Molecular mechanisms of G0/G1 cell-cycle arrest and apoptosis induced by terfenadine in human cancer cells

Terfenadine (TF), a highly potent histamine H1 receptor antagonist, has been shown to exert no significant central nervous system side effects in clinically effective doses. In this study, we demonstrated that TF induced significant growth inhibition of human cancer cells, including Hep G2, HT 29, and COLO 205 cells, through induction of G(0)/G(1) phase cell-cycle arrest. The minimal dose of TF induced significant G(0)/G(1) arrest in these cells was 1-3 microM. The protein levels of p53, p21/Cip1, and p27/Kip1 were significantly elevated, whereas the kinase activities of cyclin-dependent kinase 2 (CDK2) and CDK4 were inhibited simultaneously in the TF-treated cells. On the other hand, significant apoptosis, but not G(0)/G(1) arrest, was induced in the HL 60 (p53-null) or Hep 3B (with deleted p53) cells when treated with TF (3-5 microM). To clarify the roles of p21/Cip1 and p27/Kip1 protein expression, which was involved in G(0)/G(1) arrest and apoptosis induced by TF in human cancer cells, antisense oligodeoxynucleotides (ODNs) specific to p21/Cip1 and p27/Kip1 were used, and the expression of the p21/Cip1 and p27/Kip1 were monitored by immunoblotting analysis. Our data demonstrated that the percentage of the apoptotic cells detected by annexin V/PI analysis in the TF-treated group was clearly attenuated by pretreatment with p27/Kip1-specific ODNs. These results indicated that p27/Kip1 (but not p21/Cip1) protein indeed played a critical role in the TF-induced apoptosis. We also demonstrated that the TF-induced G(0)/G(1) cell-cycle arrest effect was not reversed by TF removal, and this growth inhibition lasted for at least 7 d. Importantly, the occurrence of apoptosis and cell growth arrest was not observed in the TF-treated normal human fibroblast, even at a dose as high as 25 microM. Our study showed the molecular mechanisms for TF-induced cell growth inhibition and the occurrence of apoptosis in human cancer cells.     

p21(Waf1/Cip1) and p53 are downstream effectors of protein kinase C delta in tumor suppression and differentiation in human colon cancer cells

We have previously demonstrated that the delta isoform of protein kinase C (PKCdelta) is importantly involved in cell growth inhibition and tumor suppression in colon cancer cells. To investigate further the activity and mechanism of action of PKCdelta, we have retrovirally transduced a PKCdelta cDNA in HCT116 human colon cancer cells. PKCdelta-overexpressing cells (HCT116/PKCdelta) were growth-inhibited, showed marked morphologic changes and underwent multinucleation and phenotypic changes characteristic of mitotic catastrophe. Compared to controls, HCT116/PKCdelta cells showed a highly attenuated tumorigenic profile and poor anchorage-independent growth. In addition, transfected cells established junction-coordinated intercellular communications, expressed cell surface microvilli and overexpressed the colon differentiation marker alkaline phosphatase. HCT116/PKCdelta cells also produced the 89 kDa, carboxy-terminal catalytic domain of PARP. In HCT116/PKCdelta cells, p21(Waf1/Cip1) and p53 were transiently upregulated for 48 hr after PKCdelta transduction. In a p21 null subline of HCT116 cells (HCT116/p21null), overexpression of PKCdelta did not affect tumorigenicity or differentiation, indicating that p21 is essential for the antitumorigenic activity of PKCdelta. Similarly, overexpression of PKCdelta caused no significant phenotypic changes in HCT116/E6 cells, an HCT116 subline in which the p53 protein is downregulated by the human papillomavirus E6 gene product. We conclude that overexpression of PKCdelta in human colon cancer cells induces multiple antineoplastic effects that depend on the activities of p21(Waf1/Cip1) and p53.     

Antitumor effects of novel compound,guttiferone K,on colon cancer by p21Waf1/Cip1‐mediated G0/G1 cell cycle arrest and apoptosis

Low selectivity is one of the major problems of currently used anticancer drugs, therefore, there is a high demand for novel, selective antitumor agents. In this study, the anticancer effects and mechanisms of guttiferone K (GUTK), a novel polyprenylated acylphloroglucinol derivative isolated from Garcinia cowa Roxb., were examined for its development as a novel drug targeting colon cancer. GUTK concentration‐ and time‐dependently reduced the viability of human colon cancer HT‐29 cells (IC₅₀ value 5.39 ± 0.22 μM) without affecting the viability of normal human colon epithelial CCD 841 CoN cells and induced G₀/G₁ cell cycle arrest in HT‐29 cells by down‐regulating cyclins D1, D3 and cyclin‐dependent kinases 4 and 6, while selectively restoring p21Waf1/Cip1 and p27Kip1 to levels comparable to those observed in normal colon cells, without affecting their levels in normal cells. GUTK (10.0 μM) induced cleavage of PARP, caspases‐3, ‐8 and ‐9 and chromatin condensation to stimulate caspase‐3‐mediated apoptosis. The addition of a JNK inhibitor, SP600125, partially reversed GUTK‐induced caspase‐3 activity, indicating the possible involvement of JNK in GUTK‐induced apoptosis. Furthermore, GUTK (10 mg/kg, i.p.) significantly decreased the tumor volume in a syngeneic colon tumor model when used alone or in combination with 5‐fluorouracil without toxicity to the mice. Immunohistochemical staining of the tumor sections revealed a mechanism involving an increase in cleaved caspase‐3 and a decrease in cell proliferation marker Ki‐67. Our results support GUTK as a promising novel, potent and selective antitumor drug candidate for colon cancer.     

Up-regulation of p21WAF1/Cip1 by saRNA induces G1-phase arrest and apoptosis in T24 human bladder cancer cells

Very recent studies have reported that chemically synthesized small duplex RNAs complementary to the promoters of target genes can activate gene expression in different cancer cell lines. Such dsRNA have been referred to as saRNA for small activating RNA. The present study was conducted to evaluate the potential of p21(WAF1/Cip1) (p21) induction by small activating RNA targeting the p21 promoter in the treatment of bladder cancer. Using T24 human bladder cancer cells, we found that p21 saRNA caused dose- and time-dependent inhibition of cell proliferation and viability which was associated with induced G1-phase cell cycle arrest and apoptosis. The decreased anti-apoptotic protein Bcl-xL and activation of caspase-3 and PARP also supported the efficacy of the treatment. These data suggest that up-regulation of p21 by saRNA may be an effective way for treating human bladder and other types of cancers.     

Urinary Bladder Lesions after the Chernobyl Accident: Immunohistochemical Assessment of p53, Proliferating Cell Nuclear Antigen, Cyclin D1 and p21WAF1/Cip1

During the 11-year period subsequent to the Chernobyl accident, the incidence of urinary bladder cancer in Ukraine has increased from 26.2 to 36.1 per 100,000 population. Cesium-137 (¹³⁷Cs) accounts for 80–90% of the incorporated radioactivity in this population, which has been exposed to long-term, low-dose ionizing radiation, and 80% of the more labile pool of cesium is excreted via the urine. The present study was performed to evaluate the histopathological features and the immunohistochemical status of p53, p21^WAF1/Cip1, cyclin D1 and PCNA (proliferating cell nuclear antigen) in urinary bladder mucos a of 55 males (49-92 years old) with benign prostatic hyperplasia who underwent surgery in Kiev, Ukraine, in 1995 and 1996. Group I (28 patients) inhabiting radiocontaminated areas of the country, group II (17 patients) from Kiev city with less radiocontamination and a control group III (10 patients) living in so-called "clean" areas of Ukraine were compared. In groups I and II, an increase in multiple areas of moderate or severe dysplasia or carcinoma in situ was seen in 42 (93%) of 45 cases. In addi tion, two small transitional cell carcinomas were found in one patient in each of groups I and II. Nuclear accumulation of p53, PCNA, cyclin D1, and to a lesser extent p21^WAF1/Cip1, was significantly increased in both groups I and II as compared with the control group III, indicating possible transformation events or enhancement of repair activities, that may precede the defect in the regulatory pathway itself, at least in the G1 phase of the cell cycle. Our results suggest that early malignant transformation is taking place in the bladder urothelium of people in the radiocontaminated areas of Ukraine and that this could possibly lead sometime in the future to an increased incidence of urinary bladder cancer.     

Interferon Regulatory Factor 1 (IRF-1) induces p21(WAF1/CIP1) dependent cell cycle arrest and p21(WAF1/CIP1) independent modulation of survivin in cancer cells

We have shown that the ectopic expression of Interferon Regulatory Factor 1 (IRF-1) results in human cancer cell death accompanied by the down-regulation of the Inhibitor of Apoptosis Protein (IAP) survivin and the induction of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1). In this report, we investigated the direct role of p21 in the suppression of survivin. We show that IRF-1 down-regulates cyclin B1, cdc-2, cyclin E, E2F1, Cdk2, Cdk4, and results in p21-mediated G1 cell cycle arrest. Interestingly, while p21 directly mediates G1 cell cycle arrest, IRF-1 or other IRF-1 signaling pathways may directly regulate survivin in human cancer cells.     

Involvement of Ras/Raf-1/ERK actions in the magnolol-induced upregulation of p21 and cell-cycle arrest in colon cancer cells

Previously, we showed that magnolol induces cell-cycle arrest in cultured colon and liver cancer cells through an upregulation of the p21 protein. The aim of this study was to delineate the molecular mechanism underlying this magnolol-induced increase of p21 protein. Thus our RT-PCR analysis demonstrated that the mRNA levels of p21 were increased at 1 h after magnolol treatment and sustained for at least 24 h. The p21 promoter activity was also increased by magnolol treatment. Western blot analysis demonstrated that treatment of COLO-205 cells with magnolol increased the levels of phosphorylation of extracellular signal-regulated kinase (ERK). Pretreatment of the cells with PD98059 abolished the magnolol-induced upregulation of p21 protein, suggesting the involvement of an ERK pathway in the magnolol-induced upregulation of p21 in COLO-205 cells. Ras inhibitor peptide abolished the magnolol-induced increase of phosphorylated ERK protein levels, increase of p21 protein, and decrease of thymidine incorporation. Moreover, treatment of COLO-205 with magnolol increased the phosphorylated Raf-1 protein (the Ras target molecule). Pretreatment of the cells with Raf-1 inhibitor reversed the magnolol-induced decrease in thymidine incorporation. Treatment of the cells with CaM kinase inhibitor, but not protein kinase A (PKA) inhibitor or phosphatidylinosital 3-kinase (PI3K) inhibitor, abolished the magnolol-induced activation of ERK and decrease of thymidine incorporation. Taken together, our results suggest that magnolol activates ERK phosphorylation through a Ras/Raf-1-mediated pathway. Subsequently, p21 expression is increased, and finally thymidine incorporation is decreased.     

A p21(waf1)-independent pathway for inhibitory phosphorylation of cyclin-dependent kinase p34(cdc2) and concomitant G(2)/M arrest by the chemopreventive flavonoid apigenin.

Apigenin, a nonmutagenic flavonoid, has been shown to inhibit ultraviolet light-induced skin tumorigenesis when topically applied to mouse skin. Our previous studies have shown that apigenin treatment of cultured mouse keratinocytes induces G(2)/M arrest accompanied by an increase in p53 protein stability and expression of p21(waf1). In this study, we determined whether the G(2)/M arrest induced by apigenin was dependent upon the presence of the cyclin dependent kinase inhibitor p21(waf1). We exposed WWT.8 (p21(waf1) wild-type) and WKO.16 (p21(waf1) null) mouse keratinocytes to various doses of apigenin for 24 h and observed G(2)/M arrest in both cell lines, thereby establishing that the apigenin-induced G(2)/M arrest was p21(waf1) independent. A 4-h treatment with apigenin induced increases in p53 protein level by sixfold and tenfold in the WWT.8 p21(waf1) wild-type cells and WKO.16 p21(waf1) null cells, respectively. After 24 h in WWT.8 cells, p21(waf1) protein also was induced in a dose-dependent manner, but it was not expressed in WKO.16 keratinocytes. We then measured the effect of apigenin treatment on the mammalian homologue of the yeast cdc2 gene (p34(cdc2)) cyclin-dependent kinase and cyclin B1 (cycB1), because these proteins complex to regulate G(2)/M progression. Apigenin treatment decreased the protein level of p34(cdc2), and p34(cdc2) kinase activity was inhibited in both p21(waf1)(+/+) and p21(waf1)(-/-) cell lines by approximately 40%. The inhibition of p34(cdc2) kinase activity by apigenin treatment correlated with increasing levels of p34(cdc2) phosphorylation at Tyr15, a site in the p34(cdc2) kinase that undergoes inhibitory phosphorylation by Wee1 kinase. Apigenin treatment also had no effect on the protein level or activity of the competing phosphatase, cdc25c, which dephosphorylates p34(cdc2) kinase at Tyr15. Apigenin had little effect on the accumulation of cycB1 protein. These results supported the conclusion that G(2)/M arrest induced by apigenin was accompanied by inhibition of the p34(cdc2) cyclin-dependent kinase protein level and activity in a p21(waf1)-independent manner.     

Caffeic acid phenethyl ester induced cell cycle arrest and growth inhibition in androgen-independent prostate cancer cells via regulation of Skp2, p53, p21Cip1 and p27Kip1

Prostate cancer (PCa) patients receiving the androgen ablation therapy ultimately develop recurrent castration-resistant prostate cancer (CRPC) within 1–3 years. Treatment with caffeic acid phenethyl ester (CAPE) suppressed cell survival and proliferation via induction of G1 or G2/M cell cycle arrest in LNCaP 104-R1, DU-145, 22Rv1, and C4–2 CRPC cells. CAPE treatment also inhibited soft agar colony formation and retarded nude mice xenograft growth of LNCaP 104-R1 cells. We identified that CAPE treatment significantly reduced protein abundance of Skp2, Cdk2, Cdk4, Cdk7, Rb, phospho-Rb S807/811, cyclin A, cyclin D1, cyclin H, E2F1, c-Myc, SGK, phospho-p70S6kinase T421/S424, phospho-mTOR Ser2481, phospho-GSK3α Ser21, but induced p21^Cip1, p27^Kip1, ATF4, cyclin E, p53, TRIB3, phospho-p53 (Ser6, Ser33, Ser46, Ser392), phospho-p38 MAPK Thr180/Tyr182, Chk1, Chk2, phospho-ATM S1981, phospho-ATR S428, and phospho-p90RSK Ser380. CAPE treatment decreased Skp2 and Akt1 protein expression in LNCaP 104-R1 tumors as compared to control group. Overexpression of Skp2, or siRNA knockdown of p21^Cip1, p27^Kip1, or p53 blocked suppressive effect of CAPE treatment. Co-treatment of CAPE with PI3K inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 or Bcl-2 inhibitor ABT737 showed synergistic suppressive effects. Our finding suggested that CAPE treatment induced cell cycle arrest and growth inhibition in CRPC cells via regulation of Skp2, p53, p21^Cip1, and p27^Kip1.     

p21~(WAF1/CIP1)在乳腺癌中的表达及意义

目的　探讨p2 1WAF1/CIP1蛋白在乳腺癌中表达的临床意义。方法　运用免疫组化SP法半定量检测p2 1蛋白在癌旁正常乳腺组织、乳腺癌组织中的表达。结果　p2 1蛋白表达位于细胞核 ,呈棕黄色。在 2 0例癌旁正常乳腺组织中 ,无p2 1蛋白表达。在 69例乳腺癌组织中有 3 0例p2 1蛋白阳性表达。在乳腺癌组织中 ,随组织学分级升高 ,p2 1阳性率下降 (P <0 0 5 ) ,随临床分期升高 ,p2 1阳性率下降 (P <0 0 5 )。有淋巴结转移组p2 1阳性率低于无淋巴结转移组 (P <0 0 5 )。p2 1蛋白阳性表达者术后 5年无瘤生存率高于p2 1蛋白阴性者术后 5年无瘤生存率 (P <0 0 5 )。结论　p2 1蛋白可用来评估乳腺癌细胞分化情况及转移潜能 ,可判断乳腺癌患者预后。     

Relative non-steroidal anti-inflammatory drug (NSAID) antiproliferative activity is mediated through p21-induced G1 arrest and E2F inhibition

This study was performed to compare the relative antineoplastic activity of 10 different non-steroidal anti-inflammatory drugs (NSAIDs) in clinical use, and to investigate the underlying mechanisms of this activity in a squamous cell carcinoma of the head and neck model (SCCHN). A standard 5-day MTT assay was used to calculate IC(50) values in UM-SCC-1 cells for 10 NSAIDs, including celecoxib, rofecoxib, sulindac sulfide, sulindac sulfone, indomethacin, ketoprofen, flurbiprofen, naproxen, piroxicam, and aspirin. Celecoxib, a COX-2 specific inhibitor, was by far the most potent NSAID, with an IC(50) of 39.9 +/- 1.1 microM, followed by sulindac sulfide (116.5 +/- 2.34 microM). Celecoxib and sulindac sulfide also induced more activation of caspase-3 than any other NSAID. Cell cycle analysis showed that celecoxib and sulindac sulfide both induced a 3-fold increase in G(1) phase distribution, and this correlated with strong induction of p21(waf1/cip1), inhibition of cyclin D1, and hypophosphorylation of Rb. Celecoxib and sulindac sulfide treatment induced strong downstream inhibition of E2F transactivating activity as determined by a luciferase reporter assay. These data demonstrate the wide range of activity of various NSAID agents, and reveal a mechanism of action through cell cycle inhibition and induction of apoptosis.     

Inhibitor of cyclooxygenase-2 induces cell-cycle arrest in the epithelial cancer cell line via up-regulation of cyclin dependent kinase inhibitor p21

Cyclooxygenase-2 is the rate-limiting enzyme in synthesis of prostaglandins and other eicosanoids. Prior reports have shown that inhibition of cyclooxygenase-2 activity, either by selective inhibitors or by antisense oligonucleotide, results in suppression of growth of squamous cell carcinoma cell lines which express high cyclooxygenase-2 levels, such as NA, a cell line established from a squamous cell carcinoma of the tongue. To investigate the mechanisms by which cyclooxygenase-2 inhibitors suppressed growth of these cells, the effects of NS-398, the selective cyclooxygenase-2 inhibitor, on cell-cycle distribution were examined. NS-398 induced G0/G1 cell-cycle arrest in NA cells which expressed cyclooxygenase-2. G0/G1 arrest induced by NS-398 was accompanied by up-regulation of cyclin-dependent kinase inhibitor p21, but not by up-regulation of the other cyclin-dependent kinase inhibitors. Transfection with p21 antisense oligonucleotide inhibited cell-cycle arrest induced by NS-398. Accumulation in G0/G1 was also observed in NA cells transfected with cyclooxygenase-2 antisense oligonucleotide. On the other hand, NS-398-treated NA cells showed a loss of plasma membrane asymmetry, a marker of early events in apoptosis. However, NS-398 did not induce other morphological and biochemical changes related to apoptotic cell death. These results suggest that cyclooxygenase-2 inhibitor induces G0/G1 cell-cycle arrest in NA cells by up-regulation of p21. Our results also suggest that NS-398 is not sufficient to complete the whole process of apoptosis in NA cells, although it induces an early event in apoptosis.     

DNA damage uncouples the mitogenic response to IGF-I in MCF-7 malignant breast cancer cells by switching the roles of PI3 kinase and p21WAF1/Cip1

In addition to its mitogenic and survival actions, recent evidence indicates that IGF-I can enhance DNA repair, implying IGF activity may limit the efficacy of many therapeutic strategies that rely on induction of DNA damage. Although the individual pathways by which DNA damage and IGF-I act are well understood, the cross-talk between these signaling events is not well defined. We examined the effects of DNA damage on the IGF-I response of MCF-7 breast cancer cells. Cells were exposed to the UV mimetic, 4-NQO, or the gamma-irradiation mimetic and chemotherapeutic drug, bleomycin; cellular proliferation was assessed by cell counting, tritiated thymidine incorporation and FACS cell cycle analysis. Although IGF-I acutely suppressed the p53 response to both agents, it subsequently enhanced the chronic increase in p53 and p21(WAF1/Cip1), resulting in cell cycle arrest; however, no apoptosis was observed. Use of specific inhibitors demonstrated that PI3 kinase was utilized with p38 MAPK to induce the p53 response to DNA damage, but was also utilized by IGF-I to diminish the acute p53 response. In addition, p21WAF1/Cip1 was increased by IGF-I, which has previously been shown to contribute to the mitogenic response. Here we demonstrate that with DNA damage IGF-I can also enhance the chronic p53-dependent increase in p21WAF1/Cip1, culminating in growth arrest. Overall, we have shown that PI3 kinase and p21WAF1/Cip1 play dual roles in mediating the mitogenic response to IGF-I, but these are both switched by cellular DNA damage to mediate a growth arrest.     

Tetrazolium violet induces G0/G1 arrest and apoptosis in brain tumor cells

Summary Tetrazolium violet (TV), a potent anticancer agent, has been shown to induce cell growth-inhibition in tumor cells. However, the related mechanism has not been revealed yet. In this report we assessed the influence of TV on cell growth and cell cycle in brain tumor cells. Treatment of C6 tumor cells with TV (5–15 μM for 24–72 h) resulted in a growth inhibition in a dose and time-dependent manner and G0/G1 phase arrest, determined by flow cytometry analysis. These effects were accompanied by apoptosis other than necrosis, evidenced by nuclear condensation, terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay and trypan blue exclusion assay plus lactate dehydrogenase (LDH) release assay. Treatment of cells with TV at 15 μM for 24 h resulted in an increase in the activity of caspase-3, evidenced by colorimetric assay, and a dramatic up-regulation of p53, accompanied with a significant increase of Bax/Bcl-2 ratio, as evidenced by immunofluorescence assay. These results suggest that TV induces growth inhibition of C6 cells through p53-midiated apoptotic pathway and G0/G1 checkpoint mechanism. Although detailed mechanisms remain to be explored, selective blockage of tumor cells in G0/G1 phase accompanied by p53-associated apoptosis makes tetrazolium violet a promising anticancer agent, meriting further investigations.     

N-myc downstream regulated gene1/Cap43 overexpression suppresses tumor growth by hepatic cancer cells through cell cycle arrest at the G0/G1 phase

N-myc downstream regulated gene-1 (NDRG1)/Cap43 regulates tumor growth and metastasis in various carcinomas. In this study we examined whether and how NDRG1/Cap43 modulates tumor growth by human hepatocellular carcinoma (HCC) cells. NDRG1/Cap43 cDNA was used to transfect HCC cell lines (KIM-1), and stable transfectants overexpressing NDRG1/Cap43 (KIM-1/Cap43) were obtained. Cell cycle analysis showed that KIM-1/Cap43 cells were arrested in the G0/G1 phase. Western blot analysis demonstrated an increase in p21 in KIM-1/Cap43 cells in culture under full confluency as compared with KIM-1/Mock. When KIM-1 cells, which are very low in NDRG1/Cap43 expression, were treated with mimosine, a G0/G1 cell cycle blocker, expression of NDRG1/Cap43 was induced in a dose dependent manner, together with p21 induction and CDK4 reduction. In vivo, KIM-1/Cap43 cells showed markedly decreased tumor growth rates compared with those of KIM-1/Mock. Immunohistochemical staining demonstrated markedly higher p21 labeling index in the KIM-1/Cap43 tumor than KIM-1/Mock tumor, and lower CDK4 and Ki-67 labeling index in the KIM-1/Cap43 than KIM-1/Mock. In order to confirm suppressive effects of NDRG1/Cap43, we further established a stable transfectant expressing NDRG1/Cap43 (HAK-1B/Cap43) using another HCC cell line, HAK-1B. Western blot analysis demonstrated an increase in p21 and a decrease in CDK4 in HAK-1B/Cap43 cells in culture under full confluency as compared with HAK-1B/Mock. HAK-1B/Cap43 also showed decreased tumor growth rates as compared with its control counterpart in vivo. NDRG1/Cap43 overexpression thus induced cell cycle arrest at the G0/G1 phase accompanied by increased p21 and decreased CDK4 expression in HCC cells. NDRG1/Cap43 might play a key role in the cell cycle control of G0/G1 in HCC cells.     

Lycorine hydrochloride inhibits melanoma cell proliferation,migration and invasion via down-regulating p21Cip1/WAF1

Lycorine hydrochloride (LH) is an active ingredient sourced from the medicinal herb Lycoris radiata. Previous studies have suggested that LH exerts tumor suppression activity in several human cancers. However, the anti-cancer effect of LH in melanoma and the potential molecular mechanisms still need to be further studied. p21^Cip1/WAF1, unlike its traditional cyclin-dependent kinase (CDK) inhibitor role, is believed to act as an oncogene under certain cellular conditions. In this research, an increased expression of p21^Cip1/WAF1 was found in human melanoma tissues and positively related to the tumor invasion depth. High level of p21^Cip1/WAF1 was found to correlate with bad outcomes of melanoma patients by Kaplan-Meier survival analysis. Functional experiments demonstrated that the proliferation, migration and invasion ability of A375 and MV3 melanoma cells was powerfully inhibited by LH through inducing S phase cell cycle arrest and regulating epithelial-mesenchymal transition (EMT). In NOD/SCID mice model, LH effectively inhibited the xenograft tumor growth and lung metastasis of A375 cells. Further research revealed that LH reduced p21^Cip1/WAF1 protein by accelerating its ubiquitination. Importantly, the LH-induced suppression of cell proliferation and metastasis was rescued by p21^Cip1/WAF1 overexpression, both in vitro an in vivo. Taken together, LH, which suppresses the proliferation and metastasis of melanoma cells via down-regulating p21^Cip1/WAF1, is expected to be developed as an effective medicine for melanoma therapy.