Influence of cigarette smoking on biventricular systolic function independent of respiratory function: a cross-sectional study.

BACKGROUND: Cigarette smoking harms nearly every organ, including the heart and lungs. A comprehensive assessment of both cardiac and respiratory function is necessary for evaluating the direct effects of tobacco on the heart. However, few previous studies examining the effects of cigarette smoking on cardiac function included an assessment of lung function. This cross-sectional study investigated the influence of cigarette smoking on cardiac function, independent of respiratory function. METHODS: We retrospectively reviewed the medical records of 184 consecutive cases that underwent both spirometry and transthoracic echocardiography around the same time (within 1 month) in one hospital from April 2019 to March 2020. Participants were classified into three groups based on lifetime smoking exposure (pack-years): non-smoker (n = 49), low exposure (1-20 pack-years, n = 40), and high exposure (≥ 20 pack years, n = 95). Multiple linear regression analysis was used to assess the relationship among cigarette smoking, and cardiac and respiratory functions. The relationship between selected dependent variables and lifetime pack-years was assessed in two models with multiple linear regression analysis. Model 1 was adjusted for age and male sex; and Model 2 was adjusted for Model 1 plus forced expiratory volume percentage in 1 s and forced vital capacity percentage. RESULTS: Compared with the non-smokers, the participants with high smoking exposure had lower left ventricular (LV) systolic function and larger LV size. Multiple linear regression analysis revealed a negative association of cumulative lifetime pack-years with LV and right ventricular (RV) systolic functions, even after adjustment for age, sex, and spirometric parameters (forced expiratory volume percentage in 1 s and forced vital capacity percentage). Meanwhile, there was no significant association of smoking exposure with LV diastolic function (E/e' and E/A) and RV diastolic function (e't and e't/a't). CONCLUSIONS: Cumulative smoking exposure was associated with a negative effect on biventricular systolic function in patients with relatively preserved cardiac function, independent of respiratory function.

Cardiac events in Patients in their forties with Kawasaki disease and regression of coronary artery aneurysms.

Over a 50-year period from the first description of Kawasaki disease, we encountered three male patients with a history of Kawasaki disease, who had their first cardiac events in their forties. They were considered to have almost normal coronary arteries in the coronary angiograms when they were children and adolescents. They had no follow-up examinations after 20 years old. The 1st patient had an acute myocardial infarction, and the 2nd was a new appearance of coronary aneurysm and stenotic lesions with coronary artery calcification. The 3rd patient had unexpected sudden death. The interval from the onset of Kawasaki disease to the cardiac events ranged from 37 to 38 years. In the former two patients, coronary artery lesions could not be evaluated immediately after Kawasaki disease. Although the 3rd patient had bilateral medium-sized coronary artery aneurysms, his coronary aneurysms regressed 1 year after acute Kawasaki disease. The intimal thickening at a previous coronary aneurysm at the age of 19 was mild. The patients with regressed coronary aneurysms were asymptomatic for about 40 years after Kawasaki disease, prior to their cardiac events. Coronary artery calcification of the proximal portion of the major coronary arteries was a predictable marker in such patients. To prevent serious cardiac events in middle-aged adult patients, reevaluation of coronary artery lesions and restarting of anti-thrombotic therapy are needed. We must be aware that there are some differences in the clinical course and time of cardiac events between patients with giant aneurysms and those with medium aneurysms.

A new class of pentapeptide KISS1 receptor agonists with hypothalamic-pituitary-gonadal axis activation.

The kisspeptin (Kp, Kp-54, metastin)/KISS1R system plays crucial roles in regulating the secretion of gonadotropin-releasing hormone. Continuous administration of nonapeptide Kp analogs caused plasma testosterone depletion, whereas bolus administration caused strong plasma testosterone elevation in male rats. To develop a new class of small peptide drugs, we focused on stepwise N-terminal truncation of Kp analogs and discovered potent pentapeptide analogs. Benzoyl-Phe-azaGly-Leu-Arg(Me)-Trp-NH2 (16) exhibited high agonist activity for KISS1R and excellent metabolic stability in rat serum. A single injection of a 4-pyridyl analog (19) at the N-terminus of 16 into male Sprague Dawley rats caused a robust increase in plasma luteinizing hormone levels, but unlike continuous administration of nonapeptide Kp analogs, continuous administration of 19 maintained moderate testosterone levels in rats. These results indicated that small peptide drugs can be successfully developed for treating sex hormone deficiency.

Prolyl isomerase Pin1 is required sperm production by promoting mitosis progression of spermatogonial stem cells.

A prolyl isomerase Pin1 deficient (Pin1-/-) male mice had severe testicular atrophy. We investigated the function of Pin1 in spermatogenesis by analyzing the Pin1-/- mice at reproductive age. Pin1-/- mice had lessαPLZF positive spermatogonia (undifferentiated spermatogonia) than wild type (WT). Nevertheless, the Pin1-/- testis contained approximately the same number of GFRα1 positive spermatogonia (SSCs in steady state) as the WT testis. Furthermore, degeneration of the spermatogenia appeared in seminiferous tubules of 10 months old Pin1-/- mouse testis, and abnormal shape GFRα1 positive spermatogonia were observed. In Pin1-/- spermatogonia, the ratio of the phospho-histone H3 positive cells (mitotic cells) in GFRα1-positive spermatogonia was higher than that of WT. These results suggest that Pin1 promotes the progression of the mitotic cell cycle of SSC in steady-state, which is required for the sperm production from SSCs.

Distribution of Kawasaki Disease Coronary Artery Aneurysms and the Relationship to Coronary Artery Diameter.

We investigated how the diameter of coronary artery aneurysm (CAA) relates to the distribution immediately after Kawasaki disease (KD). Two hundred and four pts (155 males and 49 females) who had undergone selective coronary angiography (CAGs) less than 100 days after the onset of KD were studied. We measured the maximum diameter of each artery segment in the initial CAGs. We analyzed the relationship between the maximum diameters and the distribution of CAA. We divided the patients into four groups based on the maximum CAA diameter in each patient (large(L) ≥8 mm, medium(M) ≥6 and <8 mm, small(S) ≥4 and <6 mm, very small(VS) <4 mm) and counted the affected segments. There were 87, 61, 36, and 20 patients in groups L, M, S, VS, respectively. The number of segments with CAA in each group was L 6 ± 2, M 4 ± 2, S 2 ± 2, VS 2 ± 1. The number of affected segments in L was significantly more than M, and a large value for L indicated that involvement was significantly more likely to be bilateral. The larger the maximum diameter of CAA, the more extensive disease involvement and the more likely to be bilateral. A large maximum CAA can also indicate coronary involvement in the longitudinal directions. It is an important charcteristic in distribution of CAA caused by KD vasculitis.

Deguelin Potentiates Apoptotic Activity of an EGFR Tyrosine Kinase Inhibitor (AG1478) in PIK3CA-Mutated Head and Neck Squamous Cell Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is known to be intrinsically resistant to inhibitors for epidermal growth factor receptor (EGFR). Until now, clinical outcomes for HNSCC using EGFR inhibitors as single agents have yielded disappointing results. Here, we aimed to study whether combinatorial treatment using AG1478 (EGFR tyrosine kinase inhibitor) and deguelin, which is a rotenoid isolated from the African plant Mundulea sericea, could enhance the anti-tumor effects of AG1478 in HNSCC. For Ca9-22 cells with EGFR, KRAS, and PIK3CA wild types, AG1478 alone suppressed both phosphorylated levels of ERK and AKT and induced apoptosis. On the contrary, for HSC-4 cells with EGFR and KRAS wild types, and a PIK3CA mutant, AG1478 alone did not suppress the phosphorylated level of AKT nor induce apoptosis, while it suppressed ERK phosphorylation. Forced expression of constitutively active PIK3CA (G1633A mutation) significantly reduced the apoptotic effect of AG1478 on the PIK3CA wild-type Ca9-22 cells. When HSC-4 cells with the PIK3CA G1633A mutation were treated with a combination of AG1478 and deguelin, combination effects on apoptosis induction were observed through the inhibition of the AKT pathway. These results suggest that the combination of EGFR tyrosine kinase inhibitor with deguelin is a potential therapeutic approach to treat PIK3CA-mutated HNSCC.

Assessment of age-specific safety of laparoscopic surgery in elderly patients with ovarian tumors.

AIM: We assessed the age-specific safety of laparoscopic surgery in elderly patients with ovarian tumors. MATERIAL AND METHODS: We performed a retrospective analysis of 55 elderly patients treated by laparoscopic salpingo-oophorectomy under the diagnosis of an ovarian tumor between January 2009 and December 2014. We divided patients into three groups: "young-elderly" (aged 65-74), "old-elderly" (aged 75-84), and "super-elderly" (aged 85-105) and assessed clinical characteristics, surgical results and postoperative course. Statistical significance of categorical variables was examined by the Student's t-test, Mann-Whitney U test, or Fisher's exact test. Multiple regression analysis was used for multivariate analysis. RESULTS: Of a total of 55 patients who underwent laparoscopic surgery, there were 36 patients in the young-elderly group, 17 in the old-elderly group, and two in the super-elderly group. Statistical analysis was performed between the young-elderly and the old-elderly groups because of the small number in the super-elderly group. More frequent comorbidities were found in the patients in the old-elderly than in the young-elderly group (Fisher's exact test, P = 0.007). There were no significant differences in operative time, estimated blood loss and postoperative hospital stay between the young-elderly and old-elderly groups. Intraoperative complications only occurred in the young-elderly group. Postoperative complications occurred in all groups. CONCLUSIONS: Although patients in the old-elderly group had a significantly higher risk for surgery, they had equivalent surgical results to the young-elderly group for laparoscopic salpingo-oophorectomy.

Acidic extracellular pH promotes epithelial mesenchymal transition in Lewis lung carcinoma model.

BACKGROUND: Epithelial mesenchymal transition (EMT) is thought to be an essential feature of malignant tumor cells when they spread into the stroma. Despite the extracellular acidity of tumor tissues, the effect of acidic extracellular pH (pH e ) on EMT in carcinoma models, including the Lewis lung carcinoma (LLC) model, remains unclear. METHODS: High and low metastatic LLC variants were generated by repeated tail vein injection of metastatic cells. DMEM/F12 medium, which has been supplemented with 15 mM HEPES, 4 mM phosphoric acid, and 1 g/L NaHCO3 and adjusted to the desire pH with HCl or NaOH, was used for cell culture. EMT marker gene expression was determined by quantitative reverse transcription-polymerase chain reaction. Migration and invasion activities were analyzed by wound healing assay and the Boyden chamber assay through Matrigel®, respectively. RESULTS: Low metastatic variant LLCm1 cells showed a cobble-stone like morphology at pH e 7.4. At pH e 6.8, however, their morphology became fibroblastic, similar in shape to high metastatic variant LLCm4 cells. Steady state levels of matrix metalloproteinase-9 (Mmp9) mRNA were induced by acidic pH e , maximizing at pH 6.8, with the levels of Mmp9 mRNA higher in LLCm4 than in LLCm1 cells. Both variants showed decreased levels of E-cadherin and increased levels of vimentin at pH e 6.8. Acidic pH e also induced expression of mRNAs encoding the E-cadherin repressors, Zeb2, Twist1 and Twist2, as well as enhancing cell motility and in vitro invasion through Matrigel®. CONCLUSIONS: Acidic pH e can induce EMT in some types of carcinoma.

Design, synthesis, and biological activities of 1-aryl-1,4-diazepan-2-one derivatives as novel triple reuptake inhibitors.

A novel series of triple reuptake inhibitors were explored by ligand-based drug design. A cyclic structure was designed from cyclopropane derivative 5 using the core structure of reported monoamine reuptake inhibitors, leading to the formation of the 1-aryl-1,4-diazepan-2-one derivative 23j-S. Compound 23j-S was shown to act as a potent TRI with an excellent ADME-Tox profile. Oral administration of 23j-S significantly enhanced norepinephrine, dopamine, and serotonin levels in the mouse prefrontal cortex and showed significant antidepressant-like activity in tail suspension tests in mouse.

[CT, MRI].

Coronary arterial lesions (CAL) due to Kawasaki disease (KD) should be monitored by conventional coronary angiography(CAG) throughout their lives. However, recently multi detector row computed tomography(MDCT) and magnetic resonance coronary angiography (MRCA) have developed remarkably. We performed MRCA in 1,200 cases of KD (age at 4 month to 41 years: medium 8 years) since 1999. MR myocardial imaging is also performed simultaneously in patients with severe stenotic lesions. Noninvasive MRCA is useful to monitor CAL in young children. MDCT is efficient at screening of CAL and at making a diagnosis of no acute coronary syndromes in adults with chest pain. These examinations are very useful to decrease CAG to the least times as possible.

Study on cellular uptake of a hydrophobic near-infrared-absorbing diradical-platinum(II) complex solubilized by albumin using hyperspectral imaging, spectrophotometry, and spectrofluorimetry.

Owing to its biopenetrability and minimal invasiveness, near-infrared (NIR) light in the region between 700-1100 nm has attracted attention in cancer diagnosis and therapy. Our group previously reported that the hydrophobic diradical-platinum(II) complex PtL2 is a promising agent for cancer photothermal therapy (L = 3,5-dibromo-1,2-diiminobenzosemiquinonate radical). Because PtL2 does not fluoresce, its intercellular uptake of PtL2 cannot be observed with a fluorescence microscope. In this study, we clarified the uptake and intracellular behavior of PtL2 solubilized by bovine serum albumin (BSA) using hyperspectral imaging enabling spectrophotometric analysis of the image. The spectral changes in the obtained images indicated that the internalization of PtL2 was followed by crystallization of the complex during the long incubation period (> 4 h). Additionally, the binding constant Kb = 5.91 × 104 M-1 could be estimated upon fluorescence quenching analysis of BSA upon binding of PtL2; Kb is two orders of magnitude smaller than that of albumin-common drugs. Considering the small Kb and low solubility of PtL2 in water, we ultimately proposed the internalization path and fate of PtL2 in the cell: release of PtL2 from BSA near cellular membranes and subsequent cellular uptake via membrane permeation followed by saturation, resulting in crystallization.

Different effects of crizotinib treatment in two non-small cell lung cancer patients with SDC4::ROS1 fusion variants.

The possibility of stratifying patients according to differences in ROS proto-oncogene 1 (ROS1) fusion partners has been discussed. This study aimed to clarify the clinicopathological differences between two SDC4::ROS1 positive NSCLC cases who had different responses to crizotinib. Cytology and pathology samples from two NSCLC cases with SDC4::ROS1 who were diagnosed and treated with crizotinib at Nihon University Itabashi Hospital were obtained. Case 1 has been well-controlled with crizotinib for over 5 years, but case 2 was worse and overall survival was 19 months. Sequencing analysis of ROS1 fusion genes was performed by reverse-transcription-PCR and Sanger's sequencing methods. In addition, thyroid transcription factor (TTF)-1, ROS-1, Ki67, and phosphorylated extracellular signal-regulated kinase (pERK)1/2 expression were investigated using immunohistochemistry. Sequencing analysis showed SDC4 exon2::ROS1 exon 32 (exon33 deleted) in case 1, and coexistence of SDC4 exon2::ROS1 exon 34 and SDC4 exon2::ROS1 exon35 in case 2. The Ki67 index was not different, but ROS1 and pERK1/2 expression levels tended to be higher in the tumor cells of case 2 than in case 1. Therapeutic response to crizotinib and patients' prognosis in ROS1 rearranged NSCLC may be related to the activation of ROS1 signaling, depending on ROS1 and pERK1/2 overexpression status, even if the ROS1 fusion partner is the same.

The biological effects and thermal degradation of NPB-22, a synthetic cannabinoid.

PURPOSE: NPB-22 (quinolin-8-yl 1-pentyl-1H-indazole-3-carboxylate), Adamantyl-THPINACA (N-(1-adamantantyl)-1-[(tetrahydro-2H-pyran-4-yl)methyl]-1H-indazole-3-carboxamide), and CUMYL-4CN-B7AICA (1-(4-cyanobutyl)-N-(2-phenylpropan-2-yl)-1H- pyrrolo[2,3-b]pyridine-3-carboxamide), synthetic cannabinoids were evaluated in terms of CB1 (cannabinoid receptor type 1) and CB2 (cannabinoid receptor type 2) activities, and their biological effects when inhaled similar to cigarettes were examined. METHODS: The half maximal effective concentration values of the aforementioned synthetic cannabinoids at the CB1 and CB2 were investigated using [35S]guanosine-5'-O-(3-thio)-triphosphate binding assays. In addition, their biological effects were evaluated using the inhalation exposure test with mice. The smoke generated was recovered by organic solvents in the midget impingers, and the thermal degradation compounds of the smoke components were identified and quantified using a liquid chromatography-photo diode array detector. RESULTS: NPB-22 and Adamantyl-THPINACA had equivalent CB1 activity in in vitro assays. Meanwhile, NPB-22 had a weaker biological effect on some items on the inhalation exposure test than Adamantyl-THPINACA. When analyzing organic solvents in the midget impingers, it was revealed that NPB-22 was degraded to 8-quinolinol and pentyl indazole 3-carboxylic acid by combustion. In addition, these degradation compounds did not have CB1 activity. CONCLUSION: It was estimated that the biological effects of NPB-22 on the inhalation exposure test weakened because it underwent thermal degradation by combustion, and the resultant degradation compounds did not have any CB1 activity in vitro.

Acidic extracellular microenvironment and cancer.

Acidic extracellular pH is a major feature of tumor tissue, extracellular acidification being primarily considered to be due to lactate secretion from anaerobic glycolysis. Clinicopathological evidence shows that transporters and pumps contribute to H+ secretion, such as the Na+/H+ exchanger, the H+-lactate co-transporter, monocarboxylate transporters, and the proton pump (H+-ATPase); these may also be associated with tumor metastasis. An acidic extracellular pH not only activates secreted lysosomal enzymes that have an optimal pH in the acidic range, but induces the expression of certain genes of pro-metastatic factors through an intracellular signaling cascade that is different from hypoxia. In addition to lactate, CO2 from the pentose phosphate pathway is an alternative source of acidity, showing that hypoxia and extracellular acidity are, while being independent from each other, deeply associated with the cellular microenvironment. In this article, the importance of an acidic extracellular pH as a microenvironmental factor participating in tumor progression is reviewed.

Novel triple reuptake inhibitors with low risk of CAD associated liabilities: design, synthesis and biological activities of 4-[(1S)-1-(3,4-dichlorophenyl)-2-methoxyethyl]piperidine and related compounds.

A novel triple reuptake inhibitor with low potential of liabilities associated with cationic amphiphilic drug (CAD) was identified following an analysis of existing drugs. Low molecular weight (MW < ca. 300), low aromatic ring count (number = 1) and reduced lipophilicity (ClogP < 3.5) were hypothesized to be key factors to avoid the CAD associated liabilities (CYP2D6 inhibition, hERG inhibition and phospholipidosis). Based on the hypothesis, a series of piperidine compounds was designed with consideration of the common characteristic features of CNS drugs. Optimization of the side chain by adjusting overall lipophilicity suggested that incorporation of a methoxymethyl group could provide compounds with a balance of both potent reuptake inhibition and low liability potential. Compound (S)-3a showed a potent antidepressant-like effect in the mice tail suspension test (MED = 10 mg/kg, p.o.), proportional monoamine transporter occupancies and enhancement of monoamine concentrations in mouse prefrontal cortex.

SPARC is a decoy counterpart for c­Fos and is associated with osteoblastic differentiation of bone marrow stromal cells by inhibiting adipogenesis.

Secreted protein acidic and rich in cysteine (SPARC), also called basement­membrane protein 40 or osteonectin, is a matricellular protein that is abundant not only in bone tissue as a non­collagenous protein but is also ubiquitously expressed in non­calcified tissue. SPARC is located intracellularly and disruption of the Sparc gene has been reported to reduce bone formation and increase fat tissue; however, the mechanism by which SPARC inhibits adipogenesis remains unclear. The present study evaluated the intracellular function of SPARC in adipogenesis using the bone marrow stromal cell line ST2. When ST2 cells with low SPARC production were cloned, intrinsic activator protein­1 (AP­1) activity was markedly higher, mineralized nodule formation was significantly lower and lipid accumulation was significantly increased compared with in the parental ST2 cells. Forced expression of secreted SPARC with the signal peptide­coding sequences of wild­type Sparc or preprotrypsin in SPARC­low ST2 cells significantly reduced AP­1 transcription activity; however, these reductions were not observed in the absence of signal peptide sequences. Recombinant SPARC, produced using Brevibacillus brevis, specifically bound to c­Fos but not c­Jun and inhibited the binding of c­Fos/c­Jun to a TPA­response element sequence. These data suggested that SPARC was incorporated into the cells from the extracellular spaces and serves an intracellular role as a decoy counterpart for c­Fos, as well as being associated with osteoblastogenesis through the inhibition of adipogenesis. These findings may provide new insights into regenerative medicine.

MRI Contrasting Agent Based on Mn-MOF-74 Nanoparticles with Coordinatively Unsaturated Sites.

PURPOSE: The development of magnetic resonance imaging (MRI) contrasting agents (CAs) that are safer and have a higher relaxivity than Gd(III)-based agents is a significant research topic. Herein, we propose the use of a Mn-based metal organic framework (MOF), Mn-MOF-74, characterized by a safe paramagnetic center, a coordinatively unsaturated site (CUS) for aquation, and a long rotational correlation time, endowing high relaxivity. Furthermore, biocompatibility and delivery to the tumor are generally expected for MOFs that are obtainable in the nanometer size range. PROCEDURE: Drop-wise mixing of 2,5-dihydroxyterephthalic acid (DHTP) and Mn(II) acetate yielded Mn-MOF-74 with a diameter of < 150 nm, which was then modified with 1-fivefold higher amounts of poly(ethylene glycol) (M.W. = 5000) to afford MOFs stably dispersed in water for at least 24 h. RESULTS: The longitudinal and transverse relaxivity of the PEG-modified MOF was in the range of r1 = 8.08-13.5 and r2 = 32.7-46.8 mM-1 s-1, respectively (1.0 T, 23.7-23.9 °C), being larger than those of typical Gd(III)- and Mn(II)-based CAs of single-nuclear metal complexes. The in vivo imaging of a tumor-bearing mouse clearly showed that the tumor could be readily recognized due to signal enhancement (117%) in T1-weighted images, whereas other tissues showed small signal changes. CONCLUSIONS: These results suggest that PEG-Mn-MOF-74 can be passively delivered to tumors and can act as a high-relaxivity T1 agent.

Selective crystallization of dysprosium complex from neodymium/dysprosium mixture enabled by cooperation of coordination and crystallization.

Designing a molecular-level Ln3+ separation system remains a challenge for developing next-generation separation methodologies. Herein, we report crystallization-based Nd3+/Dy3+ separation using a tripodal Schiff base ligand. Highly selective crystallization of the Dy3+ complex was enabled by cooperation between the coordination and crystallization processes.

Emergence of the super antenna effect in mixed crystals of ytterbium and lutetium complexes showing near-infrared luminescence.

The synthesis of luminescent molecular crystalline materials requires a good understanding of the luminescence properties of crystals in which many molecules are densely packed. Previously, we studied the near-infrared (NIR) luminescence of a trivalent ytterbium (Yb(iii)) complex with a Schiff base ligand, tris[2-(5-methylsalicylideneimino)ethyl]amine (H3L). Herein, we extended our study on the Yb complex (YbL) to enhance and understand its solid-state luminescence via mixed crystallization with the lutetium complex (LuL). We prepared (YbL) x (LuL)1-x mixed crystals (x = 0.01, 0.05, 0.1, 0.2, 0.3, 0.5, and 0.7) and studied their NIR luminescence properties. The NIR luminescence intensity per Yb(iii) ion for (YbL)0.01(LuL)0.99 was determined to be two orders of magnitude larger than that for YbL. The excitation spectral shape of (YbL)0.01(LuL)0.99 was different from the absorption spectral shape of YbL but similar to that of LuL. We attribute this observation to the emergence of an intermolecular energy-migration path. In the mixed crystals, LuL molecules acted as a light-harvesting super antenna for Yb(iii) luminescence. Decay measurements of the NIR luminescence for (YbL) x (LuL)1-x with x > 0.2 showed mono-exponential decay, while (YbL) x (LuL)1-x with x < 0.1 showed a grow-in component, which reflected the lifetime of the intermediate state for energy migration. The decay lifetime values tended to increase with decreasing x, suggesting that Yb(iii) isolation resulted in a reduction in concentration quenching. We propose that the luminescence enhancement in the highly Yb-diluted conditions was mainly caused by an increase in the super antenna effect.

Identification of novel functional organic anion-transporting polypeptide 1B3 polymorphisms and assessment of substrate specificity.

OBJECTIVE: The uptake carrier organic anion-transporting polypeptide 1B3 (OATP1B3, gene SLCO1B3) is involved in the hepatic clearance of xenobiotics including statins, taxanes, and mycophenolic acid. We thought to assess the SLCO1B3 coding region for yet unidentified polymorphisms and to analyze their functional relevance. METHODS: We used DNA of ethnically diverse individuals for polymerase chain reaction, and determined polymorphisms by sequencing or temperature-dependent capillary electrophoresis. We then created variant OATP1B3 expression plasmids by site-directed mutagenesis, which were transiently expressed and functionally characterized in human cervical carcinoma (HeLa) cells using radiolabeled substrates. RESULTS: We identified six nonsynonymous polymorphisms including novel variants such as 439A>G (Thr147Ala), 767G>C (Gly256Ala), 1559A>C (His520Pro), and 1679T>C (Val560Ala). Allelic frequencies occurred to be ethnicity-dependent, with the latter observed only in African-Americans (3.6%). After expression in HeLa cells, His520Pro, Val560Ala, and Met233Ile or Met233Ile_Ser112Ala haplotype variants showed decreased uptake activity compared with wild type for cholecystokinin-8 and rosuvastatin, but not for atorvastatin. Kinetic cholecystokinin-8 analysis showed reduced Vmax without altering Km. His520Pro and Val560Ala exhibited decreased total and plasma membrane protein expressions. Val560 mapped onto a structural model of OATP1B3 showed that this is a key region for substrate-transporter interaction. His520 resides in a predicted extracellular region thought to be critical to the pH-dependent component of OATP1B3 activity. Loss of activity at pH 7.4 and 8.0 relative to pH 6.5 was significantly greater for the Pro520 variant. CONCLUSION: OATP1B3 polymorphisms that result in altered expression, substrate specificity, and pH-dependent activity may be of potential relevance to hepatic clearance of substrate drugs in vivo.