Overcome Doxorubicin Resistance Research Articles

Pyrrolidine dithiocarbamate reverses Bcl-xL-mediated apoptotic resistance to doxorubicin by inducing paraptosis.

Elevated Bcl-xL expression in cancer cells contributes to doxorubicin (DOX) resistance, leading to failure in chemotherapy. In addition, the clinical use of high-dose doxorubicin (DOX) in cancer therapy has been limited by issues with cardiotoxicity and hepatotoxicity. Here, we show that co-treatment with pyrrolidine dithiocarbamate (PDTC) attenuates DOX-induced apoptosis in Chang-L liver cells and human hepatocytes, but overcomes DOX resistance in Bcl-xL-overexpressing Chang-L cells and several hepatocellular carcinoma (HCC) cell lines with high Bcl-xL expression. Additionally, combined treatment with DOX and PDTC markedly retarded tumor growth in a Huh-7 HCC cell xenograft tumor model, compared to either mono-treatment. These results suggest that DOX/PDTC co-treatment may provide a safe and effective therapeutic strategy against malignant hepatoma cells with Bcl-xL-mediated apoptotic defects. We also found that induction of paraptosis, a cell death mode that is accompanied by dilation of the endoplasmic reticulum and mitochondria, is involved in this anti-cancer effect of DOX/PDTC. The intracellular glutathione levels were reduced in Bcl-xL-overexpressing Chang-L cells treated with DOX/PDTC, and DOX/PDTC-induced paraptosis was effectively blocked by pretreatment with thiol-antioxidants, but not by non-thiol antioxidants. Collectively, our results suggest that disruption of thiol homeostasis may critically contribute to DOX/PDTC-induced paraptosis in Bcl-xL-overexpressing cells.

Recombinant methioninase in combination with doxorubicin (DOX) overcomes first-line DOX resistance in a patient-derived orthotopic xenograft nude-mouse model of undifferentiated spindle-cell sarcoma

We have previously established a patient-derived orthotopic xenograft (PDOX) model of undifferentiated spindle cell sarcoma (USCS). Recombinant methioninase (rMETase) has previously demonstrated efficacy in PDOX mouse models of human cancers. In the present study, we determined if rMETase in combination with doxorubicin (DOX) can overcome first-line DOX resistance in a PDOX models of USCS. The USCS PDOX mouse models were randomized into the following groups when tumor volume reached 100 mm3: G1, control without treatment; G2, doxorubicin (DOX) (3 mg/kg, intraperitoneal [i.p.] injection, weekly, for 2 weeks); G3, rMETase (100 units/mouse, i.p., daily, for 2 weeks); G4, DOX (3 mg/kg, i.p., weekly, for 2 weeks) combined with rMETase (100 units/mouse, i.p., daily, for 2 weeks). Tumor size and body weight were measured twice a week. On day 14 after initiation, the USCS PDOX tumor sizes were (G1): 360 ± 85 mm3; DOX (G2): 355 ± 111 mm3, p = .927; rMETase (G3): 182 ± 57 mm3, p = .0003; DOX + rMETase (G4): 134 ± 29 mm3, p = .00001. These results indicate that rMETase can overcome USCS resistance to DOX, which is first line therapy for this disease. The body weight of treated mice was not significantly different in any group. The present results demonstrate the power of the PDOX model to identify effective therapy for recalcitrant cancer and the potential of rMETase to overcome DOX resistance.

Autophagy and doxorubicin resistance in cancer.

Doxorubicin (DOX), also known as adriamycin, is a DNA topoisomerase II inhibitor and belongs to the family of anthracycline anticancer drugs. DOX is used for the treatment of a wide variety of cancer types. However, resistance among cancer cells has emerged as a major barrier to effective treatment using DOX. Currently, the role of autophagy in cancer resistance to DOX and the mechanisms involved have become one of the areas of intense investigation. More and more preclinical data are being obtained on reversing DOX resistance through modulation of autophagy as one of the promising therapeutic strategies. This review summarizes the recent advances in autophagy-targeting therapies that overcome DOX resistance from in-vitro studies to animal models for exploration of novel delivery systems. In-depth understanding of the mechanisms of autophagy regulation in relation to DOX resistance and development of molecularly targeted autophagy-modulating agents will provide a promising therapeutic strategy for overcoming DOX resistance in cancer treatment.

Synergistic combination chemotherapy using carrier-free celastrol and doxorubicin nanocrystals for overcoming drug resistance.

A key challenge of chemotherapy in clinical treatments is multidrug resistance (MDR), which mainly arises from drug efflux-induced tumor cell survival. Thus, it is necessary to provide biocompatible chemotherapeutics to improve drug accumulation in MDR cells. Herein, two clinical small molecular drugs, celastrol (CST) and doxorubicin (DOX), were self-assembled into carrier-free and biocompatible nanoparticles (CST/DOX NPs) via a simple and green precipitation method for synergistic combination chemotherapy to overcome DOX resistance. These spherical CST/DOX NPs can improve the water-solubility of CST, reduce the dosage of DOX, and therefore significantly enhance cellular drug accumulation by activating heat shock factor 1 (HSF-1) and inhibiting NF-κB to depress P-gp expression, which results in apoptosis and autophagy of DOX resistant cells through the ROS/JNK signaling pathway. Finally, synergistic combination chemotherapy was attained in both MCF-7/MDR cells and 3D multicellular tumor spheroids. Thus, CST/DOX NPs provide an alternative for overcoming drug resistance in future clinical applications.

HSA-based multi-target combination therapy: regulating drugs’ release from HSA and overcoming single drug resistance in a breast cancer model

Multi-drug delivery systems, which may be promising solution to overcome obstacles, have limited the clinical success of multi-drug combination therapies to treat cancer. To this end, we used three different anticancer agents, Cu(BpT)Br, NAMI-A, and doxorubicin (DOX), to build human serum albumin (HSA)-based multi-drug delivery systems in a breast cancer model to investigate the therapeutic efficacy of overcoming single drug (DOX) resistance to cancer cells in vivo, and to regulate the drugs’ release from HSA. The HSA complex structure revealed that NAMI-A and Cu(BpT)Br bind to the IB and IIA sub-domain of HSA by N-donor residue replacing a leaving group and coordinating to their metal centers, respectively. The MALDI-TOF mass spectra demonstrated that one DOX molecule is conjugated with lysine of HSA by a pH-sensitive linker. Furthermore, the release behavior of three agents form HSA can be regulated at different pH levels. Importantly, in vivo results revealed that the HSA–NAMI-A–Cu(BpT)Br–DOX complex not only increases the targeting ability compared with a combination of the three agents (the NAMI-A/Cu(BpT)Br/DOX mixture), but it also overcomes DOX resistance to drug-resistant breast cancer cell lines.

Silybin counteracts doxorubicin resistance by inhibiting GLUT1 expression

Despite significant advances in the diagnosis and treatment of cancer, the development of drug resistance still remains one of the principal causes that hampers the effectiveness of the therapy. Emerging evidences support the idea that the dysregulated metabolism could be related to drug resistance. The major goal of this study was to target cancer metabolic pathways using new pharmacological approaches coming from natural sources in order to possibly prevent or overcome this phenomenon. Firstly, the metabolic profile of human colorectal adenocarcinoma cells sensitive (LoVo WT) and resistant to doxorubicin (LoVo DOX) was delineated demonstrating that resistant cells remodel their metabolism toward a glycolytic phenotype. In particular it was observed that doxorubicin-resistant cancer cells exhibit an increased dependency from glucose for their survival, associated with overexpression of the glycolytic pathway. Moreover, both GLUT1 mRNA and protein expression significantly increased in LoVo DOX cells. Given the results about the metabolic profile, silybin, modulator of GLUTs, was selected as potential candidate to overcome doxorubicin resistance and, intriguingly, data revealed not only that silybin is more active in resistant cells than in wild type cells, but also that the combined treatment with doxorubicin and silybin presents a synergistic effect in LoVo DOX cells. Although many unanswered questions still remain about the molecular mechanism of silybin, these data suggest that targeting GLUTs may be a good strategy to restore doxorubicin sensitivity and elude drug resistance.

Tumor-homing, pH- and ultrasound-responsive polypeptide-doxorubicin nanoconjugates overcome doxorubicin resistance in cancer therapy

Nanomedicines hold promise in overcoming drug resistance in cancer therapy, but the in vivo therapeutic efficacy is limited by their inefficient tumor targeting, poor tumor penetration, low cellular uptake and insufficient drug release. Here we report tumor-homing, pH- and ultrasound-responsive polypeptide-doxorubicin nanoconjugates for overcoming doxorubicin resistance. These nanoconjugates show accelerated cellular uptake and doxorubicin release and thus enhanced cytotoxicity to doxorubicin-resistant cancer cells when exposed to ultrasound. In a doxorubicin-resistant breast cancer mouse model, they exhibited improved tumor accumulation and penetration following exposure to ultrasound. More importantly, they displayed significantly improved in vivo anticancer efficacy without appreciable side effects post ultrasound irradiation. These findings suggest that these nanoconjugates are promising as a new class of intelligent nanomedicines for overcoming drug resistance in cancer therapy.

Mitochondria and Nucleus Dual Delivery System To Overcome DOX Resistance.

Doxorubicin (DOX) is a broad-spectrum chemotherapy drug to treat tumors. However, severe side effects and development of DOX resistance hinder its clinical application. In order to overcome DOX resistance, DOX/TPP-DOX@Pasp-hyd-PEG-FA micelles were prepared by using newly synthesized comb-like amphiphilic material Pasp-hyd-PEG-FA. Drug released in vitro from micelles showed a pH-dependent manner. DOX/TPP-DOX@Pasp-hyd-PEG-FA induced more apoptosis in KB cell and MCF-7/ADR cell than DOX@Pasp-hyd-PEG-FA. Confocal laser scanning microscopy experiment indicated that DOX/TPP-DOX@Pasp-hyd-PEG-FA delivered TPP-DOX and DOX to the nucleus and mitochondria of the tumor cell simultaneously. Thus, DOX/TPP-DOX@Pasp-hyd-PEG-FA could significantly damage the mitochondrial membrane potential. DOX/TPP-DOX@Pasp-hyd-PEG-FA markedly shrinked the tumor volume in tumor-bearing nude mice grafted with MCF-7/ADR cell as compared with the same dose of free DOX. DOX was mainly accumulated in tumor tissue after DOX/TPP-DOX@Pasp-hyd-PEG-FA was injected to tumor-bearing nude mice by tail vein. After free DOX was injected to tumor-bearing nude mice by tail vein, DOX widely distributed through the whole body. Therefore, mitochondria and nucleus dual delivery system has potential in overcoming DOX resistance.

Berberine Enhances Chemosensitivity and Induces Apoptosis Through Dose-orchestrated AMPK Signaling in Breast Cancer.

Breast cancer is the most common malignancy in women. Although personalized or targeting molecular cancer therapy is more popular up to now, the cytotoxicity chemotherapy for patients with advanced breast cancer is considered as the alternative option. However, chemoresistance is still the common and critical limitation for breast cancer treatment. Berberine, known as AMPK activator, has shown multiple activities including antitumor effect. In this study, we investigate the chemosensitive effect of different dosages berberine on drug-resistant human breast cancer MCF-7/MDR cell in vitro and in vivo, and the mechanisms underlying AMPK activation on Doxorubicin (DOX) chemosensitivity. Our results showed that berberine could overcome DOX resistance in dose-orchestrated manner: On one hand, low-dose berberine can enhance DOX sensitivity in drug-resistance breast cancer cells through AMPK-HIF-1α-P-gp pathway. On the other hand, high-dose berberine alone directly induces apoptosis through the AMPK-p53 pathway with the independence of HIF-1α expression. Taken together, our findings demonstrate that berberine sensitizes drug-resistant breast cancer to DOX chemotherapy and directly induces apoptosis through the dose-orchestrated AMPK signaling pathway in vitro and in vivo. Berberine appears to be a promising chemosensitizer and chemotherapeutic drug for breast cancer treatment.

Doxorubicin and resveratrol co-delivery nanoparticle to overcome doxorubicin resistance.

With the extensive application of doxorubicin (DOX), DOX resistance has become one of the main obstacles to the effective treatment of breast cancer. In this paper, DOX and resveratrol (RES) were co-encapsulated in a modified PLGA nanoparticle (NPS) to overcome the DOX resistance. CLSM results indicated that DOX and RES were simultaneously delivered into the nucleus of DOX-resistant human breast cancer cells by DOX/RES-loaded NPS. Consequently, DOX/RES-loaded NPS showed significant cytotoxicity on MDA-MB-231/ADR cells and MCF-7/ADR cells. Furthermore, DOX/RES-loaded NPS could overcome DOX resistance by inhibiting the expression of drug resistance-related protein such as P-gp, MRP-1 and BCRP, and induce apoptosis through down-regulating the expression of NF-κB and BCL-2. In tumor-bearing mice, DOX/RES-loaded NPS mainly delivered DOX and RES to tumor tissue. Compared with free DOX, DOX/RES-loaded NPS significantly inhibited the DOX-resistant tumor growth in tumor-bearing mice without causing significant systemic toxicity. In a word, DOX/RES-loaded NPS could overcome the DOX resistance and had the potential in the treatment of DOX-resistant breast cancer.

Abstract 2168: Optimizing doxorubicin derivatives delivery using temperature sensitive biopolymers in multidrug resistant breast cancer cells

Abstract The anticancer agent doxorubicin is an anthracycline compound that shows high potency in treating cancer, and it is one of the most widely used chemotherapeutics. However, efficiency of doxorubicin treatment is limited by low blood plasma solubility, poor blood pharmacokinetics, and non-selective cell killing that results in serious toxicity to healthy tissues. Additionally, cancer cells often develop drug resistance, which is a significant limiting factor to the drug's effectiveness. Motivated by these problems, we have designed a drug delivery system that can specifically deliver drug to the tumor site while overcoming doxorubicin resistance in breast cancer cell lines. This drug delivery system consists of: ELP - Elastin like polypeptide, CPP - Cell penetrating peptide, a cleavable linker – to enable doxorubicin release in the targeted low pH environment, and a derivative of the anticancer agent Doxorubicin (modified by a 6-maleimidocaproyl moiety for conjugation to a terminal cysteine residue on ELP). ELP is thermally responsive and improves the complex's pharmacokinetics by prolonging its clearance rate while the CPP mediates cellular uptake of large macromolecules. The linker is an acid sensitive amino acid sequence (Gly-Phe-Leu-Gly) that serves as a substrate for lysosomal enzymes. In this study, we compared cytotoxicity of a cleavable (cDox) and non-cleavable (ncDox) doxorubicin derivative delivered by ELP biopolymer in the breast cancer cell lines MCF-7 and MCF7/ADR (doxorubicin resistant cell line). We showed that cDox had two fold higher cytotoxicity than ncDox in both cell lines. When ncDox, however, was conjugated to the ELP biopolymer containing a lysosomally degradable GFLG spacer, the drug delivery construct was equally cytotoxic to both sensitive and resistant cell lines, indicating that the construct delivers doxorubicin into cells by a mechanism that bypasses doxorubicin resistance. Confocal fluorescence microscopy showed that after two hours of exposure to the doxorubicin derivatives, cDox was predominantly localized in the nucleus in both cell lines. However, ncDox was localized in the plasma membrane and cytosol. Intracellular doxorubicin accumulation examined by flow cytometry indicated 3 fold higher uptake of ELP-cDox in sensitive MCF 7 cells compared to resistant MCF7 ADR cells. Cellular uptake of ELP-cDox was further enhanced two fold when conjugated with CPP. In conclusion, our current results indicate that ELP-doxorubicin conjugates may successfully overcome drug resistance in breast cancer cells, providing a promising approach for the use of chemotherapeutic agents such as doxorubicin in patients with drug resistant breast cancers. Citation Format: Sonja Dragojevic, Jung Su Ryu, Felix Kratz, Drazen Raucher. Optimizing doxorubicin derivatives delivery using temperature sensitive biopolymers in multidrug resistant breast cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2168.

PI3K/Akt/mTOR activation by suppression of ELK3 mediates chemosensitivity of MDA-MB-231 cells to doxorubicin by inhibiting autophagy

Drug resistance in breast cancer remains a major obstacle of clinical therapy. We found that suppression of ELK3 in the triple negative breast cancer cell line MDA-MB-231 impaired autophagy and led to a hypersensitive response to doxorubicin treatment. In ELK3-knockdown MDA-MB-231 cells (ELK3 KD), autophagy was not activated under starvation conditions, which is a major stimulus of autophagy activation. We revealed that activation of the PI3K/Akt pathway was the main cause of impaired autophagy in ELK3 KD. Our results suggest that targeting ELK3 may be a potential approach to overcome doxorubicin resistance in breast cancer therapeutics.

MiR-122 Reverses the Doxorubicin-Resistance in Hepatocellular Carcinoma Cells through Regulating the Tumor Metabolism.

Doxorubicin (DOX) is one of the most commonly used anticancer drugs in the treatment of hepatoma. However, acquired drug resistance is one of the major challenges for the chemotherapy. In this study, a down-regulation of miR-122 was observed in doxorubicin-resistant Huh7 (Huh7/R) cells compared with its parental Huh7 cells, suggesting miR-122 is associated with the chemoresistance. Meanwhile, luciferase reporter assay proved that the PKM2 is the target of miR-122, and we reported that the glucose metabolism is significantly up-regulated in Huh7/R cells. Importantly, overexpression of miR-122 in Huh7/R cells reversed the doxorubicin-resistance through the inhibition of PKM2, inducing the apoptosis in doxorubicin-resistant cancer cells. Thus, this study revealed that the dysregulated glucose metabolism contributes to doxorubicin resistance, and the inhibition of glycolysis induced by miR-122 might be a promising therapeutic strategy to overcome doxorubicin resistance in hepatocellular carcinoma.

Overcoming doxorubicin resistance of cancer cells by Cas9-mediated gene disruption.

In this study, Cas9 system was employed to down-regulate mdr1 gene for overcoming multidrug resistance of cancer cells. Disruption of the MDR1 gene was achieved by delivery of the Cas9-sgRNA plasmid or the Cas9-sgRNA ribonucleoprotein complex using a conventional gene transfection agent and protein transduction domain (PTD). Doxorubicin showed considerable cytotoxicity to the drug-resistant breast cancer cells pre-treated with the RNA-guided endonuclease (RGEN) systems, whereas virtually non-toxic to the untreated cells. The potency of drug was enhanced in the cells treated with the protein-RNA complex as well as in those treated with plasmids, suggesting that mutation of the mdr1 gene by intracellular delivery of Cas9-sgRNA complex using proper protein delivery platforms could recover the drug susceptibility. Therefore, Cas9-mediated disruption of the drug resistance-related gene can be considered as a promising way to overcome multidrug resistance in cancer cells.

Liposome armed with herpes virus-derived gH625 peptide to overcome doxorubicin resistance in lung adenocarcinoma cell lines.

New delivery systems including liposomes have been developed to circumvent drug resistance. To enhance the antitumor efficacy of liposomes encapsulating anti-cancer agents, we used liposomes externally conjugated to the 20 residue peptide gH625. Physicochemical characterization of the liposome system showed a size of 140 nm with uniform distribution and high doxorubicin encapsulation efficiency. We evaluated the effects of increasing concentrations of liposomes encapsulating Doxo (LipoDoxo), liposomes encapsulating Doxo conjugated to gH625 (LipoDoxo-gH625), empty liposomes (Lipo) or free Doxo on growth inhibition of either wild type (A549) or doxorubicin-resistant (A549 Dx) human lung adenocarcinoma. After 72 h, we found that the growth inhibition induced by LipoDoxo-gH625 was higher than that caused by LipoDoxo with an IC50 of 1 and 0.3 μM in A549 and A549 Dx cells, respectively. The data on cell growth inhibition were paralleled by an higher oxidative stress and an increased uptake of Doxo induced by LipoDoxo-gH625 compared to LipoDoxo, above all in A549 Dx cells. Cytometric analysis showed that the antiproliferative effects of each drug treatment were mainly due to the induction of apoptosis. In conclusion, liposomes armed with gH625 are able to overcome doxorubicin resistance in lung adenocarcinoma cell lines.

Free DOX and chitosan-N-arginine conjugate stabilized indocyanine green nanoparticles for combined chemophotothermal therapy

Indocyanine green (ICG) is a FDA-approved near-infrared (NIR) cyanine dye used in medical diagnostics. However, the utility of ICG remains limited by its unstable optical property, and concentration-dependent aggregation and precipitation. A chitosan–arginine conjugate (CS-N-Arg) was developed to increase the stability of ICG in physiological buffer saline via formation of strong electrostatic interactions between ICG and CS-N-Arg. The CS-N-Arg/ICG complex prevented ICG from aggregation and precipitation, thus it could serve as a theranostic nanomaterial for image-guided photothermal cancer therapy. The CS-N-Arg/ICG NPs showed excellent photostability, clear fluorescent images, and rapid temperature rise under laser irradiation. Cell viability assay indicated that CS-N-Arg/ICG NPs could efficiently suppress the growth of doxorubicin (DOX) resistant breast cancer cell (MCF-7/ADR cells) under NIR photothermal treatments. In combination of DOX with CS-N-Arg/ICG NPs, a combined effect was observed in MCF-7/ADR breast cancer cells due to dual hyperthermia and chemical therapeutic effects. The present observations suggest that CS-N-Arg/ICG NPs can effectively deliver ICG molecules to MCF-7/ADR breast cancer cells and overcome DOX resistance in the cells by hyperthermia.

Src/STAT3-dependent heme oxygenase-1 induction mediates chemoresistance of breast cancer cells to doxorubicin by promoting autophagy.

Chemotherapeutic resistance in breast cancer, whether acquired or intrinsic, remains a major clinical obstacle. Thus, increasing tumor cell sensitivity to chemotherapeutic agents will be helpful in improving the clinical management of breast cancer. In the present study, we found an induction of HO-1 expression in doxorubicin (DOX)-treated MDA-MB-231 human breast adenocarcinoma cells, which showed insensitivity to DOX treatment. Knockdown HO-1 expression dramatically upregulated the incidence of MDA-MB-231 cell death under DOX treatment, indicating that HO-1 functions as a critical contributor to drug resistance in MDA-MB-231 cells. We further observed that DOX exposure induced a cytoprotective autophagic flux in MDA-MB-231 cells, which was dependent on HO-1 induction. Moreover, upregulation of HO-1 expression required the activation of both signal transducer and activator of transcription (STAT)3 and its upstream regulator, protein kinase Src. Abrogating Src/STAT3 pathway activation attenuated HO-1 and autophagy induction, thus increasing the chemosensitivity of MDA-MB-231 cells. Therefore, we conclude that Src/STAT3-dependent HO-1 induction protects MDA-MB-231 breast cancer cells from DOX-induced death through promoting autophagy. In the following study, we further demonstrated the contribution of Src/STAT3/HO-1/autophagy pathway activation to DOX resistance in another breast cancer cell line, MDA-MB-468, which bears a similar phenotype to MDA-MB-231 cells. Therefore, activation of Src/STAT3/HO-1/autophagy signaling pathway might play a general role in protecting certain subtypes of breast cancer cells from DOX-induced cytotoxicity. Targeting this signaling event may provide a potential approach for overcoming DOX resistance in breast cancer therapeutics.

Resistant mechanisms of doxorubicin

Doxorubicin is widely used in chemotherapy of breast cancer and many other cancers.How ever,doxorubicin resistance restricts its use to some extent.The mechanisms of doxorubicin resistance may involve transport protein,apoptosis protein,DNA repair function,enzyme and other factors.Researches about reversing doxorubicin resistance are ongoing and provide some effective therapeutic regimens to overcome doxorubicin resistance clinically.But the specific mechanism of doxorubicin resistance is yet to be elucidated.It is expected that more reasonable and effective counter-measures will be put forward to improve the theraputical effect of doxorubicin. Key words: Drug resistance, neoplasm; Doxorubicin

Doxorubicin Loading, Release, and Stability of Polyamidoamine Dendrimer-Coated Magnetic Nanoparticles

Nanotechnology is a promising alternative to overcome the limitations of classical chemotherapy. As a novel approach, dendrimer-coated magnetic nanoparticles (DcMNPs) maintain suitable drug delivery system because of their buildup of functional groups, symmetry perfection, nanosize, and internal cavities. They can also be targeted to the tumor site in a magnetic field. The aim of this study is to obtain an effective targeted delivery system for doxorubicin, using polyamidoamine (PAMAM) DcMNPs. Different generations (G2 , G3 , G4 , and G7 ) of PAMAM DcMNPs were synthesized. Doxorubicin loading, release, and stability efficiencies in these nanoparticles (NPs) were studied. The results showed that low-generation NPs obtained in this study have pH-sensitive drug release characteristics. G4 DcMNP, which releases most of the drug in lower pH, seems to be the most suitable generation for efficient Doxorubicin delivery. Furthermore, application of doxorubicin-loaded G4 DcMNPs may help to overcome doxorubicin resistance in MCF-7 cells. On the contrary, G2 and G3 DcMNPs would be suitable for the delivery of drugs such as vinca alkaloids (Johnson IS, Armstrong JG, Gorman M, Burnett JP. 1963. Cancer Res 23:1390-1427.) and taxenes (Clarke SJ, Rivory LP. 1999. Clin Pharmacokinet 36(2):99-114.), which show their effects in cytoplasm. The results of this study can provide new insights in the development of pH-sensitive targeted drug delivery systems to overcome drug resistance during cancer therapy.

Targeted delivery of doxorubicin through conjugation with EGF receptor–binding peptide overcomes drug resistance in human colon cancer cells

Induction of multidrug resistance by doxorubicin (DOX), together with non-specific toxicities, has restricted DOX-based chemotherapy. Recently, we demonstrated that DOX conjugated with an EGF receptor-binding peptide (DOX-EBP) had enhanced anticancer efficacy and reduced systemic toxicity when targeting EGF receptor-overexpressing tumours. Here we investigated whether DOX-EBP is able to overcome drug resistance and the underlying molecular mechanisms. DOX-resistant SW480/DOX cells were derived from non-resistant SW480 cells by stepwise exposure to increasing concentrations of DOX, and P-glycoprotein overexpression induced by DOX was confirmed by Western blotting. Cytotoxicity and intracellular distribution of drugs were evaluated by MTT assay and fluorescence microscopy respectively. EGF receptor-mediated endocytosis was determined in EGF receptor and endocytosis inhibition assays. Drug accumulation in tumour cells and murine xenografts was determined by HPLC. The cytotoxicity and accumulation of DOX-EBP in SW480/DOX cells were almost the same as in SW480 cells, but those of free DOX were reduced. DOX-EBP accumulation was prevented by inhibitors of both EGF receptors and endocytosis, suggesting EGF receptors mediate endocytotic uptake. Tumour accumulation of DOX-EBP was significantly higher than free DOX in mice, and the levels of DOX-EBP were similar in DOX-resistant and non-resistant tumour tissues. Importantly, DOX-EBP, but not free DOX, was effective at inhibiting solid tumour growth and increased survival rate in both sensitive and resistant models. DOX-EBP can overcome DOX resistance of tumour cells and increase in vivo antitumour efficacy. Therefore, it has the potential to be a potent therapeutic agent for treating drug-resistant cancers.