The plant alkaloid cryptolepine induces p21WAF1/CIP1 and cell cycle arrest in a human osteosarcoma cell line

Abstract

Abstract. We previously established a bioassay method to screen for compounds that activate the promoter activity of p21 WAF1/CIP1 , a potent inhibitor of cyclin-dependent kinases, in a p53-independent manner. As an activator of p21 WAF1/CIP1promoter activity, we isolated cryptolepine (CLP: 5-methyl indolo (2,3b)-quiniine), an indoloquinoline alkaloid, from the traditional Ayurvedic medicinal plant Sida cordifolia. We show here that CLP induces the expression of p21 WAF1/CIP1with growth arrest in p53-mutated human osteosarcoma MG63 cells. Four micromolar of CLP completely inhibited the growth of MG63 cells and caused G 2 /M-phase arrest. CLP up-regulated the expression of p21 WAF1/CIP1 at both mRNA and protein levels in a dose-dependent manner. Using several mutant p21 WAF1/CIP1 promoter constructs, we found that the CLP-responsive element is an Sp1 site at -82 relative to the transcription start site of the p21 WAF1/CIP1 promoter. These findings suggest that CLP arrests the growth of MG63 cells by activating the p21 WAF1/CIP1 promoter through the specific Sp1 site in a p53-independent manner. In addition, CLPmediated cell cycle arrest was reduced by the knockout of the p21 WAF1/CIP1 gene in human colon cancer HCT116 cells, suggesting that the cell cycle arrest by CLP was at least partially mediated through the induction of p21expression. Although we need further study of chemotherapeutic effect in vivo, these results raise the possibility that CLP might be a suitable chemotherapeutic agent for treatment of osteosarcoma. IntroductionOsteosarcoma is the most common form of primary malignant bone tumor that mainly occurs in juvenile patients. Although the development of effective adjuvant regimen of chemotherapy has substantially improved the prognosis of patients (1-4), >20% of patients still die as a result of tumor metastasis (5-10). One of the most serious causes of therapeutic failure is the resistance of the tumor cells to chemotherapeutic agents (11,12). To overcome the drug resistance, new anti-tumor agents or chemicals and new anti-tumor therapeutic approaches need to be developed. p21 WAF1/CIP1 is a member of the cyclin-dependent kinase inhibitors and induces G 1 -and G 2 /M-phase cell cycle arrest (13-16). p21 WAF1/CIP1 induces differentiation of both normal and transformed cells and suppresses the growth of malignant cells, in vitro and in vivo (17,18 WAF1/CIP1 is an attractive molecular target to suppress cell growth even in p53-mutated cancer cells and p21 WAF1/CIP1 -inducing agents by a p53-independent pathway might be effective for chemotherapy of osteosarcoma with poor prognosis.Recently, we established a bioassay method using p53-negative human osteosarcoma MG63 cells to screen for compounds that activate the p21 WAF1/CIP1 promoter activity in a p53-independent manner (23). As an activator of p21 WAF1/CIP1 promoter activity, we isolated cryptolepine (CLP: 5-methyl indolo (2,3b)-quiniine), an indoloquinoline alkaloid, from the traditional Ayurvedic medicinal plant Sida cordifolia. CLP was first is...

Anticancer Effect of Celecoxib via COX-2 Dependent and Independent Mechanisms in Human Gastric Cancers Cells

“…The p21 gene expression product, p21 protein, is an inhibitor of cyclin cell-cycle dependent kinase. Recent studies have shown that cyclins and their related cyclin-dependent kinases (CDKs) play important roles in the control of the cell cycle [25,26]. Another study investigated the expression of cell-cycle mediators in ovarian cancer cells after transfection with p21 and p53, and found that cell growth was inhibited after transfection [27].…”

Characterization of the cytotoxic activity of the indoloquinoline alkaloid cryptolepine in human tumour cell lines and primary cultures of tumour cells from patients

“…Although cryptolepine has not been comprehensively studied with respect to anti-cancer activity, in vitro studies in tumour cell lines have found it to intercalate into DNA, inhibit topoisomerase II, induce p21WAF1/CIP1 and thereby elicit cell cycle arrest and apoptotic cell death [10,11].…”

Total synthesis of natural products containing benzofuran rings

“…315,316 The total synthesis of 336 was achieved as illustrated in Scheme 72. The total synthesis started from coupling of 1,5-naphthalenediol mono-methoxymethyl (MOM) ether 329 with bromobenzoquinone 330 mediated by K 2 CO 3 in DMSO 317 to afford a C-O coupling product.…”