Accumulation Of Doxorubicin Research Articles

Cathepsin G-Induced Cell Aggregation of Breast Cancer MCF-7 Decreases Doxorubicin Sensitivity in a Hypoxia-Inducible Factor-Independent Mechanism.

Solid tumors habitually harbor regions with insufficient oxygen away from vasculature. Hypoxia is an important factor that confers malignant phenotypes like chemoresistance to tumor cells. We have demonstrated that cathepsin G (CG) stimulates cell aggregation in breast cancer MCF-7 cells by activating insulin-like growth factor-1 signaling. We investigated whether cancer cell aggregates induced by CG acquire hypoxia-dependent chemoresistance. Pimonidazole staining and hypoxia-inducible factor (HIF)-1α and -2α expression indicated that the core of the cell aggregates was hypoxic. Electrophoretic mobility shift and reporter assays showed that the CG-induced cell aggregates displayed transcriptional activity through HIF-responsive elements. Moreover, HIF target genes PGK1 and SLC2A1 demonstrated upregulated expression in CG-induced cell aggregates, indicating that the aggregates expressed functional HIF. Doxorubicin (DXR)-induced cytotoxicity was significantly lower in the cell aggregates induced by CG compared with monolayer cells under normoxia. Unexpectedly, the upregulation of P-glycoprotein expression, which is reported to be a HIF-target gene, and decreasing intracellular accumulation of DXR was not detected in the cell aggregates as opposed to in monolayer cells under normoxia. Additionally, reduction of DXR sensitivity in the aggregates was not suppressed by treatment with the HIF inhibitor, YC-1 and HIF-1α small interfering RNA (siRNA). Therefore, we conclude that cell aggregation induced by CG decreases DXR sensitivity via a HIF-independent mechanism.

Inhibition of Autophagy Promotes the Elimination of Liver Cancer Stem Cells by CD133 Aptamer-Targeted Delivery of Doxorubicin.

Doxorubicin is the most frequently used chemotherapeutic agent for the treatment of hepatocellular carcinoma. However, one major obstacle to the effective management of liver cancer is the drug resistance derived from the cancer stem cells. Herein, we employed a CD133 aptamer for targeted delivery of doxorubicin into liver cancer stem cells to overcome chemoresistance. Furthermore, we explored the efficacy of autophagy inhibition to sensitize liver cancer stem cells to the treatment of CD133 aptamer-doxorubicin conjugates based on the previous observation that doxorubicin contributes to the survival of liver cancer stem cells by activating autophagy. The kinetics and thermodynamics of aptamer-doxorubicin binding, autophagy induction, cell apoptosis, and self-renewal of liver cancer stem cells were studied using isothermal titration calorimetry, Western blot analysis, annexin V assay, and tumorsphere formation assay. The aptamer-cell binding andintracellular accumulation of doxorubicin were quantified via flow cytometry. CD133 aptamer-guided delivery of doxorubicin resulted in a higher doxorubicin concentration in the liver cancer stem cells. The combinatorial treatment strategy of CD133 aptamer-doxorubicin conjugates and an autophagy inhibitor led to an over 10-fold higher elimination of liver cancer stem cells than that of free doxorubicin in vitro. Future exploration of cancer stem cell-targeted delivery of doxorubicin in conjunction with autophagy inhibition in vivo may well lead to improved outcomes in the treatment of hepatocellular carcinoma.

Doxorubicin causes ferroptosis and cardiotoxicity by intercalating into mitochondrial DNA and disrupting Alas1-dependent heme synthesis.

Clinical use of doxorubicin (DOX) is limited because of its cardiotoxicity, referred to as DOX-induced cardiomyopathy (DIC). Mitochondria-dependent ferroptosis, which is triggered by iron overload and excessive lipid peroxidation, plays a pivotal role in the progression of DIC. Here, we showed that DOX accumulated in mitochondria by intercalating into mitochondrial DNA (mtDNA), inducing ferroptosis in an mtDNA content-dependent manner. In addition, DOX disrupted heme synthesis by decreasing the abundance of 5'-aminolevulinate synthase 1 (Alas1), the rate-limiting enzyme in this process, thereby impairing iron utilization, resulting in iron overload and ferroptosis in mitochondria in cultured cardiomyocytes. Alas1 overexpression prevented this outcome. Administration of 5-aminolevulinic acid (5-ALA), the product of Alas1, to cultured cardiomyocytes and mice suppressed iron overload and lipid peroxidation, thereby preventing DOX-induced ferroptosis and DIC. Our findings reveal that the accumulation of DOX and iron in mitochondria cooperatively induces ferroptosis in cardiomyocytes and suggest that 5-ALA can be used as a potential therapeutic agent for DIC.

Inhibiting the Activity of ABCG2 by KU55933 in Colorectal Cancer.

Therapeutic resistance is a frequent problem of cancer treatment and a leading cause of mortality in patients with metastatic colorectal cancer (CRC). Recent insight into the mechanisms that confer multidrug resistance has elucidated that the ATP-binding cassette (ABC) superfamily G member 2 (ABCG2) assists cancer cells in escaping therapeutic stress caused by toxic chemotherapy. Therefore, it is necessary to develop ABCG2 inhibitors. In the present study, we investigated the inhibitory effect of KU55933 on ABCG2 in CRC. The cytotoxicity assay and drug accumulation assay were used to examine the inhibitory effect of KU55933 on ABCG2. The protein expressions were detected by Western blot assay. The docking assay was performed to predict the binding site and intermolecular interactions between KU55933 and ABCG2. KU55933 was more potent than the known ABCG2 inhibitor fumitremorgin C to enhance the sensitivity of mitoxantrone and doxorubicin and the intracellular accumulation of mitoxantrone, doxorubicin and rhodamine 123 inside CRC cells with ABCG2 overexpression. Moreover, KU55933 did not affect the protein level of ABCG2. Furthermore, the docking data showed that KU55933 was tightly located in the drug-binding pocket of ABCG2. In summary, our data presented that KU55933 could effectively inhibit the drug pump activity of ABCG2 in colorectal cancer, which is further supported by the predicted model that showed the hydrophobic interactions of KU55933 within the drug-binding pocket of ABCG2. KU55933 can potently inhibit the activity of ABCG2 in CRC.

Enhancing Cancer Cell Membrane Permeability by Application of Tumor Treating Fields (TTFields)

<h3>Purpose/Objective(s)</h3> Tumor Treating Fields (TTFields) are alternating electric fields that display anti-mitotic effects on cancer cells. TTFields therapy is approved for treatment of glioblastoma (GBM) and unresectable malignant pleural mesothelioma. It has recently been shown that treatment of GBM cells with TTFields increases cell membrane permeability. The current study aimed to further explore this effect, testing the potential of TTFields to facilitate cellular accumulation of the anticancer agent doxorubicin (DOX) <i>in vitro</i> and <i>in vivo</i>. <h3>Materials/Methods</h3> Breast mammary carcinoma (4T1), breast adenocarcinoma (MCF-7), uterine sarcoma (MES-SA), and lung fibroblast (MRC-5) cell lines were treated with TTFields for 72 h at an intensity of 1.7 V/cm RMS across a range of frequencies (100-400 kHz). Membrane permeability was determined by 7-aminoactinomycin D (7-AAD) intracellular accumulation, and cytotoxicity was examined by cell counts, both measured by flow cytometry. TTFields at the frequency inducing highest permeability were applied together with DOX to DOX-sensitive and matched DOX-resistant 4T1 cells, followed by flow cytometry measurements of intracellular DOX accumulation and cytotoxicity. Mice orthotopically inoculated with 4T1 cells were treated with TTFields for 72 h, with DOX injected 24 h before end of treatment. DOX florescence was measured using flow cytometry for detection at the single-cell level and <i>in vivo</i> imaging system (IVIS) for whole tumor assessment. <h3>Results</h3> While TTFields increased 7-AAD intracellular accumulation in 4T1, MCF-7, and MES-SA cancer cells, there was no such effect when TTFields were applied to the non-cancer MRC-5 cells. Maximal permeability was induced in 4T1 cells with 300 kHz TTFields, whereas highest TTFields-induced cytotoxicity was observed at 150 kHz. TTFields application allowed for DOX accumulation in DOX-resistant cells to the same extent as in DOX-sensitive cells. TTFields delivery also sensitized both cell types to DOX, with cytotoxicity observed at low drug concentrations. <i>In vivo</i>, DOX accumulation was elevated by 2- to 3-fold in tumors isolated from mice treated with TTFields relative to control mice. <h3>Conclusion</h3> TTFields elevated the permeability of 4T1 breast cancer cells, allowing for enhanced intracellular accumulation of DOX and improved drug efficacy, even in cells resistant to DOX. TTFields-induced accumulation of DOX was also demonstrated <i>in vivo</i>.

Palbociclib enhances the effect of doxorubicin-induced apoptosis in activated B-cell-like diffuse large B-cell lymphoma cells.

Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma around the world. While R-CHOP has significantly improved patient outcomes, a subset of patients still has poor outcome. Here, the oncogenic roles of cyclin dependent kinase 4/6 (CDK4/6)-Cyclin D (CCND) signaling axis in DLBCL and its potential mechanism were investigated to explore the possibility of targeting CDK4/6-CCND signaling axis for DLBCL therapy. The transcription levels, functional enrichment analysis, mutation analysis, and prognostic values were performed via the Oncomine, GEPIA, UALCAN, cBioPortal, and Metascape and GenomicScape databases. Expression of CDK4/6-CCND signaling axis in DLBCL patients and DLBCL cell lines was evaluated by qRT-PCR. Additionally, the impact of CDK4/6-CCND signaling axis on cell viability and apoptosis in DLBCL cell lines were evaluated in vitro . The transcription levels of CDK4/6-CCND signaling were increased in DLBCL patients. Meanwhile, in Gene Expression Omnibus dataset, the expression of CDK4 and CCND2 was higher in ABC-DLBCL, whereas the expression of CCND1 and CCND3 was higher in GCB-DLBCL. Moreover, according to the results of qRT-PCR, the expression of CDK4/6-CCND signaling axis in ABC-DLBCL cell line is higher than that in GCB-DLBCL cell lines. Prognostic analysis indicated that upregulation of CDK4, CCND2, and CCND3 was significantly associated with poor survival. Cell function experiments showed that palbociclib could enhance the apoptosis-promoting and cell viability-inhibiting effects of doxorubicin on ABC-DLBCL (SU-DHL-2) cells. Doxorubicin accumulation experiment showed that palbociclib promoted doxorubicin accumulation in ABC-DLBCL cells. Additionally, Western blot analysis demonstrated that palbociclib prevented antiapoptotic protein BCL2 expression in ABC-DLBCL cell line. Our study provides novel insights into targeted therapies for ABC-DLBCL patients.

Discovery of a novel highly potent and low-toxic jatrophane derivative enhancing the P-glycoprotein-mediated doxorubicin sensitivity of MCF-7/ADR cells

Use of novel modulators targeting P-glycoprotein (P-gp, ABCB1 transporter) is among the most accepted strategies for overcoming multidrug resistance in cancer chemotherapy. In the current study, we pursued our structure-activity relationship studies of jatrophane derivatives by structural modification of compound 1, a natural jatrophane isolated from Euphorbia. sororia A. Nine compounds exhibited higher reversal activity in P-gp/ABCB1-mediated MCF-7/ADR cells than verapamil (VRP). The cytotoxicity and doxorubicin (DOX) intracellular accumulation effects of jatrophane derivatives were assessed in normal HEK293T cells and DOX-resistant MCF-7/ADR cells. The most potent compound 17 merits multiple activities, including (1) high efficiency (EC50 = 182.17 ± 32.67 nM) in reversing P-gp-mediated resistance to DOX, low cytotoxicity, and a high therapeutic index; and (2) increasing the accumulation of Rhodamine123 (Rho123) and DOX in a dose-dependent manner compared to verapamil in MCF-7/ADR cells. Our results indicated that the reversal activity of 17 was due to the stimulation of the P-gp ATPase activity instead of the direct inhibition of P-gp protein expression. A docking study demonstrated that 17 has a high binding affinity toward the DOX recognition site of P-gp. This resulted in 17 enhancing the sensitivity of DOX to MCF-7/ADR cells by stimulating P-gp ATPase activity, increasing intracellular DOX and Rho123 concentrations, inhibiting the phosphoinositide 3-kinase/serine-threonine kinase mediated by DOX and further reducing the expression of P-gp. This study provides a promising P-gp inhibitor for reversing multidrug resistance and provides a basis for further research.

Beta-naphthoflavone and doxorubicin synergistically enhance apoptosis in human lung cancer cells by inducing doxorubicin accumulation, mitochondrial ROS generation, and JNK pathway signaling

Doxorubicin is one of the most effective chemotherapeutic agents available for treating various cancers, including lung cancer—the leading cause of cancer death in both men and women. However, its clinical application has been impeded by severe adverse effects, notably cardiotoxicity. Development of cellular resistance to doxorubicin is another major obstacle that must be overcome for broader application of the drug. In the present study, we examined the therapeutic potential of beta-naphthoflavone (BNF), a synthetic derivative of a naturally occurring flavonoid, in combination with doxorubicin for the treatment of lung cancer. Among our novel observations were that BNF enhances the efficacy of doxorubicin by inducing doxorubicin accumulation, mitochondrial ROS generation, and JNK pathway signaling in lung cancer cells. These combined effects were also evident in many other cancer cell types. BNF further exhibited synergistic induction of apoptosis in lung cancer cells when combined with several other cancer drugs, including irinotecan, cisplatin, and 5-fluorouracil. Our results suggest that BNF can be developed as a promising adjuvant agent for enhancing the efficacy of doxorubicin.

Liposome-mediated PD-L1 multivalent binding promotes the lysosomal degradation of PD-L1 for T cell-mediated antitumor immunity

Immune checkpoint blockade (ICB) has shown remarkable therapeutic efficacy in a variety of cancers. However, patients exhibit unexpectedly low response rates to ICB therapy owing to the unwanted recycling and cellular abundance of PD-L1. Herein, rational design of PD-L1 multivalent binding liposome is investigated through PEGylated liposomes incorporating different ratios of PD-L1 binding peptide. Liposomes incorporating 10 mol% PD-L1 binding peptides (10-PD-L1-Lipo) promote the multivalent binding with PD-L1 on tumor cell surface, which is endocytosed for its trafficking toward the lysosomes instead of the recycling endosomes. Thereby, 10-PD-L1-Lipo leads to a significant PD-L1 degradation that prevents its recycling and cellular abundance compared to anti-PD-L1 antibody, disrupting immune escape mechanism of tumor cells and enhancing T cell-mediated antitumor immunity. Moreover, a clinically applicable doxorubicin (DOX) liposomal formulation is established via drug encapsulation into 10-PD-L1-Lipo. The resulting DOX-PD-L1-Lipo primes tumors via immunogenic chemotherapy by preferential DOX accumulation by the EPR effect and overcomes PD-L1 abundance induced following chemotherapy through multivalent binding-mediated PD-L1 degradation. As a result, the synergistic immunogenic chemotherapy and multivalent binding-mediated PD-L1 degradation by DOX-PD-L1-Lipo show significantly enhanced antitumor efficacy and immune responses in colon tumor models. Collectively, this study suggests the rationally designed PEGylated liposomes to promote PD-L1 multivalent binding providing a new route for safe and more effective ICB therapy.

The role of size in PEGylated liposomal doxorubicin biodistribution and anti-tumour activity.

The size of nanoliposome‐encapsulated drugs significantly affects their therapeutic efficacy, biodistribution, targeting ability, and toxicity profile for the cancer treatment. In the present study, the biodistribution and anti‐tumoral activity of PEGylated liposomal Doxorubicin (PLD) formulations with different sizes were investigated. First, 100, 200, and 400 nm PLDs were prepared by remote loading procedure and characterised for their size, zeta potential, encapsulation efficacy, and release properties. Then, in vitro cellular uptake and cytotoxicity were studied by flow cytometry and MTT assay, and compared with commercially available PLD Caelyx®. In vivo studies were applied on BALB/c mice bearing C26 colon carcinoma. The cytotoxicity and cellular uptake tests did not demonstrate any statistically significant differences between PLDs. The biodistribution results showed that Caelyx® and 100 nm liposomal formulations had the most doxorubicin (Dox) accumulation in the tumour tissue and, as a result, considerably suppressed tumour growth compared with 200 and 400 nm PLDs. In contrast, larger nanoparticles (200 and 400 nm formulations) had more accumulation in the liver and spleen. This study revealed that 90 nm Caelyx® biodistribution profile led to the stronger anti‐tumour activity of the drug and hence significant survival extension, and showed the importance of vesicle size in the targeting of nanoparticles to the tumour microenvironment for the treatment of cancer.

Alpha-Lipoic Acid Protects Against Doxorubicin-Induced Cardiotoxicity by Regulating Pyruvate Dehydrogenase Kinase 4.

As a widely used anti-tumor anthracycline, the accumulation of Doxorubicin (DOX) in body causes irreparable cardiomyocyte damage and therefore is limited in clinical application. Strategies to prevent from DOX-associated cardiotoxicity are urgent for patients who undergo DOX-based chemotherapy. Since oxidative stress injury being the major reason for myocardial toxicity of DOX, here we demonstrated that, Alpha-lipoic acid (ALA), which is a reductive agent, plays a cardioprotective role in attenuating DOX-induced cardiotoxicity by inhibiting pyruvate dehydrogenase kinase 4 (PDK4) expression. In vivo, the beneficial effect of ALA was evidenced by increased survival rate, mechanical contraction, and oxidative phosphorylation, while decreased reactive oxidative species (ROS) and apoptosis. In vitro, PDK4 overexpression remarkably increased DOX-induced apoptosis and ROS production in H9C2 cells. Notably, the protective effect of ALA was abrogated by PDK4 overexpression. We further used PDK4 knockout mice to identify the role of PDK4 in DOX-induced cardiotoxicity. Results elicited that PDK4 deficiency showed a consistent effect in protecting DOX cardiotoxicity as ALA treatment, which was evidenced by restored redox homeostasis and mitochondrial metabolism, finally inhibited myocardial injury. In conclusion, the cardioprotective role of ALA against DOX cardiotoxicity was dependent on PDK4-mediated regulation of oxidative stress and mitochondria metabolism.

Targeting TPC2 sensitizes acute lymphoblastic leukemia cells to chemotherapeutics by impairing lysosomal function

Despite novel therapy regimens and extensive research, chemoresistance remains a challenge in leukemia treatment. Of note, recent studies revealed lysosomes as regulators of cell death and chemotherapy response, suggesting this organelle is a novel target for chemosensitization. Interestingly, drug-resistant VCR-R CEM acute lymphoblastic leukemia (ALL) cells have an increased expression of the lysosomal cation channel Two-Pore-Channel 2 (TPC2) compared to drug-naïve CCRF-CEM ALL cells. Concurrently, knockout (KO) of TPC2 sensitized drug-resistant VCR-R CEM cells to treatment with cytostatics. The chemosensitizing effect could be confirmed in several cell lines as well as in heterogeneous, patient-derived xenograft ALL cells, using the pharmacological TPC2 inhibitors naringenin and tetrandrine. We reveal that a dual mechanism of action mediates chemo sensitization by loss of lysosomal TPC2 function. First, because of increased lysosomal pH, lysosomal drug sequestration is impaired, leading to an increased nuclear accumulation of doxorubicin and hence increased DNA damage. Second, lysosomes of TPC2 KO cells are more prone to lysosomal damage as a result of morphological changes and dysregulation of proteins influencing lysosomal stability. This leads to induction of lysosomal cell death (LCD), evident by increased cathepsin B levels in the cytosol, truncation of pro-apoptotic Bid, as well as the reversibility of cell death by co-treatment with the cathepsin B inhibitor CA-074Me in TPC2 KO cells. In summary, this study establishes TPC2 as a novel, promising, druggable target for combination therapy approaches in ALL to overcome chemoresistance, which could be exploited in the clinic in the future. Additionally, it unravels LCD signaling as an important death-inducing component upon loss of TPC2 function.

Ketoconazole Reverses Imatinib Resistance in Human Chronic Myelogenous Leukemia K562 Cells.

Chronic myeloid leukemia (CML) is a hematologic disorder characterized by the oncogene BCR-ABL1, which encodes an oncoprotein with tyrosine kinase activity. Imatinib, a BCR-ABL1 tyrosine kinase inhibitor, performs exceptionally well with minimal toxicity in CML chemotherapy. According to clinical trials, however, 20–30% of CML patients develop resistance to imatinib. Although the best studied resistance mechanisms are BCR-ABL1-dependent, P-glycoprotein (P-gp, a drug efflux transporter) may also contribute significantly. This study aimed to establish an imatinib-resistant human CML cell line, evaluate the role of P-gp in drug resistance, and assess the capacity of ketoconazole to reverse resistance by inhibiting P-gp. The following parameters were determined in both cell lines: cell viability (as the IC50) after exposure to imatinib and imatinib + ketoconazole, P-gp expression (by Western blot and immunofluorescence), the intracellular accumulation of a P-gp substrate (doxorubicin) by flow cytometry, and the percentage of apoptosis (by the Annexin method). In the highly resistant CML cell line obtained, P-gp was overexpressed, and the level of intracellular doxorubicin was low, representing high P-gp activity. Imatinib plus a non-toxic concentration of ketoconazole (10 μM) overcame drug resistance, inhibited P-gp overexpression and its efflux function, increased the intracellular accumulation of doxorubicin, and favored greater apoptosis of CML cells. P-gp contributes substantially to imatinib resistance in CML cells. Ketoconazole reversed CML cell resistance to imatinib by targeting P-gp-related pathways. The repurposing of ketoconazole for CML treatment will likely help patients resistant to imatinib.

Abstract 1801: Application of Tumor Treating Fields (TTFields) to cancer cells enhances their membrane permeability

Abstract INTRODUCTION: Tumor Treating Fields (TTFields) are alternating electric fields at intermediate frequencies that exert anti-mitotic effects on cancerous cells. TTFields therapy is approved in several territories for treatment of glioblastoma (GBM) and unresectable malignant pleural mesothelioma. Recently, membrane permeability of GBM cells has been found to be increased in response to TTFields application. The current study aimed to further explore this effect, testing the potential of TTFields to facilitate cellular accumulation of the anticancer agent doxorubicin (DOX) in breast carcinoma cells. METHODS: 4T1 breast mammary carcinoma cells were treated with TTFields (1.7 V/cm RMS) for 72 h across a frequency range (50-500 kHz). Cytotoxicity was examined by cell counts, and permeability determined by 7-aminoactinomycin D (7-AAD) intracellular accumulation, both measured by flow cytometry. Next, TTFields at the frequency inducing highest permeability was applied to chemotherapy-sensitive and matched chemotherapy-resistant cells. Intracellular accumulation of DOX and drug-induced cytotoxicity were measured by flow cytometry. In vivo validation was performed by 72 h delivery of TTFields at the frequency of maximal permeability to mice orthotopically inoculated with 4T1 cells and injected with DOX 24 h before treatment cessation. DOX florescence was measured using in vivo imaging system (IVIS) for whole tumor assessment and flow cytometry for detection at the single-cell level. RESULTS: While highest TTFields-induced cytotoxicity was observed at 150 kHz, 7-AAD intracellular accumulation was maximal at 300 kHz. When TTFields were delivered concomitant with DOX, the drug accumulated to the same extent in chemotherapy-resistant cells as in chemotherapy-sensitive cells. Application of TTFields also sensitized both cell types to DOX, with cytotoxicity observed at low drug concentrations. Furthermore, 2- to 3-fold higher DOX accumulation was seen in tumors isolated from mice treated with TTFields relative to control. CONCLUSIONS: Permeability of 4T1 breast cancer cells was elevated by TTFields, allowing enhanced intracellular accumulation of DOX and improving drug efficacy, even in chemotherapy-resistant cells. Increased cellular accumulation of DOX was also demonstrated in vivo. Citation Format: Bella Koltun, Tali Voloshin, Tal Kan, Lilach Koren, Yaara Porat, Alexandra Volodin, Noa Kaynan, Anat Klein-Goldberg, Rom Paz, Boris Brant, Yiftah Barsheshet, Efrat Zemer-Tov, Adi Haber, Moshe Giladi, Uri Weinberg, Yoram Palti. Application of Tumor Treating Fields (TTFields) to cancer cells enhances their membrane permeability [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1801.

Induction of P-glycoprotein expression by dandelion in tumor and heart tissues: Impact on the anti-tumor activity and cardiotoxicity of doxorubicin

BackgroundPreviously, we have investigated the anti-tumor activity and mechanism through which dandelion acts against triple-negative breast cancer (TNBC). However, traditional Chinese medicine is mostly accepted as an adjunct therapy during chemotherapy in clinical practice. So far, little is known about the effects of dandelion in conjunction with chemotherapeutic drugs. PurposeTo investigate the effects of dandelion on the anti-tumor activity and cardiotoxicity of doxorubicin (DOX), and to further explore the molecular mechanisms through which these effects occur. Study designAt the beginning of this study, dandelion was observed to alleviate DOX-induced cardiotoxicity and reduce the anti-tumor activity of DOX. Subsequently, we investigated whether the resistance to DOX mediated by P-glycoprotein was involved in the above effects. MethodsThe cardioprotective effect of dandelion was investigated on DOX-treated mice by histological analysis, myocardial enzyme assays, and an untargeted metabolomics study based on LC-Q-TOF/MS. TNBC cell lines and 4T1 tumor-bearing mice were employed to investigate the combined anti-tumor activity. Laser scanning confocal microscope and a flow cytometry analysis were employed to measure the intracellular accumulation of DOX. A specific, sensitive, and rapid LC–MS/MS method was developed to detect the efflux of DOX from cells. Expression of P-glycoprotein in mouse tumor and heart tissues was detected via Western blotting analysis. ResultsDandelion was found to significantly alleviate DOX-induced cardiotoxicity, as was evidenced by improved cardiomyocyte morphology, decreased LDH and CK-MB release, and adjusted metabolic biomarker levels. However, in vitro and in vivo studies showed that dandelion could reduce the anti-tumor activity of DOX. This counteraction was achieved by activating of the drug efflux transporter P-glycoprotein, thereby promoting the efflux of DOX from cells and reducing the intracellular accumulation of DOX. Moreover, the activation of P-glycoprotein by dandelion in mouse heart tissue was also observed, thus suggesting that the decrease of cardiac DOX accumulation plays an important role in the cardioprotective effect of dandelion. ConclusionDandelion can activate the P-glycoprotein in heart and tumor tissues, which ameliorates DOX-induced cardiotoxicity but attenuates DOX cytotoxicity toward TNBC. Our findings have important implications for the correct clinical use of dandelion.

Tumor-associated macrophage membrane-camouflaged pH-responsive polymeric micelles for combined cancer chemotherapy-sensitized immunotherapy

The low immunogenicity and tumor immunosuppressive microenvironment (TIM) are two major obstacles for cancer immunotherapy. Synergistically immunogenic cell death induction and tumor-associated macrophages depletion could perfectly overcome this limitation. Herein, a tumor-associated macrophage (TAMs) membrane-camouflaged pH-responsive doxorubicin (DOX) loaded hyaluronic acid (HA)-g-poly (histidine) polymeric micelles (DHP@M2) was fabricated for the first time. DHP@M2 could effectively accumulated into tumor regions via TAMs membrane mediated immune camouflage. In acidic tumor microenvironment, particle size of DHP was enlarged due to decrease hydrophobic interaction of inner core, which caused a “membrane escape effect” to expose inner HA residue. Together high expression of α4β1 integrin, DHP@M2 could reach CD44/VCAM-1 dual-targetability to facilitate intracellular DOX accumulation for efficient ICD induction. Meanwhile, TAMs membrane could absorb colony stimulating factor 1(CSF1) through high expression of its receptor (CSF1R) on TAMs membrane to deplete TAMs in tumor tissues and relieved TIM. This strategy could efficiently induce cytotoxic T lymphocyte (CTLs) infiltration for antitumor immune response and inhibit tumor progression in 4T1 tumor bearing Balb/c mice. Therefore, DHP@M2 is suitable for cancer chemotherapy-sensitized immunotherapy.

Live Macrophage-Delivered Doxorubicin-Loaded Liposomes Effectively Treat Triple-Negative Breast Cancer.

Triple-negative breast cancer is often aggressive and resistant to various cancer therapies, especially corresponding targeted drugs. It is shown that targeted delivery of chemotherapeutic drugs to tumor sites could enhance treatment outcome against triple-negative breast cancer. In this study, we exploited the active tumor-targeting capability of macrophages by loading doxorubicin-carrying liposomes on their surfaces via biotin-avidin interactions. Compared with conventional liposomes, this macrophage-liposome (MA-Lip) system further increased doxorubicin accumulation in tumor sites, penetrated deeper into tumor tissue, and enhanced antitumor immune response. As a result, the MA-Lip system significantly lengthened the survival rate of 4T1 cell-bearing mice with low toxicity. Besides, the MA-Lip system used highly biocompatible and widely approved materials, which ensured its long-term safety. This study provides a system for triple-negative breast cancer treatment and offers another macrophage-based strategy for tumor delivery.

Dual-targeted and controlled release delivery of doxorubicin to breast adenocarcinoma: In vitro and in vivo studies

Doxorubicin (DOX) is a chemotherapeutic drug that belongs to the anthracyclines family. Cardiotoxicity is one of the main limiting factor of prescribing DOX. To reduce its side effects and enhance the drug delivery to the targeted tissues, we aimed to establish a new targeted and controlled release drug delivery system for treatment of breast cancer. In this article, we tried to synthesize a new nanoplatform consisted of DOX conjugate with hydrazide and disulfide bonds to the hyaluronic acid (HA). Firstly, 4,4′-Dithiodibutyric acid (DTBH) was conjugated with HA. Then, 3-aminophenyl boronic acid monohydrate (APBA) was conjugated with DTBH-HA. Subsequently, DOX was added to DTBH-HA-APBA. HA is a natural polymer with the ability to target CD44, a cell surface adhesion receptor, which are highly overexpressed on the surface of variety of cancer cells. Other targeting agent, APBA can target sialic acid on the cancer cells surface and improve the tumor uptake. Formation of The DTBH-HA-APBA conjugate was confirmed by proton nuclear magnetic resonance (1H NMR) spectroscopy. Scanning emission electron microscopy (SEM) images of the DOX-DTBH-HA-APBA displayed a spherical shape with an average diameter of about 70 nm. In vitro drug release study showed considerably different release pattern of DOX from the formulation at acidic pH (5.4) which was higher than normal pH (7.4). Cellular uptake and cellular cytotoxicity analysis were examined in human breast adenocarcinoma cell line (MCF-7) and mouse breast cancer cells (4T1) as positive cell lines and Chinese Hamster Ovary cells (CHO) as negative cell line. Results confirmed that there is a remarkable difference between dual-targeted (DOX-DTBH-HA-APBA) and single targeted (DOX-DTBH-HA) formulations in both positive cell lines regarding internalization and cytotoxicity. In vivo studies indicated that dual-targeted formulation has the best efficacy with minimum side effects in mouse model. Fluorescence imaging of organs revealed that DOX-DTBH-HA-APBA showed greater DOX accumulation compared with DOX-DTBH-HA and free DOX in tumor site. Also, pathological evaluation indicated that there is no observable cardiotoxicity with final formulation.

Novel Epoxides of Soloxolone Methyl: An Effect of the Formation of Oxirane Ring and Stereoisomerism on Cytotoxic Profile, Anti-Metastatic and Anti-Inflammatory Activities In Vitro and In Vivo.

It is known that epoxide-bearing compounds display pronounced pharmacological activities, and the epoxidation of natural metabolites can be a promising strategy to improve their bioactivity. Here, we report the design, synthesis and evaluation of biological properties of αO-SM and βO-SM, novel epoxides of soloxolone methyl (SM), a cyanoenone-bearing derivative of 18βH-glycyrrhetinic acid. We demonstrated that the replacement of a double-bound within the cyanoenone pharmacophore group of SM with α- and β-epoxide moieties did not abrogate the high antitumor and anti-inflammatory potentials of the triterpenoid. It was found that novel SM epoxides induced the death of tumor cells at low micromolar concentrations (IC50(24h) = 0.7–4.1 µM) via the induction of mitochondrial-mediated apoptosis, reinforced intracellular accumulation of doxorubicin in B16 melanoma cells, probably by direct interaction with key drug efflux pumps (P-glycoprotein, MRP1, MXR1), and the suppressed pro-metastatic phenotype of B16 cells, effectively inhibiting their metastasis in a murine model. Moreover, αO-SM and βO-SM hampered macrophage functionality in vitro (motility, NO production) and significantly suppressed carrageenan-induced peritonitis in vivo. Furthermore, the effect of the stereoisomerism of SM epoxides on the mentioned bioactivities and toxic profiles of these compounds in vivo were evaluated. Considering the comparable antitumor and anti-inflammatory effects of SM epoxides with SM and reference drugs (dacarbazine, dexamethasone), αO-SM and βO-SM can be considered novel promising antitumor and anti-inflammatory drug candidates.

Intratumoral Pi deprivation benefits chemoembolization therapy via increased accumulation of intracellular doxorubicin

It is a decade-long controversy that transarterial chemoembolization (TACE) has definite priority over transarterial embolization (TAE) in treating patients with hepatocellular carcinoma (HCC), since HCC cells are regularly resistant to chemotherapy by enhanced expression of proteins that confer drug resistance, and ABC transporters pump the intracellular drug out of the cell. We addressed this issue by modulating the chemo-environment. In an animal model, sevelamer, a polymeric phosphate binder, was introduced as an embolic agent to induce intratumoral inorganic phosphate (Pi) starvation, and trans-arterially co-delivered with doxorubicin (DOX). The new type of TACE was named as DOX-TASE. This Pi-starved environment enhanced DOX tumoral accumulation and retention, and DOX-TASE thereby induced more severe tumor necrosis than that induced by conventional TACE (C-TACE) and drug-eluting bead TACE (D-TACE) at the same dose. In vitro tests showed that Pi starvation increased the cellular accumulation of DOX in an irreversible manner and enhanced cytotoxicity and cell apoptosis by suppressing the expression of ABC transporters (P-glycoprotein (P-gp), BCRP, and MRP1) and the production of intracellular ATP. Our results are indicative of an alternative interventional therapy combining chemotherapy with embolization more effectively.