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1.
Sci Rep ; 13(1): 21572, 2023 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-38062130

RESUMO

Osteosarcoma is rare but is the most common bone tumor. Diagnostic tools such as magnetic resonance imaging development of chemotherapeutic agents have increased the survival rate in osteosarcoma patients, although 5-year survival has plateaued at 70%. Thus, development of new treatment approaches is needed. Here, we report that IL-17, a proinflammatory cytokine, increases osteosarcoma mortality in a mouse model with AX osteosarcoma cells. AX cell transplantation into wild-type mice resulted in 100% mortality due to ectopic ossification and multi-organ metastasis. However, AX cell transplantation into IL-17-deficient mice significantly prolonged survival relative to controls. CD4-positive cells adjacent to osteosarcoma cells express IL-17, while osteosarcoma cells express the IL-17 receptor IL-17RA. Although AX cells can undergo osteoblast differentiation, as can patient osteosarcoma cells, IL-17 significantly inhibited that differentiation, indicating that IL-17 maintains AX cells in the undifferentiated state seen in malignant tumors. By contrast, IL-17RA-deficient mice transplanted with AX cells showed survival comparable to wild-type mice transplanted with AX cells. Biopsy specimens collected from osteosarcoma patients showed higher expression of IL-17RA compared to IL-17. These findings suggest that IL-17 is essential to maintain osteosarcoma cells in an undifferentiated state and could be a therapeutic target for suppressing tumorigenesis.


Assuntos
Neoplasias Ósseas , Osteossarcoma , Humanos , Camundongos , Animais , Receptores de Interleucina-17/metabolismo , Interleucina-17/genética , Interleucina-17/metabolismo , Osteossarcoma/patologia , Diferenciação Celular , Neoplasias Ósseas/patologia
2.
PLoS One ; 18(11): e0293944, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37939095

RESUMO

When ruptured, ligaments and tendons have limited self-repair capacity and rarely heal spontaneously. In the knee, the Anterior Cruciate Ligament (ACL) often ruptures during sports activities, causing functional impairment and requiring surgery using tendon grafts. Patients with insufficient time to recover before resuming sports risk re-injury. To develop more effective treatment, it is necessary to define mechanisms underlying ligament repair. For this, animal models can be useful, but mice are too small to create an ACL reconstruction model. Thus, we developed a transgenic rat model using control elements of Scleraxis (Scx), a transcription factor essential for ligament and tendon development, to drive GFP expression in order to localize Scx-expressing cells. As anticipated, Tg rats exhibited Scx-GFP in ACL during developmental but not adult stages. Interestingly, when we transplanted the flexor digitorum longus (FDP) tendon derived from adult Scx-GFP+ rats into WT adults, Scx-GFP was not expressed in transplanted tendons. However, tendons transplanted from adult WT rats into Scx-GFP rats showed upregulated Scx expression in tendon, suggesting that Scx-GFP+ cells are mobilized from tissues outside the tendon. Importantly, at 4 weeks post-surgery, Scx-GFP-expressing cells were more frequent within the grafted tendon when an ACL remnant was preserved (P group) relative to when it was not (R group) (P vs R groups (both n = 5), p<0.05), and by 6 weeks, biomechanical strength of the transplanted tendon was significantly increased if the remnant was preserved (P vsR groups (both n = 14), p<0.05). Scx-GFP+ cells increased in remnant tissue after surgery, suggesting remnant tissue is a source of Scx+ cells in grafted tendons. We conclude that the novel Scx-GFP Tg rat is useful to monitor emergence of Scx-positive cells, which likely contribute to increased graft strength after ACL reconstruction.


Assuntos
Lesões do Ligamento Cruzado Anterior , Reconstrução do Ligamento Cruzado Anterior , Humanos , Adulto , Ratos , Animais , Camundongos , Ligamento Cruzado Anterior/cirurgia , Tendões/cirurgia , Lesões do Ligamento Cruzado Anterior/cirurgia , Articulação do Joelho/cirurgia
3.
Sci Rep ; 13(1): 20019, 2023 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-37973808

RESUMO

Lumbar spinal stenosis (LSS) is a degenerative disease characterized by intermittent claudication and numbness in the lower extremities. These symptoms are caused by the compression of nerve tissue in the lumbar spinal canal. Ligamentum flavum (LF) hypertrophy and spinal epidural lipomatosis in the spinal canal are known to contribute to stenosis of the spinal canal: however, detailed mechanisms underlying LSS are still not fully understood. Here, we show that surgically harvested LFs from LSS patients exhibited significantly increased thickness when transthyretin (TTR), the protein responsible for amyloidosis, was deposited in LFs, compared to those without TTR deposition. Multiple regression analysis, which considered age and BMI, revealed a significant association between LF hypertrophy and TTR deposition in LFs. Moreover, TTR deposition in LF was also significantly correlated with epidural fat (EF) thickness based on multiple regression analyses. Mesenchymal cell differentiation into adipocytes was significantly stimulated by TTR in vitro. These results suggest that TTR deposition in LFs is significantly associated with increased LF hypertrophy and EF thickness, and that TTR promotes adipogenesis of mesenchymal cells. Therapeutic agents to prevent TTR deposition in tissues are currently available or under development, and targeting TTR could be a potential therapeutic approach to inhibit LSS development and progression.


Assuntos
Ligamento Amarelo , Estenose Espinal , Humanos , Estenose Espinal/complicações , Ligamento Amarelo/metabolismo , Pré-Albumina/metabolismo , Canal Medular/metabolismo , Hipertrofia/metabolismo , Vértebras Lombares/metabolismo
4.
Toxicol In Vitro ; 93: 105691, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37660997

RESUMO

Severe diarrhea is a common side effect of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). We aimed to evaluate the risk of EGFR-TKI-induced diarrhea using spheroids of human and monkey crypt-derived intestinal stem cells. Intestinal spheroids exhibited higher toxic susceptibility to EGFR-TKIs than Caco-2 cells. As concentration of EGFR-TKIs increased, cellular ATP first decreased relative to the control condition, followed by an increase in LDH release, in contrast with their simultaneous changes with traditional cytotoxic anticancer drugs. The toxic sensitivity of spheroids to various EGFR-TKIs corresponded to clinical diarrhea incidence. Afatinib, a second-generation EGFR-TKI, exhibited higher toxic sensitivity compared with the first-generation ones, corresponding to the clinical evidence that afatinib-induced diarrhea is almost inevitable and severe. By contrast, the third-generation osimertinib, which reduces the risk of diarrhea, showed mitigated cytotoxicity compared with afatinib. For irreversible EGFR-TKIs, the decreased ATP level persisted or its recovery was delayed even after drug removal compared with reversible ones. Furthermore, the highest drug accumulation in spheroids (TKIspheroids) and inhibition potency against EGFR (TKIspheroids/Ki) were observed for afatinib. This system would be useful for predicting the risk of EGFR-TKI-induced diarrhea; moreover, on-target cytotoxicity against intestinal stem cells might contribute to clinically observed diarrhea.


Assuntos
Antineoplásicos , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Animais , Afatinib/toxicidade , Afatinib/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Inibidores de Proteínas Quinases/toxicidade , Haplorrinos/metabolismo , Células CACO-2 , Receptores ErbB/metabolismo , Mutação , Antineoplásicos/farmacologia , Diarreia/induzido quimicamente , Trifosfato de Adenosina
5.
Radiol Case Rep ; 18(9): 3093-3100, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37404221

RESUMO

Focal nodular hyperplasia (FNH) or FNH-like lesions of the liver are benign lesions that can be mostly diagnosed by hepatobiliary phase gadoxetic acid-enhanced magnetic resonance imaging (MRI). Accurate imaging diagnosis is based on the fact that most FNHs or FNH-like lesions show characteristic hyper- or isointensity on hepatobiliary phase images. We report a case of an FNH-like lesion in a 73-year-old woman that mimicked a malignant tumor. Dynamic contrast-enhanced computed tomography (CT) and MRI using gadoxetic-acid revealed an ill-defined nodule showing early enhancement in the arterial phase and gradual and prolonged enhancement in the portal and equilibrium/transitional phases. Hepatobiliary phase imaging revealed inhomogeneous hypointensity, accompanied by a slightly isointense area compared to the background liver. Angiography-assisted CT showed a portal perfusion defect of the nodule, inhomogeneous arterial blood supply in the early phase, and less internal enhancement in the late phase, accompanied by irregularly shaped peritumoral enhancement. No central stellate scar was identified in any of the images. Imaging findings could not exclude the possibility of hepatocellular carcinoma, but the nodule was pathologically diagnosed as an FNH-like lesion by partial hepatectomy. In the present case, an unusual inhomogeneous hypointensity on hepatobiliary phase imaging made it difficult to diagnose the FNH-like lesions.

6.
Drug Metab Dispos ; 51(9): 1136-1144, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37142427

RESUMO

Prediction of intestinal absorption of drugs in humans is one of the critical elements in the development process for oral drugs. However, it remains challenging, because intestinal absorption of drugs is influenced by multiple factors, including the function of various metabolic enzymes and transporters, and large species differences in drug bioavailability hinder the prediction of human bioavailability directly from in vivo animal experiments. For the screening of intestinal absorption properties of drugs, a transcellular transport assay with Caco-2 cells is still routinely used by pharmaceutical companies because of its convenience, but the predictability of the fraction of the oral dose that goes to the portal vein of metabolic enzyme/transporter substrate drugs was not always good because the cellular expression of metabolic enzymes and transporters is different from that in the human intestine. Recently, various novel in vitro experimental systems have been proposed such as the use of human-derived intestinal samples, transcellular transport assay with induced pluripotent stem-derived enterocyte-like cells, or differentiated intestinal epithelial cells derived from intestinal stem cells at crypts. Crypt-derived differentiated epithelial cells have an excellent potential to characterize species differences and regional differences in intestinal absorption of drugs because a unified protocol can be used for the proliferation of intestinal stem cells and their differentiation into intestinal absorptive epithelial cells regardless of the animal species and the gene expression pattern of differentiated cells is maintained at the site of original crypts. The advantages and disadvantages of novel in vitro experimental systems for characterizing intestinal absorption of drugs are also discussed. SIGNIFICANCE STATEMENT: Among novel in vitro tools for the prediction of human intestinal absorption of drugs, crypt-derived differentiated epithelial cells have many advantages. Cultured intestinal stem cells are rapidly proliferated and easily differentiated into intestinal absorptive epithelial cells simply by changing the culture media. A unified protocol can be used for the establishment of intestinal stem cell culture from preclinical species and humans. Region-specific gene expression at the collection site of crypts can be reproduced in differentiated cells.


Assuntos
Células Epiteliais , Intestinos , Animais , Humanos , Células CACO-2 , Células Epiteliais/metabolismo , Células-Tronco , Proteínas de Membrana Transportadoras/metabolismo , Diferenciação Celular , Absorção Intestinal , Mucosa Intestinal/metabolismo
7.
Radiol Phys Technol ; 16(2): 325-337, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37097551

RESUMO

The Japan Association of Radiological Technologists (JART) and the Japan Medical Imaging and Radiological Systems Industries Association jointly conducted a nationwide survey to reveal the current situation of diagnostic displays in Japan using a questionnaire on the performance and quality control (QC) of diagnostic displays for mammography and common use. The questionnaire for radiological technologists (RTs) was distributed via email to 4519 medical facilities throughout Japan, where RTs affiliated with JART were employed; 613 (13.6%) facilities responded. Diagnostic displays with suitable maximal luminance (500 cd/m2 or higher for mammography and 350 cd/m2 or higher for common use) and resolution (5 megapixels for mammography) have been widely used. However, while 99% of the facilities recognized the necessity of QC, only approximately 60% implemented it. This situation arose due to several barriers to QC implementation, such as insufficient devices, time, staff, knowledge, and the recognition of QC as a duty. The implementation of QC can lead to the avoidance of incidents or accidents caused by a decrease in luminance, variation in luminance response, and the influence of ambient light. Moreover, the barriers discouraging the implementation of QC are mainly related to a lack of human resources and budgets. Therefore, to popularize the QC of diagnostic displays in all facilities, it is crucial to identify countermeasures to eliminate these barriers and to continue positive actions for popularization.


Assuntos
Mamografia , Radiologia , Humanos , Japão , Controle de Qualidade , Radiologia/métodos , Inquéritos e Questionários
8.
Clin Transl Sci ; 15(6): 1519-1531, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35421902

RESUMO

The accurate prediction of OATP1B-mediated drug-drug interactions (DDIs) is challenging for drug development. Here, we report a physiologically-based pharmacokinetic (PBPK) model analysis for clinical DDI data generated in heathy subjects who received oral doses of cyclosporin A (CysA; 20 and 75 mg) as an OATP1B inhibitor, and the probe drugs (pitavastatin, rosuvastatin, and valsartan). PBPK models of CysA and probe compounds were combined assuming inhibition of hepatic uptake of endogenous coproporphyrin I (CP-I) by CysA. In vivo Ki of unbound CysA for OATP1B (Ki,OATP1B ), and the overall intrinsic hepatic clearance per body weight of CP-I (CLint,all,unit ) were optimized to account for the CP-I data (Ki,OATP1B , 0.536 ± 0.041 nM; CLint,all,unit , 41.9 ± 4.3 L/h/kg). DDI simulation using Ki,OATP1B reproduced the dose-dependent effect of CysA (20 and 75 mg) and the dosing interval (1 and 3 h) on the time profiles of blood concentrations of pitavastatin and rosuvastatin, but DDI simulation using in vitro Ki,OATP1B failed. The Cluster Gauss-Newton method was used to conduct parameter optimization using 1000 initial parameter sets for the seven pharmacokinetic parameters of CP-I (ß, CLint, all , Fa Fg , Rdif , fbile , fsyn , and vsyn ), and Ki,OATP1B and Ki,MRP2 of CysA. Based on the accepted 546 parameter sets, the range of CLint, all and Ki,OATP1B was narrowed, with coefficients of variation of 12.4% and 11.5%, respectively, indicating that these parameters were practically identifiable. These results suggest that PBPK model analysis of CP-I is a promising translational approach to predict OATP1B-mediated DDIs in drug development.


Assuntos
Coproporfirinas , Modelos Biológicos , Interações Medicamentosas , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado , Rosuvastatina Cálcica
9.
Clin Pharmacol Ther ; 111(6): 1315-1323, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35292967

RESUMO

This study was designed to assess the quantitative performance of endogenous biomarkers for organic anion transporting polypeptide (OATP) 1B1/1B3-mediated drug-drug interactions (DDIs). Ten healthy volunteers orally received OATP1B1/1B3 probe cocktail (0.2 mg pitavastatin, 1 mg rosuvastatin, and 2 mg valsartan) and an oral dose of cyclosporin A (CysA, 20 mg and 75 mg) separated by a 1-hour interval (20 mg (-1 hour), and 75 mg (-1 hour)). CysA 75 mg was also given with a 3-hour interval (75 mg (-3 hours)) to examine the persistence of OATP1B1/1B3 inhibition. The area under the plasma concentration-time curve ratios (AUCRs) were 1.63, 3.46, and 2.38 (pitavastatin), 1.39, 2.16, and 1.81 (rosuvastatin), and 1.42, 1.77, and 1.85 (valsartan), at 20 mg, 75 mg (-1 hour) and 75 mg (-3 hours) of CysA, respectively. CysA effect on OATP1B1/1B3 was unlikely to persist at the dose examined. Among 26 putative OATP1B1/1B3 biomarkers evaluated, AUCR and maximum concentration ratio (Cmax R) of CP-I showed the highest Pearson's correlation coefficient with CysA AUC (0.94 and 0.93, respectively). Correlation between AUCR of pitavastatin, and Cmax R or AUCR of CP-I were consistent between this study and our previous study using rifampicin as an OATP1B1/1B3 inhibitor. Nonlinear regression analysis of AUCR-1 of pitavastatin and CP-I against CysA Cmax yielded Ki,OATP1B1/1B3,app (109 ± 35 and 176 ± 42 nM, respectively), similar to the Ki ,OATP1B1/1B3 estimated by our physiologically-based pharmacokinetic model analysis described previously (107 nM). The endogenous OATP1B1/1B3 biomarkers, particularly Cmax R and AUCR of CP-I, corroborates OATP1B1/1B3 inhibition and yields valuable information that improve accurate DDI predictions in drug development, and enhance our understanding of interindividual variability in the magnitude of DDIs.


Assuntos
Ciclosporina , Transportadores de Ânions Orgânicos , Ciclosporina/farmacologia , Interações Medicamentosas , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado , Rosuvastatina Cálcica/farmacocinética , Membro 1B3 da Família de Transportadores de Ânion Orgânico Carreador de Soluto , Valsartana
10.
J Surg Case Rep ; 2022(2): rjac020, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35154639

RESUMO

Neuroendocrine carcinomas (NECs) arising from the extrahepatic bile duct (EHBD) are extremely rare, and their preoperative diagnosis is difficult. A small number of resected cases of EHBD NECs has been reported, and their prognosis is usually poor. A 62-year-old man presented with obstructive jaundice and liver disease. Radiological imaging revealed wall thickness and stricture of the distal common bile duct (CBD); however, lymph node or distant metastasis was not detected. Adenocarcinoma was detected on biopsy, and surgery was performed with a preoperative diagnosis of cholangiocarcinoma of the distal CBD. Pathological examination revealed adenocarcinoma of the CBD mucosa (20%) and NEC of the CBD wall (80%). The final pathological diagnosis was small-cell NEC of the EHBD. His post-operative course was good, and there was no recurrence for 4 months after surgery. Herein, we report a case of resected EHBD NEC and a literature review.

11.
Drug Metab Dispos ; 50(3): 204-213, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34992074

RESUMO

This study aimed to demonstrate the usefulness of human jejunal spheroid-derived differentiated intestinal epithelial cells as a novel in vitro model for clarifying the impact of intestinal drug-metabolizing enzymes and transporters on the intestinal absorption of substrate drugs in humans. Three-dimensional human intestinal spheroids were successfully established from surgical human jejunal specimens and expanded for a long period using L-WRN-conditioned medium, which contains Wnt3a, R-spondin 3, and noggin. The mRNA expression levels of intestinal pharmacokinetics-related genes in the human jejunal spheroid-derived differentiated intestinal epithelial cells were drastically increased over a 5-day period after seeding compared with those in human jejunal spheroids and were approximately the same as those in human jejunal tissue over a culture period of at least 13 days. Activities of typical drug-metabolizing enzymes [cytochrome P450 (CYP) 3A, CYP2C9, uridine 5'-diphospho-glucuronosyltransferase 1A, and carboxylesterase 2] and uptake/efflux transporters [peptide transporter 1/solute carrier 15A1], P-glycoprotein, and breast cancer resistance protein) in the differentiated cells were confirmed. Furthermore, intestinal availability (Fg) values estimated from the apical-to-basolateral permeation clearance across cell monolayer showed a good correlation with the in vivo Fg values in humans for five CYP3A substrate drugs (Fg range, 0.35-0.98). In conclusion, the functions of major intestinal drug-metabolizing enzymes and transporters could be maintained in human jejunal spheroid-derived differentiated intestinal epithelial cells. This model would be useful for the quantitative evaluation of the impact of intestinal drug-metabolizing enzymes and transporters on the intestinal absorption of substrate drugs in humans. SIGNIFICANCE STATEMENT: Limited information is available regarding the quantitative prediction of the impact of drug-metabolizing enzymes and transporters on the human intestinal absorption of substrates using in vitro assays with differentiated cells derived from human intestinal spheroids/organoids. This study confirmed the functions of typical drug-metabolizing enzymes and transporters in human jejunal spheroid-derived differentiated intestinal epithelial cells and demonstrated that intestinal availability (Fg) estimated from apical-to-basolateral permeation clearance across cell monolayers showed a good correlation with in vivo human Fg for CYP3A substrates.


Assuntos
Mucosa Intestinal , Proteínas de Neoplasias , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Células Epiteliais/metabolismo , Humanos , Absorção Intestinal , Mucosa Intestinal/metabolismo , Proteínas de Neoplasias/metabolismo
12.
Stem Cell Reports ; 16(2): 295-308, 2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33513361

RESUMO

We aimed to establish an in vitro differentiation procedure to generate matured small intestinal cells mimicking human small intestine from human-induced pluripotent stem cells (iPSCs). We previously reported the efficient generation of CDX2-expressing intestinal progenitor cells from embryonic stem cells (ESCs) using 6-bromoindirubin-3'-oxime (BIO) and (3,5-difluorophenylacetyl)-L-alanyl-L-2-phenylglycine tert-butyl ester (DAPT) to treat definitive endodermal cells. Here, we demonstrate the generation of enterocyte-like cells by culturing human iPSC-derived intestinal progenitor cells on a collagen vitrigel membrane (CVM) and treating cells with a simple maturation medium containing BIO, DMSO, dexamethasone, and activated vitamin D3. Functional tests further confirmed that these iPSC-derived enterocyte-like cells exhibit P-gp- and BCRP-mediated efflux and cytochrome P450 3A4 (CYP3A4)-mediated metabolism. We concluded that hiPS cell-derived enterocyte-like cells can be used as a model for the evaluation of drug transport and metabolism studies in the human small intestine.


Assuntos
Técnicas de Cultura de Células/métodos , Enterócitos/citologia , Enterócitos/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Intestino Delgado/citologia , Intestino Delgado/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Adulto , Diferenciação Celular , Linhagem Celular , Células Cultivadas , Colágeno/metabolismo , Meios de Cultura , Citocromo P-450 CYP3A/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/metabolismo , Adulto Jovem
13.
Drug Metab Dispos ; 49(1): 84-93, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33087448

RESUMO

Intestinal permeability is a critical factor for orally administered drugs. It can be facilitated by uptake transporters or limited by efflux transporters and metabolic enzymes in the intestine. The present study aimed to characterize the Ussing chamber system incorporating human intestinal tissue as an in vitro model for investigating the impact of intestinal uptake/efflux transporters on the intestinal absorption of substrate drugs in humans. We confirmed the functions of major intestinal uptake/efflux drug transporters in freshly isolated human jejunum sections by demonstrating a significant decrease in the mucosal uptake of cefadroxil (peptide transporter 1) and methotrexate (proton-coupled folate transporter), mucosal-to-serosal permeability of ribavirin (concentrative nucleoside transporters/equilibrative nucleoside transporters), and serosal-to-mucosal permeability of P-glycoprotein and breast cancer resistance protein substrates in the presence of their typical inhibitors. The mucosal-to-serosal apparent permeability coefficients (Papp) of 19 drugs, including substrates of drug transporters and cytochrome P450 3A, ranged from 0.60 × 10-6 to 29 × 10-6 cm/s and showed a good correlation with reported fraction of an oral dose that enters the gut wall and passes into the portal circulation with escaping intestinal metabolism (FaFg) values in humans. Furthermore, the Papp values for cefadroxil, methotrexate, and ribavirin in the presence of the corresponding transporter inhibitors underestimated the FaFg of these drugs, which clearly showed that intestinal uptake transporters facilitate their intestinal absorption in humans. In conclusion, the functions of major intestinal uptake/efflux drug transporters could be maintained in freshly isolated human jejunum sections. The Ussing chamber system incorporating human intestinal tissue would be useful for evaluating the impact of intestinal uptake/efflux transporters on the intestinal absorption of various types of drugs in humans. SIGNIFICANCE STATEMENT: Although previous studies have predicted the intestinal absorption of drugs in humans using the Ussing chamber system incorporating human intestinal tissue, there is little systematic information about drug transport mediated by multiple transporters in this system. We confirmed the functions of major intestinal uptake/efflux transporters in freshly isolated human jejunum sections and demonstrated that the mucosal-to-serosal apparent permeability coefficient of various types of drugs showed a good correlation with reported human FaFg values.


Assuntos
Absorção Intestinal/fisiologia , Mucosa Intestinal , Jejuno , Proteínas de Membrana Transportadoras/metabolismo , Preparações Farmacêuticas/metabolismo , Administração Oral , Transporte Biológico , Humanos , Mucosa Intestinal/enzimologia , Mucosa Intestinal/metabolismo , Jejuno/metabolismo , Jejuno/patologia , Circulação Hepática/fisiologia , Permeabilidade , Farmacocinética
14.
Jpn J Radiol ; 39(2): 118-142, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32840742

RESUMO

A variety of neoplastic and non-neoplastic lesions of the pancreas can present with a predominantly cystic architecture. These lesions are increasingly being detected as incidental findings on routine cross-sectional imaging following technological advances in these techniques and their widespread use. The different histopathological behaviors show various common and uncommon imaging findings, and some cases show similar appearance in spite of different histopathology. Each lesion requires specific management because of the differing risk of progression to malignancy, and an accurate imaging diagnosis is crucial. The typical imaging characteristics that differentiate pancreatic cystic lesions have been well described and fully summarized. However, in addition to a small percentage of cases that shows uncommon imaging findings, a substantial percentage of cystic lesions shows overlapping imaging findings that can lead to radiological misdiagnosis. For appropriate diagnosis and optimal treatment strategy, it is important to know the uncommon and overlapping imaging findings of these lesions, in addition to familiarity with the typical aspects. In this article, we reconfirm the well-known characteristic imaging features of pancreatic cystic lesions and present several diagnostically challenging cases, focusing on the uncommon and overlapping imaging findings.


Assuntos
Pâncreas/diagnóstico por imagem , Cisto Pancreático/diagnóstico por imagem , Neoplasias Pancreáticas/diagnóstico por imagem , Diagnóstico Diferencial , Feminino , Humanos , Imageamento por Ressonância Magnética , Masculino , Pâncreas/patologia , Cisto Pancreático/patologia , Neoplasias Pancreáticas/patologia , Tomografia Computadorizada por Raios X
15.
J Pharm Sci ; 110(1): 376-387, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33122051

RESUMO

Hepatic uptake clearance has been measured in suspended human hepatocytes (SHH). Plated human hepatocytes (PHH) after short-term culturing are increasingly employed to study hepatic transport driven mainly by its higher throughput. To know pros/cons of both systems, the hepatic uptake clearances of several organic anion transporting polypeptide 1B substrates were compared between PHH and SHH by determining the initial uptake velocities or through dynamic model-based analyses. For cerivastatin, pitavastatin and rosuvastatin, initial uptake clearances (PSinf) obtained using PHH were comparable to those using SHH, while cell-to-medium concentration (C/M) ratios were 2.7- to 5.4-fold higher. For pravastatin and dehydropravastatin, hydrophilic compounds with low uptake/cellular binding, their PSinf and C/M ratio in PHH were 1.8- to 3.2-fold lower than those in SHH. These hydrophilic substrates are more prone to wash-off during the uptake study using PHH, which may explain the apparently lower uptake than SHH. The C/M ratios obtained using PHH were stable over an extended time, making PHH suitable to estimate the C/M ratios and hepatocyte-to-medium unbound concentration ratios (Kp,uu). In conclusion, PHH is useful in evaluating hepatic uptake/efflux clearances and Kp,uu of OATP1B substrates in a high-throughput manner, however, a caution is warranted for hydrophilic drugs with low uptake/cellular binding.


Assuntos
Hepatócitos , Transportadores de Ânions Orgânicos , Transporte Biológico , Hepatócitos/metabolismo , Humanos , Fígado/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Pravastatina/metabolismo
16.
AAPS J ; 22(6): 133, 2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-33063163

RESUMO

Suspended human hepatocytes (SHH) have long been used in assessing hepatic drug uptake, while plated human hepatocytes in short-term monolayer culture (PHH) have gained use in recent years. This study aimed to cross-evaluate SHH and PHH in measuring the hepatic uptake mediated by organic anion transporting polypeptide 1Bs (OATP1Bs). We compared the time courses of cell-to-medium (C/M) concentration ratios and initial uptake clearance values of the OATP1B substrates (pitavastatin, rosuvastatin, cerivastatin, pravastatin, dehydropravastatin, and SC-62807) between SHH and PHH. For all compounds except cerivastatin, the C/M ratios in SHH displayed an apparent overshoot (an initial increase followed by a decrease) during the 180-min uptake experiment, but not in PHH. Based on the literature evidence suggesting the possible internalization of OATP1Bs in primary hepatocytes, separate experiments measured the drug uptake after varying lengths of pre-incubation in the drug-free medium. The initial uptake clearances of pitavastatin and rosuvastatin declined in SHH beyond an apparent threshold time of 20-min drug-free pre-incubation, but not in PHH. Kinetic modeling quantitatively captured the decline in the active uptake clearance in SHH, and more than half of the active uptake clearances of pitavastatin and rosuvastatin were prone to loss during the 180-min uptake experiment. These results suggested a partial, time-delayed loss of the functional OATP1Bs in SHH upon prolonged incubation. Our results indicate that PHH is more appropriate for experiments where a prolonged incubation is required, such as estimation of unbound hepatocyte-to-medium concentration ratio (Kp,uu) at the steady-state.


Assuntos
Hepatócitos/enzimologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacocinética , Transportador 1 de Ânion Orgânico Específico do Fígado/metabolismo , Adulto , Células Cultivadas , Criança , Meios de Cultura/análise , Meios de Cultura/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Eliminação Hepatobiliar , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/análise , Masculino , Modelos Biológicos , Cultura Primária de Células/métodos
17.
J Ovarian Res ; 13(1): 47, 2020 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-32336272

RESUMO

BACKGROUND: Ovarian cancer (OC) is a leading cause of cancer-related death in women, and thus an accurate diagnosis of the predisposition and its early detection is necessary. The aims of this study were to determine whether serum exosomal microRNA-34a (miR-34a) in ovarian cancer could be used as a potential biomarker. METHODS: Exosomes from OC patients' serum were collected, and exosomal miRNAs were extracted. The relative expression of miR-34a was calculated from 58 OC samples by quantitative real-time polymerase chain reaction. RESULTS: Serum exosomal miR-34a levels were significantly increased in early-stage OC patients compared with advanced-stage patients. Its levels were significantly lower in patients with lymph node metastasis than in those with no lymph node metastasis. Furthermore, its levels in the recurrence group were significantly lower than those in the recurrence-free group. CONCLUSIONS: Serum exosomal miR-34a could be a potential biomarker for improving the diagnostic efficiency of OC.


Assuntos
Biomarcadores Tumorais/sangue , Carcinoma Epitelial do Ovário/sangue , Carcinoma Epitelial do Ovário/genética , Exossomos/genética , MicroRNAs/sangue , Neoplasias Ovarianas/sangue , Neoplasias Ovarianas/genética , Adulto , Idoso , Carcinoma Epitelial do Ovário/patologia , Exossomos/ultraestrutura , Feminino , Humanos , Microscopia Eletrônica de Transmissão , Pessoa de Meia-Idade , Neoplasias Ovarianas/patologia , Ovário/patologia
18.
Mol Pharm ; 17(6): 1884-1898, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32271581

RESUMO

We developed a practical synthetic method for fluorine-18 (18F)-labeled pitavastatin ([18F]PTV) as a positron emission tomography (PET) tracer to assess hepatobiliary transporter activity and conducted a PET scan as a preclinical study for proof-of-concept in rats. This method is a one-pot synthesis involving aromatic 18F-fluorination of an arylboronic acid ester followed by deprotection under acidic conditions, which can be reproduced in general clinical sites equipped with a standard radiolabeling system due to the simplified procedure. PET imaging confirmed that intravenously administered [18F]PTV was rapidly accumulated in the liver and gradually transferred into the intestinal lumen through the bile duct. Radiometabolite analysis showed that [18F]PTV was metabolically stable, and 80% of the injected dose was detected as the unchanged form in both blood and bile. We applied integration plot analysis to assess tissue uptake clearance (CLuptake, liver and CLuptake, kidney) and canalicular efflux clearance (CLint, bile), and examined the effects of inhibitors on membrane transport. Treatment with rifampicin, an organic anion transporting polypeptide inhibitor, significantly reduced CLuptake, liver and CLuptake, kidney to 44% and 64% of control, respectively. In contrast, Ko143, a breast cancer resistance protein inhibitor, did not affect CLuptake, liver but significantly reduced CLint, bile to 39% of control without change in [18F]PTV blood concentration. In addition, we found decreased CLuptake, liver and increased CLint, bile in Eisai hyperbilirubinemic rats in response to altered expression levels of transporters. We expect that [18F]PTV can be translated into clinical application, as our synthetic method does not need special apparatus in the radiolabeling system and PET scan with [18F]PTV can quantitatively evaluate transporter activity in vivo.


Assuntos
Radioisótopos de Flúor/química , Quinolinas/química , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/efeitos dos fármacos , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Western Blotting , Cromatografia em Camada Fina , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Estrutura Molecular , Proteínas de Neoplasias/efeitos dos fármacos , Proteínas de Neoplasias/metabolismo , Transportadores de Ânions Orgânicos/efeitos dos fármacos , Transportadores de Ânions Orgânicos/metabolismo , Tomografia por Emissão de Pósitrons , Ratos , Ratos Sprague-Dawley , Rifampina/química
19.
Oncol Lett ; 19(3): 1872-1880, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32194682

RESUMO

CD24, which is upregulated in several human malignancies, is related to Epithelial-mesenchymal-transition (EMT) and has characteristics of cancer stem-like cells, especially in cisplatin-resistant ovarian carcinoma cells. Drug delivery systems represent a promising therapeutic approach for diseases with treatment resistance, and the present study investigated a novel CD24-targeted drug delivery system for advanced ovarian carcinoma. We produced liposomal cisplatin with a red fluorescent substance - cyanine 5.5 (GL-CDDP-Cy5.5). In order to target CD24-positive cells, an anti-CD24 monoclonal antibody was modified to the above drug (CD24-GL-CDDP-Cy5.5). Specific uptake of CD24-GL-CDDP-Cy5.5 was confirmed using a therapeutically resistant ovarian cancer cell line, Caov-3 cells. Antitumor effects of CD24-GL-CDDP-Cy5.5 were then evaluated in Caov-3 ×enograft mice. CD24-GL-CDDP-Cy5.5 showed more specific uptake by flow cytometry than GL-CDDP-Cy5.5. In xenograft mice, GL-CDDP-Cy5.5 and CD24-GL-CDDP-Cy5.5 treatment had significantly higher platinum concentration in disseminated tumor cells than cisplatin (P<0.05). Moreover, CD24-GL-CDDP-Cy5.5 suppressed tumor growth and prolonged survival time compared with other treatments. Median survival times of the control, cisplatin, GL-CDDP-Cy5.5 and CD24-GL-CDDP-Cy5.5 groups were 37, 36, 46 and 54 days after inoculation, respectively. Immunohistochemical analysis showed that CD24-GL-CDDP-Cy5.5 treatment, compared with GL-CDDP-Cy5.5, decreased the number of CD24-positive cells and suppressed the EMT phenomenon significantly (P<0.05). The present study demonstrated that CD24-GL-CDDP-Cy5.5, compared with other treatments, improved therapeutic efficacy. The present results suggested the potential for targeting anticancer therapeutics for CD24-positive cells to prevent disease progression.

20.
Drug Metab Dispos ; 48(5): 387-394, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32114508

RESUMO

Paclitaxel has been considered to cause OATP1B-mediated drug-drug interactions at therapeutic doses; however, its clinical relevance has not been demonstrated. This study aimed to elucidate in vivo inhibition potency of paclitaxel against OATP1B1 and OATP1B3 using endogenous OATP1B biomarkers. Paclitaxel is an inhibitor of OATP1B1 and OATP1B3, with Ki of 0.579 ± 0.107 and 5.29 ± 3.87 µM, respectively. Preincubation potentiated its inhibitory effect on both OATP1B1 and OATP1B3, with Ki of 0.154 ± 0.031 and 0.624 ± 0.183 µM, respectively. Ten patients with non-small cell lung cancer who received 200 mg/m2 of paclitaxel by a 3-hour infusion were recruited. Plasma concentrations of 10 endogenous OATP1B biomarkers-namely, coproporphyrin I, coproporphyrin III, glycochenodeoxycholate-3-sulfate, glycochenodeoxycholate-3-glucuronide, glycodeoxycholate-3-sulfate, glycodeoxycholate-3-glucuronide, lithocholate-3-sulfate, glycolithocholate-3-sulfate, taurolithocholate-3-sulfate, and chenodeoxycholate-24-glucuronide-were determined in the patients with non-small cell lung cancer on the day before paclitaxel administration and after the end of paclitaxel infusion for 7 hours. Paclitaxel increased the area under the plasma concentration-time curve (AUC) of the endogenous biomarkers 2- to 4-fold, although a few patients did not show any increment in the AUC ratios of lithocholate-3-sulfate, glycolithocholate-3-sulfate, and taurolithocholate-3-sulfate. Therapeutic doses of paclitaxel for the treatment of non-small cell lung cancer (200 mg/m2) will cause significant OATP1B1 inhibition during and at the end of the infusion. This is the first demonstration that endogenous OATP1B biomarkers could serve as surrogate biomarkers in patients. SIGNIFICANCE STATEMENT: Endogenous biomarkers can address practical and ethical issues in elucidating transporter-mediated drug-drug interaction (DDI) risks of anticancer drugs clinically. We could elucidate a significant increment of the plasma concentrations of endogenous OATP1B biomarkers after a 3-hour infusion (200 mg/m2) of paclitaxel, a time-dependent inhibitor of OATP1B, in patients with non-small cell lung cancer. The endogenous OATP1B biomarkers are useful to assess the possibility of OATP1B-mediated DDIs in patients and help in appropriately designing a dosing schedule to avoid the DDIs.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Transportador 1 de Ânion Orgânico Específico do Fígado/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Paclitaxel/farmacologia , Membro 1B3 da Família de Transportadores de Ânion Orgânico Carreador de Soluto/antagonistas & inibidores , Idoso , Área Sob a Curva , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/metabolismo , Carboplatina/farmacologia , Carboplatina/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/patologia , Interações Medicamentosas , Feminino , Células HEK293 , Humanos , Infusões Intravenosas , Transportador 1 de Ânion Orgânico Específico do Fígado/genética , Transportador 1 de Ânion Orgânico Específico do Fígado/metabolismo , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Paclitaxel/uso terapêutico , Polimorfismo de Nucleotídeo Único , Estudos Prospectivos , Proteínas Recombinantes/metabolismo , Membro 1B3 da Família de Transportadores de Ânion Orgânico Carreador de Soluto/metabolismo , Resultado do Tratamento
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