [69]. Overexpression of p21 is able to induce cell differentiation in a variety of normal and tumor cells mediated by the induction of cell cycle exit [8]. Supporting its positive role in differentiation of normal cells, p21 promotes/establishes differentiation of murine oligodendrocytes [70], normal human peripheral blood monocytes [63], human and mouse endomitotic megakaryocytes [71], normal erythroid progenitors including reprogramming of murine erythroleukemia cells MEL [72] and murine C2C12 skeletal muscle cells [73,74,75]. Whereas muscular differentiation is impaired in p21-knockout mice [76], murine p21 expression correlates with cell cycle arrest and terminal differentiation of multiple cell lineages including skeletal muscle, cartilage, skin and nasal epithelium during mouse embryogenesis and adult tissue independent of p53 [77]. A rapid and transient increase of p21 was also observed in a temperature-sensitive fetal human intestinal (tsFHI) epithelial cell line, where it is involved in early stages of differentiation [78]. Furthermore, p21 has been shown to have an important but quite contradictory role in murine adipocyte differentiation: its knockdown in 3T3-L1 cells or its ablation in MEFs reduces adipocyte differentiation [79], suggestive of its pro-adipogenic role, while mice lacking p21 display adipocyte hyperplasia and obesity [80], implicative of its anti-adipogenic function. Moreover, murine stabilized nuclear p21 conveyed via phosphorylation at Thr-55 by murine protein serine-threonine kinase 38 (MPK38) inhibits adipocyte differentiation [81]. In line with this negative role, p21 inhibits differentiation of 32Dcl3 murine granulocytes [64]. It is strongly downregulated during differentiation of primary osteoblasts and osteoblastic cell lines derived from p21 null mice [82] and its expression decreases in terminally differentiated primary keratinocytes of mice [83]. Interestingly, p21’s expression elevates at the onset of keratinocyte differentiation [84] as reported for chondrogenic differentiation, in which p21 was increased at early stages of chondrogenesis and degraded in the later stages of murine ATDC5 cell differentiation [85].
Moreover, p21’s cytoplasmic localization plays a positive role for normal cell differentiation: the cytoplasmic residing of p21 was observed in mature human monocytes [63] and rat neurons [86], inhibiting apoptosis [63] or promoting neurite outgrowth through loss of stress fiber formation [86], respectively. Additionally, cytoplasmic translocation of p21 correlates with rat pancreatic myofibroblast to fibroblast conversion and protects against apoptosis [87]. During murine C2C12 myoblast differentiation, the appearance of the apoptosis-resistant phenotype is correlated with p21 induction [74] and its cytoplasmic localization in mature myofibrils through phosphorylation at Ser-153 [75]. These data strongly highlight the notion that p21 is tightly involved in normal cell differentiation, whether it confers a positive or negative role depends on a number of factors including its expression level, postmodification, cell type, differentiation stage and cell microenvironment (Table 2).
High attention has been paid to the link between p21 and differentiation/renewal of stem/progenitor cells. Interestingly, the reprogramming efficiency was highly increased upon depletion of p53 or p21 in MEFs [65,66]. The p53-p21-axis thus acts as a barrier and protector of induced pluripotent stem (iPS) cell generation since its silencing facilitates the reprogramming even in the presence of DNA damage [65,66,67]. In leukemic stem cells p21 prevents DNA-damage accumulation and exhaustion [88]. Increased p21 accelerates the differentiation of murine embryonic stem cells (ECSs) by repressing the pluripotency factor sex determining region Y-box 2 (SOX2) [89]. p21 is also indispensable for maintaining self-renewal of quiescent hematopoietic stem cells [90], keratinocyte stem cells [91] and neuronal stem cells [92] of mice. Therefore, inhibition of p21 results in the depletion of leukemic or hematopoietic stem cells, which is, depending on the cellular context, either positive or negative for the malignant progression [8]. Overall, basal p21 expression is required for stem cell maintenance protecting the cells from exhaustion, whereas high levels of p21 trigger differentiation and limit self-renewal of adult stem cells [93]. In addition, an up-regulation of cytoplasmic p21 accompanies differentiation of proliferating murine trophoblastic stem cells into giant cells, which are resistant to DNA damage induced apoptosis [94]. In contrast, studies have shown that human mesenchymal stem/progenitor cells (MSC) deficient in p21 have increased potential to differentiate into adipocytes, osteoblasts or chondrocytes [95,96,97], partially explained by a reduction of their replicative senescence [96,97]. Altogether, p21 is tightly linked to differentiation of various stem/progenitor cells (Table 2).
p21 is also involved in human cancer cell differentiation and development. In particular, epithelial–mesenchymal transition (EMT) and its reverse form mesenchymal–epithelial transition (MET) are crucial differential processes in cancer progression. While the activation of EMT permits cancer cells to acquire migratory, invasive and stem-like properties, MET is associated with the tumor-initiating ability required for metastatic colonization [98]. Markedly, p21 has been shown to attenuate EMT in cell lines and in mouse models: p21 inhibits EMT in colorectal cancer cells and normal human mammary epithelial cells (MCF10A) either in complex with zinc finger E-box binding homeobox 1 (ZEB1) through distinct miRNA clusters [99] or in response to transforming growth factor-β (TGF-β) [100]. p21 represses Ras- and c-Myc-dependent stem cell renewal and breast tumor EMT in mice [101]. Consistently, repression of p21 promotes EMT in response to long non-coding RNA plasmacytoma variant translocation 1 (PVT1) in distinct triple-negative breast cancer cell lines [102] as well as in cooperation with p53-upregulated modulator of apoptosis (PUMA) in MCF10A cells [103]. Silencing of p21 enhances the features of EMT in MEC cell lines, which are immortalized from the peripheral blood of a patient with B-chronic lymphocytic leukemia [101].
In further supporting p21’s role in cancer cell differentiation, the expression of p21 increases in various human malignant cell lines including lymphoma-derived cell line U937 [104,105,106], p53-null promyelocytic human leukemia (HL) cells HL-60 [107,108,109,110], acute myeloid leukemia (ML) cell lines ML-1, ML-2 and ML-3 [111], megakaryoblastic leukemia cell line CMK [112], epithelial colorectal adenocarcinoma cells Caco-2 [113], neuroblastoma cell line SH-SY5Y [114,115] and chondrosarcoma cells SW1353 [116], sometimes associated with its cytoplasmic localization [63,117]. In support of this, increased p21 was also observed in primary tumors from lung adenocarcinomas [118] and squamous cell carcinomas of the larynx [119]. Interestingly, p53-independent p21-overexpression is correlated with head and neck cancer differentiation [120]. In contrast, there is also evidence showing that p21 has an inhibitory role in cancer cell differentiation expanding its “antagonistic duality” [121] to this field. Stably overexpressed p21 displays decreased sensitivity to differentiation induction of human colon cancer cell line HT29 [122].
Loss of differentiation control can lead to a variety of diseases including cancer [68]. The entire evidence indicates that p21 plays a crucial role in differentiation of various normal or malignant cells and tissues but its impact relies on the cell type and the stage of differentiation. Of note, in the vast majority of studies, p21 has a positive role in differentiation; it initiates and/or is increased during cellular differentiation, partially by inhibition of apoptosis favoring cell survival. Unfortunately, it is not always clear whether the cells are terminally differentiated, in a quiescent or senescent state, since all these states are preceded by a G1 arrest mediated by p21 induction.

2.3. p21 in Migration and Invasion

Cell migration is a tightly integrated process orchestrating embryonic morphogenesis and contributing to tissue homeostasis [123]. Defects in cell motility are associated with pathological processes including metastasis responsible for over 90% of cancer associated deaths [124]. The role of p21 in cell motility is multifaceted depending on the cellular context. p21’s contribution to cell motility ascribes largely to its cytoplasmic localization, induced predominantly by phosphorylation of its distinct sites via various kinases [3,13]. Cytoplasmic p21 elicits an inhibition of the Ras homolog gene family, member A (RhoA)/Rho-associated, coiled-coil containing protein kinase (ROCK)/LIM domain kinase (LIMK)-pathway by binding to ROCK, increases cofilin expression and cytoskeleton remodeling due to a loss of actin stress fibers favoring cell motility in Ras-transformed MEFs [60,61]. The cytoplasmic accumulation of p21 requires firstly the oncogene Ras inducing its transcription [125] and secondly the phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB, also named Akt)-pathway for its cytoplasmic retention [60]. Inhibition of PI3K, an upstream kinase of PKB/Akt mainly responsible for phosphorylation and cytoplasmic residing of p21 [3,126], causes the restoration of cofilin phosphorylation and a loss of cytoplasmic p21 [60]. Cytoplasmic p21 is also involved in neuronal differentiation regulating Rho-induced actin remodeling, which induces neurite outgrowth through suppression of actin stress fibers [86]. Interestingly, cytoplasmic p21 is targeted and degraded by Cullin-2 RING ubiquitin ligase with a leucine rich repeat 1 protein (CRL2LRR1) acting as a critical regulator of cell motility promoting a non-motile, stationary cell state by preventing p21 from inhibiting the RhoA/ROCK/LIMK pathway [127]. Knocking down Cullin-2 or LRR1 selectively elevates the protein level of p21 reducing stress fibers and cell-cell connections and raising F-actin on the cell periphery and cell motility in HeLa cells [127]. These morphological adjustments are suppressed by co-depletion of p21 [127]. Moreover, the accumulation of cytoplasmic p21 and p27 was linked to the oncogenic transformation of cells by activated Ras [60,128] or HER2/neu [129,130,131]. Importantly, cytoplasmic p21 has an oncogenic role in promoting mammary tumorigenesis and metastasis downregulating E-cadherin in vivo [132].
Oppositely, the complex of cytoplasmic p21 together with ectopic expressed cytosol-resided p53 (mutation in its nuclear localization sequence) suppresses cell invasion due to favoring apoptotic induction by targeting B-cell lymphoma 2 family proteins in non-small cell lung cancer cell line H1299 [133]. It has also been shown that the complex of p21 with wild type p53 induces Slug degradation further suppressing invasion [134]. In consistence with this observation, mutant p53 leads to Slug accumulation and increased cancer cell invasiveness [135].
p21 functioning as transcription factor/co-factor is essential for TGF-β mediated breast cancer cell migration and invasion, whereas its gene silencing blocked the tumor invasion in a mammary fat pad xenograft mouse model and various triple negative breast cancer cell lines, without alterations in cell growth and proliferation [136]. In this study high p21 expression was correlated with poor overall and distant metastasis free survival of breast cancer patients promoting migration/invasion at the transcriptional level [136]. Moreover, the complex of nuclear p21 and cyclin D1 is involved in actin remodeling of TGF-β-induced cell migration [137]. In accordance with this, another study with a breast cancer mouse model has shown that invasion is accompanied by an upregulation of p21 pointing to its role in a “reciprocal switching between proliferation and invasion” [138]. Together with p16, p21 promotes tumor growth in mice by enhancing the chemotaxis of monocytic myeloid-derived suppressor cells as observed with double-knockout mice [139]. Suppression of p21 compromises the migration and invasion capability of various trophoblastic and cancer cell lines mediated by, at least partially, a reduction of the extracellular signal-regulated kinase 3, matrix metalloproteinase 2 and tissue inhibitor of metalloproteinases 2 [140]. p21 knockout mice evince a dramatic suppression of metastasis, independent of tumor growth and rescuable by re-expression of p21 [138]. The long non-coding RNA small nucleolar RNA host gene 20 (SNHG20) represses p21 in non-small lung cell cancer cells A549 facilitating proliferation and migration [141]. Nuclear p21 inhibits but cytoplasmic p21 promotes migration and invasion in gastric cancer cell line AGS [142]. Recently, Galanos et al. utilized inducible p21-expressing p53-null osteosarcoma Saos2 cells and Li-Fraumeni patient-derived fibroblasts to study the impact of its sustained expression [62]. After 10 days induction, a population of cells bypassed senescence with increased genomic instability, chemoresistance and aggressiveness ascribed to enhanced invasiveness, partially explained by transcriptional upregulation of matrix metallopeptidases and “stemness-like features” [62]. The sustained p21-expression provokes radiation sensitive 52 (Rad52)-dependent error-prone double strand break repair promoting genomic instability [143].
The role of p21 in migration/invasion is associated with the environmental and cellular context, which is not unexpected, considering the reciprocal impact between versatile factors and p21.

3. Translating in Tumor Therapy

3.1. p21 and Its Deregulation in Cancer

p21 is often deregulated in human cancer. Depending on the cellular context, p21 promotes or inhibits tumorigenesis, extensively summarized by Abbas and Dutta [2]. p21 has been regarded as a tumor suppressor by regulating the cell cycle and maintaining genomic stability [2,3,4]. Though mutations in the p21 gene are regarded as extremely rare [4], the data from The Cancer Genome Atlas suggest that p21 is not infrequently mutated in bladder cancer (Philip Abbosh, Fox Chase Cancer Center) [144]. Some epigenetic alterations have also been reported [15]. Hypermethylation and silencing of p21 were observed in non-small cell lung [145] and prostate [146] cancer cells, high grade breast cancer [147] and acute lymphoblastic leukemia associated with poor prognosis [148]. Epigenetically silencing through long non-coding RNAs happens frequently [149] and was recently detected in cholangiocarcinoma [150]. Additionally, diverse single nucleotide polymorphisms of p21 are reported to influence the risk of developing cancers including esophageal [151], colorectal and estrogen-related cancer [152], associated with a higher risk of second primary malignancies in head and neck carcinoma [153].
p21 is often decreased in human cancer through loss of functional p53 or hyperactive oncogenes like c-Myc: p53 mutations have been discovered in more than 50% of human malignancies [154], whereas c-Myc, p21’s transcriptional repressor [32], is often overexpressed in human cancer [155]. Both drive p21 downregulation affecting its tumor suppressive activity and being linked to poor prognosis of patients with colorectal cancer, non-small-cell lung carcinoma, breast, gastric and ovarian cancers [2] as well as pancreatic cancer [156]. In consistent with these observations, p21 (over)expression has been correlated with a favorable prognosis in tonsillar carcinoma, gastric cancer, cervical adenocarcinoma, pancreatic cancer, as well as laryngeal and oral carcinoma [2].
In line with p21’s “antagonistic duality” [51], its overexpression leading to its oncogenic activity is found in a variety of human cancers including breast cancer, renal cell carcinoma, testicular cancer, hepatocellular carcinoma, multiple myeloma, gliomas, prostate cancer, cervical carcinoma, ovarian cancer, acute myeloid cancer, esophageal squamous cell carcinoma and soft tissue sarcomas [2,157]. Cytoplasmic p21 is observed in breast cancer [158,159] and hepatocellular carcinoma [160]. Recently, the role of p21 in gastric cancer samples was investigated and linked to its localization [142]. High cytoplasmic p21 level was positively associated with advanced tumor/lymph node/metastasis (TNM) stage, invasion depth, lymph node metastasis, distant metastasis and shorter overall survival of over 800 patients [142]. Cytoplasmic expression of p21 is acting as an oncogene with “anti-apoptotic gain-of-function” pointing to a role in tumorigenesis [161,162]. This residing appears frequently in human tumors, which was linked to aggressiveness as well as poor prognosis [2,4,14], and affected the response to chemo- and radiotherapy leading to drug resistance [15]. Taken together, the upregulation of p21 and its frequently cytoplasmic relocation correlate positively with poor prognosis, tumor grade, invasiveness and/or drug resistance. Despite the contradictory roles of p21 in the development of human cancer, increased cytoplasmic p21 is generally regarded as being tumor-promoting.

3.2. Lessons from Mouse Studies

Although p21 knockout mice were initially described to remain tumor-free until the age of seven months [163], a subsequent study showed cancer development at an average age of 16 month [164]. Evidence for classifying p21 as tumor suppressor came from additional deletion mouse studies. p21 knockout increased the rate of tumorigenesis in retinoblastoma protein [165] and adenomatous polyposis coli-haploinsufficient [166] and p18-deficient [167] transgenic mice, as detailed in [2]. Loss of p21 together with a mutation in p53 (p53R172P, preventing cells from apoptosis) accelerated the tumor onset [25]. In the context of mouse mammary tumor virus (MMTV)-Ras, p21 knockout also increased the tumor onset [168].
Conversely, p21 depletion in a c-Myc background decreased the overall tumor onset [168], and it reduced radiation-induced tumor development in wild type [164] and ataxia-telangiectasia mutated-deficient mice [169]. The absence of p21 resulted in a significant extension of the lifespan of p53-null and p53-haploinsufficient mice [170]. This effect can be attributed exclusively to a decrease in the incidence of spontaneous and radiation-induced thymic lymphomas [170] supportive of an oncogenic role of p21 in thymic lymphoma. Moreover, Cdkn1a;Puma;Noxa triple-knockout mice did not develop spontaneous tumors until the age of 500 days, suggesting the existence of additional pathways serving as mediators of p53-driven tumor suppression [171]. In sum, loss of p21 has differential effects on tumorigenesis based on the specific cellular context and the genetic background.

3.3. Considering p21 in Cancer Therapy

Given p21’s “antagonistic duality” [51] in various cellular processes (Table 1), it is obvious that p21 can have a dual role in tumor development and progression relying on the cancer type, the p53 status and the used chemotherapeutics. It can serve as a biomarker for specific therapies or prognosis, partially depending on its subcellular localization. In fact, the induction of p21 has been used as a drug response parameter [16].
Simply interfering with p21 as anti-cancer therapy bears risks and undesired side effects. First, increasing p21 can cause senescence, a supposed permanent growth arrest [172], which was believed to be only tumor suppressive terminating tumor regression, and is now regarded as a tumor promoter [173]. Senescent cells secrete numerous soluble factors promoting tissue repair, invasiveness of neighboring cells, chronic inflammation and tumor progression [174], and contribute to the escape of drug-induced apoptosis [175]. We examined the therapeutic potential of p21 in the context of Poloxin, a well-studied Polo-like kinase 1 (Plk1) inhibitor [176,177]. Plk1, a highly conserved serine/threonine kinase with critical roles during mitosis, is overexpressed in various tumor entities serving as a poor prognostic marker [178] and is thus considered as a promising target for molecular cancer therapy [179]. Cancer cells without p21 showed a stronger mitotic arrest accompanied by proliferation inhibition, more DNA damage and apoptosis induction upon Poloxin treatment relative to cancer cells with functional p21 and p53, which displayed a cytoplasmic re-localization and an anti-apoptotic feature [176]. Interestingly, long-term treatment (four days) of HCT116 p21+/+ cells with Poloxin led to senescence, whereas strong apoptosis induction was observed in cells lacking p21 [48]. Similar effects were detected in HCT116 cells treated with low doses of the anti-cancer drug camptothecin for four days [180]. Conversely, there are studies where overexpressed p21 enhanced the apoptotic response upon cisplatin treatment [181,182,183]. Remarkably, cellular senescence contributes to therapy resistance [184] and an aggressive tumor relapse by undergoing an epigenetic reprogramming of senescent cells into a stemness-like state [173,185]. In support of this observation, sustained expression of p21 exhibits oncogenic functions in a p53-null background leading to escaping senescence and chemoresistance [62]. Cells bypassing senescence display an increased genomic instability pointing again to p21’s “two-faced involvement” as genome guardian versus genomic instability mediator [186]. This duality is commonly attributed to the cellular or environmental context in which tumors develop. The mechanistic basis underlying such context-dependent phenomena remains to be defined in most cases, and its elucidation is essential for both understanding cell biology and the rational design of cancer therapy [187]. Thus, for therapeutic approaches, simply increasing p21 may not be beneficial and could provoke opposite undesirable/unintended outcomes.
Second, considering p21’s role in the cell cycle, stem cell differentiation and EMT of tumor cells, depleting p21 may result in either tumor suppressive or oncogenic effects depending on the cellular context. Cancer stem cells have been suggested to promote tumorigenesis as “seeds for metastasis” [188]. p21 is indispensable for maintaining self-renewal of leukemia stem cells [88], and it is able to inhibit oncogene-induced EMT and breast tumor stem cells in transgenic mice [101]. In a study with five patient-derived glioma stem cell-enriched cell lines, the authors have reported that p21 and p27 operate both as tumor suppressors, limiting cell proliferation, but also as oncogenes, conferring cell resistance to DNA damage and developing drug resistance [189]. Further investigations are mandatory to delineate if suppression of p21 serves as a therapeutic target, though its antisense therapy radiosensitizes human colon cancer cells [190] and glioma cells [191] to apoptosis.
Third, targeting cytoplasmic p21 is highly promising, as it contributes to tumor cell survival by inhibiting apoptosis, promoting cell migration and causing drug resistance [3,14,16,157]. Cytoplasmic expression of p21, which is common in human tumors, is linked to aggressiveness and poor prognosis [2,4]. The cytoplasmic residing of p21 is provoked by different kinases at distinct phosphorylation sites (Thr-57 [192], Ser-130 [192], Thr-145 [126], Ser-146 [193] and Ser-153 [75,194]), as summarized in [3]. Accordingly, increased cytoplasmic p21 due to Akt phosphorylation at Thr-145 favors tumor progression, drug resistance and poor prognosis [126,195,196,197]. Cytoplasmic re-localization of p21 by Akt-induced phosphorylation in HER2/neu overexpressing cells was correlated to apoptosis resistance [126]. Blocking the Akt pathway results in nuclear localization of p21 restoring its function as cell cycle inhibitor [126]. A recent study has shown that phosphorylation of p21 at Thr-145 and Ser-146 by Akt enhances cell survival contributing to taxol resistance in glioblastoma cell lines [195]. Cytoplasmic p21 is also associated with poor response to tamoxifen treatment in breast cancer cells MCF7 [198]. Further strong evidence comes from a recent investigation of the impact of cytoplasmic p21 on tumorigenesis in vivo [132]: MMTV/neu mice expressing cytoplasmic p21 (T145D, mimicking the Akt phosphorylation) in the mammary epithelium had an accelerated tumor onset as well as enhanced lung metastasis indicating an oncogenic role of p21 [132]. Moreover, cytoplasmic p21 induced by Akt-dependent phosphorylation enhances chemoresistance in response to doxorubicin in triple-negative breast cancer cells SUM159 [196]. Increased cytoplasmic p21 is related to cisplatin resistance in testicular and ovarian cancer [199,200] and to failure of paclitaxel treatment in human nasal squamous carcinoma RPMI-2650 [197]. Additionally, cytoplasmic p21 mediates 5-fluorouracil resistance in colorectal cancer cells [201]. A model for cancer aggressiveness are the human mantle cell lymphoma cell lines REC1, G519 and JVM2, where p21 increases its level along with disease progression and is localized to the cytoplasm [202]. Interestingly, cytoplasmic p21 is locally degraded by CRL2LRR1 and the LRR1 gene is located at chromosome 14q21.3, which is frequently lost in metastatic cancer including breast cancer [127,203]. Accordingly, cytoplasmic localization of p21 is associated with anti-apoptotic properties and therapy resistance, which leads to a worse overall survival of patients suggesting p21’s prognostic importance.
Collectively, cytoplasmic localization of p21 could be a reliable biomarker and a promising intervention target. General targeting p21 will be associated with undesirable effects, regarding its multifaceted functions in stress-responsive senescence induction, apoptosis control, cell cycle regulation, genome integrity and stem cell maintenance.

3.4. Concepts of Targeting p21

Given its differential expression and localization in human cancer, p21 could be targeted by three approaches [16]: first, increasing p21 expression, with histone deacetylase inhibitors [204] or through restoring p53 activity [205,206] as well as interfering with p21’s degradation using proteasome inhibitors [207], promotes its function as tumor suppressor and inhibits tumor growth. Second, down-regulation of p21 could be considered in malignant cells with highly expressed p21, especially with sustained p53-independent expression. Techniques of choice are antisense oligonucleotides (ASOs) [14], synthetic miRNA mimics [208,209] or prospectively a targeted therapy using the clustered regularly interspaced short palindromic repeats (CRISPR) technique [210]. ASOs targeting p21 has been already applied in breast cancer [211,212], colorectal cancer [190], glioma [191], myeloid leukemia [110] and renal cell carcinoma cell lines [213] showing growth arrest and/or apoptosis. Given their lacking efficacy and delivery options, transient expression and off-target effects, it is difficult to translate ASOs or siRNA based therapies into the clinic [157,214]. Novel iron chelators are also able to decrease p21 expression with anti-proliferative effects [215]. Few inhibitors targeting p21 are known including the Cdk inhibitor butyrolactone [216] and the small-molecule LLW10 [217], both induce p21’s proteasomal degradation. Due to the poor specificity targeting p21, these inhibitors are probably not the best choice for a clinical application. Alternatively, a possible interfering strategy would be to reduce its nuclear oncogenic properties by blocking its PCNA binding with small molecules like T2AA [218], by blocking error-prone DNA repair [219] or by the targeted elimination of therapy-resistant senescent cells with senotherapeutics including senolytics and senoptotics [220] (Figure 1A). Third, another promising alternative is to target cytoplasmic p21 or interfering with its nuclear-cytoplasmic shuttling (Figure 1B).
Currently, the best approach would be to target cytoplasmic p21, to inhibit its anti-apoptotic properties and to sensitize chemotherapy resistant cancer cells, as demonstrated in renal cell carcinoma [157] or breast cancer [14]. Interestingly, the small molecule UC2288 [221], which was synthesized based on the chemical model of sorafenib, the multi-kinase inhibitor [222], attenuates p21 at the level of transcription and posttranscription, and subsequently decreases cytoplasmic p21 [157]. Given that its cytoplasmic localization has so far been mainly attributed to its phosphorylation by Akt kinase [3], Akt inhibitors would be a good choice facilitating p21’s nuclear relocation and sensitizing cells to chemotherapeutics like cisplatin [199]. Identifying specific inhibitors to selectively block the nuclear-cytoplasmic-shuttling of p21 or to promote its cytoplasmic degradation would be also excellent desirable targeting strategies. However tissue- and cell-type specificity could complicate a general therapeutic strategy. Nevertheless, targeting p21’s sustained high expression (Figure 1A) and its cytoplasmic localization (Figure 1B) could be beneficial for cancer intervention. Yet, caution should be taken in terms of its localization, the p53 status and/or other potential genetic lesions affecting p21.

4. Conclusions

Being known over 25 years, p21 is still a challenging and fascinating protein. Rather than simply being a tumor suppressor, p21 also has oncogenic potential, demonstrated particularly by its sustained expression and its cytoplasmic localization, leading to chemoresistance and tumor heterogeneity, and reflecting its dual functionality depending on the cellular and environmental context. A great body of clinical, preclinical and cell-based studies provides insight into its molecular understanding, very often accompanied by inconsistency and divergence. While p21 conveys its multifaceted roles and impacts various fundamental cellular activities including cell differentiation and migration, its own regulation is dynamically affected by intra- and extracellular events. p21’s expression and localization influence the cellular response to external stimuli including chemotherapeutics. These agents, in turn, impact p21’s expression, localization, postmodification, stability and functions. Due to its functional “duality” [51] and its “intrinsically unstructured” [7] feature, it is still a long way to go to uncover the various faces of p21 in normal and malignant cells. Further investigations are required to entirely elucidate its roles under distinct cellular and environmental circumstances with different genetic backgrounds and to define its application as a prognostic marker and a therapeutic target in individual tumor entities. Despite this, p21 already shows a great potential by providing additional prognostic information for the selection of patients for an adequate anti-cancer therapy. Many questions are however still open. To finally define p21 as an trustworthy intervention target is “a work in progress” [144].

Author Contributions

Conceptualization, N.-N.K.; writing—original draft preparation, N.-N.K.; writing—review and editing, J.Y., F.L. and N.-N.K.; visualization, J.Y. and N.-N.K.; funding acquisition, N.-N.K.

Funding

This research was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation)—project number 390921723.

Acknowledgments

We apologize to colleagues whose work could not be included due to space limitations.

Conflicts of Interest

The authors declare no conflict of interest. The funders had no role in the writing of the manuscript.

References

  1. Xiong, Y.; Hannon, G.J.; Zhang, H.; Casso, D.; Kobayashi, R.; Beach, D. p21 is a universal inhibitor of cyclin kinases. Nature 1993, 366, 701–704. [Google Scholar] [CrossRef] [PubMed]
  2. Abbas, T.; Dutta, A. p21 in cancer: Intricate networks and multiple activities. Nat. Rev. Cancer 2009, 9, 400–414. [Google Scholar] [CrossRef] [PubMed]
  3. Kreis, N.N.; Louwen, F.; Yuan, J. Less understood issues: p21(Cip1) in mitosis and its therapeutic potential. Oncogene 2015, 34, 1758–1767. [Google Scholar] [CrossRef] [PubMed]
  4. Roninson, I.B. Oncogenic functions of tumour suppressor p21(Waf1/Cip1/Sdi1): Association with cell senescence and tumour-promoting activities of stromal fibroblasts. Cancer Lett. 2002, 179, 1–14. [Google Scholar] [CrossRef]
  5. Follis, A.V.; Galea, C.A.; Kriwacki, R.W. Intrinsic protein flexibility in regulation of cell proliferation: Advantages for signaling and opportunities for novel therapeutics. Adv. Exp. Med. Biol. 2012, 725, 27–49. [Google Scholar] [CrossRef] [PubMed]
  6. Lacy, E.R.; Filippov, I.; Lewis, W.S.; Otieno, S.; Xiao, L.; Weiss, S.; Hengst, L.; Kriwacki, R.W. p27 binds cyclin-CDK complexes through a sequential mechanism involving binding-induced protein folding. Nat. Struct. Mol. Biol. 2004, 11, 358–364. [Google Scholar] [CrossRef] [PubMed]
  7. Yoon, M.K.; Mitrea, D.M.; Ou, L.; Kriwacki, R.W. Cell cycle regulation by the intrinsically disordered proteins p21 and p27. Biochem. Soc. Trans. 2012, 40, 981–988. [Google Scholar] [CrossRef] [PubMed]
  8. Warfel, N.A.; El-Deiry, W.S. p21WAF1 and tumourigenesis: 20 years after. Curr. Opin. Oncol. 2013, 25, 52–58. [Google Scholar] [CrossRef] [PubMed]
  9. Gartel, A.L.; Tyner, A.L. Transcriptional regulation of the p21((WAF1/CIP1)) gene. Exp. Cell Res. 1999, 246, 280–289. [Google Scholar] [CrossRef] [PubMed]
  10. Gartel, A.L.; Radhakrishnan, S.K. Lost in transcription: p21 repression, mechanisms, and consequences. Cancer Res. 2005, 65, 3980–3985. [Google Scholar] [CrossRef] [PubMed]
  11. Jung, Y.S.; Qian, Y.; Chen, X. Examination of the expanding pathways for the regulation of p21 expression and activity. Cell. Signal. 2010, 22, 1003–1012. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  12. Starostina, N.G.; Kipreos, E.T. Multiple degradation pathways regulate versatile CIP/KIP CDK inhibitors. Trends. Cell. Biol. 2012, 22, 33–41. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  13. Child, E.S.; Mann, D.J. The intricacies of p21 phosphorylation: Protein/protein interactions, subcellular localization and stability. Cell Cycle 2006, 5, 1313–1319. [Google Scholar] [CrossRef] [PubMed]
  14. Weiss, R.H. p21Waf1/Cip1 as a therapeutic target in breast and other cancers. Cancer. Cell. 2003, 4, 425–429. [Google Scholar] [CrossRef] [Green Version]
  15. Abukhdeir, A.M.; Park, B.H. P21 and p27: Roles in carcinogenesis and drug resistance. Expert Rev. Mol. Med. 2008, 10, e19. [Google Scholar] [CrossRef] [PubMed]
  16. Stivala, L.A.; Cazzalini, O.; Prosperi, E. The cyclin-dependent kinase inhibitor p21CDKN1A as a target of anti-cancer drugs. Curr. Cancer Drug Targets 2012, 12, 85–96. [Google Scholar] [CrossRef] [PubMed]
  17. El-Deiry, W.S.; Tokino, T.; Velculescu, V.E.; Levy, D.B.; Parsons, R.; Trent, J.M.; Lin, D.; Mercer, W.E.; Kinzler, K.W.; Vogelstein, B. WAF1, a potential mediator of p53 tumor suppression. Cell 1993, 75, 817–825. [Google Scholar] [CrossRef]
  18. Harper, J.W.; Adami, G.R.; Wei, N.; Keyomarsi, K.; Elledge, S.J. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 1993, 75, 805–816. [Google Scholar] [CrossRef]
  19. Noda, A.; Ning, Y.; Venable, S.F.; Pereira-Smith, O.M.; Smith, J.R. Cloning of senescent cell-derived inhibitors of DNA synthesis using an expression screen. Exp. Cell Res. 1994, 211, 90–98. [Google Scholar] [CrossRef]
  20. Jiang, H.; Lin, J.; Su, Z.Z.; Herlyn, M.; Kerbel, R.S.; Weissman, B.E.; Welch, D.R.; Fisher, P.B. The melanoma differentiation-associated gene mda-6, which encodes the cyclin-dependent kinase inhibitor p21, is differentially expressed during growth, differentiation and progression in human melanoma cells. Oncogene 1995, 10, 1855–1864. [Google Scholar]
  21. Gu, Y.; Turck, C.W.; Morgan, D.O. Inhibition of CDK2 activity in vivo by an associated 20K regulatory subunit. Nature 1993, 366, 707–710. [Google Scholar] [CrossRef] [PubMed]
  22. Romanov, V.S.; Rudolph, K.L. p21 shapes cancer evolution. Nature Cell. Biol. 2016, 18, 722–724. [Google Scholar] [CrossRef] [PubMed]
  23. Kreis, N.N.; Friemel, A.; Zimmer, B.; Roth, S.; Rieger, M.A.; Rolle, U.; Louwen, F.; Yuan, J. Mitotic p21Cip1/CDKN1A is regulated by cyclin-dependent kinase 1 phosphorylation. Oncotarget 2016, 7, 50215–50228. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  24. Kreis, N.N.; Sanhaji, M.; Rieger, M.A.; Louwen, F.; Yuan, J. p21Waf1/Cip1 deficiency causes multiple mitotic defects in tumor cells. Oncogene 2014, 33, 5716–5728. [Google Scholar] [CrossRef] [PubMed]
  25. Barboza, J.A.; Liu, G.; Ju, Z.; El-Naggar, A.K.; Lozano, G. p21 delays tumor onset by preservation of chromosomal stability. Proc. Natl. Acad. Sci. USA 2006, 103, 19842–19847. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  26. Suzuki, H.; Yabuta, N.; Okada, N.; Torigata, K.; Aylon, Y.; Oren, M.; Nojima, H. Lats2 phosphorylates p21/CDKN1A after UV irradiation and regulates apoptosis. J. Cell Sci. 2013, 126, 4358–4368. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  27. Besson, A.; Dowdy, S.F.; Roberts, J.M. CDK inhibitors: Cell cycle regulators and beyond. Dev. Cell. 2008, 14, 159–169. [Google Scholar] [CrossRef]
  28. Waga, S.; Hannon, G.J.; Beach, D.; Stillman, B. The p21 inhibitor of cyclin-dependent kinases controls DNA replication by interaction with PCNA. Nature 1994, 369, 574–578. [Google Scholar] [CrossRef]
  29. Engeland, K. Cell cycle arrest through indirect transcriptional repression by p53: I have a DREAM. Cell Death Differ. 2018, 25, 114–132. [Google Scholar] [CrossRef]
  30. Chang, B.D.; Watanabe, K.; Broude, E.V.; Fang, J.; Poole, J.C.; Kalinichenko, T.V.; Roninson, I.B. Effects of p21Waf1/Cip1/Sdi1 on cellular gene expression: Implications for carcinogenesis, senescence, and age-related diseases. Proc. Natl. Acad. Sci. USA 2000, 97, 4291–4296. [Google Scholar] [CrossRef]
  31. Ferrandiz, N.; Caraballo, J.M.; Garcia-Gutierrez, L.; Devgan, V.; Rodriguez-Paredes, M.; Lafita, M.C.; Bretones, G.; Quintanilla, A.; Munoz-Alonso, M.J.; Blanco, R.; et al. p21 as a transcriptional co-repressor of S-phase and mitotic control genes. PLoS ONE 2012, 7, e37759. [Google Scholar] [CrossRef] [PubMed]
  32. Mitchell, K.O.; El-Deiry, W.S. Overexpression of c-Myc inhibits p21WAF1/CIP1 expression and induces S-phase entry in 12-O-tetradecanoylphorbol-13-acetate (TPA)-sensitive human cancer cells. Cell Growth Differ. 1999, 10, 223–230. [Google Scholar] [PubMed]
  33. Kitaura, H.; Shinshi, M.; Uchikoshi, Y.; Ono, T.; Iguchi-Ariga, S.M.; Ariga, H. Reciprocal regulation via protein-protein interaction between c-Myc and p21(cip1/waf1/sdi1) in DNA replication and transcription. J. Biol. Chem. 2000, 275, 10477–10483. [Google Scholar] [CrossRef] [PubMed]
  34. Waldman, T.; Kinzler, K.W.; Vogelstein, B. p21 is necessary for the p53-mediated G1 arrest in human cancer cells. Cancer Res. 1995, 55, 5187–5190. [Google Scholar] [PubMed]
  35. Bunz, F.; Dutriaux, A.; Lengauer, C.; Waldman, T.; Zhou, S.; Brown, J.P.; Sedivy, J.M.; Kinzler, K.W.; Vogelstein, B. Requirement for p53 and p21 to sustain G2 arrest after DNA damage. Science 1998, 282, 1497–1501. [Google Scholar] [CrossRef] [PubMed]
  36. Barr, A.R.; Cooper, S.; Heldt, F.S.; Butera, F.; Stoy, H.; Mansfeld, J.; Novak, B.; Bakal, C. DNA damage during S-phase mediates the proliferation-quiescence decision in the subsequent G1 via p21 expression. Nat. Commun. 2017, 8, 14728. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  37. LaBaer, J.; Garrett, M.D.; Stevenson, L.F.; Slingerland, J.M.; Sandhu, C.; Chou, H.S.; Fattaey, A.; Harlow, E. New functional activities for the p21 family of CDK inhibitors. Genes. Dev. 1997, 11, 847–862. [Google Scholar] [CrossRef] [PubMed]
  38. Cheng, M.; Olivier, P.; Diehl, J.A.; Fero, M.; Roussel, M.F.; Roberts, J.M.; Sherr, C.J. The p21(Cip1) and p27(Kip1) CDK ’inhibitors’ are essential activators of cyclin D-dependent kinases in murine fibroblasts. EMBO J. 1999, 18, 1571–1583. [Google Scholar] [CrossRef] [PubMed]
  39. Dash, B.C.; El-Deiry, W.S. Phosphorylation of p21 in G2/M promotes cyclin B-Cdc2 kinase activity. Mol. Cell. Biol. 2005, 25, 3364–3387. [Google Scholar] [CrossRef]
  40. Pan, Z.Q.; Reardon, J.T.; Li, L.; Flores-Rozas, H.; Legerski, R.; Sancar, A.; Hurwitz, J. Inhibition of nucleotide excision repair by the cyclin-dependent kinase inhibitor p21. J. Biol. Chem. 1995, 270, 22008–22016. [Google Scholar] [CrossRef]
  41. Soria, G.; Speroni, J.; Podhajcer, O.L.; Prives, C.; Gottifredi, V. p21 differentially regulates DNA replication and DNA-repair-associated processes after UV irradiation. J. Cell Sci. 2008, 121, 3271–3282. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  42. Podust, V.N.; Podust, L.M.; Goubin, F.; Ducommun, B.; Hubscher, U. Mechanism of inhibition of proliferating cell nuclear antigen-dependent DNA synthesis by the cyclin-dependent kinase inhibitor p21. Biochemistry 1995, 34, 8869–8875. [Google Scholar] [CrossRef] [PubMed]
  43. Adimoolam, S.; Lin, C.X.; Ford, J.M. The p53-regulated cyclin-dependent kinase inhibitor, p21 (cip1, waf1, sdi1), is not required for global genomic and transcription-coupled nucleotide excision repair of UV-induced DNA photoproducts. J. Biol. Chem. 2001, 276, 25813–25822. [Google Scholar] [CrossRef] [PubMed]
  44. Wani, M.A.; Wani, G.; Yao, J.; Zhu, Q.; Wani, A.A. Human cells deficient in p53 regulated p21(waf1/cip1) expression exhibit normal nucleotide excision repair of UV-induced DNA damage. Carcinogenesis 2002, 23, 403–410. [Google Scholar] [CrossRef]
  45. McDonald, E.R., 3rd; Wu, G.S.; Waldman, T.; El-Deiry, W.S. Repair Defect in p21 WAF1/CIP1 -/- human cancer cells. Cancer Res. 1996, 56, 2250–2255. [Google Scholar] [PubMed]
  46. Stivala, L.A.; Riva, F.; Cazzalini, O.; Savio, M.; Prosperi, E. p21(waf1/cip1)-null human fibroblasts are deficient in nucleotide excision repair downstream the recruitment of PCNA to DNA repair sites. Oncogene 2001, 20, 563–570. [Google Scholar] [CrossRef] [PubMed]
  47. Mauro, M.; Rego, M.A.; Boisvert, R.A.; Esashi, F.; Cavallo, F.; Jasin, M.; Howlett, N.G. p21 promotes error-free replication-coupled DNA double-strand break repair. Nucleic Acids. Res. 2012, 40, 8348–8360. [Google Scholar] [CrossRef] [Green Version]
  48. Cazzalini, O.; Scovassi, A.I.; Savio, M.; Stivala, L.A.; Prosperi, E. Multiple roles of the cell cycle inhibitor p21(CDKN1A) in the DNA damage response. Mutat. Res. 2010, 704, 12–20. [Google Scholar] [CrossRef]
  49. Zhang, Y.; Fujita, N.; Tsuruo, T. Caspase-mediated cleavage of p21Waf1/Cip1 converts cancer cells from growth arrest to undergoing apoptosis. Oncogene 1999, 18, 1131–1138. [Google Scholar] [CrossRef] [Green Version]
  50. Jin, Y.H.; Yoo, K.J.; Lee, Y.H.; Lee, S.K. Caspase 3-mediated cleavage of p21WAF1/CIP1 associated with the cyclin A-cyclin-dependent kinase 2 complex is a prerequisite for apoptosis in SK-HEP-1 cells. J. Biol. Chem. 2000, 275, 30256–30263. [Google Scholar] [CrossRef]
  51. Gartel, A.L.; Tyner, A.L. The role of the cyclin-dependent kinase inhibitor p21 in apoptosis. Mol. Cancer Ther. 2002, 1, 639–649. [Google Scholar] [PubMed]
  52. Gartel, A.L. The conflicting roles of the cdk inhibitor p21(CIP1/WAF1) in apoptosis. Leuk. Res. 2005, 29, 1237–1238. [Google Scholar] [CrossRef] [PubMed]
  53. Zheng, K.; He, Z.; Kitazato, K.; Wang, Y. Selective Autophagy Regulates Cell Cycle in Cancer Therapy. Theranostics 2019, 9, 104–125. [Google Scholar] [CrossRef]
  54. Capparelli, C.; Chiavarina, B.; Whitaker-Menezes, D.; Pestell, T.G.; Pestell, R.G.; Hulit, J.; Ando, S.; Howell, A.; Martinez-Outschoorn, U.E.; Sotgia, F.; et al. CDK inhibitors (p16/p19/p21) induce senescence and autophagy in cancer-associated fibroblasts, “fueling” tumor growth via paracrine interactions, without an increase in neo-angiogenesis. Cell Cycle 2012, 11, 3599–3610. [Google Scholar] [CrossRef] [PubMed]
  55. Mohapatra, P.; Preet, R.; Das, D.; Satapathy, S.R.; Choudhuri, T.; Wyatt, M.D.; Kundu, C.N. Quinacrine-mediated autophagy and apoptosis in colon cancer cells is through a p53- and p21-dependent mechanism. Oncol. Res. 2012, 20, 81–91. [Google Scholar] [CrossRef]
  56. Fujiwara, K.; Daido, S.; Yamamoto, A.; Kobayashi, R.; Yokoyama, T.; Aoki, H.; Iwado, E.; Shinojima, N.; Kondo, Y.; Kondo, S. Pivotal role of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 in apoptosis and autophagy. J. Biol. Chem. 2008, 283, 388–397. [Google Scholar] [CrossRef] [PubMed]
  57. McConnell, B.B.; Starborg, M.; Brookes, S.; Peters, G. Inhibitors of cyclin-dependent kinases induce features of replicative senescence in early passage human diploid fibroblasts. Curr. Biol. 1998, 8, 351–354. [Google Scholar] [CrossRef] [Green Version]
  58. Fang, L.; Igarashi, M.; Leung, J.; Sugrue, M.M.; Lee, S.W.; Aaronson, S.A. p21Waf1/Cip1/Sdi1 induces permanent growth arrest with markers of replicative senescence in human tumor cells lacking functional p53. Oncogene 1999, 18, 2789–2797. [Google Scholar] [CrossRef] [Green Version]
  59. Wang, Y.; Blandino, G.; Givol, D. Induced p21waf expression in H1299 cell line promotes cell senescence and protects against cytotoxic effect of radiation and doxorubicin. Oncogene 1999, 18, 2643–2649. [Google Scholar] [CrossRef] [Green Version]
  60. Lee, S.; Helfman, D.M. Cytoplasmic p21Cip1 is involved in Ras-induced inhibition of the ROCK/LIMK/cofilin pathway. J. Biol. Chem. 2004, 279, 1885–1891. [Google Scholar] [CrossRef]
  61. Denicourt, C.; Dowdy, S.F. Cip/Kip proteins: More than just CDKs inhibitors. Genes. Dev. 2004, 18, 851–855. [Google Scholar] [CrossRef] [PubMed]
  62. Galanos, P.; Vougas, K.; Walter, D.; Polyzos, A.; Maya-Mendoza, A.; Haagensen, E.J.; Kokkalis, A.; Roumelioti, F.M.; Gagos, S.; Tzetis, M.; et al. Chronic p53-independent p21 expression causes genomic instability by deregulating replication licensing. Nat. Cell Biol. 2016, 18, 777–789. [Google Scholar] [CrossRef]
  63. Asada, M.; Yamada, T.; Ichijo, H.; Delia, D.; Miyazono, K.; Fukumuro, K.; Mizutani, S. Apoptosis inhibitory activity of cytoplasmic p21(Cip1/WAF1) in monocytic differentiation. EMBO J. 1999, 18, 1223–1234. [Google Scholar] [CrossRef] [PubMed]
  64. Ghanem, L.; Steinman, R.A. p21Waf1 inhibits granulocytic differentiation of 32Dcl3 cells. Leuk. Res. 2006, 30, 1285–1292. [Google Scholar] [CrossRef] [PubMed]
  65. Kawamura, T.; Suzuki, J.; Wang, Y.V.; Menendez, S.; Morera, L.B.; Raya, A.; Wahl, G.M.; Izpisua Belmonte, J.C. Linking the p53 tumour suppressor pathway to somatic cell reprogramming. Nature 2009, 460, 1140–1144. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  66. Hong, H.; Takahashi, K.; Ichisaka, T.; Aoi, T.; Kanagawa, O.; Nakagawa, M.; Okita, K.; Yamanaka, S. Suppression of induced pluripotent stem cell generation by the p53-p21 pathway. Nature 2009, 460, 1132–1135. [Google Scholar] [CrossRef] [PubMed]
  67. Liu, Y.; Hoya-Arias, R.; Nimer, S.D. The role of p53 in limiting somatic cell reprogramming. Cell. Res. 2009, 19, 1227–1228. [Google Scholar] [CrossRef] [Green Version]
  68. Ruijtenberg, S.; van den Heuvel, S. Coordinating cell proliferation and differentiation: Antagonism between cell cycle regulators and cell type-specific gene expression. Cell Cycle 2016, 15, 196–212. [Google Scholar] [CrossRef] [Green Version]
  69. Dutto, I.; Tillhon, M.; Cazzalini, O.; Stivala, L.A.; Prosperi, E. Biology of the cell cycle inhibitor p21(CDKN1A): Molecular mechanisms and relevance in chemical toxicology. Arch. Toxicol. 2015, 89, 155–178. [Google Scholar] [CrossRef]
  70. Zezula, J.; Casaccia-Bonnefil, P.; Ezhevsky, S.A.; Osterhout, D.J.; Levine, J.M.; Dowdy, S.F.; Chao, M.V.; Koff, A. p21cip1 is required for the differentiation of oligodendrocytes independently of cell cycle withdrawal. EMBO Rep. 2001, 2, 27–34. [Google Scholar] [CrossRef]
  71. Baccini, V.; Roy, L.; Vitrat, N.; Chagraoui, H.; Sabri, S.; Le Couedic, J.P.; Debili, N.; Wendling, F.; Vainchenker, W. Role of p21(Cip1/Waf1) in cell-cycle exit of endomitotic megakaryocytes. Blood 2001, 98, 3274–3282. [Google Scholar] [CrossRef]
  72. Papetti, M.; Wontakal, S.N.; Stopka, T.; Skoultchi, A.I. GATA-1 directly regulates p21 gene expression during erythroid differentiation. Cell Cycle 2010, 9, 1972–1980. [Google Scholar] [CrossRef] [PubMed]
  73. Guo, K.; Wang, J.; Andres, V.; Smith, R.C.; Walsh, K. MyoD-induced expression of p21 inhibits cyclin-dependent kinase activity upon myocyte terminal differentiation. Mol. Cell. Biol. 1995, 15, 3823–3829. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  74. Wang, J.; Walsh, K. Resistance to apoptosis conferred by Cdk inhibitors during myocyte differentiation. Science 1996, 273, 359–361. [Google Scholar] [CrossRef] [PubMed]
  75. Mercer, S.E.; Ewton, D.Z.; Deng, X.; Lim, S.; Mazur, T.R.; Friedman, E. Mirk/Dyrk1B mediates survival during the differentiation of C2C12 myoblasts. J. Biol. Chem. 2005, 280, 25788–25801. [Google Scholar] [CrossRef] [PubMed]
  76. Chinzei, N.; Hayashi, S.; Ueha, T.; Fujishiro, T.; Kanzaki, N.; Hashimoto, S.; Sakata, S.; Kihara, S.; Haneda, M.; Sakai, Y.; et al. P21 deficiency delays regeneration of skeletal muscular tissue. PLoS ONE 2015, 10, e0125765. [Google Scholar] [CrossRef]
  77. Parker, S.B.; Eichele, G.; Zhang, P.; Rawls, A.; Sands, A.T.; Bradley, A.; Olson, E.N.; Harper, J.W.; Elledge, S.J. p53-independent expression of p21Cip1 in muscle and other terminally differentiating cells. Science 1995, 267, 1024–1027. [Google Scholar] [CrossRef]
  78. Tian, J.Q.; Quaroni, A. Involvement of p21(WAF1/Cip1) and p27(Kip1) in intestinal epithelial cell differentiation. Am. J. Physiol. 1999, 276, C1245–C1258. [Google Scholar] [CrossRef]
  79. Inoue, N.; Yahagi, N.; Yamamoto, T.; Ishikawa, M.; Watanabe, K.; Matsuzaka, T.; Nakagawa, Y.; Takeuchi, Y.; Kobayashi, K.; Takahashi, A.; et al. Cyclin-dependent kinase inhibitor, p21WAF1/CIP1, is involved in adipocyte differentiation and hypertrophy, linking to obesity, and insulin resistance. J. Biol. Chem. 2008, 283, 21220–21229. [Google Scholar] [CrossRef]
  80. Naaz, A.; Holsberger, D.R.; Iwamoto, G.A.; Nelson, A.; Kiyokawa, H.; Cooke, P.S. Loss of cyclin-dependent kinase inhibitors produces adipocyte hyperplasia and obesity. FASEB J. 2004, 18, 1925–1927. [Google Scholar] [CrossRef]
  81. Seong, H.A.; Ha, H. Thr55 phosphorylation of p21 by MPK38/MELK ameliorates defects in glucose, lipid, and energy metabolism in diet-induced obese mice. Cell Death Dis. 2019, 10, 380. [Google Scholar] [CrossRef] [PubMed]
  82. Bellosta, P.; Masramon, L.; Mansukhani, A.; Basilico, C. p21(WAF1/CIP1) acts as a brake in osteoblast differentiation. J. Bone Miner. Res. 2003, 18, 818–826. [Google Scholar] [CrossRef] [PubMed]
  83. Di Cunto, F.; Topley, G.; Calautti, E.; Hsiao, J.; Ong, L.; Seth, P.K.; Dotto, G.P. Inhibitory function of p21Cip1/WAF1 in differentiation of primary mouse keratinocytes independent of cell cycle control. Science 1998, 280, 1069–1072. [Google Scholar] [CrossRef] [PubMed]
  84. Missero, C.; Calautti, E.; Eckner, R.; Chin, J.; Tsai, L.H.; Livingston, D.M.; Dotto, G.P. Involvement of the cell-cycle inhibitor Cip1/WAF1 and the E1A-associated p300 protein in terminal differentiation. Proc. Natl. Acad. Sci. USA 1995, 92, 5451–5455. [Google Scholar] [CrossRef] [PubMed]
  85. Negishi, Y.; Ui, N.; Nakajima, M.; Kawashima, K.; Maruyama, K.; Takizawa, T.; Endo, H. p21Cip-1/SDI-1/WAF-1 gene is involved in chondrogenic differentiation of ATDC5 cells in vitro. J. Biol. Chem. 2001, 276, 33249–33256. [Google Scholar] [CrossRef] [PubMed]
  86. Tanaka, H.; Yamashita, T.; Asada, M.; Mizutani, S.; Yoshikawa, H.; Tohyama, M. Cytoplasmic p21(Cip1/WAF1) regulates neurite remodeling by inhibiting Rho-kinase activity. J. Cell. Biol. 2002, 158, 321–329. [Google Scholar] [CrossRef] [PubMed]
  87. Manapov, F.; Muller, P.; Rychly, J. Translocation of p21(Cip1/WAF1) from the nucleus to the cytoplasm correlates with pancreatic myofibroblast to fibroblast cell conversion. Gut 2005, 54, 814–822. [Google Scholar] [CrossRef]
  88. Viale, A.; De Franco, F.; Orleth, A.; Cambiaghi, V.; Giuliani, V.; Bossi, D.; Ronchini, C.; Ronzoni, S.; Muradore, I.; Monestiroli, S.; et al. Cell-cycle restriction limits DNA damage and maintains self-renewal of leukaemia stem cells. Nature 2009, 457, 51–56. [Google Scholar] [CrossRef]
  89. Yamamizu, K.; Schlessinger, D.; Ko, M.S. SOX9 accelerates ESC differentiation to three germ layer lineages by repressing SOX2 expression through P21 (WAF1/CIP1). Development 2014, 141, 4254–4266. [Google Scholar] [CrossRef] [Green Version]
  90. Cheng, T.; Rodrigues, N.; Shen, H.; Yang, Y.; Dombkowski, D.; Sykes, M.; Scadden, D.T. Hematopoietic stem cell quiescence maintained by p21cip1/waf1. Science 2000, 287, 1804–1808. [Google Scholar] [CrossRef]
  91. Topley, G.I.; Okuyama, R.; Gonzales, J.G.; Conti, C.; Dotto, G.P. p21(WAF1/Cip1) functions as a suppressor of malignant skin tumor formation and a determinant of keratinocyte stem-cell potential. Proc. Natl. Acad. Sci. USA 1999, 96, 9089–9094. [Google Scholar] [CrossRef] [PubMed]
  92. Kippin, T.E.; Martens, D.J.; van der Kooy, D. p21 loss compromises the relative quiescence of forebrain stem cell proliferation leading to exhaustion of their proliferation capacity. Genes. Dev. 2005, 19, 756–767. [Google Scholar] [CrossRef] [Green Version]
  93. Ju, Z.; Choudhury, A.R.; Rudolph, K.L. A dual role of p21 in stem cell aging. Ann. N. Y. Acad. Sci. 2007, 1100, 333–344. [Google Scholar] [CrossRef] [PubMed]
  94. De Renty, C.; DePamphilis, M.L.; Ullah, Z. Cytoplasmic localization of p21 protects trophoblast giant cells from DNA damage induced apoptosis. PLoS ONE 2014, 9, e97434. [Google Scholar] [CrossRef] [PubMed]
  95. Masson, A.O.; Hess, R.; O’Brien, K.; Bertram, K.L.; Tailor, P.; Irvine, E.; Ren, G.; Krawetz, R.J. Increased levels of p21((CIP1/WAF1)) correlate with decreased chondrogenic differentiation potential in synovial membrane progenitor cells. Mech. Ageing Dev. 2015, 149, 31–40. [Google Scholar] [CrossRef] [PubMed]
  96. Yew, T.L.; Chiu, F.Y.; Tsai, C.C.; Chen, H.L.; Lee, W.P.; Chen, Y.J.; Chang, M.C.; Hung, S.C. Knockdown of p21(Cip1/Waf1) enhances proliferation, the expression of stemness markers, and osteogenic potential in human mesenchymal stem cells. Aging Cell. 2011, 10, 349–361. [Google Scholar] [CrossRef] [PubMed]
  97. Cheng, H.; Qiu, L.; Ma, J.; Zhang, H.; Cheng, M.; Li, W.; Zhao, X.; Liu, K. Replicative senescence of human bone marrow and umbilical cord derived mesenchymal stem cells and their differentiation to adipocytes and osteoblasts. Mol. Biol. Rep. 2011, 38, 5161–5168. [Google Scholar] [CrossRef] [PubMed]
  98. Liao, T.T.; Yang, M.H. Revisiting epithelial-mesenchymal transition in cancer metastasis: The connection between epithelial plasticity and stemness. Mol. Oncol. 2017, 11, 792–804. [Google Scholar] [CrossRef]
  99. Li, X.L.; Hara, T.; Choi, Y.; Subramanian, M.; Francis, P.; Bilke, S.; Walker, R.L.; Pineda, M.; Zhu, Y.; Yang, Y.; et al. A p21-ZEB1 complex inhibits epithelial-mesenchymal transition through the microRNA 183-96-182 cluster. Mol. Cell. Biol. 2014, 34, 533–550. [Google Scholar] [CrossRef]
  100. Bachman, K.E.; Blair, B.G.; Brenner, K.; Bardelli, A.; Arena, S.; Zhou, S.; Hicks, J.; De Marzo, A.M.; Argani, P.; Park, B.H. p21(WAF1/CIP1) mediates the growth response to TGF-beta in human epithelial cells. Cancer Biol. Ther. 2004, 3, 221–225. [Google Scholar] [CrossRef]
  101. Liu, M.; Casimiro, M.C.; Wang, C.; Shirley, L.A.; Jiao, X.; Katiyar, S.; Ju, X.; Li, Z.; Yu, Z.; Zhou, J.; et al. p21CIP1 attenuates Ras- and c-Myc-dependent breast tumor epithelial mesenchymal transition and cancer stem cell-like gene expression in vivo. Proc. Natl. Acad. Sci. USA 2009, 106, 19035–19039. [Google Scholar] [CrossRef] [PubMed]
  102. Wang, L.; Wang, R.; Ye, Z.; Wang, Y.; Li, X.; Chen, W.; Zhang, M.; Cai, C. PVT1 affects EMT and cell proliferation and migration via regulating p21 in triple-negative breast cancer cells cultured with mature adipogenic medium. Acta Biochim. Biophys. Sin. 2018, 50, 1211–1218. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  103. Zhang, Y.; Yan, W.; Jung, Y.S.; Chen, X. PUMA Cooperates with p21 to Regulate Mammary Epithelial Morphogenesis and Epithelial-To-Mesenchymal Transition. PLoS ONE 2013, 8, e66464. [Google Scholar] [CrossRef] [PubMed]
  104. Liu, M.; Lee, M.H.; Cohen, M.; Bommakanti, M.; Freedman, L.P. Transcriptional activation of the Cdk inhibitor p21 by vitamin D3 leads to the induced differentiation of the myelomonocytic cell line U937. Genes Dev. 1996, 10, 142–153. [Google Scholar] [CrossRef] [PubMed]
  105. Biggs, J.R.; Kudlow, J.E.; Kraft, A.S. The role of the transcription factor Sp1 in regulating the expression of the WAF1/CIP1 gene in U937 leukemic cells. J. Biol. Chem. 1996, 271, 901–906. [Google Scholar] [CrossRef]
  106. Casini, T.; Pelicci, P.G. A function of p21 during promyelocytic leukemia cell differentiation independent of CDK inhibition and cell cycle arrest. Oncogene 1999, 18, 3235–3243. [Google Scholar] [CrossRef] [Green Version]
  107. Jiang, H.; Lin, J.; Su, Z.Z.; Collart, F.R.; Huberman, E.; Fisher, P.B. Induction of differentiation in human promyelocytic HL-60 leukemia cells activates p21, WAF1/CIP1, expression in the absence of p53. Oncogene 1994, 9, 3397–3406. [Google Scholar]
  108. Schwaller, J.; Koeffler, H.P.; Niklaus, G.; Loetscher, P.; Nagel, S.; Fey, M.F.; Tobler, A. Posttranscriptional stabilization underlies p53-independent induction of p21WAF1/CIP1/SDI1 in differentiating human leukemic cells. J. Clin. Investig. 1995, 95, 973–979. [Google Scholar] [CrossRef]
  109. Yoshida, K.; Murohashi, I.; Hirashima, K. p53-independent induction of p21 (WAF1/CIP1) during differentiation of HL-60 cells by tumor necrosis factor alpha. Int. J. Hematol. 1996, 65, 41–48. [Google Scholar] [CrossRef]
  110. Freemerman, A.J.; Vrana, J.A.; Tombes, R.M.; Jiang, H.; Chellappan, S.P.; Fisher, P.B.; Grant, S. Effects of antisense p21 (WAF1/CIP1/MDA6) expression on the induction of differentiation and drug-mediated apoptosis in human myeloid leukemia cells (HL-60). Leukemia 1997, 11, 504–513. [Google Scholar] [CrossRef] [Green Version]
  111. Ullmannova, V.; Stockbauer, P.; Hradcova, M.; Soucek, J.; Haskovec, C. Relationship between cyclin D1 and p21(Waf1/Cip1) during differentiation of human myeloid leukemia cell lines. Leuk. Res. 2003, 27, 1115–1123. [Google Scholar] [CrossRef]
  112. Matsumura, I.; Ishikawa, J.; Nakajima, K.; Oritani, K.; Tomiyama, Y.; Miyagawa, J.; Kato, T.; Miyazaki, H.; Matsuzawa, Y.; Kanakura, Y. Thrombopoietin-induced differentiation of a human megakaryoblastic leukemia cell line, CMK, involves transcriptional activation of p21(WAF1/Cip1) by STAT5. Mol. Cell. Biol. 1997, 17, 2933–2943. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  113. Gartel, A.L.; Serfas, M.S.; Gartel, M.; Goufman, E.; Wu, G.S.; el-Deiry, W.S.; Tyner, A.L. p21 (WAF1/CIP1) expression is induced in newly nondividing cells in diverse epithelia and during differentiation of the Caco-2 intestinal cell line. Exp. Cell Res. 1996, 227, 171–181. [Google Scholar] [CrossRef]
  114. Poluha, W.; Poluha, D.K.; Chang, B.; Crosbie, N.E.; Schonhoff, C.M.; Kilpatrick, D.L.; Ross, A.H. The cyclin-dependent kinase inhibitor p21 (WAF1) is required for survival of differentiating neuroblastoma cells. Mol. Cell. Biol. 1996, 16, 1335–1341. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  115. Liu, Y.; Encinas, M.; Comella, J.X.; Aldea, M.; Gallego, C. Basic helix-loop-helix proteins bind to TrkB and p21(Cip1) promoters linking differentiation and cell cycle arrest in neuroblastoma cells. Mol. Cell. Biol. 2004, 24, 2662–2672. [Google Scholar] [CrossRef] [PubMed]
  116. Hiraoka, K.; Zenmyo, M.; Komiya, S.; Kawabata, R.; Yokouchi, M.; Suzuki, R.; Hamada, T.; Kato, S.; Nagata, K. Relationship of p21 (waf1/cip1) and differentiation in chondrosarcoma cells. Virchows Arch. 2002, 440, 285–290. [Google Scholar] [CrossRef] [PubMed]
  117. Asada, M.; Ohmi, K.; Delia, D.; Enosawa, S.; Suzuki, S.; Yuo, A.; Suzuki, H.; Mizutani, S. Brap2 functions as a cytoplasmic retention protein for p21 during monocyte differentiation. Mol. Cell. Biol. 2004, 24, 8236–8243. [Google Scholar] [CrossRef] [PubMed]
  118. Takeshima, Y.; Yamasaki, M.; Nishisaka, T.; Kitaguchi, S.; Inai, K. p21WAF1/CIP1 expression in primary lung adenocarcinomas: Heterogeneous expression in tumor tissues and correlation with p53 expression and proliferative activities. Carcinogenesis 1998, 19, 1755–1761. [Google Scholar] [CrossRef] [PubMed]
  119. Nadal, A.; Jares, P.; Cazorla, M.; Fernandez, P.L.; Sanjuan, X.; Hernandez, L.; Pinyol, M.; Aldea, M.; Mallofre, C.; Muntane, J.; et al. p21WAF1/Cip1 expression is associated with cell differentiation but not with p53 mutations in squamous cell carcinomas of the larynx. J. Pathol. 1997, 183, 156–163. [Google Scholar] [CrossRef]
  120. Van Oijen, M.G.; Tilanus, M.G.; Medema, R.H.; Slootweg, P.J. Expression of p21 (Waf1/Cip1) in head and neck cancer in relation to proliferation, differentiation, p53 status and cyclin D1 expression. J. Oral. Pathol. Med. 1998, 27, 367–375. [Google Scholar] [CrossRef]
  121. Gartel, A.L. Inducer and inhibitor: “antagonistic duality” of p21 in differentiation. Leuk. Res. 2006, 30, 1215–1216. [Google Scholar] [CrossRef] [PubMed]
  122. Yamamoto, H.; Soh, J.W.; Shirin, H.; Xing, W.Q.; Lim, J.T.; Yao, Y.; Slosberg, E.; Tomita, N.; Schieren, I.; Weinstein, I.B. Comparative effects of overexpression of p27Kip1 and p21Cip1/Waf1 on growth and differentiation in human colon carcinoma cells. Oncogene 1999, 18, 103–115. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  123. Ridley, A.J.; Schwartz, M.A.; Burridge, K.; Firtel, R.A.; Ginsberg, M.H.; Borisy, G.; Parsons, J.T.; Horwitz, A.R. Cell migration: Integrating signals from front to back. Science 2003, 302, 1704–1709. [Google Scholar] [CrossRef] [PubMed]
  124. Chaffer, C.L.; Weinberg, R.A. A perspective on cancer cell metastasis. Science 2011, 331, 1559–1564. [Google Scholar] [CrossRef] [PubMed]
  125. Gartel, A.L.; Najmabadi, F.; Goufman, E.; Tyner, A.L. A role for E2F1 in Ras activation of p21(WAF1/CIP1) transcription. Oncogene 2000, 19, 961–964. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  126. Zhou, B.P.; Liao, Y.; Xia, W.; Spohn, B.; Lee, M.H.; Hung, M.C. Cytoplasmic localization of p21Cip1/WAF1 by Akt-induced phosphorylation in HER-2/neu-overexpressing cells. Nature Cell. Biol. 2001, 3, 245–252. [Google Scholar] [CrossRef] [PubMed]
  127. Starostina, N.G.; Simpliciano, J.M.; McGuirk, M.A.; Kipreos, E.T. CRL2(LRR-1) targets a CDK inhibitor for cell cycle control in C. elegans and actin-based motility regulation in human cells. Dev. Cell. 2010, 19, 753–764. [Google Scholar] [CrossRef] [PubMed]
  128. Liu, X.; Sun, Y.; Ehrlich, M.; Lu, T.; Kloog, Y.; Weinberg, R.A.; Lodish, H.F.; Henis, Y.I. Disruption of TGF-beta growth inhibition by oncogenic ras is linked to p27Kip1 mislocalization. Oncogene 2000, 19, 5926–5935. [Google Scholar] [CrossRef]
  129. Xia, W.; Chen, J.S.; Zhou, X.; Sun, P.R.; Lee, D.F.; Liao, Y.; Zhou, B.P.; Hung, M.C. Phosphorylation/cytoplasmic localization of p21Cip1/WAF1 is associated with HER2/neu overexpression and provides a novel combination predictor for poor prognosis in breast cancer patients. Clin. Cancer Res. 2004, 10, 3815–3824. [Google Scholar] [CrossRef]
  130. Winters, Z.E.; Leek, R.D.; Bradburn, M.J.; Norbury, C.J.; Harris, A.L. Cytoplasmic p21WAF1/CIP1 expression is correlated with HER-2/neu in breast cancer and is an independent predictor of prognosis. Breast Cancer Res. 2003, 5, R242–R249. [Google Scholar] [CrossRef]
  131. Zhao, H.; Faltermeier, C.M.; Mendelsohn, L.; Porter, P.L.; Clurman, B.E.; Roberts, J.M. Mislocalization of p27 to the cytoplasm of breast cancer cells confers resistance to anti-HER2 targeted therapy. Oncotarget 2014, 5, 12704–12714. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  132. Cheng, X.; Xia, W.; Yang, J.Y.; Hsu, J.L.; Chou, C.K.; Sun, H.L.; Wyszomierski, S.L.; Mills, G.B.; Muller, W.J.; Yu, D.; et al. Activation of p21(CIP1/WAF1) in mammary epithelium accelerates mammary tumorigenesis and promotes lung metastasis. Biochem. Biophys. Res. Commun. 2010, 403, 103–107. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  133. Kim, E.M.; Jung, C.H.; Kim, J.; Hwang, S.G.; Park, J.K.; Um, H.D. The p53/p21 Complex Regulates Cancer Cell Invasion and Apoptosis by Targeting Bcl-2 Family Proteins. Cancer Res. 2017, 77, 3092–3100. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  134. Kim, J.; Bae, S.; An, S.; Park, J.K.; Kim, E.M.; Hwang, S.G.; Kim, W.J.; Um, H.D. Cooperative actions of p21WAF1 and p53 induce Slug protein degradation and suppress cell invasion. EMBO Rep. 2014, 15, 1062–1068. [Google Scholar] [CrossRef] [PubMed]
  135. Wang, S.P.; Wang, W.L.; Chang, Y.L.; Wu, C.T.; Chao, Y.C.; Kao, S.H.; Yuan, A.; Lin, C.W.; Yang, S.C.; Chan, W.K.; et al. p53 controls cancer cell invasion by inducing the MDM2-mediated degradation of Slug. Nature Cell. Biol. 2009, 11, 694–704. [Google Scholar] [CrossRef]
  136. Dai, M.; Al-Odaini, A.A.; Arakelian, A.; Rabbani, S.A.; Ali, S.; Lebrun, J.J. A novel function for p21Cip1 and acetyltransferase p/CAF as critical transcriptional regulators of TGFbeta-mediated breast cancer cell migration and invasion. Breast Cancer Res. 2012, 14, R127. [Google Scholar] [CrossRef] [PubMed]
  137. Dai, M.; Al-Odaini, A.A.; Fils-Aime, N.; Villatoro, M.A.; Guo, J.; Arakelian, A.; Rabbani, S.A.; Ali, S.; Lebrun, J.J. Cyclin D1 cooperates with p21 to regulate TGFbeta-mediated breast cancer cell migration and tumor local invasion. Breast Cancer Res. 2013, 15, R49. [Google Scholar] [CrossRef]
  138. Qian, X.; Hulit, J.; Suyama, K.; Eugenin, E.A.; Belbin, T.J.; Loudig, O.; Smirnova, T.; Zhou, Z.N.; Segall, J.; Locker, J.; et al. p21CIP1 mediates reciprocal switching between proliferation and invasion during metastasis. Oncogene 2013, 32, 2292–2303. [Google Scholar] [CrossRef]
  139. Okuma, A.; Hanyu, A.; Watanabe, S.; Hara, E. p16(Ink4a) and p21(Cip1/Waf1) promote tumour growth by enhancing myeloid-derived suppressor cells chemotaxis. Nat. Commun. 2017, 8, 2050. [Google Scholar] [CrossRef]
  140. Kreis, N.N.; Friemel, A.; Ritter, A.; Roth, S.; Rolle, U.; Louwen, F.; Yuan, J. Function of p21 (Cip1/Waf1/CDKN1A) in Migration and Invasion of Cancer and Trophoblastic Cells. Cancers 2019, 11, 989. [Google Scholar] [CrossRef]
  141. Chen, Z.; Chen, X.; Chen, P.; Yu, S.; Nie, F.; Lu, B.; Zhang, T.; Zhou, Y.; Chen, Q.; Wei, C.; et al. Long non-coding RNA SNHG20 promotes non-small cell lung cancer cell proliferation and migration by epigenetically silencing of P21 expression. Cell Death Dis. 2017, 8, e3092. [Google Scholar] [CrossRef] [PubMed]
  142. Huang, Y.; Wang, W.; Chen, Y.; Huang, Y.; Zhang, J.; He, S.; Tan, Y.; Qiang, F.; Li, A.; Roe, O.D.; et al. The opposite prognostic significance of nuclear and cytoplasmic p21 expression in resectable gastric cancer patients. J. Gastroenterol. 2014, 49, 1441–1452. [Google Scholar] [CrossRef] [PubMed]
  143. Galanos, P.; Pappas, G.; Polyzos, A.; Kotsinas, A.; Svolaki, I.; Giakoumakis, N.N.; Glytsou, C.; Pateras, I.S.; Swain, U.; Souliotis, V.L.; et al. Mutational signatures reveal the role of RAD52 in p53-independent p21-driven genomic instability. Genome Biol. 2018, 19, 37. [Google Scholar] [CrossRef] [PubMed]
  144. El-Deiry, W.S. p21(WAF1) Mediates Cell-Cycle Inhibition, Relevant to Cancer Suppression and Therapy. Cancer Res. 2016, 76, 5189–5191. [Google Scholar] [CrossRef] [PubMed]
  145. Teramen, H.; Tsukuda, K.; Tanaka, N.; Ueno, T.; Kubo, T.; Ando, M.; Soh, J.; Asano, H.; Pass, H.I.; Toyooka, S.; et al. Aberrant methylation of p21 gene in lung cancer and malignant pleural mesothelioma. Acta. Med. Okayama 2011, 65, 179–184. [Google Scholar] [CrossRef]
  146. Bott, S.R.; Arya, M.; Kirby, R.S.; Williamson, M. p21WAF1/CIP1 gene is inactivated in metastatic prostatic cancer cell lines by promoter methylation. Prostate Cancer Prostatic Dis. 2005, 8, 321–326. [Google Scholar] [CrossRef] [PubMed]
  147. Askari, M.; Sobti, R.C.; Nikbakht, M.; Sharma, S.C. Aberrant promoter hypermethylation of p21 (WAF1/CIP1) gene and its impact on expression and role of polymorphism in the risk of breast cancer. Mol. Cell. Biochem. 2013, 382, 19–26. [Google Scholar] [CrossRef]
  148. Roman-Gomez, J.; Castillejo, J.A.; Jimenez, A.; Gonzalez, M.G.; Moreno, F.; Rodriguez Mdel, C.; Barrios, M.; Maldonado, J.; Torres, A. 5′ CpG island hypermethylation is associated with transcriptional silencing of the p21(CIP1/WAF1/SDI1) gene and confers poor prognosis in acute lymphoblastic leukemia. Blood 2002, 99, 2291–2296. [Google Scholar] [CrossRef]
  149. Morlando, M.; Fatica, A. Alteration of Epigenetic Regulation by Long Noncoding RNAs in Cancer. Int. J. Mol. Sci. 2018, 19, 570. [Google Scholar] [CrossRef]
  150. Yu, Y.; Zhang, M.; Wang, N.; Li, Q.; Yang, J.; Yan, S.; He, X.; Ji, G.; Miao, L. Epigenetic silencing of tumor suppressor gene CDKN1A by oncogenic long non-coding RNA SNHG1 in cholangiocarcinoma. Cell Death Dis. 2018, 9, 746. [Google Scholar] [CrossRef]
  151. Yang, W.; Qi, Q.; Zhang, H.; Xu, W.; Chen, Z.; Wang, L.; Wang, Y.; Dong, X.; Jiao, H.; Huo, Z. p21 Waf1/Cip1 polymorphisms and risk of esophageal cancer. Ann. Surg. Oncol. 2010, 17, 1453–1458. [Google Scholar] [CrossRef] [PubMed]
  152. Ma, H.; Zhou, Z.; Wei, S.; Wei, Q. Association between p21 Ser31Arg polymorphism and cancer risk: A meta-analysis. Chin. J. Cancer 2011, 30, 254–263. [Google Scholar] [CrossRef] [PubMed]
  153. Wang, Z.; Sturgis, E.M.; Zhang, F.; Lei, D.; Liu, Z.; Xu, L.; Song, X.; Wei, Q.; Li, G. Genetic variants of p27 and p21 as predictors for risk of second primary malignancy in patients with index squamous cell carcinoma of head and neck. Mol. Cancer 2012, 11, 17. [Google Scholar] [CrossRef] [PubMed]
  154. Perri, F.; Pisconti, S.; Della Vittoria Scarpati, G. P53 mutations and cancer: A tight linkage. Ann. Transl. Med. 2016, 4, 522. [Google Scholar] [CrossRef] [PubMed]
  155. Chen, H.; Liu, H.; Qing, G. Targeting oncogenic Myc as a strategy for cancer treatment. Signal. Transduct. Target. Ther. 2018, 3, 5. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  156. Sun, Y.; Yang, S.; Sun, N.; Chen, J. Differential expression of STAT1 and p21 proteins predicts pancreatic cancer progression and prognosis. Pancreas 2014, 43, 619–623. [Google Scholar] [CrossRef] [PubMed]
  157. Liu, R.; Wettersten, H.I.; Park, S.H.; Weiss, R.H. Small-molecule inhibitors of p21 as novel therapeutics for chemotherapy-resistant kidney cancer. Future Med. Chem. 2013, 5, 991–994. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  158. Winters, Z.E.; Hunt, N.C.; Bradburn, M.J.; Royds, J.A.; Turley, H.; Harris, A.L.; Norbury, C.J. Subcellular localisation of cyclin B, Cdc2 and p21(WAF1/CIP1) in breast cancer. association with prognosis. Eur. J. Cancer 2001, 37, 2405–2412. [Google Scholar] [CrossRef]
  159. Ping, B.; He, X.; Xia, W.; Lee, D.F.; Wei, Y.; Yu, D.; Mills, G.; Shi, D.; Hung, M.C. Cytoplasmic expression of p21CIP1/WAF1 is correlated with IKKbeta overexpression in human breast cancers. Int. J. Oncol. 2006, 29, 1103–1110. [Google Scholar]
  160. Shiraki, K.; Wagayama, H. Cytoplasmic p21(WAF1/CIP1) expression in human hepatocellular carcinomas. Liver Int. 2006, 26, 1018–1019. [Google Scholar] [CrossRef]
  161. Blagosklonny, M.V. Are p27 and p21 cytoplasmic oncoproteins? Cell Cycle 2002, 1, 391–393. [Google Scholar] [CrossRef] [PubMed]
  162. Coqueret, O. New roles for p21 and p27 cell-cycle inhibitors: A function for each cell compartment? Trends Cell. Biol. 2003, 13, 65–70. [Google Scholar] [CrossRef]
  163. Deng, C.; Zhang, P.; Harper, J.W.; Elledge, S.J.; Leder, P. Mice lacking p21CIP1/WAF1 undergo normal development, but are defective in G1 checkpoint control. Cell 1995, 82, 675–684. [Google Scholar] [CrossRef]
  164. Martin-Caballero, J.; Flores, J.M.; Garcia-Palencia, P.; Serrano, M. Tumor susceptibility of p21(Waf1/Cip1)-deficient mice. Cancer Res. 2001, 61, 6234–6238. [Google Scholar] [PubMed]
  165. Brugarolas, J.; Bronson, R.T.; Jacks, T. p21 is a critical CDK2 regulator essential for proliferation control in Rb-deficient cells. J. Cell. Biol. 1998, 141, 503–514. [Google Scholar] [CrossRef] [PubMed]
  166. Yang, W.C.; Mathew, J.; Velcich, A.; Edelmann, W.; Kucherlapati, R.; Lipkin, M.; Yang, K.; Augenlicht, L.H. Targeted inactivation of the p21(WAF1/cip1) gene enhances Apc-initiated tumor formation and the tumor-promoting activity of a Western-style high-risk diet by altering cell maturation in the intestinal mucosal. Cancer Res. 2001, 61, 565–569. [Google Scholar]
  167. Franklin, D.S.; Godfrey, V.L.; O’Brien, D.A.; Deng, C.; Xiong, Y. Functional collaboration between different cyclin-dependent kinase inhibitors suppresses tumor growth with distinct tissue specificity. Mol. Cell. Biol. 2000, 20, 6147–6158. [Google Scholar] [CrossRef] [PubMed]
  168. Bearss, D.J.; Lee, R.J.; Troyer, D.A.; Pestell, R.G.; Windle, J.J. Differential effects of p21(WAF1/CIP1) deficiency on MMTV-ras and MMTV-myc mammary tumor properties. Cancer Res. 2002, 62, 2077–2084. [Google Scholar]
  169. Wang, Y.A.; Elson, A.; Leder, P. Loss of p21 increases sensitivity to ionizing radiation and delays the onset of lymphoma in atm-deficient mice. Proc. Natl. Acad. Sci. USA 1997, 94, 14590–14595. [Google Scholar] [CrossRef]
  170. De la Cueva, E.; Garcia-Cao, I.; Herranz, M.; Lopez, P.; Garcia-Palencia, P.; Flores, J.M.; Serrano, M.; Fernandez-Piqueras, J.; Martin-Caballero, J. Tumorigenic activity of p21Waf1/Cip1 in thymic lymphoma. Oncogene 2006, 25, 4128–4132. [Google Scholar] [CrossRef]
  171. Valente, L.J.; Gray, D.H.; Michalak, E.M.; Pinon-Hofbauer, J.; Egle, A.; Scott, C.L.; Janic, A.; Strasser, A. p53 efficiently suppresses tumor development in the complete absence of its cell-cycle inhibitory and proapoptotic effectors p21, Puma, and Noxa. Cell. Rep. 2013, 3, 1339–1345. [Google Scholar] [CrossRef] [PubMed]
  172. Hayflick, L. The Limited in Vitro Lifetime of Human Diploid Cell Strains. Exp. Cell Res. 1965, 37, 614–636. [Google Scholar] [CrossRef]
  173. Milanovic, M.; Yu, Y.; Schmitt, C.A. The Senescence-Stemness Alliance—A Cancer-Hijacked Regeneration Principle. Trends Cell. Biol. 2018, 28, 1049–1061. [Google Scholar] [CrossRef] [PubMed]
  174. Rodier, F.; Campisi, J. Four faces of cellular senescence. J. Cell. Biol. 2011, 192, 547–556. [Google Scholar] [CrossRef] [PubMed]
  175. Gartel, A.L. p21(WAF1/CIP1) and cancer: A shifting paradigm? Biofactors 2009, 35, 161–164. [Google Scholar] [CrossRef] [PubMed]
  176. Kreis, N.N.; Louwen, F.; Zimmer, B.; Yuan, J. Loss of p21Cip1/CDKN1A renders cancer cells susceptible to Polo-like kinase 1 inhibition. Oncotarget 2015, 6, 6611–6626. [Google Scholar] [CrossRef] [PubMed]
  177. Yuan, J.; Sanhaji, M.; Kramer, A.; Reindl, W.; Hofmann, M.; Kreis, N.N.; Zimmer, B.; Berg, T.; Strebhardt, K. Polo-box domain inhibitor poloxin activates the spindle assembly checkpoint and inhibits tumor growth in vivo. Am. J. Pathol. 2011, 179, 2091–2099. [Google Scholar] [CrossRef]
  178. Strebhardt, K.; Ullrich, A. Targeting polo-like kinase 1 for cancer therapy. Nat. Rev. Cancer 2006, 6, 321–330. [Google Scholar] [CrossRef]
  179. Louwen, F.; Yuan, J. Battle of the eternal rivals: Restoring functional p53 and inhibiting Polo-like kinase 1 as cancer therapy. Oncotarget 2013, 4, 958–971. [Google Scholar] [CrossRef]
  180. Han, Z.; Wei, W.; Dunaway, S.; Darnowski, J.W.; Calabresi, P.; Sedivy, J.; Hendrickson, E.A.; Balan, K.V.; Pantazis, P.; Wyche, J.H. Role of p21 in apoptosis and senescence of human colon cancer cells treated with camptothecin. J. Biol. Chem. 2002, 277, 17154–17160. [Google Scholar] [CrossRef]
  181. Kondo, S.; Barna, B.P.; Kondo, Y.; Tanaka, Y.; Casey, G.; Liu, J.; Morimura, T.; Kaakaji, R.; Peterson, J.W.; Werbel, B.; et al. WAF1/CIP1 increases the susceptibility of p53 non-functional malignant glioma cells to cisplatin-induced apoptosis. Oncogene 1996, 13, 1279–1285. [Google Scholar] [PubMed]
  182. Lincet, H.; Poulain, L.; Remy, J.S.; Deslandes, E.; Duigou, F.; Gauduchon, P.; Staedel, C. The p21(cip1/waf1) cyclin-dependent kinase inhibitor enhances the cytotoxic effect of cisplatin in human ovarian carcinoma cells. Cancer Lett. 2000, 161, 17–26. [Google Scholar] [CrossRef]
  183. Kreis, N.N.; Sommer, K.; Sanhaji, M.; Kramer, A.; Matthess, Y.; Kaufmann, M.; Strebhardt, K.; Yuan, J. Long-term downregulation of Polo-like kinase 1 increases the cyclin-dependent kinase inhibitor p21(WAF1/CIP1). Cell Cycle 2009, 8, 460–472. [Google Scholar] [CrossRef] [PubMed]
  184. Gordon, R.R.; Nelson, P.S. Cellular senescence and cancer chemotherapy resistance. Drug Resist. Updat. 2012, 15, 123–131. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  185. Milanovic, M.; Fan, D.N.Y.; Belenki, D.; Dabritz, J.H.M.; Zhao, Z.; Yu, Y.; Dorr, J.R.; Dimitrova, L.; Lenze, D.; Monteiro Barbosa, I.A.; et al. Senescence-associated reprogramming promotes cancer stemness. Nature 2018, 553, 96–100. [Google Scholar] [CrossRef] [PubMed]
  186. Georgakilas, A.G.; Martin, O.A.; Bonner, W.M. p21: A Two-Faced Genome Guardian. Trends Mol. Med. 2017, 23, 310–319. [Google Scholar] [CrossRef] [PubMed]
  187. Evangelou, K.; Galanos, P.; Gorgoulis, V.G. The Janus face of p21. Mol. Cell. Oncol. 2016, 3, e1215776. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  188. Medema, J.P. Cancer stem cells: The challenges ahead. Nature Cell. Biol. 2013, 15, 338–344. [Google Scholar] [CrossRef]
  189. Morris-Hanon, O.; Furmento, V.A.; Rodriguez-Varela, M.S.; Mucci, S.; Fernandez-Espinosa, D.D.; Romorini, L.; Sevlever, G.E.; Scassa, M.E.; Videla-Richardson, G.A. The Cell Cycle Inhibitors p21(Cip1) and p27(Kip1) Control Proliferation but Enhance DNA Damage Resistance of Glioma Stem Cells. Neoplasia 2017, 19, 519–529. [Google Scholar] [CrossRef]
  190. Tian, H.; Wittmack, E.K.; Jorgensen, T.J. p21WAF1/CIP1 antisense therapy radiosensitizes human colon cancer by converting growth arrest to apoptosis. Cancer Res. 2000, 60, 679–684. [Google Scholar]
  191. Kokunai, T.; Urui, S.; Tomita, H.; Tamaki, N. Overcoming of radioresistance in human gliomas by p21WAF1/CIP1 antisense oligonucleotide. J. Neurooncol. 2001, 51, 111–119. [Google Scholar] [CrossRef] [PubMed]
  192. Hwang, C.Y.; Lee, C.; Kwon, K.S. Extracellular signal-regulated kinase 2-dependent phosphorylation induces cytoplasmic localization and degradation of p21Cip1. Mol. Cell. Biol. 2009, 29, 3379–3389. [Google Scholar] [CrossRef] [PubMed]
  193. Zhang, Y.; Wang, Z.; Magnuson, N.S. Pim-1 kinase-dependent phosphorylation of p21Cip1/WAF1 regulates its stability and cellular localization in H1299 cells. Mol. Cancer Res. 2007, 5, 909–922. [Google Scholar] [CrossRef] [PubMed]
  194. Rodriguez-Vilarrupla, A.; Jaumot, M.; Abella, N.; Canela, N.; Brun, S.; Diaz, C.; Estanyol, J.M.; Bachs, O.; Agell, N. Binding of calmodulin to the carboxy-terminal region of p21 induces nuclear accumulation via inhibition of protein kinase C-mediated phosphorylation of Ser153. Mol. Cell. Biol. 2005, 25, 7364–7374. [Google Scholar] [CrossRef] [PubMed]
  195. Li, Y.; Dowbenko, D.; Lasky, L.A. AKT/PKB phosphorylation of p21Cip/WAF1 enhances protein stability of p21Cip/WAF1 and promotes cell survival. J. Biol. Chem. 2002, 277, 11352–11361. [Google Scholar] [CrossRef]
  196. Vincent, A.J.; Ren, S.; Harris, L.G.; Devine, D.J.; Samant, R.S.; Fodstad, O.; Shevde, L.A. Cytoplasmic translocation of p21 mediates NUPR1-induced chemoresistance: NUPR1 and p21 in chemoresistance. FEBS Lett. 2012, 586, 3429–3434. [Google Scholar] [CrossRef] [PubMed]
  197. Heliez, C.; Baricault, L.; Barboule, N.; Valette, A. Paclitaxel increases p21 synthesis and accumulation of its AKT-phosphorylated form in the cytoplasm of cancer cells. Oncogene 2003, 22, 3260–3268. [Google Scholar] [CrossRef] [Green Version]
  198. Perez-Tenorio, G.; Berglund, F.; Esguerra Merca, A.; Nordenskjold, B.; Rutqvist, L.E.; Skoog, L.; Stal, O. Cytoplasmic p21WAF1/CIP1 correlates with Akt activation and poor response to tamoxifen in breast cancer. Int. J. Oncol. 2006, 28, 1031–1042. [Google Scholar] [CrossRef]
  199. Koster, R.; di Pietro, A.; Timmer-Bosscha, H.; Gibcus, J.H.; van den Berg, A.; Suurmeijer, A.J.; Bischoff, R.; Gietema, J.A.; de Jong, S. Cytoplasmic p21 expression levels determine cisplatin resistance in human testicular cancer. J. Clin. Investig. 2010, 120, 3594–3605. [Google Scholar] [CrossRef] [Green Version]
  200. Xia, X.; Ma, Q.; Li, X.; Ji, T.; Chen, P.; Xu, H.; Li, K.; Fang, Y.; Weng, D.; Weng, Y.; et al. Cytoplasmic p21 is a potential predictor for cisplatin sensitivity in ovarian cancer. BMC Cancer 2011, 11, 399. [Google Scholar] [CrossRef]
  201. Maiuthed, A.; Ninsontia, C.; Erlenbach-Wuensch, K.; Ndreshkjana, B.; Muenzner, J.K.; Caliskan, A.; Husayn, A.P.; Chaotham, C.; Hartmann, A.; Vial Roehe, A.; et al. Cytoplasmic p21 Mediates 5-Fluorouracil Resistance by Inhibiting Pro-Apoptotic Chk2. Cancers 2018, 10, 373. [Google Scholar] [CrossRef] [PubMed]
  202. Zaidi, A.R.S.; Dresman, S.; Burt, C.; Rule, S.; McCallum, L. Molecular signatures for CCN1, p21 and p27 in progressive mantle cell lymphoma. J. Cell Commun. Signal. 2018. [Google Scholar] [CrossRef]
  203. Richard, F.; Pacyna-Gengelbach, M.; Schluns, K.; Fleige, B.; Winzer, K.J.; Szymas, J.; Dietel, M.; Petersen, I.; Schwendel, A. Patterns of chromosomal imbalances in invasive breast cancer. Int. J. Cancer 2000, 89, 305–310. [Google Scholar] [CrossRef]
  204. Ocker, M.; Schneider-Stock, R. Histone deacetylase inhibitors: Signalling towards p21cip1/waf1. Int. J. Biochem. Cell Biol. 2007, 39, 1367–1374. [Google Scholar] [CrossRef] [PubMed]
  205. Shen, H.; Maki, C.G. Pharmacologic activation of p53 by small-molecule MDM2 antagonists. Curr. Pharm. Des. 2011, 17, 560–568. [Google Scholar] [CrossRef]
  206. Martinez, J.D. Restoring p53 tumor suppressor activity as an anticancer therapeutic strategy. Future Oncol. 2010, 6, 1857–1862. [Google Scholar] [CrossRef] [Green Version]
  207. Rastogi, N.; Mishra, D.P. Therapeutic targeting of cancer cell cycle using proteasome inhibitors. Cell. Div. 2012, 7, 26. [Google Scholar] [CrossRef] [PubMed]
  208. Ivanovska, I.; Ball, A.S.; Diaz, R.L.; Magnus, J.F.; Kibukawa, M.; Schelter, J.M.; Kobayashi, S.V.; Lim, L.; Burchard, J.; Jackson, A.L.; et al. MicroRNAs in the miR-106b family regulate p21/CDKN1A and promote cell cycle progression. Mol. Cell. Biol. 2008, 28, 2167–2174. [Google Scholar] [CrossRef]
  209. Van Beijnum, J.R.; Giovannetti, E.; Poel, D.; Nowak-Sliwinska, P.; Griffioen, A.W. miRNAs: Micro-managers of anticancer combination therapies. Angiogenesis 2017, 20, 269–285. [Google Scholar] [CrossRef]
  210. Ghosh, D.; Venkataramani, P.; Nandi, S.; Bhattacharjee, S. CRISPR-Cas9 a boon or bane: The bumpy road ahead to cancer therapeutics. Cancer Cell Int. 2019, 19, 12. [Google Scholar] [CrossRef]
  211. Weiss, R.H.; Marshall, D.; Howard, L.; Corbacho, A.M.; Cheung, A.T.; Sawai, E.T. Suppression of breast cancer growth and angiogenesis by an antisense oligodeoxynucleotide to p21(Waf1/Cip1). Cancer Lett. 2003, 189, 39–48. [Google Scholar] [CrossRef]
  212. Fan, Y.; Borowsky, A.D.; Weiss, R.H. An antisense oligodeoxynucleotide to p21(Waf1/Cip1) causes apoptosis in human breast cancer cells. Mol. Cancer Ther. 2003, 2, 773–782. [Google Scholar] [PubMed]
  213. Park, S.H.; Park, J.Y.; Weiss, R.H. Antisense attenuation of p21 sensitizes kidney cancer to apoptosis in response to conventional DNA damaging chemotherapy associated with enhancement of phospho-p53. J. Urol. 2008, 180, 352–360. [Google Scholar] [CrossRef] [PubMed]
  214. Burnett, J.C.; Rossi, J.J.; Tiemann, K. Current progress of siRNA/shRNA therapeutics in clinical trials. Biotechnol. J. 2011, 6, 1130–1146. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  215. Moussa, R.S.; Park, K.C.; Kovacevic, Z.; Richardson, D.R. Ironing out the role of the cyclin-dependent kinase inhibitor, p21 in cancer: Novel iron chelating agents to target p21 expression and activity. Free Radic. Biol. Med. 2019, 133, 276–294. [Google Scholar] [CrossRef]
  216. Sax, J.K.; Dash, B.C.; Hong, R.; Dicker, D.T.; El-Deiry, W.S. The cyclin-dependent kinase inhibitor butyrolactone is a potent inhibitor of p21 (WAF1/CIP1 expression). Cell Cycle 2002, 1, 90–96. [Google Scholar]
  217. Park, S.H.; Wang, X.; Liu, R.; Lam, K.S.; Weiss, R.H. High throughput screening of a small molecule one-bead-one-compound combinatorial library to identify attenuators of p21 as chemotherapy sensitizers. Cancer Biol. Ther. 2008, 7, 2015–2022. [Google Scholar] [CrossRef] [Green Version]
  218. Punchihewa, C.; Inoue, A.; Hishiki, A.; Fujikawa, Y.; Connelly, M.; Evison, B.; Shao, Y.; Heath, R.; Kuraoka, I.; Rodrigues, P.; et al. Identification of small molecule proliferating cell nuclear antigen (PCNA) inhibitor that disrupts interactions with PIP-box proteins and inhibits DNA replication. J. Biol. Chem. 2012, 287, 14289–14300. [Google Scholar] [CrossRef]
  219. Hengel, S.R.; Spies, M.A.; Spies, M. Small-Molecule Inhibitors Targeting DNA Repair and DNA Repair Deficiency in Research and Cancer Therapy. Cell Chem. Biol. 2017, 24, 1101–1119. [Google Scholar] [CrossRef] [Green Version]
  220. Myrianthopoulos, V.; Evangelou, K.; Vasileiou, P.V.S.; Cooks, T.; Vassilakopoulos, T.P.; Pangalis, G.A.; Kouloukoussa, M.; Kittas, C.; Georgakilas, A.G.; Gorgoulis, V.G. Senescence and senotherapeutics: A new field in cancer therapy. Pharmacol. Ther. 2019, 193, 31–49. [Google Scholar] [CrossRef]
  221. Inoue, H.; Hwang, S.H.; Wecksler, A.T.; Hammock, B.D.; Weiss, R.H. Sorafenib attenuates p21 in kidney cancer cells and augments cell death in combination with DNA-damaging chemotherapy. Cancer Biol. Ther. 2011, 12, 827–836. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  222. Wettersten, H.I.; Hee Hwang, S.; Li, C.; Shiu, E.Y.; Wecksler, A.T.; Hammock, B.D.; Weiss, R.H. A novel p21 attenuator which is structurally related to sorafenib. Cancer Biol. Ther. 2013, 14, 278–285. [Google Scholar] [CrossRef] [PubMed] [Green Version]
Cancers 11 01220 g001 550
Figure 1. Schematic illustration delineating potential anti-cancer concepts by targeting p21. (A) Targeting oncogenic sustained p53-independent p21 nuclear expression. (B) Targeting accumulated p21 in the cytoplasm. Abbreviations: ASO, antisense oligonucleotides; Cdk, cyclin-dependent kinase; CRL2, Cullin-2 RING ubiquitin-ligase; CRISPR, clustered regularly interspaced short palindromic repeats; LRR1, leucine rich repeat 1; miRNA, microRNA; PCNA, proliferating cell nuclear antigen; p-p21, phospho-p21.
Figure 1. Schematic illustration delineating potential anti-cancer concepts by targeting p21. (A) Targeting oncogenic sustained p53-independent p21 nuclear expression. (B) Targeting accumulated p21 in the cytoplasm. Abbreviations: ASO, antisense oligonucleotides; Cdk, cyclin-dependent kinase; CRL2, Cullin-2 RING ubiquitin-ligase; CRISPR, clustered regularly interspaced short palindromic repeats; LRR1, leucine rich repeat 1; miRNA, microRNA; PCNA, proliferating cell nuclear antigen; p-p21, phospho-p21.
Cancers 11 01220 g001
Table
Table 1. Dual behavior of p21 is highly dependent on its subcellular localization (nucleus or cytoplasm). Nuclear p21 acts mainly as a tumor suppressor and cytoplasmic p21 as oncogene (* representative references are shown).
Table 1. Dual behavior of p21 is highly dependent on its subcellular localization (nucleus or cytoplasm). Nuclear p21 acts mainly as a tumor suppressor and cytoplasmic p21 as oncogene (* representative references are shown).
Function of p21LocalizationTumor Suppressor/OncogeneReferences *
Apoptosis inhibition/activationBothOncogene/Tumor suppressor[51,52]
Autophagy induction/inhibitionCytoplasm?[54,56]
Cell cycle arrestNucleusTumor suppressor[1,18]
Cell cycle progressionCytoplasmOncogene[37,39]
Chromosomal stabilityNucleusTumor suppressor[23,24,25]
DifferentiationBothBoth[63,64]
DNA synthesis inhibitionNucleusTumor suppressor[28]
DNA repair inhibition/activationNucleusBoth[40,47]
Maintaining stem cell renewalNucleusBoth[65,66,67]
Migration, cytoskeletal dynamicsCytoplasmOncogene[60,61]
Re-replicationNucleusOncogene[62]
Senescence induction/escapeNucleusBoth[57,62]
Transcriptional co-factorNucleusTumor suppressor[29,30,31]
Table
Table 2. Overview of p21’s function in normal cell differentiation.
Table 2. Overview of p21’s function in normal cell differentiation.
Cell Line/SystemAssociation of DifferentiationReferences
Erythroid progenitorsp21 promotes differentiation of normal erythroid progenitors[72]
Human MSCsp21 deficiency causes increased differentiation capacity[95,96,97]
Human PBMsIncreased cytoplasmic p21 during differentiation[63]
MEFs (iPS)p21 depletion induces reprogramming of differentiated cells[65,66,67]
MegakaryocytesHigh p53-independent p21 level in endomitotic megakaryocytes[71]
Mice (in vivo)p21 expression correlates with differentiation[76,77]
Murine pre-adipocytesp21 has a contradictory role in adipocyte differentiation[79,80,81]
Murine ATDC5p21-upregulation in early skeletal cartilage differentiation stages[85]
Murine C2C12Induction of (cytoplasmic) p21 correlates with an apoptosis-resistant phenotype of differentiating myoblasts[73,74,75]
Murine 32Dcl3p21 inhibits differentiation of granulocytes[64]
Murine ESCsIncreased p21 in ESCs accelerates differentiation into endothelial cells, hepatocytes and neurons by repressing SOX2[89]
Murine keratinocytesDecreased p21 in terminally differentiated primary keratinocytes but increased p21 at the onset of differentiation[83,84]
Murine oligodendrocytesIncreased p21 is required for establishment of differentiation[70]
Murine osteoblastsStrongly downregulated p21 during differentiation[82]
Murine trophoblastic stem cellsIncrease of cytoplasmic p21 during differentiation into apoptosis-resistant trophoblast giant cells[94]
Rat neuronsNeurite outgrowth and branching of hippocampal neurons by cytoplasmic p21[86]
Rat pancreatic myofibroblastsTranslocation of p21 from the nucleus to the cytoplasm correlates with pancreatic myofibroblast to fibroblast cell conversion[84]
tsFHIp21 is involved in early differentiation stages in human fetal intestinal epithelial cell line[78]
Abbreviations: ESCs, embryonic stem cells; iPS, induced pluripotent stem cells; MEFs, mouse embryonic fibroblasts; MSCs, mesenchymal stem cells; PBMs, peripheral blood monocytes; SOX2, sex determining region Y-box 2; tsFHI, temperature-sensitive fetal human intestinal.

Share and Cite

MDPI and ACS Style

Kreis, N.-N.; Louwen, F.; Yuan, J. The Multifaceted p21 (Cip1/Waf1/CDKN1A) in Cell Differentiation, Migration and Cancer Therapy. Cancers 2019, 11, 1220. https://doi.org/10.3390/cancers11091220

AMA Style

Kreis N-N, Louwen F, Yuan J. The Multifaceted p21 (Cip1/Waf1/CDKN1A) in Cell Differentiation, Migration and Cancer Therapy. Cancers. 2019; 11(9):1220. https://doi.org/10.3390/cancers11091220

Chicago/Turabian Style

Kreis, Nina-Naomi, Frank Louwen, and Juping Yuan. 2019. "The Multifaceted p21 (Cip1/Waf1/CDKN1A) in Cell Differentiation, Migration and Cancer Therapy" Cancers 11, no. 9: 1220. https://doi.org/10.3390/cancers11091220

APA Style

Kreis, N. -N., Louwen, F., & Yuan, J. (2019). The Multifaceted p21 (Cip1/Waf1/CDKN1A) in Cell Differentiation, Migration and Cancer Therapy. Cancers, 11(9), 1220. https://doi.org/10.3390/cancers11091220

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop