[Uric Acid Metabolism, Uric Acid Transporters and Dysuricemia].

Serum urate levels are determined by the balance between uric acid production and uric acid excretion capacity from the kidneys and intestinal tract. Dysuricemia, including hyperuricemia and hypouricemia, develops when the balance shifts towards an increase or a decrease in the uric acid pool. Hyperuricemia is mostly a multifactorial genetic disorder involving several disease susceptibility genes and environmental factors. Hypouricemia, on the other hand, is caused by genetic abnormalities. The main genes involved in dysuricemia are xanthine oxidoreductase, an enzyme that produces uric acid, and the urate transporters urate transporter 1/solute carrier family 22 member 12 (URAT1/SLC22A12), glucose transporter 9/solute carrier family 2 member 9 (GLUT9/SLC2A9) and ATP binding cassette subfamily G member 2 (ABCG2). Deficiency of xanthine oxidoreductase results in xanthinuria, a rare disease with marked hypouricemia. Xanthinuria can be due to a single deficiency of xanthine oxidoreductase or in combination with aldehyde oxidase deficiency as well. The latter is caused by a deficiency in molybdenum cofactor sulfurase, which is responsible for adding sulphur atoms to the molybdenum cofactor required for xanthine oxidoreductase and aldehyde oxidase to exert their action. URAT1/SLC22A12 and GLUT9/SLC2A9 are involved in urate reabsorption and their deficiency leads to renal hypouricemia, a condition that is common in Japanese due to URAT1/SLC22A12 deficiency. On the other hand, ABCG2 is involved in the secretion of urate, and many Japanese have single nucleotide polymorphisms that result in its reduced function, leading to hyperuricemia. In particular, severe dysfunction of ABCG2 leads to hyperuricemia with reduced extrarenal excretion.

Purpurogallin carboxylic acid exhibits synergistic effects with 5­fluorouracil on liver cancer cells in vitro by targeting ABCG2.

Purpurogallin carboxylic acid (PCA) is a natural phenol compound derived from Macleaya microcarpa (Maxim.) Fedde, which exerts particular antioxidant and anti-inflammatory capacities. However, the effects and mechanisms of PCA on liver cancer cells remain unknown. Therefore, network pharmacology and computer virtual docking were used to identify the target-proteins of PCA. In addition, surface plasmon resonance, protease activity and rhodamine excretion assays were carried out to evaluate the effects of PCA on the activity of ATP binding cassette subfamily G member 2 (ABCG2). The synergistic effects of PCA and 5-fluorouracil (5-FU) on liver cancer cell proliferation, cell cycle arrest, colony formation and spheroid formation abilities in vitro were determined by Cell Counting Kit-8 (CCK-8) assay, flow cytometry, western blot analysis, colony formation and spheroid formation assays, respectively. ABCG2 was identified as a potential target of PCA, with a high docking score. The equilibrium dissociation constant of PCA for ABCG2 protein was 1.84 µM, while the median inhibitory concentration of this protein was 3.09 µM. In addition, the results demonstrated that PCA could significantly reduce the drug efflux capacity of liver cancer cells. CCK-8 assays revealed that liver cancer cell treatment with 10 µM PCA and 10 µM 5-FU exhibited the most potent synergistic effects on liver cancer cell proliferation at 48 h. Additionally, cell co-treatment with PCA and 5-FU also significantly attenuated the colony and spheroid formation abilities of liver cancer cells in vitro, while it promoted their arrest at the G1 phase of the cell cycle. Furthermore, ABCG2 silencing in liver cancer cells notably abrogated the synergistic effects of PCA and 5-FU. In conclusion, the present study demonstrated that PCA exhibited synergistic effects with 5-FU on liver cancer cells in vitro via targeting ABCG2. Therefore, PCA combined with 5-FU may be a potential strategy for liver cancer therapy.

Effectiveness of lapatinib for enhancing 5-aminolevulinic acid-mediated protoporphyrin IX fluorescence and photodynamic therapy in human cancer cell lines with varied ABCG2 activities.

5-Aminolevulinic acid (ALA) is a prodrug for protoporphyrin IX (PpIX)-mediated photodynamic therapy (PDT) and fluorescence-guided tumor surgery. We previously reported that lapatinib, a repurposed ABCG2 inhibitor, enhanced ALA-induced PpIX fluorescence and PDT by blocking ABCG2-mediated PpIX efflux. In the present study, we evaluated how the variation in ABCG2 activities/protein levels affected tumor cell response to the enhancement of PpIX/PDT by lapatinib and Ko143, an ABCG2 tool inhibitor. ABCG2 activities and protein levels were determined in a panel of human cancer cell lines. Effects of lapatinib and Ko143 on enhancing ALA-PpIX fluorescence and PDT were evaluated and correlated with tumor cell ABCG2 activities. We found that both lapatinib and Ko143 enhanced ALA-PpIX fluorescence and PDT in a dose-dependent manner, although lapatinib exhibited lower efficacy and potency than Ko143 in nearly all cancer cell lines. The EC50 of ABCG2 inhibitors for enhancing ALA-PpIX and PDT had a positive correlation with tumor cell ABCG2 activities, indicating that tumor cell lines with lower ABCG2 activities were more sensitive to ABCG2 inhibitors for PpIX/PDT enhancement. Our results suggest that, for optimal therapeutic enhancement, the dose of ABCG2 inhibitors needs to be tailored based on the ABCG2 expression/activity in tumors.

Exposure to anticancer drugs modulates the expression of ACSL4 and ABCG2 proteins in adrenocortical carcinoma cells.

Adrenocortical carcinoma (ACC) is a rare and malignant disease, with more than 50 % of patients developing hormone-secreting tumors. These tumors are genetically heterogeneous and potentially lethal, as metastasis is often underway at the time of diagnosis. While chemoresistance can be multifactorial, Acyl CoA synthetase 4 (ACSL4) is known to contribute to the generation of highly aggressive cellular phenotypes, while increased expression and activity of multidrug transporters such as ATP-binding cassette subfamily G member 2 (ABCG2) are known to play a key role. Therefore, the objective of this work was to determine changes in the expression of ACSL4 and ABCG2 in ACC cell lines after exposure to antitumor drugs. Bioinformatics analysis of public database GSE140818 revealed higher ACSL4 and ABCG2 expression in HAC15 cells resistant to mitotane when compared to wild type cells. In addition, our studies revealed an increase in ACSL4 and ABCG2 expression in lowly aggressive H295R cells undergoing early treatment with non-lethal concentrations of mitotane, doxorubicin and cisplatin. Comparable results were obtained in lowly aggressive breast cancer cells MCF-7. The increase in ACSL4 and ABCG2 expression favored tumor cell viability, proliferation and compound efflux, an effect partially offset by ACSL4 and ABCG2 inhibitors. These results provide relevant data on the undesired molecular effects of antitumor drugs and may fuel future studies on patients' early response to antitumor treatment.

Targeting ABCG2 transporter to enhance 5-aminolevulinic acid for tumor visualization and photodynamic therapy.

5-Aminolevulinic acid (ALA) has been approved by the U. S. FDA for fluorescence-guided resection of high-grade glioma and photodynamic therapy (PDT) of superficial skin precancerous and cancerous lesions. As a prodrug, ALA administered orally or topically is metabolized in the heme biosynthesis pathway to produce protoporphyrin IX (PpIX), the active drug with red fluorescence and photosensitizing property. Preferential accumulation of PpIX in tumors after ALA administration enables the use of ALA for PpIX-mediated tumor fluorescence diagnosis and PDT, functioning as a photo-theranostic agent. Extensive research is currently underway to further enhance ALA-mediated PpIX tumor disposition for better tumor visualization and treatment. Particularly, the discovery of PpIX as a specific substrate of ATP binding cassette subfamily G member 2 (ABCG2) opens the door to therapeutic enhancement with ABCG2 inhibitors. Studies with human tumor cell lines and human tumor samples have demonstrated ABCG2 as an important biological determinant of reduced ALA-PpIX tumor accumulation, inhibition of which greatly enhances ALA-PpIX fluorescence and PDT response. These studies strongly support targeting ABCG2 as an effective therapeutic enhancement approach. In this review, we would like to summarize current research of ABCG2 as a drug efflux transporter in multidrug resistance, highlight previous works on targeting ABCG2 for therapeutic enhancement of ALA, and provide future perspectives on how to translate this ABCG2-targeted therapeutic enhancement strategy from bench to bedside.

5-Aminolevulinic Acid as a Theranostic Agent for Tumor Fluorescence Imaging and Photodynamic Therapy.

5-Aminolevulinic acid (ALA) is a naturally occurring amino acid synthesized in all nucleated mammalian cells. As a porphyrin precursor, ALA is metabolized in the heme biosynthetic pathway to produce protoporphyrin IX (PpIX), a fluorophore and photosensitizing agent. ALA administered exogenously bypasses the rate-limit step in the pathway, resulting in PpIX accumulation in tumor tissues. Such tumor-selective PpIX disposition following ALA administration has been exploited for tumor fluorescence diagnosis and photodynamic therapy (PDT) with much success. Five ALA-based drugs have now received worldwide approval and are being used for managing very common human (pre)cancerous diseases such as actinic keratosis and basal cell carcinoma or guiding the surgery of bladder cancer and high-grade gliomas, making it the most successful drug discovery and development endeavor in PDT and photodiagnosis. The potential of ALA-induced PpIX as a fluorescent theranostic agent is, however, yet to be fully fulfilled. In this review, we would like to describe the heme biosynthesis pathway in which PpIX is produced from ALA and its derivatives, summarize current clinical applications of ALA-based drugs, and discuss strategies for enhancing ALA-induced PpIX fluorescence and PDT response. Our goal is two-fold: to highlight the successes of ALA-based drugs in clinical practice, and to stimulate the multidisciplinary collaboration that has brought the current success and will continue to usher in more landmark advances.

NRBP1 modulates uric acid transporter ABCG2 expression by activating the Wnt/B-catenin pathway in HK-2 cells / NRBP1 regula la expresión del transportador de ácido úrico ABCG2 en las células HK-2 mediante la activación de la vía Wnt/B-catenina

Background: Nuclear receptor binding protein 1 (NRBP1) and ATP-binding cassette subfamily G member 2 (ABCG2) was the gout risk gene and high-capacity urate exporter respectively. However, the relationship between NRBP1 and ABCG2 and the underlying molecular mechanism contributing to these associations are unknown. Methods: Firstly, the efficiency of the overexpression and knockdown of NRBP1 was confirmed by western blot. Next, the effect of NRBP1 overexpression and knockdown on the expression of ABCG2, organic anion transporter 1 (OAT1), glucose transporter 9 (GLUT9) and urate transporter 1 (URAT1) was detected by qRT-PCR and western blot. At the same time, the cellular location of ABCG2 and its expression after NRBP1 overexpression and knockdown was tested by immunofluorescence (IF) staining. Then, the mechanism of NRBP1 modulates ABCG2 expression was evaluated by western blot with or without the β-catenin inhibitor (21H7). Results: The lentivirus system was used to generate stable NRBP1 overexpression, while the plasmids carrying a NRBP1 siRNA was generated to knockdown NRBP1 expression in HK-2 cells. Meanwhile, the overexpression of NRBP1 significantly decreased the mRNAs and proteins expression of GLUT9 and URAT1, while the knockdown of NRBP1 increased the mRNAs and proteins expression of ABCG2 significantly. In addition, the NRBP1 modulates the expression of ABCG2 was by ctivating the Wnt/β-catenin pathway in HK-2 cells according to the IF and western blot results. Conclusion: Taken together, our study demonstrated that NRBP1 inhibition played an essential role in attenuating hyperuricemia and gout by upregulation of ABCG2 via Wnt/β-catenin signaling pathway in HK-2 cells. (AU)

Antecedentes: La proteína de unión al receptor nuclear 1 (NRBP1) y el miembro G de la subclase ATP binding Box 2 (ABCG2) son los genes de riesgo de gota y los genes de salida de urato de alto rendimiento, respectivamente. Sin embargo, se desconoce la relación entre NRBP1 y ABCG2, y los posibles mecanismos moleculares que conducen a estas asociaciones. Métodos: En primer lugar, la sobreexpresión y el knockout de NRBP1 fueron confirmados por Western-blot. Los efectos de la sobreexpresión y knockout de NRBP1 en la expresión de ABCG2, transportador de aniones orgánicos 1 (OAT1), transportador de glucosa 9 (GLUT9) y transportador de ácido úrico 1 (URAT1) fueron detectados por qRT-PCR y Western-blot. Mientras tanto, la localización y expresión de ABCG2 después de la sobreexpresión y knockout de NRBP1 fueron detectadas por inmunofluorescencia (IF). Luego, el efecto regulador de NRBP1 sobre la expresión de ABCG2 fue estudiado por Western-blot y comparado con el inhibidor de la β-catenina (21H7). Resultados: El sistema lentiviral indujo una sobreexpresión estable de NRBP1, mientras que el plásmido portador de SiRNA NRBP1 inhibió la expresión de NRBP1 en las células HK-2. Mientras tanto, la sobreexpresión de NRBP1 redujo significativamente la expresión de ARNm y proteínas de GLUT9 y URAT1, mientras que el knockout de NRBP1 aumentó significativamente la expresión de ARNm y proteínas de ABCG2. Además, de acuerdo con los resultados de IF y Western-blot, NRBP1 regula la expresión de ABCG2 activando la vía Wnt/β-catenina en las células HK-2. Conclusión: La inhibición del NRBP1 aumenta la regulación de ABCG2 a través de la vía de señalización Wnt/β-catenina, que desempeña un papel importante en la reducción de la hiperuricemia y la gota. (AU)

NRBP1 modulates uric acid transporter ABCG2 expression by activating the Wnt/ß-catenin pathway in HK-2 cells.

BACKGROUND: Nuclear receptor binding protein 1 (NRBP1) and ATP-binding cassette subfamily G member 2 (ABCG2) was the gout risk gene and high-capacity urate exporter respectively. However, the relationship between NRBP1 and ABCG2 and the underlying molecular mechanism contributing to these associations are unknown. METHODS: Firstly, the efficiency of the overexpression and knockdown of NRBP1 was confirmed by western blot. Next, the effect of NRBP1 overexpression and knockdown on the expression of ABCG2, organic anion transporter 1 (OAT1), glucose transporter 9 (GLUT9) and urate transporter 1 (URAT1) was detected by qRT-PCR and western blot. At the same time, the cellular location of ABCG2 and its expression after NRBP1 overexpression and knockdown was tested by immunofluorescence (IF) staining. Then, the mechanism of NRBP1 modulates ABCG2 expression was evaluated by western blot with or without the ß-catenin inhibitor (21H7). RESULTS: The lentivirus system was used to generate stable NRBP1 overexpression, while the plasmids carrying a NRBP1 siRNA was generated to knockdown NRBP1 expression in HK-2 cells. Meanwhile, the overexpression of NRBP1 significantly decreased the mRNAs and proteins expression of GLUT9 and URAT1, while the knockdown of NRBP1 increased the mRNAs and proteins expression of ABCG2 significantly. In addition, the NRBP1 modulates the expression of ABCG2 was by ctivating the Wnt/ß-catenin pathway in HK-2 cells according to the IF and western blot results. CONCLUSION: Taken together, our study demonstrated that NRBP1 inhibition played an essential role in attenuating hyperuricemia and gout by upregulation of ABCG2 via Wnt/ß-catenin signaling pathway in HK-2 cells.

Expression of BCRP/ABCG2 Protein in Invasive Breast Cancer and Response to Neoadjuvant Chemotherapy.

BACKGROUND: Breast cancer resistance protein (BCRP), or ATP-binding cassette subfamily G member 2 (ABCG2), is an ATP-binding cassette (ABC) transporter that mediates energy-dependent transport of substrate drugs out of the cell. Its overexpression may contribute to intrinsic drug resistance in vitro. However, the current literature has not yet clarified the clinical significance of BCRP/ABCG2 in invasive breast carcinoma. OBJECTIVES: The purpose of this study was to validate the expression of BCRP/ABCG2 in invasive breast carcinoma and its role in response to neoadjuvant chemotherapy. METHODS: In this study, a pretherapeutic core biopsy was performed in 222 patients. BCRP/ABCG2 expression in carcinoma tissue was measured by immunohistochemistry. BCRP/ABCG2 expression correlations with clinicopathological features, molecular subtypes, and therapy response after neoadjuvant chemotherapy were investigated. RESULTS: The results showed that BCRP/ABCG2 was expressed in different molecular subtypes. The proportions of patients with high BCRP/ABCG2 expression were similar in luminal A and luminal B tumors (luminal B, 80%; luminal A, 78%), compared with other molecular subtypes (triple-negative, 63%; HER2+, 58%, p = 0.05). BCRP/ABCG2 expression and the number of lymphatic metastases (p = 0.001) and tumor size (p = 0.011) demonstrated a statistically significant correlation. Low BCRP/ABCG2 expression was associated with an increased pathological complete response (pCR) rate of 38%, higher than the 19% in tumors with high BCRP/ABCG2 expression (p = 0.002). In multivariable analysis, BCRP/ABCG2 and hormone receptor (HR) expression were identified as independent risk factors of pCR (p = 0.003 and p = 0.013, respectively). CONCLUSIONS: BCRP/ABCG2 is highly expressed in HR-positive breast cancer. High BCRP/ABCG2 expression is associated with lymphatic metastasis, tumor size, and poor pCR. BCRP/ABCG2 may be a novel potential biomarker that can predict clinical progression and therapy response after neoadjuvant chemotherapy.

Polyoxypregnanes as safe, potent, and specific ABCB1-inhibitory pro-drugs to overcome multidrug resistance in cancer chemotherapy in vitro and in vivo.

Multidrug resistance (MDR) mediated by ATP binding cassette subfamily B member 1 (ABCB1) is significantly hindering effective cancer chemotherapy. However, currently, no ABCB1-inhibitory drugs have been approved to treat MDR cancer clinically, mainly due to the inhibitor specificity, toxicity, and drug interactions. Here, we reported that three polyoxypregnanes (POPs) as the most abundant constituents of Marsdenia tenacissima (M. tenacissima) were novel ABCB1-modulatory pro-drugs, which underwent intestinal microbiota-mediated biotransformation in vivo to generate active metabolites. The metabolites at non-toxic concentrations restored chemosensitivity in ABCB1-overexpressing cancer cells via inhibiting ABCB1 efflux activity without changing ABCB1 protein expression, which were further identified as specific non-competitive inhibitors of ABCB1 showing multiple binding sites within ABCB1 drug cavity. These POPs did not exhibit ABCB1/drug metabolizing enzymes interplay, and their repeated administration generated predictable pharmacokinetic interaction with paclitaxel without obvious toxicity in vivo. We further showed that these POPs enhanced the accumulation of paclitaxel in tumors and overcame ABCB1-mediated chemoresistance. The results suggested that these POPs had the potential to be developed as safe, potent, and specific pro-drugs to reverse ABCB1-mediated MDR. Our work also provided scientific evidence for the use of M. tenacissima in combinational chemotherapy.

A Novel 89Zr-labeled DDS Device Utilizing Human IgG Variant (scFv): "Lactosome" Nanoparticle-Based Theranostics for PET Imaging and Targeted Therapy.

"Theranostics," a new concept of medical advances featuring a fusion of therapeutic and diagnostic systems, provides promising prospects in personalized medicine, especially cancer. The theranostics system comprises a novel 89Zr-labeled drug delivery system (DDS), derived from the novel biodegradable polymeric micelle, "Lactosome" nanoparticles conjugated with specific shortened IgG variant, and aims to successfully deliver therapeutically effective molecules, such as the apoptosis-inducing small interfering RNA (siRNA) intracellularly while offering simultaneous tumor visualization via PET imaging. A 27 kDa-human single chain variable fragment (scFv) of IgG to establish clinically applicable PET imaging and theranostics in cancer medicine was fabricated to target mesothelin (MSLN), a 40 kDa-differentiation-related cell surface glycoprotein antigen, which is frequently and highly expressed by malignant tumors. This system coupled with the cell penetrating peptide (CPP)-modified and photosensitizer (e.g., 5, 10, 15, 20-tetrakis (4-aminophenyl) porphyrin (TPP))-loaded Lactosome particles for photochemical internalized (PCI) driven intracellular siRNA delivery and the combination of 5-aminolevulinic acid (ALA) photodynamic therapy (PDT) offers a promising nano-theranostic-based cancer therapy via its targeted apoptosis-inducing feature. This review focuses on the combined advances in nanotechnology and material sciences utilizing the "89Zr-labeled CPP and TPP-loaded Lactosome particles" and future directions based on important milestones and recent developments in this platform.

Polyoxypregnanes as safe, potent, and specific ABCB1-inhibitory pro-drugs to overcome multidrug resistance in cancer chemotherapy

Multidrug resistance (MDR) mediated by ATP binding cassette subfamily B member 1 (ABCB1) is significantly hindering effective cancer chemotherapy. However, currently, no ABCB1-inhibitory drugs have been approved to treat MDR cancer clinically, mainly due to the inhibitor specificity, toxicity, and drug interactions. Here, we reported that three polyoxypregnanes (POPs) as the most abundant constituents of

The ABCG2 mRNA Synthesized in vitro Promoted Uric Acid Excretion in Mice / 中国生物化学与分子生物学报

Hyperuricemia is a chronic metabolic disease caused by the accumulation of uric acid in the body caused by purine metabolism disorder. In recent years, the incidence of hyperuricemia has increased and the age of onset is showing a younger trend. Finding effective therapeutic targets and treatment methods is a hot spot of current research. The urate transporter ATP-binding cassette subfamily G member 2 (ABCG2) is mainly expressed in the kidney and promotes uric acid excretion. In this study, ABCG2 mRNA was synthesized in vitro and transfected into hyperuricemia model mice to observe its effect on mouse uric acid levels. Firstly, the DNA template of ABCG2 mRNA was chemically synthesized, and then transcribed into mRNA in vitro, followed by modification and transfection into mouse TCMK-1 renal tubular epithelial cells. Finally, the protein expression in the cells was detected by Western blot. The results showed that the amount of protein expression in TCMK-1 cells was positively correlated with the amount of transfected mRNA (P < 0. 01), indicating a successful transfection. In animal experiments, twenty-four SPF mice were randomly divided into four groups (n = 6): control group, hyperuricemia model group, benzbromarone group [20 mg/(kg•d)] and mRNA group [2 mg/(kg•3d)]. The mice have been modeled and treated for 28 days. During this period, the body weight and growth status of the mice were monitored daily. After the treatment, the levels of serum uric acid, urine uric acid, serum creatinine, blood urea nitrogen and liver xanthine oxidase were analyzed. The results showed that compared with the model group of mice, mRNA treatment can significantly reduce the levels of serum uric acid (100. 38 ± 10. 94), blood urea nitrogen (6. 30 ± 1. 10), and serum creatinine (30. 86 ± 5. 78, P<0. 05 or P<0. 01). It can also increase the level of urine uric acid (617. 48 ± 50. 34, P<0. 05) in mice and promote the excretion of uric acid. But it has no significant effect on the activity of xanthine oxidase (26. 19 ± 2. 58) in the liver. The pathological changes of mice kidney were observed by HE staining. The results showed that compared with mice in the model group, pathological damages such as renal tubular cell edema and inflammatory cell infiltration in the mRNA treatment group were significantly improved. The relative expression of mRNA in mice kidney was detected by qRT-PCR, and the protein expression of ABCG2 in mice kidney was detected by immunohistochemistry and Western blot. The results showed that the relative expression of ABCG2 mRNA and its protein were significantly up-regulated in the kidney tissues of mice in the mRNA group (P < 0. 01), indicating that the transfection was successful in vivo. In conclusion, ABCG2 mRNA synthetized and modified in vitro can be successfully expressed in hyperuricemia mice and promote excretion of uric acid and other organic ions, as well as improvement of renal injury in mice. These results provide experimental basis for the clinical application of ABCG2 as a target for the treatment of hyperuricemia related diseases.

KD025, an anti-adipocyte differentiation drug, enhances the efficacy of conventional chemotherapeutic drugs in ABCG2-overexpressing leukemia cells.

Most patients with advanced leukemia eventually die from multidrug resistance (MDR). Chemotherapy-resistant leukemia cells may lead to treatment failure and disease relapse. Overexpression of ATP-binding cassette subfamily G member 2 (ABCG2) leads to MDR, which serves as a potential biomarker and target of therapeutic intervention for leukemia cells. Targeting ABCG2 is a potential strategy for selective therapy and eradicate MDR cells, thus improving malignant leukemia treatment. KD025 (SLx-2119) is a novel Rho-associated protein kinase 2-selective inhibitor, which has been shown to inhibit adipogenesis in human adipose-derived stem cells and restore impaired immune homeostasis in autoimmunity therapy. The present study demonstrated that KD025 improved the efficacy of antineoplastic drugs in ABCG2-overexpressing leukemia cells and primary leukemia blast cells derived from patients with leukemia. Moreover, KD025 significantly inhibited the efflux of [3H]-mitoxantrone and hence accumulated higher levels of [3H]-mitoxantrone in HL60/ABCG2 cells. However, mechanistic research indicated that KD025 did not alter the protein levels and subcellular locations of ABCG2. KD025 may restrain the efflux activity of ABCG2 by obstructing ATPase activity. Taken together, KD025 can sensitize conventional antineoplastic drugs in ABCG2-overexpressing leukemia cells by blocking the pump function of ABCG2 protein. The present findings may provide a novel and useful combinational therapeutic strategy of KD025 and antineoplastic drugs for leukemia patients with ABCG2-mediated MDR.

sATP­binding cassette subfamily G member 2 enhances the multidrug resistance properties of human nasal natural killer/T cell lymphoma side population cells.

Extranodal natural killer (NK)/T cell lymphoma, nasal type (ENKL) is a rare type of non­Hodgkin's lymphoma that is associated with limited effective treatment options and unfavorable survival rate, which is partly the result of multidrug resistance (MDR). The presence of side population (SP) cells­SNK­6/ADM­SP (SSP) cells has been previously used to explore mechanisms of drug resistance. ATP­binding cassette subfamily G member 2 (ABCG2) is a gene involved in MDR and is closely associated with SPs. However, the function of ABCG2 in SSP cells is unclear. The present study verified the high expression of ABCG2 in SSP cells. The IC50 values of doxorubicin, cytarabine, cisplatin, gemcitabine and l­asparaginase were tested to evaluate drug sensitivity in SSP cells with different levels of ABCG2 expression. ABCG2 was identified as a gene promoting in MDR. ABCG2 upregulated cell proliferation, increased clonogenicity, increased invasive ability and decreased apoptosis, in vivo and in vitro, when cells were treated with gemcitabine. To conclude, ABCG2 enhanced MDR and increased the typical biological characteristics associated with cancer cells in SP cells. With further investigation of the ABCG2 gene could have the potential to reverse MDR in ENKL.

MICA enhances sensitivity to cisplatin in patients with extensive small cell lung cancer via downregulation of ABCG2.

Immunotherapy utilizing natural killer cell-activated receptor natural-killer group-2 member D ligands (NKG2DLs) has had preclinical success in the treatment of small cell lung cancer. The present study aimed to investigate the association between NKG2Ls and chemoresistance. The mRNA expression of six NKG2DLs associated with progression-free survival time (PFS) and first-line chemotherapy were assessed in the present study. Major histocompatibility complex class I polypeptide-related sequence A (MICA)-overexpressing NCI-H446 cell line was constructed, and the mRNA expression levels of 11 genes associated with chemotherapy sensitivity were determined. The results demonstrated that MICA was positively and significantly associated with PFS. Furthermore, MICA expression was 1.6 times higher in patients with prolonged PFS compared with the rapid chemoresistance group. ATP binding cassette subfamily G member 2 (ABCG2) mRNA expression was associated with chemotherapy resistance and significantly downregulated in the cell line overexpressing MICA. Moreover, following addition of nicardipine (an ABCG2 inhibitor), chemotherapeutic sensitivity increased in the MICA-overexpressing cell line. Taken together, the results of the present study suggested that MICA may enhance the chemosensitivity of patients with extensive small cell lung cancer by downregulating ABCG2.

Bioengineered miR-328-3p modulates GLUT1-mediated glucose uptake and metabolism to exert synergistic antiproliferative effects with chemotherapeutics.

MicroRNAs (miRNAs or miRs) are small noncoding RNAs derived from genome to control target gene expression. Recently we have developed a novel platform permitting high-yield production of bioengineered miRNA agents (BERA). This study is to produce and utilize novel fully-humanized BERA/miR-328-3p molecule (hBERA/miR-328) to delineate the role of miR-328-3p in controlling nutrient uptake essential for cell metabolism. We first demonstrated successful high-level expression of hBERA/miR-328 in bacteria and purification to high degree of homogeneity (>98%). Biologic miR-328-3p prodrug was selectively processed to miR-328-3p to suppress the growth of highly-proliferative human osteosarcoma (OS) cells. Besides glucose transporter protein type 1, gene symbol solute carrier family 2 member 1 (GLUT1/SLC2A1), we identified and verified large neutral amino acid transporter 1, gene symbol solute carrier family 7 member 5 (LAT1/SLC7A5) as a direct target for miR-328-3p. While reduction of LAT1 protein levels by miR-328-3p did not alter homeostasis of amino acids within OS cells, suppression of GLUT1 led to a significantly lower glucose uptake and decline in intracellular levels of glucose and glycolytic metabolite lactate. Moreover, combination treatment with hBERA/miR-328 and cisplatin or doxorubicin exerted a strong synergism in the inhibition of OS cell proliferation. These findings support the utility of novel bioengineered RNA molecules and establish an important role of miR-328-3p in the control of nutrient transport and homeostasis behind cancer metabolism.

Bioengineered miR-328-3p modulates GLUT1-mediated glucose uptake and metabolism to exert synergistic antiproliferative effects with chemotherapeutics

MicroRNAs (miRNAs or miRs) are small noncoding RNAs derived from genome to control target gene expression. Recently we have developed a novel platform permitting high-yield production of bioengineered miRNA agents (BERA). This study is to produce and utilize novel fully-humanized BERA/miR-328-3p molecule (hBERA/miR-328) to delineate the role of miR-328-3p in controlling nutrient uptake essential for cell metabolism. We first demonstrated successful high-level expression of hBERA/miR-328 in bacteria and purification to high degree of homogeneity (>98%). Biologic miR-328-3p prodrug was selectively processed to miR-328-3p to suppress the growth of highly-proliferative human osteosarcoma (OS) cells. Besides glucose transporter protein type 1, gene symbol solute carrier family 2 member 1 (GLUT1/), we identified and verified large neutral amino acid transporter 1, gene symbol solute carrier family 7 member 5 (LAT1/) as a direct target for miR-328-3p. While reduction of LAT1 protein levels by miR-328-3p did not alter homeostasis of amino acids within OS cells, suppression of GLUT1 led to a significantly lower glucose uptake and decline in intracellular levels of glucose and glycolytic metabolite lactate. Moreover, combination treatment with hBERA/miR-328 and cisplatin or doxorubicin exerted a strong synergism in the inhibition of OS cell proliferation. These findings support the utility of novel bioengineered RNA molecules and establish an important role of miR-328-3p in the control of nutrient transport and homeostasis behind cancer metabolism.

Development of precision medicine approaches based on inter-individual variability of BCRP/ABCG2.

Precision medicine is a rapidly-developing modality of medicine in human healthcare. Based on each patient׳s unique characteristics, more accurate dosages and drug selection can be made to achieve better therapeutic efficacy and less adverse reactions in precision medicine. A patient׳s individual parameters that affect drug transporter action can be used to develop a precision medicine guidance, due to the fact that therapeutic efficacy and adverse reactions of drugs can both be affected by expression and function of drug transporters on the cell membrane surface. The purpose of this review is to summarize unique characteristics of human breast cancer resistant protein (BCRP) and the genetic variability in the BCRP encoded gene ABCG2 in the development of precision medicine. Inter-individual variability of BCRP/ABCG2 can impact choices and outcomes of drug treatment for several diseases, including cancer chemotherapy. Several factors have been implicated in expression and function of BCRP, including genetic, epigenetic, physiologic, pathologic, and environmental factors. Understanding the roles of these factors in controlling expression and function of BCRP is critical for the development of precision medicine based on BCRP-mediated drug transport.

Development of precision medicine approaches based on inter-individual variability of BCRP/

Precision medicine is a rapidly-developing modality of medicine in human healthcare. Based on each patient׳s unique characteristics, more accurate dosages and drug selection can be made to achieve better therapeutic efficacy and less adverse reactions in precision medicine. A patient׳s individual parameters that affect drug transporter action can be used to develop a precision medicine guidance, due to the fact that therapeutic efficacy and adverse reactions of drugs can both be affected by expression and function of drug transporters on the cell membrane surface. The purpose of this review is to summarize unique characteristics of human breast cancer resistant protein (BCRP) and the genetic variability in the BCRP encoded gene in the development of precision medicine. Inter-individual variability of BCRP/ can impact choices and outcomes of drug treatment for several diseases, including cancer chemotherapy. Several factors have been implicated in expression and function of BCRP, including genetic, epigenetic, physiologic, pathologic, and environmental factors. Understanding the roles of these factors in controlling expression and function of BCRP is critical for the development of precision medicine based on BCRP-mediated drug transport.