ATP-binding Cassette Transporter ABCG2 Research Articles

Associations between ABCG2 gene polymorphisms and gefitinib toxicity in non-small cell lung cancer: a meta-analysis.

BackgroundGefitinib is frequently used to treat patients with non-small cell lung cancer (NSCLC) and is excreted out from cells via the ATP-binding cassette transporter ABCG2. ABCG2 gene polymorphisms have been suggested to be associated with ABCG2 protein expression and function and may influence the risk of gefitinib toxicity in NSCLC patients. Previous studies on the associations between ABCG2 gene polymorphisms and the toxicity of gefitinib in NSCLC patients have produced conflicting results. The aim of this meta-analysis was to determine whether ABCG2 gene polymorphisms are associated with the risk of gefitinib-induced toxicity in NSCLC patients.MethodsThe PubMed and EMBASE databases were searched systematically for all eligible studies. A relative risk with corresponding 95% CI was calculated to evaluate the associations between ABCG2 gene polymorphisms and gefitinib-induced toxicity.ResultsData were finally extracted from seven studies and 515 patients were found to meet the inclusion criteria of the meta-analysis. A dominant model showed that there was no significant association between the ABCG2 C421A polymorphism and the risk of gefitinib-induced toxicity, while the ABCG2 G34A polymorphism might be associated with an increased risk of skin toxicity in gefitinib therapy (relative risk =1.54, 95% CI 1.08–2.21, P=0.02). However, more reliable data are required to confirm the associations between the ABCG2 C421A and ABCG2 G34A polymorphisms and the toxicity of gefitinib in NSCLC patients.ConclusionWhile the ABCG2 C421A polymorphism might not be a reliable marker of gefitinib-related toxicity, the ABCG2 G34A genotype may be predictive of the skin toxicity of gefitinib in NSCLC patients. These conclusions need to be verified in further large-scale studies.

Elimination of stem-like cancer cell side-population by auranofin through modulation of ROS and glycolysis

Cancer side-population (SP) represents a sub-population of stem-like cancer cells that have an important role in drug resistance due to their high expression of the ATP-binding cassette transporter ABCG2 involved in drug export. Auranofin (AF), a clinical drug of gold complex that is used in treatment of rheumatoid arthritis, has been reported inducing tumor antiproliferation. However, whether AF can impact SP cells remains unclear. Our study showed that AF caused a depletion of SP cells and a downregulation of stem cell markers, and impaired their ability to form tumor colonies in vitro and incidence to develop tumors in vivo of lung cancer cells. Reactive oxygen species (ROS) had an important role in mediating AF-induced depletion of SP cells, which could be reversed by antioxidant NAC. Further study revealed that AF could also cause ATP depletion by inhibition of glycolysis. The depletion of cellular ATP might impair the function of ABCG2 pump, leading to increased drug accumulation within the cells and thus enhancing anticancer activity of chemotherapeutic agents such as adriamycin. Synergistic effect of AF and adriamycin was demonstrated both in vitro and in vivo. Simultaneous increase of ROS and inhibition of glycolysis is a novel strategy to eliminate stem-like cancer cells. Combination of AF with adriamycin seems to be promising to enhance therapeutic effectiveness.

Synthetic Analogs of Curcumin Modulate the Function of Multidrug Resistance-Linked ATP-Binding Cassette Transporter ABCG2.

Multidrug resistance (MDR) caused by the overexpression of ATP-binding cassette (ABC) transporters in cancer cells is a major obstacle in cancer chemotherapy. Previous studies have shown that curcumin, a natural product and a dietary constituent of turmeric, inhibits the function of MDR-related ABC transporters, including ABCB1, ABCC1, and especially ABCG2. However, the limited bioavailability of curcumin prevents its use for modulation of the function of these transporters in the clinical setting. In this study, we investigated the effects of 24 synthetic curcumin analogs with increased bioavailability on the transport function of ABCG2. The screening of the 24 synthetic analogs by means of flow cytometry revealed that four of the curcumin analogs (GO-Y030, GO-Y078, GO-Y168, and GO-Y172) significantly inhibited the efflux of the ABCG2 substrates, mitoxantrone and pheophorbide A, from ABCG2-overexpressing K562/breast cancer resistance protein (BCRP) cells. Biochemical analyses showed that GO-Y030, GO-Y078, and GO-Y172 stimulated the ATPase activity of ABCG2 at nanomolar concentrations and inhibited the photolabeling of ABCG2 with iodoarylazidoprazosin, suggesting that these analogs interact with the substrate-binding sites of ABCG2. In addition, when used in cytotoxicity assays, GO-Y030 and GO-Y078 were found to improve the sensitivity of the anticancer drug, SN-38, in K562/BCRP cells. Taken together, these results suggest that nontoxic synthetic curcumin analogs with increased bioavailability, especially GO-Y030 and GO-Y078, inhibit the function of ABCG2 by directly interacting at the substrate-binding site. These synthetic curcumin analogs could therefore be developed as potent modulators to overcome ABCG2-mediated MDR in cancer cells.

Peri-foci adipose-derived stem cells promote chemoresistance in breast cancer

BackgroundMesenchymal stem cells in tumor microenvironment can influence therapeutic responses in various types of cancers. For triple negative breast cancer, chemotherapy remains the mainstay of standard treatment. Our aim was to investigate the correlation between human adipose-derived stem cells (hAdSCs) and chemoresistance in triple negative breast cancer.MethodConditioned medium was collected from hAdSCs, which was isolated from breast cancer patients who had had breast mastectomy. The expression of selected CD markers was evaluated by flow cytometry to characterize hAdSCs. By array analyses of the secreted cytokines and chemokines of hAdSCs, we identified CXCL1 that mediated doxorubicin resistance and the expression of ATP-binding cassette transporters ABCG2 in TNBC. By microRNA microarray, the association between hAdSC-mediated doxorubicin resistance in TNBC was also revealed.ResultsConditioned medium collected from hAdSCs elicited doxorubicin resistance and enhanced the expression of ABCG2, which is a transporter responsible for the efflux of doxorubicin. CXCL1 secreted by hAdSCs downregulated miR-106a expression in triple negative breast cancer, and resulted in ABCG2 upregulation and doxorubicin resistance.ConclusionsOur findings suggest that CXCL1 secreted by hAdSCs elicits doxorubicin resistance through miR-106a-mediated ABCG2 upregulation in triple negative breast cancer. These findings provide a better understanding of the importance of adipose-derived stem cells in breast cancer microenvironment regarding to the development of chemoresistance and reveal the potential of discovering novel therapeutic strategies to overcome drug resistance in TNBC.

Non-small cell lung cancer PC-9 cells exhibit increased sensitivity to gemcitabine and vinorelbine upon acquiring resistance to EGFR-tyrosine kinase inhibitors.

Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (EGFR-TKIs) are widely used for the treatment of non-small cell lung cancers (NSCLCs) harboring EGFR-activating mutations. However, lung cancer cells inevitably acquire resistance to these EGFR-TKIs. The majority of patients whose lung cancer acquires resistance to EGFR-TKIs are subjected to treatment using cytotoxic agents. The present study aimed to determine if lung cancer cells acquiring resistance to EGFR-TKIs also develop altered sensitivity to cytotoxic agents. It was revealed that lung cancer cells that had developed resistance to EGFR-TKIs had increased sensitivity to gemcitabine and vinorelbine compared with EGFR-TKI naïve cells. The expression levels of ATP-binding cassette (ABC) transporter genes, including ABCC3, ABCC5 and ABCG2, were observed to be commonly repressed in EGFR-TKI-resistant cells. In addition, two cases were identified in which gemcitabine and vinorelbine exerted marked responses to lung cancers that had acquired resistance to EGFR-TKIs, even with late-line treatment. Therefore, it was proposed that gemcitabine and vinorelbine may be effective agents for patients with lung cancer previously treated with EGFR-TKIs.

DS-8201a, a new HER2-targeting antibody-drug conjugate incorporating a novel DNA topoisomerase I inhibitor, overcomes HER2-positive gastric cancer T-DM1 resistance.

Anti-HER2 therapies are beneficial for patients with HER2-positive breast or gastric cancer. T-DM1 is a HER2-targeting antibody-drug conjugate (ADC) comprising the antibody trastuzumab, a linker, and the tubulin inhibitor DM1. Although effective in treating advanced breast cancer, all patients eventually develop T-DM1 resistance. DS-8201a is a new ADC incorporating an anti-HER2 antibody, a newly developed, enzymatically cleavable peptide linker, and a novel, potent, exatecan-derivative topoisomerase I inhibitor (DXd). DS-8201a has a drug-to-antibody-ratio (DAR) of 8, which is higher than that of T-DM1 (3.5). Owing to these unique characteristics and unlike T-DM1, DS-8201a is effective against cancers with low-HER2 expression. In the present work, T-DM1-resistant cells (N87-TDMR), established using the HER2-positive gastric cancer line NCI-N87 and continuous T-DM1 exposure, were shown to be susceptible to DS-8201a. The ATP-binding cassette (ABC) transporters ABCC2 and ABCG2 were upregulated in N87-TDMR cells, but HER2 overexpression was retained. Furthermore, inhibition of ABCC2 and ABCG2 by MK571 restored T-DM1 sensitivity. Therefore, resistance to T-DM1 is caused by efflux of its payload DM1, due to aberrant expression of ABC transporters. In contrast to DM1, DXd payload of DS-8201a inhibited the growth of N87-TDMR cells in vitro. This suggests that either DXd may be a poor substrate of ABCC2 and ABCG2 in comparison to DM1, or the high DAR of DS-8201a relative to T-DM1 compensates for increased efflux. Notably, N87-TDMR xenograft tumor growth was prevented by DS-8201a. In conclusion, the efficacy of DS-8201a as a treatment for patients with T-DM1-resistant breast or gastric cancer merits investigation.

CD24 Modulates Chemosensitivity of MCF-7 Breast Cancer Cells.

The role of cluster of differentiation (CD) 24 in breast cancer remains unclear; previously, we showed that CD24 suppresses malignant phenotypes by inactivating Hedgehog signaling through signal transducer and activator of transcription (STAT) 1 inhibition. In this study, we examined how CD24 affects chemosensitivity in breast cancer cells. The CD44+CD24+ breast cancer cell line MCF-7 was transfected with CD24 with/without STAT1 siRNA, and chemosensitivity to 5-fluorouracil (5-FU) and cis-diamminedichloroplatinum (CDDP) was measured. CD24 inhibition reduced chemosensitivity to 5-FU, while STAT1 inhibition did not affect chemosensitivity to 5-FU in CD24 siRNA-transfected cells. Conversely, CD24 inhibition did not affect chemosensitivity to CDDP, while STAT1 inhibition reduced chemosensitivity to CDDP in CD24 siRNA-transfected cells. STAT1 inhibition, but not CD24 inhibition, reduced expression of the ATP-binding cassette (ABC) transporter genes, ABCB1 and ABCG2. In conclusion, CD24 inhibition may modulate chemosensitivity according to drug type, but ABC transporter expression appears not to contribute to this mechanism. This study contributes to determining the role of CD24 in breast cancer.

The Function of SDF-1-CXCR4 Axis in SP Cells-Mediated Protective Role for Renal Ischemia/Reperfusion Injury by SHH/GLI1-ABCG2 Pathway.

Renal ischemia-reperfusion (I/R) injury ranks as the primary cause of acute renal injury with severe morbidity and mortality. Side population (SP) cells have recently drawn increasing attention due to their critical role in injury repair and regeneration. Unfortunately, the underlying mechanism involved in renal I/R remains poorly elucidated. Here, pronounced increases of stromal cell-derived factor-1 (SDF-1) and its receptor CXC chemokine receptor 4 (CXCR4) were substantiated in I/R kidneys from C57BL/6 mice subjected to clamp the bilateral renal pedicles to mimic renal ischemia. Similar up-regulation of them was also determined in SP cells upon simulated ischemia/reperfusion (SI/R). In contrast to non-SP cells, SP cells exhibited higher viability, apoptosis resistance, chemotaxis, and paracrine actions following SI/R treatment, and these were further enhanced after SDF-1 stimulation. Interestingly, blocking CXCR4 signaling with AMD3100 notably ameliorated the above effects. Mechanism analysis corroborated that SDF-1/CXCR4 further induced the expression of ATP-binding cassette transporter ABCG2, an essential element for SP-mediated kidney regeneration after renal I/R injury. Moreover, AMD3100 pretreatment strikingly attenuated ABCG2 elevation in SP cells. Additionally, sonic hedgehog (SHH)-Gli 1 signaling was involved in SDF-1/CXCR4-mediated ABCG2 expression. When SP cells pretreated with AMD3100 were intravenously injected into I/R mice, SP cell-mediated decreases in blood urea nitrogen, serum creatinine, and histological score of kidney were noticeably attenuated, indicating that blocking CXCR4 pathway mitigated the therapeutic function of SP cells in renal I/R injury. Together, this research suggests that SDF-1/CXCR4 axis might act, via Shh-Gli1-ABCG2 signaling, as a positive regulator of SP cell-based therapies for renal I/R by Shh-Gli 1-ABCG2 signaling.

5-Aminolevulinic Acid Fluorescence in Detection of Peritoneal Metastases.

The value of 5-aminolevulinic acid (ALA) in fluorescence detection of peritoneal metastases and the underlying mechanisms were evaluated in patients with peritoneal surface malignancies. Oral 5-ALA was administered at a concentration of 20 mg ⁄kg body weight with 50 ml of water 2 hours prior to surgery (n=115). The diagnostic value of 5-ALA based fluorescence production was evaluated following white light inspection during prior to cytoreductive surgery and hyperthermic intraperitoneal chemotherapy. Then, peptide transporter PEPT1 (ALA influx transporter) and ATP-binding cassette transporter ABCG2 (porphyrin efflux transporter) gene expression was determined with quantitative real time (qRT)-PCR and pathological diagnoses confirmed for all tissue samples. The 5-ALA based photodynamic detection rate was 17% for appendiceal mucinous neoplasms, 54% for colorectal cancers, 33% for gastric cancers, 67% for diffuse malign peritoneal mesotheliomas, and 89% for epithelial ovarian cancer of peritoneal metastases. 5-ALA was detected in all cases of peritoneal metastases originating from cholangiocarcinomas whereas it was not able to detect any in granulosa cell and gastrointestinal stromal tumor cases. Furthermore, PEPT1 was overexpressed whereas ABCG2 expression was downregulated in tumors detected with fluorescence. 5-ALA provided 100% specificity and high sensitivity to detect peritoneal metastases in subgroups of patients with peritoneal surface mailgnancies. ALA influx transporter PEPT1 and porphyrin efflux transporter ABCG2 genes are important in tumor specific 5-ALA induced fluorescence in vivo. Further studies should clarify diagnostic utility of 5-ALA in peritoneal surface malignancies.

ABCG2 deficiency in skin impairs re-epithelialization in cutaneous wound healing.

The ATP-binding cassette transporter ABCG2 is expressed in the interfollicular epidermis and mediates the side-population phenotype in skin cells. However, the role of ABCG2 in skin is unclear. Increased expression levels of ABCG2 were found at the basal layer of transitional epidermis adjacent to cutaneous wounds in human patients, indicating that ABCG2 may be involved in regulating the wound healing process. To investigate the role of ABCG2 in cutaneous wound healing, full-thickness skin wounds were created in ABCG2 knockout (ABCG2-KO) and wild-type mice. The healing process was analysed and revealed that ABCG2 deficiency in skin results in delays in wound closure and impairments in re-epithelialization, as evidenced by reductions in both suprabasal differentiation and in p63-expressing keratinocytes migrating from transitional epidermis to epithelial tongues. The reduction in p63-expressing cells may be due to elevated levels of reactive oxygen species in ABCG2-KO epidermis, which can cause DNA damage and lead to proliferation arrest. To determine whether ABCG2 deficiency affects the potency of epidermal stem/progenitor cells (EPCs), transplantation studies were carried out, which demonstrated that ABCG2-KO EPCs display higher levels of γH2AX and lose the capacity to differentiate into suprabasal keratinocytes. A competitive repopulation assay confirmed that ABCG2 expression is critical for the proper expansion and differentiation of EPCs in cutaneous wounds. As EPCs are known to contribute to the healing of larger wounds, the current findings imply a functional role for ABCG2 in the expansion and differentiation of p63-expressing EPCs. Thus, ABCG2 deficiency in skin impairs re-epithelialization in cutaneous wound healing.

RAGE Against the ABCs.

Diabetes is an insidious disease that significantly contributes to the development of atherosclerotic cardiovascular disease (CVD). One of the major factors contributing to atherosclerosis is the accumulation of lipid-laden macrophages in the vascular wall (1). Removal of cholesterol from macrophages is essential for the regression of atherosclerotic lesions. This can be achieved by cholesterol-lowering drugs (i.e., statins) or by enhancing the reverse cholesterol transport pathway (i.e., raising HDL). Unfortunately, in the setting of diabetes, cholesterol-lowering drugs such as statins can be less effective (2), and preclinical models have revealed a major defect in lesion regression even when cholesterol levels are normalized (3–5). This suggests that a major defect in cholesterol metabolism occurs in the setting of diabetes. The Hedrick and Oram laboratories originally discovered that diabetes causes a downregulation of the ATP binding cassette transporters ABCG1 (6) and ABCA1 (7) in macrophages, promoting foam cell formation. ABCA1 and ABCG1 provide the dominant pathway of cholesterol removal from a number of cells in the body. Yvan-Charvet et al. (8) made the seminal discovery that codeletion of ABCA1/G1 results in a dramatic increase in atherosclerosis. Extensive research on the antiatherogenic role of these transporters has revealed their importance in removing cholesterol from plaque macrophages (9). However, recent discoveries have shown that ABCA1 and ABCG1 regulate the proliferation of hematopoietic stem and progenitor cells to control the abundance of blood monocytes (10). Along with promoting cholesterol efflux from macrophages, suppressing monocyte levels can directly inhibit the progression of atherosclerotic lesions, promote lesion regression, and provide beneficial outcomes after …

The E3 Ubiquitin Ligases, HUWE1 and NEDD4-1, Are Involved in the Post-translational Regulation of the ABCG1 and ABCG4 Lipid Transporters

The ATP-binding cassette transporter ABCG1 has an essential role in cellular cholesterol homeostasis, and dysregulation has been associated with a number of high burden diseases. Previous studies reported that ABCG1 is ubiquitinated and degraded via the ubiquitin proteasome system. However, so far the molecular mechanism, including the identity of any of the rate-limiting ubiquitination enzymes, or E3 ligases, is unknown. Using liquid chromatography mass spectrometry, we identified two HECT domain E3 ligases associated with ABCG1, named HUWE1 (HECT, UBA, and WWE domain containing 1, E3 ubiquitin protein ligase) and NEDD4-1 (Neural precursor cell-expressed developmentally down regulated gene 4), of which the latter is the founding member of the NEDD4 family of ubiquitin ligases. Silencing both HUWE1 and NEDD4-1 in cells overexpressing human ABCG1 significantly increased levels of the ABCG1 monomeric and dimeric protein forms, however ABCA1 protein expression was unaffected. In addition, ligase silencing increased ABCG1-mediated cholesterol export to HDL in cells overexpressing the transporter as well as in THP-1 macrophages. Reciprocally, overexpression of both ligases resulted in a significant reduction in protein levels of both the ABCG1 monomeric and dimeric forms. Like ABCG1, ABCG4 protein levels and cholesterol export activity were significantly increased after silencing both HUWE1 and NEDD4-1 in cells overexpressing this closely related ABC half-transporter. In summary, we have identified for the first time two E3 ligases that are fundamental enzymes in the post-translational regulation of ABCG1 and ABCG4 protein levels and cellular cholesterol export activity.

Allocrite Sensing and Binding by the Breast Cancer Resistance Protein (ABCG2) and P-Glycoprotein (ABCB1).

The ATP binding cassette (ABC) transporters ABCG2 and ABCB1 perform ATP hydrolysis-dependent efflux of structurally highly diverse compounds, collectively called allocrites. Whereas much is known about allocrite-ABCB1 interactions, the chemical nature and strength of ABCG2-allocrite interactions have not yet been assessed. We quantified and characterized interactions of allocrite with ABCG2 and ABCB1 using a set of 39 diverse compounds. We also investigated potential allocrite binding sites based on available transporter structures and structural models. We demonstrate that ABCG2 binds its allocrites from the lipid membrane, despite their hydrophilicity. Hence, binding of allocrite to both transporters is a two-step process, starting with a lipid-water partitioning step, driven mainly by hydrophobic interactions, followed by a transporter binding step in the lipid membrane. We show that binding of allocrite to both transporters increases with the number of hydrogen bond acceptors in allocrites. Scrutinizing the transporter translocation pathways revealed ample hydrogen bond donors for allocrite binding. Importantly, the hydrogen bond donor strength is, on average, higher in ABCG2 than in ABCB1, which explains the higher measured affinity of allocrite for ABCG2. π-π stacking and π-cation interactions play additional roles in binding of allocrite to ABCG2 and ABCB1. With this analysis, we demonstrate that these membrane-mediated weak electrostatic interactions between transporters and allocrites allow for transporter promiscuity toward allocrites. The different sensitivities of the transporters to allocrites' charge and amphiphilicity provide transporter specificity. In addition, we show that the different hydrogen bond donor strengths in the two transporters allow for affinity tuning.

ABCG2 impairs the activity of the aurora kinase inhibitor tozasertib but not of alisertib.

BackgroundRecently, we have shown that the ATP-binding cassette (ABC) transporter ABCB1 interferes with the anti-cancer activity of the pan-aurora kinase inhibitor tozasertib (VX680, MK-0457) but not of the aurora kinase A and B inhibitor alisertib (MLN8237). Preliminary data had suggested tozasertib also to be a substrate of the ABC transporter ABCG2, another ABC transporter potentially involved in cancer cell drug resistance. Here, we studied the effect of ABCG2 on the activity of tozasertib and alisertib.ResultsThe tozasertib concentration that reduces cell viability by 50 % (IC50) was dramatically increased in ABCG2-transduced UKF-NB-3ABCG2 cells (48.8-fold) compared to UKF-NB-3 cells and vector-transduced control cells. The ABCG2 inhibitor WK-X-34 reduced tozasertib IC50 to the level of non-ABCG2-expressing UKF-NB-3 cells. Furthermore, ABCG2 depletion from UKF-NB-3ABCG2 cells using another lentiviral vector expressing an shRNA against the bicistronic mRNA of ABCG2 and eGFP largely re-sensitised these cells to tozasertib. In contrast, alisertib activity was not affected by ABCG2 expression.ConclusionsTozasertib but not alisertib activity is affected by ABCG2 expression. This should be considered within the design and analysis of experiments and clinical trials investigating these compounds.Electronic supplementary materialThe online version of this article (doi:10.1186/s13104-015-1405-4) contains supplementary material, which is available to authorized users.

Curcumin Improves the Tumoricidal Effect of Mitomycin C by Suppressing ABCG2 Expression in Stem Cell-Like Breast Cancer Cells.

Cancer cells with stem cell–like properties contribute to the development of resistance to chemotherapy and eventually to tumor relapses. The current study investigated the potential of curcumin to reduce breast cancer stem cell (BCSC) population for sensitizing breast cancer cells to mitomycin C (MMC) both in vitro and in vivo. Curcumin improved the sensitivity of paclitaxel, cisplatin, and doxorubicin in breast cancer cell lines MCF-7 and MDA-MB-231, as shown by the more than 2-fold decrease in the half-maximal inhibitory concentration of these chemotherapeutic agents. In addition, curcumin sensitized the BCSCs of MCF-7 and MDA-MB-231 to MMC by 5- and 15-fold, respectively. The BCSCs could not grow to the fifth generation in the presence of curcumin and MMC. MMC or curcumin alone only marginally reduced the BCSC population in the mammospheres; however, together, they reduced the BCSC population in CD44+CD24−/low cells by more than 75% (29.34% to 6.86%). Curcumin sensitized BCSCs through a reduction in the expression of ATP-binding cassette (ABC) transporters ABCG2 and ABCC1. We demonstrated that fumitremorgin C, a selective ABCG2 inhibitor, reduced BCSC survival to a similar degree as curcumin did. Curcumin sensitized breast cancer cells to chemotherapeutic drugs by reducing the BCSC population mainly through a reduction in the expression of ABCG2.

Lymphocytes in atherosclerosis.

Atherosclerosis is the major cause of cardiovascular disease, and cardiovascular disease remains the primary cause of death in the United States. The primary complication for patients with type 2 diabetes mellitus (T2D) is cardiovascular disease.1–8 Because the incidences of T2D and obesity are drastically increasing in the United States and now worldwide, understanding how atherosclerosis is accelerated in these diseases becomes even more important to discern. Although many scientific advances in research on atherosclerosis have been made by multiple groups during the past 30 years, particularly about the function of endothelial and smooth muscle biology and macrophage foam cell formation in atherosclerotic plaque development, the roles of immune cells in regulating atherosclerosis onset and progression are still being elucidated. It was widely thought that immune cells played little part in atherogenesis until 1986, when Hansson et al9–12 reported the presence of lymphocytes within atherosclerotic lesions; yet it was many years later until it was widely accepted that immune cells play an important role in atherogenesis. Since 1986, the viewpoint has changed significantly, but details on lymphocyte subsets and mechanisms are still emerging. Moreover, controversy still exists on the roles that some lymphocyte subsets play in atherosclerosis. These discrepancies in findings are at least partly because of differences in mouse models used, diet composition, and length of time on diet for atherosclerosis measurements. However, despite a few ongoing controversies, there are many studies on immune cells in atherogenesis in mice that show clear findings.10,12–23 For example, CD4+ T lymphocytes have been shown to accelerate atherosclerosis.18,24–29 Activated CD4 T cells can be distinguished based on their phenotype, and the predominant CD4+ effector T cells studied to date in the context of atherosclerosis are Th1, …

Effects of plasma membrane ABCB6 on 5-aminolevulinic acid (ALA)-induced porphyrin accumulation in vitro: Tumor cell response to hypoxia

Currently, 5-aminolevulinic acid-based photodynamic diagnosis (ALA-PDD) is used to detect tumors during surgery and exploit tumor-specific accumulation of protoporphyrin IX (PpIX) after administration of ALA. In a recent study, we showed that the human ATP-binding cassette transporter ABCG2 plays a key role in the regulation of PpIX as a specific exporter. However, coproporphyrin III (CPIII) was also detected in urine after ALA administration in patients with tumor, indicating the presence of a CPIII transporter. We used two lines of human gastric cancer cells to measure the ALA-induced porphyrin metabolism. Intracellular and extracellular porphyrin levels and expressions of transporter were determined. In the present study, we showed that although ABCG2 did not transport CPIII, plasma membrane ABCB6 did. Moreover, under conditions of hypoxia, the expression of ABCB6 in plasma membrane was upregulated, resulting in increased extracellular CPIII concentrations. These data indicate that the expression of ABCB6 in plasma membrane is important for porphyrin accumulation after ALA administration, including hypoxic conditions.

Selection of Patients by Membrane Transporter Expressions for Aminolevulinic Acid (ALA)-Guided Photodynamic Detection of Peritoneal Metastases

Background. Photodynamic diagnosis (PDD) with 5-aminolevulinic acid (ALA) was used to detect peritoneal metastasis (PM). This study was done to verify the roles of the expressions of the peptide transporter PEPT1 and ATP-binding cassette transporter ABCG2 genes on the selection of patients in ALA PDD. Methods. The study group comprised 138 patients with PM. After oral administration of 5-ALA, PM was evaluated by PDD. Tissue protoporphyrin IX (PpIX) levels, and PEPT1/ABCG2 mRNA expressions were determined. Results. The tumor detection rate on PDD was 45.6% (63/138). PDD is a safe technique for the detection of PM from ovarian cancer, mesothelioma, and colorectal cancer. PpIX levels of ALA-PDD-positive PM were significantly higher than those of ALA-PDD-negative PM. The PpIX levels in PM that simultaneously expressed PEPT1 and ABCG2 mRNA were significantly higher than those in PM that expressed either PEPT1 or ABCG2 mRNA, as well as PM that expressed neither PEPT1 nor ABCG2 mRNA. PM with PEPT1 mRNA up-regulation showed simultaneous up-regulation of ABCG2 mRNA. PpIX accumulates in PM with PEPT1 up-regulation. At the same time, PpIX may be excreted into stromal tissue through ABCG2 transporter, resulting in the accumulation of excess PpIX in the stromal tissue near cancer cells. Conclusions. Preoperative evaluations of the expressions of the PEPT1 and ABCG2 genes by RT-PCR methods might facilitate the selection of patients most likely to benefit from ALA-PDD.

Critical role of ABCG2 in ALA-photodynamic diagnosis and therapy of human brain tumor.

Primary brain tumors occur in around 250,000 people per year globally. Survival rates in primary brain tumors depend on the type of tumor, patient's age, the extent of surgical tumor removal, and other factors. Photodynamic diagnosis (PDD) is a practical tool currently used in surgical operation of aggressive brain tumors, such as glioblastoma and meningiomas, whereas clinical application of photodynamic therapy (PDT) to brain tumor therapy has just recently started. Both PDD and PDT are achieved by a photon-induced physicochemical reaction, which is induced by the excitation of porphyrins exposed to light. In fluorescence-guided gross-total resection, PDD can be achieved by the administration of 5-aminolevulinic acid (5-ALA) as the precursor of protoporphyrin IX (PpIX). Exogenously administered ALA induces biosynthesis and accumulation of PpIX, a natural photosensitizer, in cancer cells. However, ATP-binding cassette transporter ABCG2 plays a critical role in regulating the cellular accumulation of porphyrins in cancer cells and thereby its expression and function can affect the efficacy of PDD and PDT. In response to the photoreaction of porphyrins leading to oxidative stress, the nuclear factor erythroid-derived 2-related transcription factor can transcriptionally upregulate ABCG2, which may reduce the efficacy of PDD and PDT. On the other hand, certain protein kinase inhibitors potentially enhance the efficacy of PDD and PDT by blocking ABCG2-mediated porphyrin efflux from cancer cells. In this context, it is of great interest to develop ABCG2 inhibitors that can be applied to PDD or PDT for the therapy of brain tumor and other tumors.

Expression of multidrug resistance 1 gene in association with CXCL12 in chronic myelogenous leukaemia

Even though the BCR-ABL tyrosine kinase inhibitor imatinib significantly improves the prognosis of chronic myelogenous leukaemia (CML) patients, drug resistance is a major obstacle to better management. We examined the interaction of recently defined bone marrow microenvironment factors CXCL12 and ATP-binding cassette (ABC) transporters in the bone marrow of CML patients in the chronic phase and blast crisis.Expression levels of mRNA extracted from frozen specimens of CML patients were measured by real-time polymerase chain reaction. The expression of the ABC transporters MDR1, ABCC1, ABCG2, and CXCL12 was significantly higher in the bone marrow samples of CML blast crisis than in those of CML chronic phase. Immunohistochemical staining for CXCL12 revealed that the proportion of CXCL12 positive reticular cell areas correlated well with the mRNA levels of CXCL12 in CML bone marrow. Finally, co-culture experiments of K562 CML cells with CXCL12 expressing mesenchymal cells (OP9 cells or human CXCL12 transfected 3T3 cells) revealed enhanced mRNA levels for MDR1 in a CXCL12 rich environment.These results suggest that imatinib treatment restores the bone marrow microenvironment in CML with the presence of CXCL12 expressing reticular cells but in turn induces the overexpression of MDR1 in haematopoietic cells due to up-regulated expression of CXCL12.