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1. A procedure for the isolation and primary culture of hepatocytes from goat and cattle is described. Hepatocyte culture performance was monitored for 51 h by measuring viability, cytochrome P-450 maintenance, dealkylation of scoparone and ethylmorphine, and glucuronidation of phenol red. 2. Culture medium composition is discussed in relation to differences between splanchnic blood composition of ruminant and monogastric animal species. Main differences are in glucose and volatile fatty acid concentrations. Modified Williams' E culture medium did not yield higher culture performance than non-modified Williams' E. 3. Coating of culture dishes with either collagen or fibronectin did not improve culture performance. 4. Williams' E, although developed for rodent cells, proves to be a suitable basal medium for ruminant hepatocytes. In this medium, culture quality is high for at least several days. 5. In cultured goat hepatocytes, biotransformation rate for scoparone amounted to 20 nmol/mg protein per h, for ethylmorphine 96 nmol/mg protein per h and for phenol red 2 nmol/mg protein per h. Biotransformation activity in cow hepatocytes is approximately half that in goat hepatocytes.

Background GLPG0634 is an orally-available, selective inhibitor of Janus kinase 1 (JAK1). JAKs are critical components in signaling pathways for a number of cytokines and growth factors, including those involved in the disease process of rheumatoid arthritis (RA). Other non-selective JAK inhibitors have shown long-term efficacy in RA trials with an early onset of action, but doses and thereby efficacy are limited by side effects. By specifically targeting JAK1, treatment with GLPG0634 is expected to result in a cleaner safety profile while maintaining the rapid onset of clinical efficacy. Objectives Evaluate the short-term efficacy and safety of GLPG0634 in RA patients with insufficient response to methotrexate (MTX) alone. Methods A double-blind, placebo-controlled Proof-of-Concept trial in patients with active RA, showing an insufficient response to MTX, was conducted. Twenty-four patients with moderate to severe disease received GLPG0634 200 mg daily, half as once-daily regimen (200 mg q.d.) and half as twice-daily regimen (100 mg b.i.d.), and 12 patients received a matching placebo for a period of four weeks, while continuing to take their stable background therapy of MTX. Results Patient and disease characteristics were comparable for all three dose groups, with DAS28 (CRP) disease activity scores between 6.3 and 6.6. In each dose group, 11 out of 12 patients were female. The patients were well exposed to GLPG0634, with the same pharmacokinetics as previously observed in healthy volunteers. GLPG0634 was found well tolerated and safe with a rapid onset and high level of efficacy. Considering the small sample size, no clinically relevant differences were observed among the 100 mg b.i.d. and the 200 mg q.d. dose regimens. GLPG0634 met the primary endpoint of significant improvement in ACR20 response rate. Overall ACR20 responses were observed in 83% of patients receiving GLPG0634 vs. 33% of patients receiving placebo (p<0.01). GLPG0634 showed impressive results in secondary efficacy endpoints, such as the DAS28 (-2.5), and in reductions of serum C-reactive protein levels (-24.4 mg/L), both significant changes vs. placebo (p<0.0001). All patients completed the trial and no treatment-emergent safety signals were reported. No severe adverse events were reported in patients receiving GLPG0634. Instead of anemia, a modest improvement in hemoglobin was observed. In contrast to observations with JAK inhibitors with other selectivity profiles, no increases in LDL/cholesterol and no liver effects (ALT/AST) were observed in this trial. Conclusions These early clinical results are the first to demonstrate that selective inhibition of JAK1 is efficacious and safe for the treatment of RA. Consistent with the lack of inhibition of JAK2, no anemia was observed. Remarkable results include the high level of response within 4 weeks and the absence of effects on LDL/lipid. An extended GLPG0634 dose-range finding trial is now being conducted to further define the optimal doses for efficacy and safety for longer term studies. D...

Hydroxylation and acetylation of sulphadimidine (SDD) and the deacetylation of N4-acetyl SDD was investigated in cultured hepatocytes from male and female rats, from male and female goats and from female sheep and cattle. Significant sex differences were observed for hydroxylation of SDD in hepatocytes from rat and goat. In goat, sheep and cow hepatocytes, the hydroxylation pathway is relatively important, whereas in rat hepatocytes, acetylation is predominant. Hepatocytes of all four species deacetylated N4-acetyl SDD. In ruminant hepatocytes, deacetylating activity was of considerable importance, whereas in rat hepatocytes, it appeared a minor pathway of metabolism. Similar to the in vivo situation, formation of N4-acetyl SDD in cultured hepatocytes results from an equilibrium of acetylation and deacetylation. A good correlation was found between results in isolated hepatocytes and previous findings in vivo, both in levels of species-related activities and in acetylation-hydroxylation ratios. In conclusion, cultured hepatocytes appear a useful in vitro model to study comparative sulfonamide metabolism.