Validation of an Isavuconazole High-Performance Liquid Chromatography Assay in Plasma for Routine Therapeutic Drug Monitoring Applications.

BACKGROUND: Isavuconazole is a triazole antifungal agent for treatment of invasive aspergillosis and mucormycosis. At present, it is unclear whether a therapeutic drug monitoring (TDM) of isavuconazole would be necessary. The aim of the investigation was to validate a high-performance liquid chromatography (HPLC) assay for routine applications. METHODS: An HPLC assay for routine determination of isavuconazole in plasma has been adapted and validated. The assay used the reagents and HPLC column provided by the ChromSystems HPLC Kit for TDM of itraconazole, posaconazole, and voriconazole. Isocratic flow rate was set at 1.0 mL/min. Detection was performed using a fluorescence detector with excitation wavelength set at 261 nm and emission wavelength set at 366 nm. RESULTS: The assay was linear between 0.15 and 10 mg/L with intraday and interday imprecision and accuracy <10% (<20% at lower limit of quantification). The method was applied to routine TDM of 7 patients after hematopoietic stem cell transplantation (n = 31 samples). In these patients, trough levels ranged from 0.45 to 3.06 mg/L (median 1.44 mg/L). CONCLUSIONS: A robust and simple HPLC assay of isavuconazole in plasma for routine TDM applications is presented.

Everolimus in combination with mycophenolate mofetil as pre- and post-transplantation immunosuppression after nonmyeloablative hematopoietic stem cell transplantation in canine littermates.

The mammalian target of rapamycin inhibitor everolimus (RAD001) is a successfully used immunosuppressant in solid-organ transplantation. Several studies have already used RAD001 in combination with calcineurin inhibitors after hematopoietic stem cell transplantation (HSCT). We investigated calcineurin inhibitor-free pre- and post-transplantation immunosuppression of RAD001 combined with mycophenolate mofetil (MMF) in a nonmyeloablative HSCT setting. After nonmyeloablative conditioning with 2 Gy total body irradiation, 8 dogs received HSCT from dog leukocyte antigen-identical siblings. Immunosuppressives were given at doses of 1.5 mg RAD001 twice daily from day -1 to +49, then tapered until day +56, and 20 mg/kg MMF from day 0 to +28, then tapered until day +42. An historical cyclosporin A (CsA)/MMF regimen was used in the control group. All dogs engrafted. Median platelet nadir amounted in all dogs to 0 × 10(9)/L (median, day +10; duration <50 × 10(9)/L, 22 days) and median leukocyte nadir was 1.0 × 10(9)/L (range, .1 to 2.5 × 10(9)/L; median, day +13). Eventually, 5 of 8 (63%) animals rejected their grafts. Two dogs died of infections on day +19 and +25. Pharmacokinetics of RAD001 and MMF showed median trough levels of 19.1 (range, 10.5 to 43.2) µg/L and .3 (.1 to 1.3) mg/L, respectively. The median area under the curve was 325 (range, 178 to 593) µg/L × hour for RAD001 and 29.6 (range, 7.9 to 40.5) ng/L × hour for MMF. All dogs developed clinically mucosal viral infections during the clinical course. Compared with the control group, the level of toxicities for RAD001/MMF increased in all qualities. Combined immunosuppression of RAD001 and MMF after nonmyeloablative HSCT is associated with significant toxicities, including a prolonged platelet recovery time as well as increased infections compared to the CsA/MMF regimen.

Everolimus in combination with cyclosporin a as pre- and posttransplantation immunosuppressive therapy in nonmyeloablative allogeneic hematopoietic stem cell transplantation.

Everolimus (RAD001) is an mTOR inhibitor that has been successfully used as an immunosuppressant in solid-organ transplantation. Data in allogeneic hematopoietic stem cell transplantation (HSCT) is limited. This study aimed to investigate pharmacokinetics, safety, and efficacy of RAD001 in a canine allogeneic HSCT model. First, pharmacokinetics of RAD001 were performed in healthy dogs in order to determine the appropriate dosing. Doses of 0.25 mg RAD001 twice daily in combination with 15 mg/kg cyclosporin A (CsA) twice daily were identified as appropriate starting doses to achieve the targeted range of RAD001 (3-8 µg/L) when orally administered. Subsequently, 10 dogs were transplanted using 2 Gy total body irradiation (TBI) for conditioning and 0.25 mg RAD001 twice daily plus 15 mg/kg CsA twice daily for pre- and posttransplantation immunosuppression. Seven of the 10 transplanted dogs were maintained at the starting RAD001 dose throughout the study. For the remaining 3 dogs, dose adjustments were necessary. RAD001 accumulation over time did not occur. All dogs initially engrafted. Five dogs eventually rejected the graft (weeks 10, 10, 13, 27, and 56). Two dogs died of pneumonia (weeks 8 and 72) but were chimeric until then. Total cholesterol rose from median 4.1 mmol/L (3.5-5.7 mmol/L) before HSCT to 6.0 mmol/l (5.0-8.5 mmol/l) at day 21 after HSCT, but remained always within normal range. Changes in creatinine and triglyceride values were not observed. Long-term engraftment rates were inferior to sirolimus/CsA and mycophenolate mofetil (MMF)/CsA regimen, respectively. RAD001/CsA caused a more pronounced reduction of platelet counts to median 2 × 10(9)/L (range: 0-21 × 10(9)/L) and longer time to platelet recovery of 21 days (range: 14-24 days) compared with MMF/CsA. CsA c(2h) levels were significantly enhanced in the RAD001/CsA regimen, but c(0h) and area under the curve from 0 to 12 hours (AUC(0-12h)) values did not differ compared with an MMF/CsA immunosuppression. In summary, immunosuppression consisting of RAD001 and CsA is well tolerated but not as efficient as with other established immunosuppressants in a canine nonmyeloablative HSCT regimen. Hence, our study does not support the application of RAD001/CsA as standard practice in this setting.

Penetration of moxifloxacin into the human pancreas following a single intravenous or oral dose.

OBJECTIVES: Failure to prevent secondary infectious complications in acute necrotizing pancreatitis (ANP) is attributable in part to the limited penetration of antimicrobial drugs. As newer quinolones are particularly attractive owing to their antimicrobial activity, for the first time we studied the penetration of moxifloxacin into pancreatic tissue in patients. PATIENTS AND METHODS: In this prospective, non-comparative clinical trial, 60 patients undergoing elective pancreas resection received a single oral or intravenous (iv) dose of 400 mg moxifloxacin for perioperative antimicrobial prophylaxis. The concentration of moxifloxacin was measured in samples taken from blood and from pancreatic tissue at the beginning and at the end of resection. RESULTS: Mean moxifloxacin concentrations in pancreatic tissue following iv or oral administration were 3.1+/-0.9 and 2.7+/-1.4 mg/kg at 3-3.7 h post-dose (first sampling) and 3.6+/-1.5 and 3.1+/-1.8 mg/kg at 4.3-5.3 h post-dose (second sampling), respectively. Corresponding mean plasma concentrations of moxifloxacin were 1.8+/-0.5 and 1.2+/-0.6 mg/L (first sampling) and 1.5+/-0.4 and 1.0+/-0.5 mg/L (second sampling), respectively. From first to second sampling, the mean tissue-to-plasma ratios varied from 1.8+/-0.6 to 2.6+/-1.2 (iv) and from 2.4+/-0.8 to 3.1+/-1.2 (oral). Pancreatic tissue concentrations of moxifloxacin exceeded the MIC90 for the relevant pathogens covered by moxifloxacin for at least 5 h after dosing. CONCLUSIONS: Moxifloxacin has been demonstrated to penetrate efficiently into human pancreatic tissue following iv or oral administration. From a pharmacological perspective, moxifloxacin appears to be promising for prophylaxis and treatment of local pancreas infections. Whether it is beneficial in the prevention and therapy of infectious complications in patients with ANP should be investigated in a controlled clinical trial.

The penetration of moxifloxacin into the pancreas of male rats in experimental acute necrotizing pancreatitis.

Infectious complications of acute necrotizing pancreatitis (ANP) determine the extent of multiorgan failure and account for 80% of deaths. Prophylactic use of antibiotics can reduce the incidence of these complications. However, the actual indication as well as choice of drug remains a controversial matter. We examined the penetration of moxifloxacin, a new broad-spectrum fluoroquinolone, in healthy and inflamed pancreatic tissue in rats after inducing ANP. The concentration of moxifloxacin in pancreatic tissue and serum was determined 10, 30, 60 and 240 min after the administration of moxifloxacin (5 mg/kg, i.v.). Mean serum concentrations 10 min after administration in rats with ANP were 1,886 ng/ml versus 1,805 ng/ml in healthy controls, and these values decreased to 350 versus 222 ng/ml, respectively, after 240 min. Corresponding concentrations in pancreatic tissue were in the mean 2-3 times higher.

Expression of multidrug resistance proteins in rat and human chronic pancreatitis.

BACKGROUND AND AIMS: The expression of the ABC-transporters MDR-1, MRP1, and MRP-2 was investigated in healthy pancreas and in chronic pancreatitis tissue samples in rats and humans to evaluate their possible involvement in a multidrug resistance of the pancreas with consequences for the pharmacologic treatment of pancreatic diseases. METHODS: Human pancreatic tissue samples of healthy tissue and chronic pancreatitis were collected during pancreas surgery. In rats, the time-course of the expression of transporter proteins was studied 14, 28, and 56 days after experimental induction of chronic pancreatitis. The expression of MDR-1, MRP-1, MRP-2, and furthermore, LRP and PAP was investigated by RT-PCR, Real Time TaqManPCR, and immunohistochemistry. RESULTS: In rat pancreas, MDR-1 (P-gp) and MRP-1 but in human pancreas MDR-1 (P-gp), MRP-1 and MRP-2 were found to be expressed. Chronic pancreatitis lead to an increased transcription of mRNA of MDR-1 (rat and human) and much lower, MRP-2 (human). CONCLUSIONS: The expression of P-gp and related transporters could have impact on the metabolism, distribution, and availability of various compounds, including drugs, in the pancreas. The results indicate that this could be more pronounced in chronic pancreatitis.

Atypical pharmacokinetics and metabolism of mycophenolic acid in a young kidney transplant recipient with impaired renal function.

A juvenile, female renal transplant recipient suffered two acute rejection episodes: the first on posttransplant day 31 while taking cyclosporine, prednisone, and mycophenolate mofetil (MMF); and the second on posttransplant day 67, when she was taking tacrolimus, prednisone, and MMF. Dosage of MMF was initially started at 2 g/d (corresponding to 600 mg MMF/m(2) twice daily.), but was reduced to 250 mg/d to 500 mg/d after severe diarrhea and a paralytic ileus on posttransplant day 16. During therapy with tacrolimus, prednisone, and MMF, predose plasma mycophenolic acid (MPA) concentrations varied from 1.1 mg/L to 8.2 mg/L (median 3.0 mg/L). On posttransplant day 91, a 12-hour pharmacokinetic profile was obtained. The concentrations of MPA and its metabolites were determined with a validated high-performance liquid chromatography (HPLC) procedure. After oral MMF (250 mg) administration, the MPA concentration showed an atypical decline from a predose concentration of 6.0 mg/L to a value of 3.8 mg/L at 75 minutes postdose, and 3.4 mg/L at 6 hours postdose, before returning to 6.0 mg/L after 12 hours. The 12-hour area under the concentration-time curve (AUC) values for MPA and its major metabolite the phenolic glucuronide MPAG were 55.1 mg.h/L and 800 mg.h/L, respectively. An unusually high concentration (12-h AUC, 165 mg.h/L) of the phenolic glucose conjugate of MPA was found. The apparent renal clearance of MPAG was only 2.2 mL/min. Her creatinine clearance was 30 mL/min. MPAG clearances have been reported to range from approximately. 5.5 mL/min to 35 mL/min at a creatinine clearance of approximately 30 mL/min in renal transplant recipients. The authors' findings suggest that conjugation and clearance of MPA through the kidney is strongly impaired in this patient. The relatively high predose MPA concentrations could result from an enhanced enterohepatic circulation of MPA and its metabolites.