Monitoring Serum Levels of Sorafenib and Its N-Oxide Is Essential for Long-Term Sorafenib Treatment of Patients with Hepatocellular Carcinoma.

Sorafenib, an oral multi-kinase inhibitor, is the final therapy prior to palliative care for advanced hepatocellular carcinoma (HCC). However, due to its adverse effects, 20% of patients must discontinue sorafenib within 1 month after first administration. To identify ways to predict the adverse effects and administer the drug for longer periods, we explored the relationship between the duration of sorafenib treatment and the pharmacokinetics of sorafenib and its major metabolite, sorafenib N-oxide. Twenty-five subjects enrolled in the study were divided into two groups: patients with dosage reduced or withdrawn due to adverse effects (n = 8), and patients with dosage maintained for 1 month after initial administration (n = 17). We evaluated early sorafenib accumulation as the area under the curve of sorafenib and sorafenib N-oxide concentrations during days 1-7 (AUC(sorafenib) and AUC(N-oxide), respectively). Inter-group comparison revealed that AUC(N-oxide) and AUC ratio (AUC(N-oxide)/AUC(sorafenib)) were significantly higher in the dosage reduction/withdrawal group (P = 0.031 and P = 0.0022, respectively). Receiver operating characteristic analysis indicated that AUC(N-oxide) and AUC ratio were reliable predictors of adverse effects. When patients were classified by cut-off points (AUC(N-oxide:) 2.0 µg ∙ day/mL, AUC ratio: 0.13), progression-free survival was significantly longer in patients with AUC(N-oxide) ≤ 2.0 µg ∙ day/mL (P = 0.0048, log-rank test). In conclusion, we recommend to simultaneously monitor serum levels of sorafenib and its N-oxide during the early stage after the first administration, which enables us to provide safe and long-term therapy for each HCC patient with sorafenib.

A quantitative HPLC-UV method for determination of serum sorafenib and sorafenib N-oxide and its application in hepatocarcinoma patients.

Sorafenib, an oral multi-kinase inhibitor, has been approved for treatment of advanced renal-cell and hepatocellular carcinoma (HCC). However, 20% of HCC patients taking sorafenib are forced to withdraw due to adverse effects within one month after administration. Orally administered sorafenib is oxidatively metabolized, predominantly by cytochrome P450 3A4 (CYP3A4), in small-intestinal mucosa or liver. We aimed to characterize the CYP3A4-mediated metabolism of sorafenib in HCC patients and explore the contribution of the major metabolite sorafenib N-oxide to adverse effects and therapeutic efficacy. We have therefore developed a method for quantitative determination of sorafenib and its N-oxide in the present study. To optimize the preanalytical procedure, we initially ascertained the solubility of the analytes. Because they are lipophilic, solvents containing more than 40% acetonitrile were required for efficient recovery. The pretreatment procedure that we ultimately developed consists of acetonitrile precipitation, followed by extraction using octadecyl silyl-silica gel to eliminate water-soluble and hydrophilic components of serum. Application of this procedure before HPLC enabled accurate and reproducible quantitation of analytes in a linear range from 0.03 to 30 µg/mL. After characterizing the peaks in the HPLC-ultraviolet chromatogram obtained from a medicated patient by LC-tandem mass spectrometry, we applied this method to HCC patients taking sorafenib, showing large inter-individual differences in the pharmacokinetic profile. In conclusion, our assay system should be useful for follow-up of patients taking sorafenib and for exploring the association between the pharmacokinetics of sorafenib and its N-oxide and the adverse effects or therapeutic efficacy.

Increased expression of (pro)renin receptor in the remnant kidneys of 5/6 nephrectomized rats.

Recent studies have revealed that (pro)renin receptor ((P)RR), a newly identified member of the renin-angiotensin system, is associated with renal organ damage. However, there is little information regarding the regulation of (P)RR expression in various pathophysiological conditions. We therefore examined the expression of (P)RR in the remnant kidneys of rats with renal mass ablation due to 5/6 nephrectomy by quantitative RT-PCR, Western blot analysis and immunohistochemistry. Expression levels of (P)RR mRNA were significantly increased in the remnant kidneys at day 56 after nephrectomy, when compared with sham operation (about 1.6-fold, P=0.001). Western blot analysis showed that expression levels of (P)RR protein were greatly increased in the remnant kidneys at day 56, compared with sham operation (about 7.9-fold, P=0.02). The renal tubular cells were immunostained with anti-(P)RR antibody in both 5/6 nephrectomized rats and sham operated rats. The glomeruli were sporadically immunostained in 5/6 nephrectomized rats, but not in sham operated rats. These findings indicate that the intra-renal (P)RR expression is increased in the remnant kidneys of 5/6 nephrectomized rats, and suggest that (P)RR may contribute to the renal injury.

Gene expression of (pro)renin receptor is upregulated in hearts and kidneys of rats with congestive heart failure.

Recent studies have revealed that (pro)renin receptor ((P)RR), a newly identified member of the renin-angiotensin system, was associated with organ damage in the kidney. However, there has been little information for (P)RR in hearts. To investigate the regulation of (P)RR in heart failure, we examined the expression of (P)RR in hearts and kidneys of rats with congestive heart failure (CHF) due to coronary ligation by quantitative RT-PCR and immunohistochemistry. Significantly increased levels of (P)RR mRNA were found in the atrium, right ventricle, non-infarcted part of left ventricle, infarcted part of left ventricle and kidney of CHF rats, when compared with sham operated rats (about 1.6-fold, 1.4-fold, 1.6-fold, 1.7-fold and 1.5-fold, respectively). Expression levels of mRNAs encoding renin and angiotensinogen in these heart and kidney tissues were also increased in the CHF rats. Immunohistochemistry showed positive (P)RR immunostaining in the myocardium, the renal tubular cells, and vascular smooth muscle and endothelial cells in the heart and the kidney. The renal tubular cells were more intensely immunostained in CHF rats than in sham operated rats. These findings suggest that the expression of (P)RR is increased in the hearts and kidneys of rats with heart failure, and that (P)RR may contribute to heart failure.

Increased gene expression of urotensin II-related peptide in the hearts of rats with congestive heart failure.

Urotensin II-related peptide (URP) is a novel endogenous ligand for urotensin II receptor (UT-R). To investigate the pathophysiological role of URP in heart failure, we examined URP, UII and UT-R expression in hearts and kidneys of rats with congestive heart failure due to coronary ligation by quantitative RT-PCR and immunocytochemistry. Significantly increased expression levels of URP mRNA were found in the atrium, the right ventricle and the infarcted part of left ventricle of heart failure rats, when compared with sham-operated rats (about 2.2-fold, 2.7-fold and 3.9-fold, respectively). Expression levels of UII mRNA in the heart were about 10% of URP mRNA, and were slightly increased only in the infarcted part of left ventricle of heart failure rats, when compared with sham-operated rats. The expression levels of UT-R mRNA were increased in the atrium of heart failure rats. There was no significant change of URP, UII and UT-R mRNA expression levels in the kidney between heart failure and sham-operated rats. The myocardium was diffusely immunostained with URP in both rats. The blood vessels in the heart were positively immunostained with URP in heart failure rats, but not in sham-operated rats, whereas they were positively immunostained with UT-R in both rats. These findings suggest that the expression of URP, UII and UT-R is enhanced in failing heart, and the UII/URP/UT-R system has important pathophysiological roles in the progression of heart failure.