Liquid Chromatography/Tandem Mass Spectrometry-Based Simultaneous Analysis of 32 Bile Acids in Plasma and Conventional Biomarker-Integrated Diagnostic Screening Model Development for Hepatocellular Carcinoma.

Imaging tests, tumor marker (TM) screening, and biochemical tests provide a definitive diagnosis of hepatocellular carcinoma (HCC). However, some patients with HCC may present TM-negative results, warranting a need for developing more sensitive and accurate screening biomarkers. Various diseases exhibit increased blood levels of bile acids, biosynthesized from cholesterol in the liver, and they have been associated with HCC. Herein, we analyzed plasma bile acids using liquid chromatography/tandem mass spectrometry and integrated them with conventional biomarkers to develop a diagnostic screening model for HCC. Plasma samples were obtained from patients diagnosed with chronic hepatitis, hepatic cirrhosis (HC), and HCC. A QTRAP 6500 mass spectrometer and a Nexera liquid chromatograph with a YMC-Triart C18 analytical column were used. The mobile phase A was a 20 mmol/L ammonium formate solution, and mobile phase B was a methanol/acetonitrile mixture (1:1, v/v) with 20 mmol/L ammonium formate. After determining the concentrations of 32 bile acids, statistical analysis and diagnostic screening model development were performed. Plasma concentrations of bile acids differed between sample groups, with significant differences observed between patients with HC and HCC. By integrating bile acid results with conventional biochemical tests, a potential diagnostic screening model for HCC was successfully developed. Future studies should increase the sample size and analyze the data in detail to verify the diagnostic efficacy of the model.

Relevance of plasma lenvatinib concentrations and endogenous urinary cytochrome P450 3A activity biomarkers in clinical practice.

Lenvatinib (LEN), a multitarget tyrosine kinase inhibitor used in various cancer treatments, is mainly metabolized by cytochrome P450 3A (CYP3A) enzymes. The importance of therapeutic drug monitoring (TDM) in patients administered LEN has been proposed. Although some biomarkers of endogenous CYP3A activity have been reported, their utility in dosage adjustments has not been well evaluated. This study investigated the correlation between plasma LEN concentrations and endogenous urinary CYP3A biomarkers in clinical practice. Concentrations of plasma LEN (N = 225) and CYP3A biomarkers (cortisol, 6ß-hydroxycortisol, deoxycholic acid, and 1ß-hydroxydeoxycholic acid) in urine (N = 214) from 20 patients (hepatocellular carcinoma, N = 6; thyroid cancer, N = 3; endometrial cancer, N = 8; and renal cell carcinoma, N = 3) collected for consultation for up to 1 year were evaluated using liquid chromatography-tandem mass spectrometry. Moreover, plasma trough LEN concentrations were predicted using a three-compartment model with linear elimination for outpatients administered LEN before sample collection. Moderate correlations were observed between the quantified actual concentrations and the predicted trough concentrations of LEN, whereas there was no correlation with endogenous urinary CYP3A biomarkers. The utility of endogenous urinary CYP3A biomarkers could not be determined. However, TDM for outpatients administered orally available medicines may be predicted using a nonlinear mixed effect model (NONMEM). This study investigated the utility of endogenous urinary CYP3A biomarkers for personalized medicine and NONMEM for predicting plasma trough drug concentrations. These findings will provide important information for further clinical investigation and detailed TDM.

The Use of an Antioxidant Enables Accurate Evaluation of the Interaction of Curcumin on Organic Anion-Transporting Polypeptides 4C1 by Preventing Auto-Oxidation.

Flavonoids have garnered attention because of their beneficial bioactivities. However, some flavonoids reportedly interact with drugs via transporters and may induce adverse drug reactions. This study investigated the effects of food ingredients on organic anion-transporting polypeptide (OATP) 4C1, which handles uremic toxins and some drugs, to understand the safety profile of food ingredients in renal drug excretion. Twenty-eight food ingredients, including flavonoids, were screened. We used ascorbic acid (AA) to prevent curcumin oxidative degradation in our method. Twelve compounds, including apigenin, daidzein, fisetin, genistein, isorhamnetin, kaempferol, luteolin, morin, quercetin, curcumin, resveratrol, and ellagic acid, altered OATP4C1-mediated transport. Kaempferol and curcumin strongly inhibited OATP4C1, and the Ki values of kaempferol (AA(-)), curcumin (AA(-)), and curcumin (AA(+)) were 25.1, 52.2, and 23.5 µM, respectively. The kinetic analysis revealed that these compounds affected OATP4C1 transport in a competitive manner. Antioxidant supplementation was determined to benefit transporter interaction studies investigating the effects of curcumin because the concentration-dependent curve evidently shifted in the presence of AA. In this study, we elucidated the food-drug interaction via OATP4C1 and indicated the utility of antioxidant usage. Our findings will provide essential information regarding food-drug interactions for both clinical practice and the commercial development of supplements.

Usefulness of the Fibrosis-4 index and alanine aminotransferase at 1 year of nucleos(t)ide analog treatment for prediction of hepatocellular carcinoma in chronic hepatitis B patients.

AIM: Nucleos(t)ide analogs do not completely prevent hepatocellular carcinoma (HCC) in chronic hepatitis B virus infection. This study aimed to evaluate the dynamics of a non-invasive liver fibrosis marker, the Fibrosis-4 (FIB-4) index, for predicting HCC development. METHODS: Among a total of 882 chronically hepatitis B virus infection-infected patients who were treated with nucleos(t)ide analogs, 472 patients without HCC history whose FIB-4 at baseline and 1 year of treatment was obtained were evaluated for the incidence of HCC. RESULTS: The median FIB-4 was 2.00 at baseline and was significantly reduced to 1.58 at 1 year (P < 0.001), but the reduction was small at 2 years or later. When a receiver operating characteristic analysis of FIB-4 was performed to predict HCC within 5 years, the area under the curve of FIB-4 at 1 year was higher than that at baseline (0.676 vs. 0.599). The HCC incidence was significantly higher in patients with FIB-4 ≥1.58 than in those with FIB-4 <1.58 (14.8% vs. 3.6% at 10 years, P < 0.001). Additionally, an abnormal alanine aminotransferase (≥31 U/L) at 1 year was an independent risk for HCC. When a fibrosis and alanine aminotransferase-1 (FAL-1) score was evaluated as an applicable number of FIB-4 ≥1.58, and alanine aminotransferase ≥31 as 0, 1, and 2, the HCC risk in patients with score 2 was significantly higher than in those with score 1 or score 0 (24.1% vs. 9.8% vs. 0.7% at 10 years, P < 0.001). CONCLUSIONS: FIB-4 ≥1.58 and alanine aminotransferase ≥31 at 1 year of nucleos(t)ide analog was an independent risk factor for HCC development, and a score using these factors stratified the risk of HCC.

Phagocytosis by macrophages promotes pancreatic ß cell mass reduction after parturition in mice.

Elucidating the mechanism(s) modulating appropriate tissue size is a critical biological issue. Pancreatic ß cells increase during pregnancy via cellular proliferation, but how ß cells promptly decrease to the original amount after parturition remains unclear. Herein, we demonstrate the role and mechanism of macrophage accumulation in this process. In the final stage of pregnancy, HTR1D signaling upregulates murine ß cell CXCL10, thereby promoting macrophage accumulation in pancreatic islets via the CXCL10-CXCR3 axis. Blocking this mechanism by administering an HTR1D antagonist or the CXCR3 antibody and depleting islet macrophages inhibited postpartum ß cell mass reduction. ß cells engulfed by macrophages increased in postpartum islets, but Annexin V administration suppressed this engulfment and the postpartum ß cell mass reduction, indicating the accumulated macrophages to phagocytose ß cells. This mechanism contributes to both maintenance of appropriate ß cell mass and glucose homeostasis promptly adapting to reduced systemic insulin demand after parturition.

Preoperative Chemotherapy Followed by Hepatectomy for Potentially Resectable UICC7 Stage IIIA, IIIB Hepatocellular Carcinoma; A Phase II Clinical Trial.

BACKGROUND: The Japanese guideline for therapeutic strategy in HCC does not recognize any benefit of preoperative chemotherapy for potentially resectable hepatocellular carcinoma (HCC), and only upfront resec tion is recommended even for an advanced HCC. Data on preoperative chemotherapy for advanced HCC is still limited. Poor prognostic factors of HCC after resection are tumor more than 5 cm in diameter, multiple lesions, and gross tumor thrombosis, which constitute UICC7 Stage IIIA and IIIB HCC. There are no prospective studies about preoperative chemotherapy in these patients. AIM: To evaluate the benefit of preoperative chemotherapy for UICC7 Stage IIIA and IIIB potentially resectable HCC. DISCUSSION: Our recent study demonstrated that the 5-year overall survival rate (OS) of patients diagnosed as UICC7 Stage IIIA and IIIB who had received upfront resection was only 16.5%. In contrast, the 5-year OS of UICC7 Stage IIIA and IIIB initially unresectable patients who had achieved conversion from unresectable to resect able status under successful hepatic infusion chemotherapy prior to resection was as high as 61.3%. Additionally, recent studies reported transarterial chemoembolization achieved outcomes comparable with those of resection. Therefore, we believe that patients with UICC7 Stage IIIA and IIIB should be considered borderline resectable. To evaluate this hypothesis we registered the present phase II clinical trial to assess the benefit of preoperative chemo therapy followed by hepatectomy in potentially resectable UICC7 Stage IIIA and IIIB HCC patients.

High throughput LC/ESI-MS/MS method for simultaneous analysis of 20 oral molecular-targeted anticancer drugs and the active metabolite of sunitinib in human plasma.

Many types of oral molecular-targeted anticancer drugs are clinically used in cancer genomic medicine. Combinations of multiple molecular-targeted anticancer drugs are also being investigated, expecting to prolong the survival of patients with cancer. Therapeutic drug monitoring of oral molecular-targeted drugs is important to ensure efficacy and safety. A liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) has been used for simultaneous determination of these drugs in human plasma. However, the sensitivity of mass spectrometers and differences in the therapeutic range of drugs have rendered the development of simultaneous LC/ESI-MS/MS methods difficult. In this study, a simultaneous quantitative method for 20 oral molecular-targeted anticancer drugs and the active metabolite of sunitinib was developed based on the results of linear range shifts of the calibration curves using four ion abundance adjustment techniques (collision energy defects, in-source collision-induced dissociation, secondary product ion selected reaction monitoring, and isotopologue selected reaction monitoring). The saturation of the detector for the seven analytes was resolved by incorporating optimal ion abundance adjustment techniques. Furthermore, the reproducibility of this method was confirmed in validation tests, and plasma from patients was measured by this method to demonstrate its usefulness in actual clinical practice. This analytical method is expected to make a substantial contribution to the promotion of personalized medicine in the future.

Significance of Desmoplastic Histopathological Growth Pattern for Colorectal Liver Metastases Treated With Preoperative Chemotherapy.

BACKGROUND/AIM: The aim of this study was to evaluate hepatectomy cases that underwent preoperative chemotherapy to examine the relationship between the development of desmoplastic histopathological growth pattern (dHGP) and prognosis and recurrence and determine whether it is useful for evaluating preoperative chemotherapy. PATIENTS AND METHODS: A total of 133 cases with hepatic metastasis for colorectal cancer that underwent surgical resection. RESULTS: Of the 102 cases that underwent preoperative chemotherapy, 34 (33%) were determined to be dHGP positive, which was statistically significantly higher than the 2 of 31 cases (6.5%) that had not undergone preoperative chemotherapy. Regarding the 5-year recurrence-free survival, the dHGP group had a value of 50.3%, whereas the non-dHGP group had a value of 7.1%. For the 5-year overall survival, the dHGP group had a better prognosis than the non-dHGP group (57.6% vs. 37.1%, respectively), with a statistically significant difference. Univariate analysis of recurrence-free survival showed that the number of tumours, the Response Evaluation Criteria in Solid Tumors, and the presence or absence of dHGP were prognostic factors, whereas multivariate analysis showed that the presence or absence of dHGP was an independent prognostic factor. Univariate analysis of the overall survival showed that the number of tumours, the Response Evaluation Criteria in Solid Tumors, and presence or absence of dHGP were prognostic factors. Multivariate analysis showed that the presence or absence of dHGP was an independent prognostic factor. CONCLUSION: dHGP is useful as a new evaluation method for evaluating the efficacy of preoperative chemotherapy.

Prepancreatic postduodenal portal vein: a case report and literature review.

BACKGROUND: Among congenital anomalies of the portal venous system, prepancreatic postduodenal portal vein (PPPV) is very rare and has only been reported to date. Herein, we report a case of PPPV identified in preoperative examinations for hepatocellular carcinoma and a literature review. CASE PRESENTATION: A 63-year-old man was admitted to our hospital for treatment of a liver tumor. After examination, he was diagnosed with hepatocellular carcinoma with a diameter of 40 mm in segment 8. Contrast-enhanced computed tomography scan showed a portal vein passing between the duodenum and pancreas, hence called PPPV. At the hepatic hilus, the portal vein branched off in a complicated course with some porto-portal communications. We determined that anatomical resection with manipulation of the hepatic hilum in this case resulted in major vascular injury. Therefore, we performed partial liver resection, and the patient was discharged uneventfully on postoperative day 14. CONCLUSIONS: Although PPPV is an extremely rare congenital vascular variant, it is important to carefully identify vascular patterns preoperatively and to recognize the possibility of such an anomaly to avoid misidentification and inadvertent injuries during surgery.

Therapeutic Drug Monitoring of Blood Sirolimus and Tacrolimus Concentrations for Polypharmacy Management in a Lymphangioleiomyomatosis Patient Taking Two Cytochrome P450 3A Inhibitors.

Patients with lymphangioleiomyomatosis (LAM) and lung transplantations are treated with multiple drugs, such as tacrolimus, mycophenolate mofetil, prednisolone, and itraconazole, for long-term suppression of rejection response and prevention of infection. Additional drugs are required when lung transplant recipients develop graft complications. Therefore, managing polypharmacy is critical because of drug-drug interactions caused by various factors, including drug-metabolizing enzymes such as cytochrome P450 3A (CYP3A). The patient was a 48-year-old woman (height 144.9 cm and weight 38.4 kg) who underwent lung transplantation for LAM. Mycophenolate mofetil, tacrolimus (target blood concentration, 4.0-8.0 ng/mL), and prednisolone were administered for immunosuppression, and itraconazole and clarithromycin were administered to manage graft infection. The patient developed unilateral lymphedema, predominantly in the left leg; therefore, sirolimus was initiated with a target blood concentration of 3.0-5.0 ng/mL. In addition to 1.0 mg/day of sirolimus, tacrolimus (0.3 mg/day), itraconazole (100 mg/day), and clarithromycin (800 mg/day) were added. Blood sirolimus concentrations ranged from 18.8 to 36.9 ng/mL on days 6 to 9; thus, treatment with sirolimus was stopped because of over-target blood concentrations. Blood concentrations of sirolimus and tacrolimus were successfully managed without adverse events using therapeutic drug monitoring (TDM) and azole anti-fungal substitution of azithromycin instead of clarithromycin although sirolimus concentration was relatively lower compared to the target range. Thereby, frequent TDM, management of polypharmacy that influences CYP3A activity, and possibly CYP3A genotyping should be appropriately conducted for personalized medicine.

Impact of Interferon on the Prognosis of Hepatitis C Virus-Related Hepatocellular Carcinoma Patients with a Sustained Virological Response -An Additional Comparison Between Preoperative and Postoperative Sustained Virological Response.

BACKGROUND: Several studies have reported that interferon (IFN) therapy improves the prognosis of patients with hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC), especially for patients who have achieved a sustained virological response (SVR). We retrospectively evaluated the clinicopathological outcomes of patients who acquired an SVR through IFN therapy pre- or post-hepatectomy for treatment naïve HCC. METHOD: Among the 305 HCV-related HCC patients entered in this study, 59 patients (SVR group) achieved an SVR after IFN therapy and received hepatectomy either after or before achieving an SVR (n=36 and n=23, respectively), while the remaining 179 patients (control group) did not receive IFN therapy, or did not achieve an SVR through IFN therapy (n=67). RESULTS: In the SVR group, the overall survival (OS) and disease-free survival (DFS) rates were significantly higher than in the control group. We evaluated the prognosis of patients with an SVR achieved pre- or post-hepatectomy separately. There were no significant differences in OS and DFS. CONCLUSION: This result suggests that the prognosis of naïve HCC may be improved by additional INF therapy to achieve SVR status after hepatectomy.

Glutathione depletion results in S-nitrosylation of protein disulfide isomerase in neuroblastoma cells.

AIMS: Protein disulfide isomerase (PDI) is an essential enzyme involved in oxidative protein folding. PDI is S-nitrosylated in the brains of Alzheimer's disease patients, and S-nitrosylated PDI is considered one of main causes of Alzheimer's disease. However, the mechanisms underlying PDI S-nitrosylation have not yet been elucidated. Because glutathione (GSH) depletion is a pathological feature of Alzheimer's disease, we investigated the effect of GSH depletion on the S-nitrosylation level of PDI. MAIN METHODS: SH-SY5Y cells, which is a human derived neuroblastoma cells, were used in this study. Glutamate and buthionine sulfoximine (BSO) were used as GSH depletors. S-nitrosylated PDI was detected by biotin-switch assay. KEY FINDINGS: GSH depletion by glutamate, a cystine/glutamate antiporter xCT inhibitor, increased S-nitrosylated PDI at C343 in SH-SY5Y cells, and induced IRE1α phosphorylation. BSO, a γ-glutamylcysteine synthetase inhibitor, also increased S-nitrosylated PDI and phosphorylated IRE1α upon GSH depletion. Because S-nitrosylated PDI at C343 is stable in neuro cells, S-nitrosylated PDI by GSH depletion progresses to neurodegeneration by the induction of endoplasmic reticulum stress via phosphorylated IRE1α signaling from the early to late stage. Furthermore, treatment with neohesperidin, but not N-acetylcysteine (NAC), improved PDI S-nitrosylation level in GSH-depleted SH-SY5Y cells because nitrosylated compound of NAC induces PDI S-nitrosylation. SIGNIFICANCE: The results of our study provide a new insight into the mechanisms of neurodegeneration, and may be useful for the development of drugs for Alzheimer's diseases.

Switching to Tenofovir Alafenamide Fumarate in Chronic Hepatitis B Patients Who Had Detectable HBV DNA during Treatment with Entecavir.

Nucleos(t)ide analogues (NAs) suppress hepatitis B virus (HBV) replication, but the risk of hepatocellular carcinoma still remains. The presence of detectable HBV DNA in the serum during NA therapies for chronic hepatitis B patients has been reported to be associated with the risk of hepatocellular carcinoma. In this study, we investigated the antiviral effect of switching from entecavir (ETV) to tenofovir alafenamide fumarate (TAF) in chronic hepatitis B patients who had detectable HBV DNA in the serum at least once within a year. Among a total of 77 cases in 7 hospitals that switched NAs from ETV to TAF, 23 patients with detectable HBV DNA in a year before switching were analyzed. When the detection frequencies of HBV DNA in the 1st and 2nd years after switching to TAF were analyzed, they were significantly lower than those in the year before switching (68.8% vs. 34.1% for the 1st year and 21.3% for the 2nd year, P < 0.001 for both). The HBsAg decline tended to be larger after switching than before (-2.5% vs. -3.0% for 1st year and -3.1% for 2nd year), but the difference was not significant. One patient died of a cardiovascular event 11 months after the treatment switch, but no adverse effects due to TAF including renal function were observed. In conclusion, it was suggested that switching from ETV to TAF might be effective to suppress the HBV DNA level further in patients whose HBV DNA is detectable, even if at a very low level.

Bile Acid-Drug Interaction via Organic Anion-Transporting Polypeptide 4C1 Is a Potential Mechanism of Altered Pharmacokinetics of Renally Excreted Drugs.

Patients with liver diseases not only experience the adverse effects of liver-metabolized drugs, but also the unexpected adverse effects of renally excreted drugs. Bile acids alter the expression of renal drug transporters, however, the direct effects of bile acids on drug transport remain unknown. Renal drug transporter organic anion-transporting polypeptide 4C1 (OATP4C1) was reported to be inhibited by chenodeoxycholic acid. Therefore, we predicted that the inhibition of OATP4C1-mediated transport by bile acids might be a potential mechanism for the altered pharmacokinetics of renally excreted drugs. We screened 45 types of bile acids and calculated the IC50, Ki values, and bile acid−drug interaction (BDI) indices of bile acids whose inhibitory effect on OATP4C1 was >50%. From the screening results, lithocholic acid (LCA), glycine-conjugated lithocholic acid (GLCA), and taurine-conjugated lithocholic acid (TLCA) were newly identified as inhibitors of OATP4C1. Since the BDI index of LCA was 0.278, LCA is likely to inhibit OATP4C1-mediated transport in clinical settings. Our findings suggest that dose adjustment of renally excreted drugs may be required in patients with renal failure as well as in patients with hepatic failure. We believe that our findings provide essential information for drug development and safe drug treatment in clinics.

Tumour Budding as an Independent Prognostic Factor for Survival in Patients With Distal Bile Duct Cancer.

BACKGROUND/AIM: Surgical resection is the standard treatment for bile duct cancer. However, even when surgical resection is possible, the 5-year survival rate is reportedly 25.0-55.0%. Therefore, bile duct cancer is associated with poor prognoses. We conducted a clinicopathological investigation, focusing on the histological phenomenon of tumour budding, which has previously been reported to be correlated with the survival of patients with a variety of cancers. PATIENTS AND METHODS: To investigate the significance of tumour budding in distal bile duct cancer, we recruited 65 patients who underwent pancreatoduodenectomy at our institution between 1995 and 2011. Tumour budding was observed and evaluated using the 'hot spot method'. The 'low' budding group comprised 0-4 cell clusters and the 'high' budding group ≥5 cell clusters. Additionally, immunostaining was performed in high-budding areas. RESULTS: Tumour budding and stage were confirmed using a Cox proportional hazards model as independent prognostic factors for overall survival (p<0.05) in all patients. There was a significant association between budding and zinc finger E-box binding homeobox 1 expression, an endothelial-mesenchymal transition-induced transcription factor. In stage II cases, the prognosis was significantly worse in the 'high' budding group compared to that in the 'low' budding group. CONCLUSION: The budding phenomenon is an independent prognostic factor for patients with distal bile duct cancer. Understanding the mechanisms underlying tumour budding in distal bile duct cancer and its relationship with poor prognoses may be useful for the development of novel treatments for this disease.

Liberal Application of Portal Vein Embolization for Right Hepatectomy Against Hepatocellular Carcinoma: Strategy to Achieve Zero Mortality for a Damaged Liver.

BACKGROUND/AIM: Right hepatectomy and extended right hepatectomy (Rt-Hr) are identified as risk factors for the development of post-hepatectomy liver failure (PHLF). Although portal vein embolization (PVE) has made it possible to safely perform extended hepatectomy, to ensure safety, in our department, PVE is performed prior to Rt-Hr for hepatocellular carcinoma (HCC) regardless of the resection rate. This study aimed to retrospectively investigate the clinical course of PVE prior to Rt-Hr for HCC cases resected in our department and the appropriateness of our policy by clarifying complications and deaths. PATIENTS AND METHODS: The target period was from 2005 to 2020. Among the HCC cases resected at our hospital, those in which PVE was performed prior to Rt-Hr were included in this study. For PHLF, the definition of the International Study Group of Liver Surgery was used. The Clavien-Dindo classification was used for postoperative complications. Perioperative mortality was defined as the overall mortality within 30 days following surgery and surgery-related deaths within 90 days following surgery. RESULTS: A total of 79 cases were included. Rt-Hr was possible in all cases after PVE and there were no cases in which serious complications occurred after PVE. PHLF was found in 14 cases (17.7%)/5 cases (6.4%)/0 cases (0%) of Grade A/B/C, respectively. Regarding postoperative complications, there were no Grade IV, and Grade IIIa/IIIb were found in 13 cases (16.5%). There were no perioperative deaths. CONCLUSION: Our department's policy of performing PVE prior to all Rt-Hr was considered to be a safe and reasonable treatment strategy.

Evaluation of a Capillary Microsampling Device for Analyzing Plasma Lenvatinib Concentration in Patients With Hepatocellular Carcinoma.

BACKGROUND: The anticancer drug, Lenvima (lenvatinib), has severe side effects. Therapeutic drug monitoring helps ensure its efficacy and safety. Regular and optimally timed blood sampling is tough, especially when lenvatinib is self-medicated. Microsampling using the easy to handle Microsampling Wing (MSW) may help circumvent this problem. However, current lenvatinib detection methods are not sensitive enough to detect its concentrations in microsamples (<50-250 µL). Thus, the aim of this study was 2-fold (1) develop an analytic method to estimate plasma lenvatinib concentrations in microsamples and (2) verify whether this method works on micro (5.6 µL) blood plasma samples obtained clinically through MSW from patients with unresectable hepatocellular carcinoma (HCC). METHODS: A simple, highly sensitive, and specific liquid chromatography-electrospray ionization tandem mass spectrometry method was developed. Using this novel protocol, the trough blood plasma concentration of lenvatinib was measured for both blood sampled conventionally and that using MSW. Thirty-five venous whole blood samples were obtained from 11 patients with HCC. Furthermore, the stability of lenvatinib in MSW samples during storage was evaluated. RESULTS: The mean plasma lenvatinib concentration estimates were not significantly different between the MSW and conventional venous blood samples. CV for interday and intraday assays was low. Up to day 5, the lenvatinib concentration in the MSW samples was 85%-115% of the initial day concentration (when stored at 25°C or 4°C). The interference of endogenous matrix components in the human plasma was low. CONCLUSIONS: These results indicate that the novel mass spectrometry protocol accurately measures lenvatinib in human plasma and is reproducible. Thus, MSW could be a useful microsampling device for lenvatinib therapeutic drug monitoring in patients with HCC when used in combination with this novel liquid chromatography-electrospray ionization tandem mass spectrometry detection method.

Binding of Citrate-Fe3+ to Plastic Culture Dishes, an Artefact Useful as a Simple Technique to Screen for New Iron Chelators.

NaCT mediates citrate uptake in the liver cell line HepG2. When these cells were exposed to iron (Fe3+), citrate uptake/binding as monitored by the association of [14C]-citrate with cells increased. However, there was no change in NaCT expression and function, indicating that NaCT was not responsible for this Fe3+-induced citrate uptake/binding. Interestingly however, the process exhibited substrate selectivity and saturability as if the process was mediated by a transporter. Notwithstanding these features, subsequent studies demonstrated that the iron-induced citrate uptake/binding did not involve citrate entry into cells; instead, the increase was due to the formation of citrate-Fe3+ chelate that adsorbed to the cell surface. Surprisingly, the same phenomenon was observed in culture wells without HepG2 cells, indicating the adsorption of the citrate-Fe3+ chelate to the plastic surface of culture wells. We used this interesting phenomenon as a simple screening technique for new iron chelators with the logic that if another iron chelator is present in the assay system, it would compete with citrate for binding to Fe3+ and prevent the formation and adsorption of citrate-Fe3+ to the culture well. This technique was validated with the known iron chelators deferiprone and deferoxamine, and with the bacterial siderophore 2,3-dihydroxybenzoic acid and the catechol carbidopa.

Non-Achievement of Alanine Aminotransferase Normalization Associated with the Risk of Hepatocellular Carcinoma during Nucleos(t)ide Analogue Therapies: A Multicenter Retrospective Study.

Patients with a chronic hepatitis B virus (HBV) infection who are treated with nucleos(t)ide analogues (NAs) are still at risk for hepatocellular carcinoma (HCC), and it has been clinically questioned whether patients with a high risk of HCC can be identified efficiently. We aimed to clarify the risk factors associated with the development of HCC during NA therapies. A total of 611 chronically HBV-infected patients without a history of HCC, who were treated with NAs for more than 6 months (median 72 months), from 2000 to 2021, were included from 16 hospitals in the Tohoku district in Japan. Incidences of HCC occurrence were analyzed with clinical factors, including on-treatment responses. Alanine aminotransferase (ALT) normalization, based on the criteria of three guidelines, was analyzed with other parameters, including the age−male−ALBI−platelets (aMAP) risk score. During the observation period, 48 patients developed HCC, and the cumulative HCC incidence was 10.6% at 10 years. Non-achievement of ALT normalization at 1 year of therapy was mostly associated with HCC development when ALT ≤ 30 U/L was used as the cut-off (cumulative incidence, 19.9% vs. 5.3% at 10 years, p < 0.001). The effectiveness of the aMAP risk score at the start of treatment was validated in this cohort. A combination of an aMAP risk score ≥ 50 and non-achievement of ALT normalization could stratify the risk of HCC significantly, and notably, there was no HCC development in 103 patients without these 2 factors. In conclusion, non-achievement of ALT normalization (≤30 U/L) at 1 year might be useful in predicting HCC during NA therapies and, in combination with the aMAP risk score, could stratify the risk more precisely.

Assessing the incidence of complications and malignancies in the long-term management of benign biliary strictures with a percutaneous transhepatic drain.

ABSTRACT: Percutaneous drainage catheters (PDCs) are required for the management of benign biliary strictures refractory to first-line endoscopic treatment. While biliary patency after PDC placement exceeds 75%, long-term catheterization is occasionally necessary. In this article, we assess the outcomes of patients at our institution who required long-term PDC placement.A single-institution retrospective analysis was performed on patients who required a PDC for 10 years or longer for the management of a benign biliary stricture. The primary outcome was uncomplicated drain management without infection or complication. Drain replacement was performed every 4 to 12 weeks as an outpatient procedure.Nine patients (three males and six females; age range of 48-96 years) required a long-term PDC; eight patients required the long-term PDC for an anastomotic stricture and one for iatrogenic bile duct stenosis. A long-term PDC was required for residual stenosis or patient refusal. Drain placement ranged from 157 to 408 months. In seven patients, intrahepatic stones developed, while in one patient each, intrahepatic cholangiocarcinoma or hepatocellular carcinoma occurred.Long-term PDC has a high rate of complications; therefore, to avoid the need for using long-term placement, careful observation or early surgical interventions are required.