Expression Of ATP-binding Cassette Transporters Research Articles

Diurnal expression of ABC and SLC transporters in jejunum is modulated by adrenalectomy

The circadian clock system drives many physiological processes, including plasma concentration of glucocorticoids and epithelial transport of some ions and nutrients. As glucocorticoids entrain the circadian rhythms in various peripheral organs, we examined whether adrenalectomy affects the expression and circadian rhythmicity of intestinal transporters of the solute carrier (SLC) and ATP-binding cassette (ABC) families, which participate in intestinal barriers for absorption of nutrients, nonnutrients and oral drugs. The rat jejunum showed rhythmic circadian profiles of Sglt1, Pept1, Nhe3, Mdr1 and Mrp2 but not Mct1, Oct1, Octn1, Oatp1, Cnt1 and Bcrp. With the exception of Pept1 and Mct1, adrenalectomy decreased the expression of all rhythmic and arrhythmic transporters including the amplitude of Sglt1 and Nhe3 rhythms but minimally affected the phases of rhythmic transporters except of Nhe3. Similarly, adrenalectomy downregulated the expression of rhythmic (Pparα, Hlf, Pgc1α) and arrhythmic (Hnf1β, Hnf4α) transcription factors, which are known to regulate the expression of transporters. We conclude that endogenous corticosteroids have a profound effect on the expression of intestinal SLC and ABC transporters and their nuclear transcription factors. The circulating corticosteroids are necessary for maintaining upregulated expression of Sglt1, Oct1, Octn1, Oatp1, Cnt1, Nhe3, Mdr1, Bcrp, Mrp2, Pparα, Pgc1α, Hnf1β, Hnf4α and Hlf and for maintaining the high amplitude of Sglt1, Nhe3, Pparα, Pgc1α and Hlf circadian rhythms. The study demonstrates that signals from the adrenal gland are necessary for maintaining the expression of arrhythmic and rhythmic intestinal transporters and that changes in the secretion of corticosteroids associated with stress might reorganize intestinal transport barriers.

Determinants of drug entry into the developing brain

Background: A major concern for clinicians in prescribing medications to pregnant women and neonates is the possibility that drugs might have damaging effects, particularly on long-term brain development. Current understanding of drug permeability at placental and blood-brain barriers during development is poor. In adults, ABC transporters limit many drugs from entering the brain; however, little is known about their function during development. Methods: The transfer of clinically relevant doses of paracetamol (acetaminophen), digoxin and cimetidine into the brain and cerebrospinal fluid (CSF) was estimated using radiolabelled drugs in Sprague Dawley rats at three developmental stages: E19, P4 and adult. Drugs were applied intraperitoneally either acutely or following chronic exposure (for five days). Entry into brain, CSF and transfer across the placenta was measured and compared to three markers (L-glucose, sucrose, glycerol) that cross barriers by “passive diffusion”. The expression of ABC transporters in the brain, choroid plexus and placenta was estimated using RT-qPCR. Results: All three drugs entered the developing brain and CSF in higher amounts than the adult brain and CSF. Comparisons with “passive” permeability markers suggested that this might be due to age-related differences in the functional capacity of ABC-efflux mechanisms. In adult animals, chronic treatment reduced digoxin (12% to 5%, p<0.01) and paracetamol (30% to 21%, p<0.05) entry compared to acute treatment, with the decrease in digoxin entry correlating with up-regulation of efflux transporter abcb1a (PGP). In fetal and newborn animals, no gene up-regulation or transfer decreases were observed. Instead, chronic paracetamol treatment resulted in increased transfer into the fetal brain (66% to 104%, p<0.001). Conclusions: These results suggest that the developing brain may be more at risk from acute drug exposure than the adult brain due to reduced efflux capacity and at greater risk from chronic treatment due to a lack of efflux mechanism regulatory capacity.

Role of ATP-binding cassette transporters in cancer initiation and progression.

ATP Binding Cassette (ABC) transporters, widely studied in cancer for their role in drug resistance, have been more recently also considered for their contribution to cancer cell biology. To date, many data provide evidences for their potential role in all the phases of cancer development from cancer susceptibility, tumor initiation, tumor progression and metastasis. Although many evidences are based on correlative analyses, data describing a direct or indirect role of ABC transporters in cancer biology are increasing. Overall, current available information suggests a relevant molecular effector role of some ABC transporters in cancer invasion and metastasis as reported in experimental tumor models. From a therapeutic point of view, due to the physiological relevant roles that ABC transporters play in the organism, the capability to selectively inhibit the function or the expression of ABC transporters in cancer stem cells or other tumor cells, represents the main challenge for researcher scientists. A detailed and updated description of the current knowledge on the role of ABC transporters in cancer biology is provided.

Determinants of drug entry into the developing brain.

Background: A major concern for clinicians in prescribing medications to pregnant women and neonates is the possibility that drugs might have damaging effects, particularly on long-term brain development. Current understanding of drug permeability at placental and blood-brain barriers during development is poor. In adults, ABC transporters limit many drugs from entering the brain; however, little is known about their function during development. Methods: The transfer of clinically relevant doses of paracetamol (acetaminophen), digoxin and cimetidine into the brain and cerebrospinal fluid (CSF) was estimated using radiolabelled drugs in Sprague Dawley rats at three developmental stages: E19, P4 and adult. Drugs were applied intraperitoneally either acutely or following chronic exposure (for five days). Entry into brain, CSF and transfer across the placenta was measured and compared to three markers (L-glucose, sucrose, glycerol) that cross barriers by "passive diffusion". The expression of ABC transporters in the brain, choroid plexus and placenta was estimated using RT-qPCR. Results: All three drugs entered the developing brain and CSF in higher amounts than the adult brain and CSF. Comparisons with "passive" permeability markers suggested that this might be due to age-related differences in the functional capacity of ABC-efflux mechanisms. In adult animals, chronic treatment reduced digoxin (12% to 5%, p<0.01) and paracetamol (30% to 21%, p<0.05) entry compared to acute treatment, with the decrease in digoxin entry correlating with up-regulation of efflux transporter abcb1a (PGP). In fetal and newborn animals, no gene up-regulation or transfer decreases were observed. Instead, chronic paracetamol treatment resulted in increased transfer into the fetal brain (66% to 104%, p<0.001). Conclusions: These results suggest that the developing brain may be more at risk from acute drug exposure than the adult brain due to reduced efflux capacity and at greater risk from chronic treatment due to a lack of efflux mechanism regulatory capacity.

Dysregulation of placental ABC transporters in a murine model of malaria-induced preterm labor

Malaria in Pregnancy (MiP) is characterized by placental accumulation of Plasmodium-infected erythrocytes, intrauterine growth restriction (IUGR) and preterm delivery (PTD). Placental ATP-binding cassette (ABC) transporters mediate the efflux of nutrients, cytokines and xenobiotics. The expression and activity of these transporters are highly responsive to infection. We hypothesized that MiP would perturb the expression of placental ABC transporters, promoting PTD. Peripheral blood, spleens, livers and placentas of pregnant mice, infected with Plasmodium berghei ANKA on gestational day (GD) 13.5, were collected and analyzed on GD18.5. The primary consequences of human MiP, including IUGR, PTD (20%) and placental inflammation, were recapitulated in our mouse model. Electron microscopy revealed attenuated presence of labyrinthine microvilli and dilated spongiotrophoblasts -granular endoplasmic reticulum cisternae. Additionally, a decrease in placental Abca1 (ABCA1), Abcb1b (P-glycoprotein), Abcb9 and Abcg2 (BCRP) expression was observed in MiP mice. In conclusion, MiP associated with PTD impairs placental ABC transporters’ expression, potentially modulating placental nutrient, environmental toxin and xenobiotic biodistribution within the fetal compartment, and may, at some degree, be involved with pregnancy outcome in MiP.

Abstract 3732: Targeted photodynamic therapy enhances the therapeutic efficacy of combination therapy (PDT and chemotherapy) on chemoresistant melanoma cells

Abstract Cutaneous melanoma remains refractory to current therapies such as chemotherapy. Failure of treatment has been attributed to enhanced self-renewal capacity and the over-expression of ABC transporters by therapy resistant populations. A need therefore exists to identify and evaluate new approaches. Alternative combinatorial approaches comprising chemotherapeutics, photodynamic therapy (PDT) and targeted-PDT have been proposed to overcome resistance. Objective This study set to investigate the efficacy of novel (targeted-PDT-IR700) and standard combinatorial effect of PDT and chemotherapy (Hypericin-PDT and DTIC) as an adjunctive combination therapy treatment to decrease cellular chemoresistance in melanoma. This was assessed in an in vitro model, to show differences in killing of melanoma cells. Methodology Single chain variable fragments (scFv) of panitumumab were cloned into a transient eukaryotic SNAP expression vector co-expressing cytosolic proteins and transfected into HEK293T cells. Purified recombinant protein was confirmed by SDS-PAGE and Western blot analyses. Benzylguanine (BG) modified IR700 was irreversibly conjugated in a 1:1 stochiometric ratio to the 1711(scFv)-SNAP using the self-labelling property of the suicide enzyme SNAP-tag. Binding activity and cytotoxicity was documented on EGFR expressing melanoma cells using flow cytometry, confocal microscopy and XTT assays. Cytotoxic activity, self-renewal capacity and ABC transporters expression (ABCB5 and ABCG2) after the combination therapy (HYP-PDT and DTIC) was determined using XTT, clonogenic assays and flow cytometry. Results Binding and internalization of purified 1711(scFv)-SNAP conjugated to BG-modified Alexa 488 or IR700 was successfully confirmed by confocal microscopy and flow cytometry. Cell viability showed cell specific cytotoxicity of the photosensitizer labeled fusion proteins upon light induction, at nanomolar concentrations which were lower than those used for the non-targeted combinatorial approach (HYP-PDT and DTIC). Moreover, DTIC resistant melanoma cells were less sensitive to therapeutic treatment than their non-resistant counterpart. This correlated with an increased self-renewal capacity and ABC transporter expression in DTIC resistant melanoma in comparison to their non-resistant counterpart. This study describes a proof of concept of selective targeting and elimination of EGFR expressing melanoma cells by targeted photoimmunotherapy using a panitumumab 1711-scFv fused to SNAP tag and conjugated to near infrared photosensitizer IR700 which improve therapeutic efficacy with lower concentration and reduced sides effect. This work highlights the efficacy of targeted PDT and suggest that it is highly applicable to improve the current clinical arsenal of therapies. Note: This abstract was not presented at the meeting. Citation Format: Lester M. Davids, Fleury Nsole Biteghe, Eden Padayachee, Stefan Barth. Targeted photodynamic therapy enhances the therapeutic efficacy of combination therapy (PDT and chemotherapy) on chemoresistant melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3732.

Multi-kinase inhibitors and cisplatin for head and neck cancer treatment in vitro.

Multidrug resistance (MDR) remains one of the major causes of suboptimal outcome following therapy in head and neck squamous cell carcinoma (HNSCC). ATP-binding cassette (ABC) transporters are overexpressed in HNSCC, which contributes to the limited effect of chemotherapeutic treatment. In addition to their named function, tyrosine kinase inhibitors (TKIs) have been revealed to impact on ABC transporter activity and expression. Therefore, the present study aimed to investigate the effects of combination therapy using different TKIs combined with cisplatin. Reverse transcription-quantitative PCR was used to characterize ABC transporter and receptor expression in 5 HNSCC cell lines treated with 3 different TKIs (pazopanib, dovitinib, nintedanib) and cisplatin. Treatment efficacy was analyzed using a crystal violet staining assay. Analysis of ABC transporter (ABCB1, ABCC1 and ABCG2) genetic alterations was performed using The Cancer Genome Atlas. Statistical analysis was conducted to evaluate the effects of mono- and combination treatment. With the exception of ABCB1, all of the investigated ABC transporters were expressed in each cell line. The additive effects of TKI + cisplatin combination treatment were observed for pazopanib in three cell lines, nintedanib in four cell lines, and were not observed for dovitinib in any of the cell lines investigated. The combination of multi-kinase inhibitors and conventional chemotherapy in HNSCC may strengthen the use of current therapeutic strategies; nintedanib appears to be the most suitable TKI for combination therapy. Further efforts are required to classify TKI efficacy with regard to cisplatin resistance.

Thymosin β10 as a predictive biomarker of response to 5-fluorouracil chemotherapy in cholangiocarcinoma

Introduction and aim. 5-Fluorouracil (5-FU) is the most commonly used chemotherapeutic drug in the treatment of cholangiocarcinoma (CCA). Since development of drug resistance to 5-FU in CCA patients is the primary cause of treatment failure, a better understanding of the mechanism of drug resistance of this cancer is essential to improve the efficacy of 5-FU in CCA therapy.Material and methods. A 5-FU resistant CCA cell line (M214-5FUR) for a comparative chemo-resistance study was established. Real time RT-PCR was used to determine gene expression levels. Cell cytotoxicity was measured by the MTT assay. Protein expression levels were detected by the immunofluorescene method.Results. It was found that 5-FU resistance was associated with the overexpression of Tβ10 in CCA cell lines. 5-FU treatment at various concentrations induced the expressions of Tβ10 and ABC transporters (ABCB1, ABCG2 ABCA3) in two CCA cell lines, KKU-M055 and KKU-M214. M214-5FUR, a 5-FU-resistant cell line, exhibited a 5-FU resistant phenotype with a 16-fold extremely high expression of Tβ10 and ABC transporters, as compared to the parental cells, KKU-M214. siRNA targeted to Tβ10 significantly reduced expression of ABC transporters tested in the M214-5FUR cells (P < 0.05).Conclusions. The present novel findingsof Tβ10 connected with drug resistance as shown in this study provides a new insight for the therapeutic value of Tβ10 as a predictive biomarker of 5-FU chemoresistance. Inhibiting Tβ10 may be a valuable adjunct for suppression of ABC transporters and sensitizing chemotherapy treatment, especially 5-FU in CCA patients.

ABCG2 expression is related to low 5-ALA photodynamic diagnosis (PDD) efficacy and cancer stem cell phenotype, and suppression of ABCG2 improves the efficacy of PDD.

Photodynamic diagnosis/therapy (PDD/PDT) are novel modalities for the diagnosis and treatment of cancer. The photosensitizer protoporphyrin IX is metabolized from 5-aminolevulinic acid (5-ALA) intracellularly, and PDD/PDT using 5-ALA have been approved in dermatologic malignancies and gliomas. However, the molecular mechanism that defines the efficacy of PDD/PDT is unknown. In this study, we analyzed the functions of ATP-binding cassette (ABC) transporters in PDD using 5-ALA. Most of the human gastrointestinal cancer line cells examined showed a homogenous staining pattern with 5-ALA, except for the pancreatic cancer line PANC-1, which showed heterogeneous staining. To analyze this heterogeneous staining pattern, single cell clones were established from PANC-1 cells and the expression of ABC transporters was assessed. Among the ABC transporter genes examined, ABCG2 showed an inverse correlation with the rate of 5-ALA-positive staining. PANC-1 clone #2 cells showed the highest level of ABCG2 expression and the lowest level of 5-ALA staining, with only a 0.6% positive rate. Knockdown of the ABCG2 gene by small interfering RNAs increased the positive rate of 5-ALA staining in PANC-1 wild-type and clone cells. Interestingly, PANC-1 clone #2 cells showed the high sphere-forming ability and tumor-formation ability, indicating that the cells contained high numbers of cancer stem cells (CSCs). Knockdown or inhibition of ABCG2 increased the rate of 5-ALA staining, but did not decrease sphere-forming ability. These results indicate that gastrointestinal cancer cell lines expressing high levels of ABCG2 are enriched with CSCs and show low rates of 5-ALA staining, but 5-ALA staining rates can be improved by inhibition of ABCG2.

Effects of histone deacetylase inhibitors on ATP-binding cassette transporters in lung cancer A549 and colorectal cancer HCT116 cells.

Histone deacetylase (HDAC) inhibitors and DNA alkylators are effective components used in combination chemotherapy. In the present study, the effects of HDAC inhibitors on the expression of ATP-binding cassette (ABC) transporters were investigated. It was observed that HDAC inhibitors induced the expression of multidrug-resistant ABC transporters differently in lung cancer A549 cells than in colorectal cancer HCT116 cells. In these two cell lines, the HDAC inhibitors suberoylanilide hydroxamic acid (SAHA) and trichostatin A (TSA) significantly increased ABCB1 expression at the mRNA and protein levels, whereas they had no evident effect on ABCG2 protein expression. SAHA and TSA decreased ABCG2 mRNA expression in A549 cells and had no evident effect on ABCG2 mRNA expression in HCT116 cells. Notably, SAHA and TSA increased the mRNA expression levels of ABCC5, ABCC6, ABCC10, ABCC11 and ABCC12, as well as the protein expression levels of ABCC2, ABCC10 and ABCC12. By contrast, these inhibitors decreased the mRNA expression levels of ABCC1, ABCC2, ABCC3 and ABCC4, as well as the expression of ABCC1 and ABCC3 proteins. Furthermore, SAHA and TSA were found to downregulate HDAC3 and HDAC4, but not HDAC1 and HDAC2. Taken together, the results suggested that HDAC inhibitors work synergistically with DNA alkylators, in part, due to the inhibitory effect of these inhibitors on ABCC1 expression, which translocates these alkylators from inside to outside of cancer cells. These results further suggested the possibility of antagonism when HDAC inhibitors are combined with anthracyclines and other ABCB1 drug ligands in chemotherapy.

MicroRNAs contribute to ATP-binding cassette transporter- and autophagy-mediated chemoresistance in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) has an elevated mortality rate, largely because of high recurrence and metastasis. Additionally, the main obstacle during treatment of HCC is that patients usually develop resistance to chemotherapy. Cancer drug resistance involves many different mechanisms, including alterations in drug metabolism and processing, impairment of the apoptotic machine, activation of cell survival signaling, decreased drug sensitivity and autophagy, among others. Nowadays, miRNAs are emerging as master regulators of normal physiology- and tumor-related gene expression. In HCC, aberrant expression of many miRNAs leads to chemoresistance. Herein, we particularly analyzed miRNA impact on HCC resistance to drug therapy. Certain miRNAs target ABC (ATP-binding cassette) transporter genes. As most of these miRNAs are downregulated in HCC, transporter levels increase and intracellular drug accumulation decrease, turning cells less sensitive to death. Others miRNAs target autophagy-related gene expression, inhibiting autophagy and acting as tumor suppressors. Nevertheless, due to its downregulation in HCC, these miRNAs do not inhibit autophagy or tumor growth and, resistance is favored. Concluding, modulation of ABC transporter and/or autophagy-related gene expression or function by miRNAs could be determinant for HCC cell survival under chemotherapeutic drug treatment. Undoubtedly, more insights on the biological processes, signaling pathways and/or molecular mechanisms regulated by miRNAs are needed. Anyway, miRNA-based therapy together with conventional chemotherapeutic drugs has a great future in cancer therapy.

MON-311 GH Action Upregulates ABC Transporters And EMT Markers In A Syngeneic Melanoma Mouse Model

Compared to many other forms of malignancies, melanoma expresses growth hormone receptors (GHRs) to a uniquely higher degree than other human cancers. Attenuating these receptors in cultured human melanoma cells resulted in decreased proliferation, and suppressed expression of ATP-binding cassette (ABC) transporters and epithelial-mesenchymal transition (EMT) markers. We hypothesize that GH promotes both the growth of mouse melanoma in vivo and the expression of ABC transporters and EMT markers. Thus, we propose that the use of a GH antagonist (GHA) could be effective at reversing these changes. To test these hypotheses, a syngeneic mouse B16-F10 melanoma model, cell proliferation assays, Western blotting, ELISAs, and real-time RT-qPCR were used. First, the high expression levels of GHR in B16-F10 cells were verified. Then, by knocking down GHR expression via small interfering RNA, the increased cell growth stimulated by GH administration was effectively reversed, independent of (insulin-like growth factor-1) IGF-1. To distinguish the effects of GH and IGF-1 on melanoma in vivo, bovine GH overexpressing transgenic mice (bGH) and GHR-knock out mice (GHR-/-) were used. Both mouse lines have significantly elevated GH, while bGH mice also exhibit high levels of IGF-1 whereas extremely low levels found in GHR-/- mice. Mouse melanoma cells were subcutaneously injected into bGH mice, GHR-/- mice, and their corresponding wild type (WT) littermate controls. Our data revealed that the ABC transporters and EMT markers in both bGH and GHR-/- mouse melanomas were significantly higher compared to their WT controls, implying that some of these effects were attributable directly to GH as opposed to IGF-1. Data analyzed from the Cancer Genome Atlas repository also suggests that human melanomas with higher GHR levels have significantly elevated expression levels of EMT markers, validating this syngeneic mouse melanoma model. On the other hand, transgenic mice that express a GHA and that have decreased GH action as well as circulating IGF-1 showed significantly decreased melanoma sizes, suggesting that targeting GHR, resulting in a compromised GH/IGF-1 axis, could delay the progression of melanoma. In summary, these results reveal the role of GH action in the upregulation of ABC transporters and EMT markers in vivo, both relevant with regards to tumor progression and poor clinical prognoses. Importantly, GH antagonism may inhibit the progression of melanoma in vivo. Studies are ongoing to determine whether GHA treatment is beneficial in the treatment of mouse melanoma in mice, and if so, blocking GH action may be considered as an anti-melanoma therapy in humans.

Developmental differences in the expression of ABC transporters at rat brain barrier interfaces following chronic exposure to diallyl sulfide

Many pregnant women and prematurely born infants require medication for clinical conditions including cancer, cardiac defects and psychiatric disorders. In adults drug transfer from blood into brain is mostly restricted by efflux mechanisms (ATP-binding cassette, ABC transporters). These mechanisms have been little studied during brain development. Here expression of eight ABC transporters (abcb1a, abcb1b, abcg2, abcc1, abcc2, abcc3, abcc4, abcc5) and activity of conjugating enzyme glutathione-s-transferase (GST) were measured in livers, brain cortices (blood-brain-barrier) and choroid plexuses (blood-cerebrospinal fluid, CSF, barrier) during postnatal rat development. Controls were compared to animals chronically injected (4 days, 200 mg/kg/day) with known abcb1a inducer diallyl sulfide (DAS). Results reveal both tissue- and age-dependent regulation. In liver abcb1a and abcc3 were up-regulated at all ages. In cortex abcb1a/b, abcg2 and abcc4/abcc5 were up-regulated in adults only, while in choroid plexus abcb1a and abcc2 were up-regulated only at P14. DAS treatment increased GST activity in livers, but not in cortex or choroid plexuses. Immunocytochemistry of ABC transporters at the CSF-brain interface showed that PGP and BCRP predominated in neuroepithelium while MRP2/4/5 were prominent in adult ependyma. These results indicate an age-related capacity of brain barriers to dynamically regulate their defence mechanisms when chronically challenged by xenobiotic compounds.

The reference liver - ABC and SLC drug transporters in healthy donor and metastatic livers.

Analysis of results and conclusions in studies dedicated to pathology of the liver are usually based on comparison of pathological liver specimens and control/reference (considered as healthy) tissues. There are two main sources of the control liver samples used as the reference livers, i.e. deceased organ donor livers and non-tumorous tissue from metastatic livers, which are also applied for drug transporter investigations. However, no information has yet been published on drug transporters in these two major types of reference livers. We explored ABC (P-gp, MRP1, MRP2, MRP3, MRP4, BCRP, BSEP) and SLC (NTCP, MCT1, OCT1, OCT3, OAT2, OATP1B1, OATP1B3, OATP2B1) family transporters expression (qPCR) and protein abundance (LC-MS/MS) in healthy donors (n = 9) and metastatic (n = 13) livers. The analysis of mRNA content revealed significant differences in ABCB11, ABCC1, ABCG2, SLC10A1, SLC16A1, SLCO1B1 and SLCO2B1 gene expression between livers from organ donors and patients who underwent surgical resection of metastatic tumors. The protein abundance of NTCP was significantly higher, whereas of P-gp significantly lower in non-tumorous tissues from metastatic livers. Greater inter-individual variability in protein abundance of all studied transporters in subjects with metastatic colon cancer was also observed. The results suggest that final conclusions in liver pathology studies may depend on the reference liver tissue used, especially in gene expression studies.

Prognostic implication of ABC transporters and cancer stem cell markers in patients with stage III colon cancer receiving adjuvant FOLFOX‑4 chemotherapy

Cancer stem cell (CSC) and ATP-binding cassette (ABC) transporters are associated with treatment resistance and outcomes of patients with cancer. The present study investigated the prognostic implications of pre-therapeutic expression of ABC transporters and CSC markers in patients with colon cancer (CC) who received adjuvant 5-fluorouracil, leucovorin and oxaliplatin combination therapy (FOLFOX-4). The immunohistochemical expression of 3 ABC transporters, including ABC subfamily C member 2 (ABCC2), ABCC3 and ABC subfamily G member 2 (ABCG2), and 3 CSC markers, including sex determining region Y-box 2 (SOX2), leucine-rich repeat-containing G protein-coupled receptor 5 and aldehyde dehydrogenase 1, were determined in 164 CC tissues from patients with stage III CC, who underwent postoperative FOLFOX-4 chemotherapy. The association between the protein expression and patients' prognoses was statistically analyzed. ABCG2 was associated with favorable overall survival rate (OS; P=0.001), and ABCC2, ABCG2 and SOX2 were associated with increased disease-free survival rate (DFS; P=0.001, 0.002 and 0.013, respectively). In multivariate analyses, ABCG2 was an independent prognostic factor for OS [hazard ratio (HR)=2.877; P=0.046], and ABCC2 and SOX2 were independent prognostic factors for DFS (HR=2.831; P=0.014; HR=2.558, P=0.020, respectively). ABCC2, ABCG2 and SOX2 may be promising prognostic markers for patients with CC receiving FOLFOX-4 therapy.

Combination of tanshinone IIA and doxorubicin possesses synergism and attenuation effects on doxorubicin in the treatment of breast cancer.

Doxorubicin (Dox) is a first-line drug for breast cancer chemotherapy. However, with the prolongation of chemotherapy cycle, breast cancer cells are increasingly tempt to resist Dox, and meanwhile, high cumulative dose of Dox brings enhancing toxic side effects, and these effects may lead to chemotherapy failure. Hence, it is necessary to search an agent in combination medication with Dox, which can not only enhance the chemosensitivity of Dox but also reduce the toxic side effects. Tanshinone IIA (Tan IIA) is reported to have antitumor activity in addition to its cardiovascular protective effects. We employed human breast cancer MCF-7 and MCF-7/dox cells in order to assess whether Tan IIA might perform such function. Our in vitro studies showed that Tan IIA could enhance the sensitivity of breast cancer cells to Dox through inhibiting the PTEN/AKT pathway and downregulating the expression of efflux ABC transporters including P-gp, BCRP, and MRP1. In addition, our in vivo studies showed Tan IIA enhanced the chemotherapeutic effect of Dox against breast cancer while reducing its toxic side effects including weight loss, myelosuppression, cardiotoxicity, and nephrotoxicity. Therefore, Tan IIA could be used as a novel agent combined with Dox in breast cancer therapy.

Antioxidant α‐mangostin downregulates ATP‐binding cassette transporters in HuT78 cells

Sézary syndrome (SS) is an aggressive form of cancer called cutaneous T‐cell lymphoma (CTCL). Patients with SS carry poor prognosis, with a median survival of approximately 40 months after diagnosis and a 5‐year survival rate below 40%. The offending neoplastic T cells for SS known as Sézary cells are aggressive and rapid growing. They are refractory to chemotherapy, and have been posting the biggest challenge for the treatment of SS. The principal mechanism by which the Sézary cells develop resistance to chemotherapeutic agents is through the overexpression of ATP‐binding cassette (ABC) transporter proteins such as P‐glycoprotein (P‐gp), multidrug‐resistance associated protein 1 (MRP1), and breast cancer resistance protein (BCRP). The expressions of P‐gp, MRP1 and BCRP genes appear to be regulated, in part, by nuclear factor E2‐related factor 2 (Nrf2), a transcriptional regulator of cellular resistance to oxidants. α‐Mangostin (α‐MGT) is a xanthone derivative that possesses antioxidant and cytotoxic activities in various types of cancers. This study is undertaken to support the hypothesis that through its antioxidant activity, α‐MGT mediates downregulation of ABC transporters expression in HuT78, the cell line isolated from a Sézary patient. The results revealed that HuT78 cells expressed P‐gp, MRP1 and BCRP at various frequencies, ranging from 31% to 78% where the expression level of MRP1 was the highest followed by that of BCRP and P‐gp. Upon exposure of HuT78 cells to 12 μM α‐MGT for 24 hours, the expression of MRP1 was decreased by 50%, BCRP was slightly downregulated, and the P‐gp was undetectable. Our preliminary data suggested that α‐MGT may be beneficial in reversing the resistant phenotype of Sézary cells for chemotherapy.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Guajadial reverses multidrug resistance by inhibiting ABC transporter expression and suppressing the PI3K/Akt pathway in drug-resistant breast cancer cells

Multidrug resistance remains a major challenge in the chemotherapy of breast cancer. Guajadial, a natural meroterpenoid, has been found to possess anti-tumor activity. However, the role of guajadial in drug resistance has not been investigated. The aim of this study was to evaluate the effect of guajadial on drug resistance in drug-resistant breast cancer cells. Cell viability was measured by MTT assay, and the IC50 values were calculated. The expression of ATP-binding cassette (ABC) transporters including P-glycoprotein (P-gp) and the breast cancer resistance protein (BCRP) was detected by qRT-PCR and western blot. The expression of Akt, p-Akt, p70S6K, and p-p70S6K was determined by western blot. We found that guajadial significantly inhibited cell viability of parental non-resistant cell line MCF-7, adriamycin (ADR)-resistant cell line MCF-7/ADR, and paclitaxel (PTX)-resistant cell line MCF-7/PTX in a dose-dependent manner. Guajadial enhanced ADR and PTX sensitivity of MCF-7/ADR and MCF-7/PTX cells, and inhibited the expression of P-gp and BCRP. Guajadial treatment resulted in an inactivation of PI3K/Akt pathway in drug-resistant MCF-7 cells. In conclusion, guajadial acted as an inhibitor of drug resistance, which might be mediated by the inhibition of ABC transporter expression and PI3K/Akt pathway in drug-resistant breast cancer cells. These findings suggested that guajadial treatment might be a new approach to overcome the drug resistance in the chemotherapy of breast cancer.

Physiological and molecular insights of bacteriocin production by Enterococcus hirae ST57ACC from Brazilian artisanal cheese.

The bacteriocinogenic Enterococcus hirae ST57ACC recently isolated from a Brazilian artisanal cheese was subjected here to additional analyses in order to evaluate its bacteriocin production and the potential influence of ABC transporter system in its expression. Besides these physiological and molecular aspects, the bacteriocin was evaluated for its cytotoxicity against HT-29. Differences in the inoculum size had no impact on the growth of E. hirae ST57ACC; however, the bacteriocin was only produced after 9h of growth when the strain was inoculated at 5% or 10% (v/v), with similar levels of bacteriocin production obtained by both conventional growth and batch fermentation. Furthermore, potential expression of ABC transporters corresponding to the bacteriocin transport and sugar metabolism was identified. In terms of adverse effects, when a semi-purified fraction of the bacteriocin and the cell-free supernatant were tested against HT-29, total cell viability was similar to observed on untreated cells, indicating the absence of cytotoxic effect. Based on the obtained results, E. hirae ST57ACC can produce its bacteriocin at industrial level by using bioreactors, its bacteriocin expression is potentially influenced by the ABC transporter system, and no cytotoxic effects were observed on HT-29 cells, indicating its potential use as a bio-preservative.

ATP-Binding Cassette transporters' gene expression in pediatric patients with acute leukemia; a comprehensive analysis of published reports through PubMed search engine

Multidrug resistance based on ABC transporters' gene expression is one of the most important health challenges through chemotherapy of patients. This resistance can cause relapse or treatment failure. The goal of this conducted study was to evaluate the results of published reports which considered ABC transporters' gene expression in pediatric patients with acute leukemia. PubMed as a free search engine was chosen. The following Mesh terms were used as: "ATP-binding cassette transporters" OR "ABC-transporters*" AND "gene expression*" AND "leukemia" OR "ALL" OR "AML" OR "acute leukemia*". Age was set as an additional filter with the age range of birth to 18 years old. Initial screening was performed according to inclusion and exclusion criteria and the quality of the selected papers was assessed. Papers categorized into three sections as: pediatric patients with ALL (6 papers from 1998-2015); pediatric patients with AML (3 papers from 1992-2011) and pediatric patients with ALL and AML (7 papers from 1992-2014). Totally 1118 patients enrolled in the searched studies (ALL and AML: 488; ALL: 405; AML: 225). The common method for evaluating gene expression of ABC transporters was RT-PCR. More than 50% of the papers showed the influence of ABC transporters' gene expression on prognosis and treatment failures of patients. Despite controversial results, the gathered information in the current report serves as a comprehensive referential resource, which can be beneficial for future planning around this title, especially in developing countries.