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Valproic acid is an anticonvulsant, which is also widely used for treating psychiatric disorders. Some clinical trials have demonstrated benefits of valproic acid augmentation therapy in schizophrenia. Interindividual variability in valproic acid dose and serum concentration may reflect functional consequences of genetic polymorphisms in genes encoding drug-metabolizing enzymes. The aim of this study was to determine the relationship between serum concentrations of valproic acid and single nucleotide polymorphisms of the cytochrome P450 (CYP) 2C19 gene in patients with schizophrenia. All patients had been receiving fixed dose of valproic acid for at least 2 weeks. The daily doses were 0.5–1.5 g. No other drugs except olanzapine were coadministered. Serum concentrations of valproic acid were measured using the ultra-high performance liquid chromatography method with mass-spectrometric detection. The CYP2C19 (CYP2C19*2 G681A rs4244285 and CYP2C19*3 G636A rs4986893) genotypes were identified by real-time PCR analyses. The mean concentration/dose ratios of valproic acid were significantly higher in patients with CYP2C19 *1/*2 genotype (P CYP2C19 *2/*3 genotype (P CYP2C12 *1/*1 genotype. The mean concentration/dose ratios of valproic acid were significantly higher in patients with 1 (P P CYP2C19 than in those without mutated alleles. And the post hoc analysis revealed that the result has acceptable statistical (power (1 – β) = 0.8486 at type I level of 0.05) to support the observed significant associations for CYP2C19 SNPs and serum C/D ratios of valproic acid. The findings of this study suggest that the genetic polymorphisms of CYP2C19 significantly affect the steady-state serum concentrations of valproic acid in Chinese Han population. The determination of the CYP2C19 genotypes may be useful for dosing adjustment in schizophrenia patients on valproic acid therapy.

The development of CRISPR-based barcoding methods creates an exciting opportunity to understand cellular phylogenies. We present a compact, tunable, high-capacity Cas12a barcoding system called dual acting inverted site array (DAISY). We combined high-throughput screening and machine learning to predict and optimize the 60-bp DAISY barcode sequences. After optimization, top-performing barcodes had ∼10-fold increased capacity relative to the best random-screened designs and performed reliably across diverse cell types. DAISY barcode arrays generated ∼12 bits of entropy and ∼66,000 unique barcodes. Thus, DAISY barcodes-at a fraction of the size of Cas9 barcodes-achieved high-capacity barcoding. We coupled DAISY barcoding with single-cell RNA-seq to recover lineages and gene expression profiles from ∼47,000 human melanoma cells. A single DAISY barcode recovered up to ∼700 lineages from one parental cell. This analysis revealed heritable single-cell gene expression and potential epigenetic modulation of memory gene transcription. Overall, Cas12a DAISY barcoding is an efficient tool for investigating cell-state dynamics.

Gene editing is a powerful tool for genome and cell engineering. Exemplified by CRISPR–Cas, gene editing could cause DNA damage and trigger DNA repair processes that are often error-prone. Such unwanted mutations and safety concerns can be exacerbated when altering long sequences. Here we couple microbial single-strand annealing proteins (SSAPs) with catalytically inactive dCas9 for gene editing. This cleavage-free gene editor, dCas9–SSAP, promotes the knock-in of long sequences in mammalian cells. The dCas9–SSAP editor has low on-target errors and minimal off-target effects, showing higher accuracy than canonical Cas9 methods. It is effective for inserting kilobase-scale sequences, with an efficiency of up to approximately 20% and robust performance across donor designs and cell types, including human stem cells. We show that dCas9–SSAP is less sensitive to inhibition of DNA repair enzymes than Cas9 references. We further performed truncation and aptamer engineering to minimize its size to fit into a single adeno-associated-virus vector for future application. Together, this tool opens opportunities towards safer long-sequence genome engineering.

The detection of intracellular nitroreductase (NTR) activity is important for the study of hypoxia in organisms. In the present study, a Rhodol-derived fluorescent chemosensor (Rhod-NO

S-nitrosylation is a posttranslational modification of protein cysteine residues leading to the formation of S-nitrosothiols and its detection is crucial to understanding of redox regulation and NO-based signaling. Prototypical detection methods for S-nitrosylation are always carried out ex situ. However, the reversible nature and the tendency of transnitrosylation highlight the necessity of its probing in intact live biological contexts. Herein we provide a fluorogenic chemical probe for the detection of S-nitrosylation in live endothelial cells. The probe is weakly emissive alone and becomes highly fluorescent only after undergoing a reaction with S-nitrosothiols in live cellular environments. This probe features high degrees of specificity and desirable sensitivity. Furthermore, it has been successfully applied to image the dynamic change of protein S-nitrosylation in live endothelial cells. The applicability of the probe in complex biological systems has been additionally verified by imaging a known target of S-nitrosylation, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in live cells. Due to the versatility exemplified, this probe holds great promise for exploring the role of protein S-nitrosylation in the pathophysiological process of a variety of vascular diseases.

miRNAs have been proposed to be key regulators of progression and metastasis in cancer. However, an understanding of their roles and molecular mechanisms is needed to provide deeper insights for better therapeutic opportunities. In this study we investigated the role and mechanism of miR-493 in the development and progression of nonsmall-cell lung cancer (NSCLC). Our data indicated that the expression of miR-493 was markedly reduced in pulmonary carcinoma. The ectopic expression of miR-493 impaired cell growth and invasion in vitro and in vivo. Mechanically, miR-493 commonly directly targeted E2F1, which resulted in a robust reduction of the expression of mRNA and protein. This effect, in turn, decreased the growth, invasion and metastasis of lung cancer cells. Our findings highlight the importance of miR-493 dysfunction in promoting tumor progression, and implicate miR-493 as a potential therapeutic target in lung cancer.

Several existing technologies enable short genomic alterations including generating indels and short nucleotide variants, however, engineering more significant genomic changes is more challenging due to reduced efficiency and precision. Here, we developed RecT Editor via Designer-Cas9-Initiated Targeting (REDIT), which leverages phage single-stranded DNA-annealing proteins (SSAP) RecT for mammalian genome engineering. Relative to Cas9-mediated homology-directed repair (HDR), REDIT yielded up to a 5-fold increase of efficiency to insert kilobase-scale exogenous sequences at defined genomic regions. We validated our REDIT approach using different formats and lengths of knock-in templates. We further demonstrated that REDIT tools using Cas9 nickase have efficient gene-editing activities and reduced off-target errors, measured using a combination of targeted sequencing, genome-wide indel, and insertion mapping assays. Our experiments inhibiting repair enzyme activities suggested that REDIT has the potential to overcome limitations of endogenous DNA repair steps. Finally, our REDIT method is applicable across cell types including human stem cells, and is generalizable to different Cas9 enzymes.

Nitroreductase (NTR) belongs to a class of flavin mononucleotide-dependent and flavin adenine dinucleotide-dependent cytoplasmic enzymes; its contents in tumor cells increase during hypoxia. The development of fluorescent probes for detection of NTR activity is of great significance for the study of the state of hypoxia in living organisms. In this paper, a red-emitting fluorescence turn-on probe EBI-NO2 was synthesized using a one-step method. The fluorescence of the probe was enhanced by 60 folds in the presence of NTR. The probe also had high selectivity towards NTR, and its detection limit was as low as 1 ng/mL. The reaction mechanism was verified using MS, molecular docking and theoretical calculations. In addition, it was successfully applied in real-time monitoring of NTR produced during growth of Escherichia coli (BL21) and in visualization of NTR in oral cancer cells (Cal-27) under hypoxia. This work provides a new imaging tool that can be applied to investigate the physiological and pathological changes in hypoxia oral cells.

This communication reports on a fluorescent probe (PPI-P) for imaging active peptidyl-prolyl cis/trans isomerases in live cells. PPI-P is capable of responding to both recombinant and cellular PPIases fluorogenically, and has been shown to specifically image active PPIases in live cells.

Here we demonstrated that chemotherapy induced 14-3-3σ expression in tongue cancer (TC) cells and overexpressed 14-3-3σ sensitized TC cells to chemotherapy especially in multidrug resistant TC (MDR-TC) cells. In agreement, 14-3-3σ knockdown enhanced resistance of TC cells to chemotherapy. Mechanically, we found 14-3-3σ physically bound to GSK3β in protein level and the binding inhibited β-catenin signaling. Coincidentally, chemotherapy as well as 14-3-3σ overexpression led to increase of GSK3β protein level. Increased GSK3β protein sensitized TC cells to chemotherapy. Moreover, deregulation of 14-3-3σ/GSK3β/β-catenin axis led to overexpressed ZEB1 in TC cells, especially in MDR-TC cells. As a negative feedback loop, ZEB1 bond to 14-3-3σ promoter to enhance promoter hypermethylation in TC cells. Promoter hypermethylation resulted into the decrease of 14-3-3σ expression. Importantly, a positive correlation was observed between 14-3-3σ and GSK3β protein expression in TC tissues from patients receiving chemotherapy. High levels of 14-3-3σ and GSK3β were associated with better prognosis in TC patients.

We found that levels of miR-491-3p were decreased in multidrug-resistant tongue cancer (TC) cells. Induction of miR-491-3p expression sensitized TC cells to chemotherapy. In agreement, functional inhibition of miR-491-3p enhanced resistance of TC cells to chemotherapy. We found that miR-491-3p directly targeted mTORC2 component Rictor and inhibited mTORC2 activity, which was increased in resistant TC cells with high p-Akt(Ser473), p-SGK1(Ser422) and p-FOXO1(Thr24) levels. Inhibition of mTORC2 activity via either Rictor knockdown or mTOR inhibitor in turn sensitized TC cells to chemotherapy. In agreement, overexpression of Rictor increased the mTORC2 activity and induced resistance of TC cells to chemotherapy. As a feedback loop, mTORC2 downregulated miR-491-3p expression by inactivating FOXO1, which otherwise would transcriptionally induce miR-491-3p expression. Levels of miR-491-3 and Rictor or mTORC2 activity negatively correlated in TC tissues. Finally, low levels of miR-491-3p and highly expressed Rictor were associated with poor prognosis in tongue cancer patients. These data provide a rationale for targeted intervention on miR-491-3p/mTORC2 axis to enhance the efficacy of chemotherapy against tongue cancer.

Dysfunction of neurovascular pericytes underlies breakdown of the blood-brain barrier, but the molecular mechanisms are largely unknown. In this study, we evaluated the role of the transient receptor potential melastatin-related 2 (TRPM2) channel and autophagy during brain pericyte injury both in vitro and in vivo.A rapid induction in autophagy in human brain vascular pericytes, in the zinc oxide nanoparticles (ZnO-NP)-induced cell stress model, was paralleled with an increase in the expression of the TRPM2-S truncated isoform, which was abolished by treatment with a nitric oxide synthase inhibitor and a peroxynitrite scavenger. Furthermore, Y1485 in the C-terminus of the TRPM2 protein was identified as the tyrosine nitration substrate by mass spectrometry. Overexpression of the Y1485S TRPM2 mutant reduced LC3-II accumulation and pericyte injury induced by ZnO-NP. Consistently, LC3-II accumulation was reduced and pericytes were better preserved in intact brain microvessels of the TRPM2 knockout mice after ZnO-NP-induced vascular injury. Innovation and Conclusions: Our present study has revealed a novel mechanism of autophagy disturbance secondary to nitrosative stress-induced tyrosine nitration of TRPM2 during pericyte injury. Antioxid. Redox Signal. 27, 1297-1316.

Epstein-Barr virus (EBV) is a human oncogenic herpesvirus associated with lymphoma and nasopharyngeal carcinoma. Because the susceptible hosts of EB virus are limited to human and cotton-top tamarins (Saguinus oedipus), there have been no appropriate animal models until the lymphoma model induced by EBV in human peripheral blood lymphocyte (hu-PBL)/SCID chimeric mice was reported. However, it is still controversial whether the EBV-associated lymphoma induced in hu-PBL/SCID mice is a monoclonal tumor. In this study, we transplanted normal human peripheral blood lymphocytes (hu-PBL) from six donors infected with EBV into SCID mice to construct hu-PBL/SCID chimeric mice. The induced tumors were found in the mediastinum or abdominal cavity of SCID mice. Microscopic observation exhibited tumor cells that were large and had a plasmablastic, centroblastic or immunoblastic-like appearance. Immunophenotyping assays showed the induced tumors were LCA-positive, CD20/CD79a-positive (markers of B cells), and CD3/CD45RO-negative (markers of T cells). A human-specific Alu sequence could be amplified by Alu-PCR. This confirmed that induced tumors were B-cell lymphomas originating from the transplanted human lymphocytes rather than mouse cells. EBER in situ hybridization detected positive signals in the nuclei of the tumor cells. Expression of EBV-encoded LMP1, EBNA-1, and EBNA-2 in the tumors was significantly positive. PCR-based capillary electrophoresis analysis of IgH gene rearrangement revealed a monoclonal peak and single amplification product in all six cases of induced tumors. This indicated that EBV can induce monoclonal proliferation of human B lymphocytes and promotes the development of lymphoma. J. Med. Virol. 88:1804-1813, 2016. © 2016 Wiley Periodicals, Inc.

Genistein (GEN) is a molecule of great interest as a potent chemopreventive agent against atherosclerosis and cancer. However, the bioavailability of GEN is very low in vivo. Our previous study showed that a GEN derivative, 7-difluoromethyl-5,4'-dimethoxygenistein (dFMGEN) has a better bioavailability than GEN in vivo. In this study, we further evaluated the efficacy of dFMGEN as a candidate for cancer therapy. We demonstrated that dFMGEN treatment decreased the viability of A549 cells in a concentration- and time-dependent manner and induced cell-cycle arrest at the G(1) phase. G(1) phase arrest was correlated with a significant reduction of Cdk4 and cyclin D1 protein level. Further studies showed that cyclin-dependent kinase (Cdk)4 and cyclin D1 protein-level decrease was caused by Cdk inhibitors p15, p21, and p27 level increase, and decreased protein level directly suppressed Rb protein phosphorylation and E2F-1 expression, then cell-cycle progression was arrested. Finally, we also found that dFMGEN has a dosage effect in suppressing tumor growth in vivo, and that dFMGEN was well tolerated by animals. In summary, our results suggest that dFMGEN has therapeutic potential for the treatment of human lung cancer.

Structural basis for site-specific reading of unmodified R2 of histone H3 tail by UHRF1 PHD finger

We have previously constructed a recombined vascular basement membrane derived multifunctional peptide (rVBMDMP) which can inhibit tumor growth. The aim of this study is to explore the effects and mechanisms of rVBMDMP on growth and motility/invasion in human A549 lung carcinoma cells. The effect of rVBMDMP on A549 cell viability was determined by MTT assay while the motility/invasion was measured by scratch and transwell assays. Molecules that responded to rVBMDMP treatment of A549 cells were explored using the high-throughput Cancer Pathway Finder cDNA Microarray. We identified 16 genes that were up-regulated, including GZMA, ITG alphaV, MCAM and Kiss1 and 21 genes that were down-regulated, including uPA, uPAR, CDC25A, IGF1 and FGF2. Selective differentially expressed genes were further analyzed by real-time quantitative PCR and Western blot analysis. These findings contribute to the understanding of the molecular mechanisms mediating rVBMDMP action, and suggest that rVBMDMP is a promising novel agent for the treatment of human lung carcinoma.

Latent membrane protein 1 (LMP1) of Epstein-Barr virus has been identified to be crucial in inducing cell transformation. However, the mechanism of LMP1-mediated epithelial cell transformation remains unclear. In this study, nasopharyngeal epithelial cells NP69 were infected with retrovirus with gene encoding wild type LMP1 or mutational LMP1 defective in binding to tumor necrosis factor receptor-associated death domain (TRADD). The NP69-LMP1(TRADD) lost some malignant phenotypes compared with the NP69-LMP1(WT). We performed proteomic approach to gain the differential protein expression profile associated with LMP1-mediated epithelial cell transformation. Furthermore, the differential expressional levels of partial identified proteins were confirmed by Western blot and real-time RT-PCR. Some were known to be related to the development of LMP1-induced transformation, and some were new LMP1-associated proteins. These data are valuable for further study of the mechanism of LMP1 in human nasopharyngeal carcinoma and provide some new clues for investigating other LMP1-associated tumors.

Background Chemoresistance is a major obstacle in successfully treating cancers, and the mechanisms responsible for drug resistance are still far from understood. Carbonic anhydrase 9 (CA9) has been shown to be upregulated in the drug-resistant tongue cancer cell line Tca8113/PYM and to be associated with drug resistance. However, the mechanisms regulating CA9 expression and its role in drug resistance remain unclear. Methods Bioinformatic and experimental analysis involving ChIP and luciferase reporter assays were used to validate Zinc finger E-box-binding homeobox 1 (ZEB1) as a transcriptional regulator of CA9. Gene expression and protein levels were evaluated by quantitative RT-PCR and western blotting, respectively. Sensitivity to chemotherapy was examined using the MTS assay and Hoechst staining and analysis caspase-3 activity to evaluate changes in apoptosis. Intracellular pH (pHi) was measured using fluorescent pH-indicator BCECF-AM. Protein expression in patient tissue samples was examined by immunohistochemistry and survival of tongue cancer patients from which these samples were derived was also analyzed. Results ZEB1 bound to the promoter of CA9 to positively regulate CA9 expression in tongue cancer cells. Knockdown of CA9 using short interfering RNA (siRNA) abolished the chemoresistance resulting from ZEB1 overexpression in Tca8113 and SCC-25 cells, and CA9 overexpression attenuated chemosensitivity induced by ZEB1 knockdown in Tca8113/PYM cells. CA9 knockdown also prevented maintenance of pHi mediated by overexpression of ZEB1 in Tca8113 and SCC-25 cells following chemotherapy, associated with increased apoptosis and caspase-3 activation. Conversely, ectopic expression of CA9 suppressed decrease in pHi mediated by ZEB1 knockdown in Tca8113/PYM cells following chemotherapy, accompanied by decreased apoptosis and caspase-3 activation. Importantly, a positive correlation was observed between ZEB1 and CA9 protein expression in tongue cancer tissues, and expression of these proteins associated with a poor prognosis for patients. Conclusion Our finding that tumor cells regulate pHi in response to chemotherapy provides new insights into mechanisms of drug resistance during cancer treatment. Identification of the ZEB1–CA9 signaling axis as a biomarker of poor prognosis in tongue cancer will be valuable in future development of therapeutic strategies aimed at improving treatment efficacy, especially in terms of drug resistance associated with this disease.

Tumor drug resistance is a major obstacle to cancer chemotherapy. We previously constructed a fusion protein based on two tumstatin-derived sequences named recombinant VBMDM (rVBMDMP). We preliminarily confirmed its inhibition of HUVEC and colon cancer cell growth. The present study further systematically observed the inhibitory effect of rVBMDMP on lung cancer cell growth and analyzed a possible mechanism to provide a theoretical basis for the development of new antitumor protein drugs. The effect of rVBMDMP on human lung adenocarcinoma (A549) and cisplatin-resistant human lung adenocarcinoma (A549/DDP) cell proliferation was evaluated by MTS assay. Hoechst 33342 staining performed together with fluorescence microscopy and immunoblot analysis were used to examine the effects of rVBMDMP on the apoptosis of A549/DDP cells. A protein phosphorylation chip was used to identify changes in rVBMDMP-induced signaling protein phosphorylation. Changes in the phosphatidylinositol 3 kinase (PI3K)/Akt signal transduction pathway and expression of multidrug resistance protein (MRP-2)-related molecules following rVBMDMP treatment in A549/DDP cells were evaluated by western blot analysis. A lung cancer xenograft model was used to evaluate the reversal effect of rVBMDMP on drug-resistance of A549/DDP cell tumors to cisplatin in vivo. The results demonstrated that rVBMDMP increased the phosphorylation of 79 signaling proteins, including focal adhesion kinase (FAK), caspase-6, Fas, FasL and FAF1 and downregulated 30 signaling proteins, including integrin αV, integrin β3, PI3K/Akt, NF-κB and MRP-2 compared with the controls. rVBMDMP also increased the sensitivity of A549 and A549/DDP cells to cisplatin and directly induced apoptosis, which may be related to MRP-2 and Bcl-2 downregulation. The effects of growth inhibition and apoptosis induction of rVBMDMP on A549/DDP cells may be related to the inhibition of integrin αVβ3 and PI3K/Akt protein phosphorylation. Finally, we observed an increase in cancer cell sensitivity to cisplatin by rVBMDMP using the A549/DDP cell xenograft model in nude mice. Our study suggests that rVBMDMP may be an effective potential chemotherapy sensitizer and may be a viable drug candidate in anticancer therapies.

Cisplatin (DDP) is the first-line chemotherapy drug widely used for the treatment of lung cancer patients, whereas the majority of cancer patients will eventually show resistance to DDP. The mechanisms responsible for DDP resistance are not fully understood. Tongue cancer resistance-associated protein 1 (TCRP1) gene was recently cloned and reported to specially mediate DDP resistance in human oral squamous cell carcinoma (OSCC) cells. However, the mechanisms of TCRP1-mediated DDP resistance are far from clear, and whether TCRP1 participates in DDP resistance in lung cancer cells remains unknown. Here, we show that TCRP1 contributes to DDP resistance in lung cancer cells. Knockdown of TCRP1 sensitizes the cells to DDP and increases the DDP-induced DNA damage. We have identified that Pol β is associated with DDP resistance, and Pol β knockdown delays the repair of DDP-induced DNA damage in A549/DDP cells. We find TCRP1 interacts with Pol β in lung cancer cells. Moreover, TCRP1 knockdown decreases the level of Pol β and increases the level of its ubiquitination. These results suggest that TCRP1 contributes to DDP resistance through the prevention of Pol β degradation in lung cancer cells. These findings provide new insights into chemoresistance and may contribute to prevention and reversal of DDP resistance in treatment of lung cancer in the future.

Cisplatin is one of the major chemotherapeutic agents used against different human cancers. A better understanding of the downstream cellular targets of cisplatin will provide information on its mechanism of action. FOXO3a is a member of the FOXO transcription factor family, which modulates the expression of genes involved in cell cycle arrest, apoptosis, and other cellular processes. In this study, we have investigated the effects of cisplatin in a panel of lung cancer cell lines. The results showed that cisplatin inhibited the proliferation of these lung cancer cell lines by inhibiting the PI3K/AKT pathway, with evidence of decreasing phosphorylation of PI3K and AKT under cisplatin treatment, and constitutively activating AKT1 could reduce cisplatin-induced cell apoptosis. More importantly, cisplatin significantly inhibited FOXO3a phosphorylation (at Thr32, AKT phosphorylation site) and induced FOXO3a nuclear accumulation, which in turn increased the expression of FOXO3a-dependent apoptotic protein Bim. Knockdown of FOXO3a expression using small interfering RNA attenuated cisplatin-induced apoptosis. Furthermore, activation of FOXO3a induced cell apoptosis irrespective of p53 status, whereas p53 may act as FOXO3a downstream molecules involved in cisplatin-induced cell apoptosis. Together, our findings suggested that FOXO3a is a relevant mediator of the cytotoxic effects of cisplatin in lung cancer cells.

Chemotherapy is not only important but also necessary for the patient of breast cancer. Breast cancer resistance protein (BCRP), an atypical drug efflux pump, mediates multidrug resistance in breast cancer. The aim of this study is to search new substrate of BCRP. The result will guide the drug selection of chemotherapy in BCRP-positive breast cancer. PA317/Tet-on/TRE-BCRP cell induced with doxycycline was used to screen the possible substrates of BCRP by MTT assay. The suspicious substrate [5-fluorouracil (5-Fu)] was further confirmed in PA317 and breast cancer cell MCF-7 by HLCP, apoptosis assay (staining and FACS) and RNAi technique. Mitoxantrone, 5-Fu, adriamycin, Methotrexate, Pirarubicin, and Etoposide were identified as substrates of BCRP. However, Paclitaxel, Vincristine, Vindesine, Mitomycin C, and cisplatin were not mediated by BCRP. 5-Fu was identified as substrate of BCRP for the first time. The further study showed that the intracellular retention dose of 5-Fu and the 5-Fu induced cellular apoptosis all decreased when BCRP highly expressed. Furthermore, 5-Fu accumulation and 5-Fu induced DNA damage increased when BCRP was silenced by RNAi in breast cancer cells. 5-Fluorouracil may be a specific substrate which can be bound by BCRP. BCRP can predict the sensitivity of breast cancer to 5-Fu. And BCRP-targeted therapy will reverse the resistance of breast cancer to 5-Fu.

Breast cancer resistance protein (BCRP) has been shown to confer multidrug resistance, but the mechanisms of its regulation are poorly understood. Here, we investigate the effects of wild-type and mutant p53, and nuclear factor kappa-B (NF-kappaB) (p50) on BCRP promoter activity in MCF-7 cells. Our results demonstrated that wild-type p53 markedly suppressed BCRP activity and enhanced the chemosensitivity of cells to mitoxantrone, whereas mutant p53 had little inhibitory effect. After inhibition of NF-kappaB, similar results were obtained. Following knockdown of endogenous p53, BCRP and p50 expressions were increased, and the chemosensitivity of the cells to mitoxantrone was decreased. We conclude that wild-type p53 acts as a negative regulator of BCRP gene transcription.