Pharmacokinetics of the Experimental Non‐Nucleosidic DNA Methyl Transferase Inhibitor N‐Phthalyl‐l‐Tryptophan (RG 108) in Rats

Abstract

DNA methyl transferase (DNMT) inhibitors can re-establish the expression of tumour suppressor genes in malignant diseases, but might also be useful in other diseases. Inhibitors in clinical use are nucleosidic cytotoxic agents that need to be integrated into the DNA of dividing cells. Here, we assessed the in vivo kinetics of a non-nucleosidic inhibitor that is potentially free of cytotoxic effects and does not require cell division. The non-specific DNMT inhibitor N-phthalyl-L-tryptophan (RG 108) was injected subcutaneously in rats. Blood was drawn 0, 0.5, 1, 2, 4, 6, 8 and 24 hr after injection and RG 108 in plasma was measured by high-performance liquid chromatography coupled to mass spectrometry. Trough levels and area under the curve (AUC) were significantly higher with multiple-dose administration and cytochrome inhibition. In this group, time to maximal plasma concentration (t max , mean AE S.D.) was 37.5 AE 15 min., terminal plasma half-life was approximately 3.7 h (60% CI: 2.1-15.6 h), maximal plasma concentration (C max ) was 61.3 AE 7.6 lM, and AUC was 200 AE 54 lmolÁh/l. RG 108 peak levels were not influenced by cytochrome inhibition or multiple-dose administration regimens. Maximal tissue levels (C max in lmol/kg) were 6.9 AE 6.7, 1.6 AE 0.4 and 3.4 AE 1.1 in liver, skeletal and heart muscle, respectively. We conclude that despite its high lipophilicity, RG 108 can be used for in vivo experiments, appears safe and yields plasma and tissue levels in the range of the described 50% inhibitory concentration of around 1 to 5 lM. RG 108 can therefore be a useful tool for in vivo DNMT inhibition.DNA methylation plays an important role in transcriptional regulation. Aberrant transcriptional regulation is an important contributing factor to the pathogenesis of many diseases from cancer to heart failure, and its restitution represents a therapeutic strategy [1,2]. This might be achieved by influencing DNA methylation with DNA methyl transferase (DNMT) inhibitors, as already established in oncology.Reactivation of hypermethylated tumour suppressor genes by DNMT inhibition has entered clinical routine for the treatment of myelodysplastic syndromes and cutaneous lymphomas [3]. Inhibition of maintenance methylation during cell division leads to the dilution of DNA methylation and in turn to a progressive reactivation of tumour suppressor genes. Several nucleosidic inhibitors of DNMTs have been developed to serve this goal [4], but the concept cannot be translated directly from malignant diseases to others involving transcriptional dysregulation. To be active, nucleosidic inhibitors such as azacytidine, decitabine and zebularine have to be integrated into the DNA [5] and require high DNA turnover. This is the case in malignantly transformed cells, but not in terminally differentiated cells like neurons or cardiac myocytes. Moreover, all these inhibitors exert myelosuppressive off-target effects [6]. As in malignant diseases, highly active compounds are needed for short-term therapy, toxic side effects ar...

Negative Evidence for a Functional Role of Neuronal DNMT3a in Persistent Pain

“…At the time we conducted our behavioral experiments, the half-life of RG108 had only been measured in vitro , where it was reported to be approximately 20 days ( Brueckner et al, 2005 ). Today, an in vivo study suggests that RG108 delivered subcutaneously may only be active for about 4 h ( Schneeberger et al, 2016 ). In our experimental context, its effects were measurable for slightly longer than that, with reduced 5-mC levels still being observed approximately 24 h after the last intrathecal injection ( Figure 6B ).…”

Senescent Markers Expressed by Periodontal Ligament-Derived Stem Cells (PDLSCs) Harvested from Patients with Periodontitis Can Be Rejuvenated by RG108

“…Since in our experimental setting, pPDLSCs almost not expressing PPARγ, significantly upregulated it after RG108 treatment, we interpreted this as a positive response leading to the restoration of stemness. According to literature data, RG 108 can be used for in vivo experiments, proving safe [ 57 ], thus this molecule could potentially be a useful tool to treat periodontitis in situ, since its injection could restore and rejuvenate PDLSCs, improving their stem cell potency and protecting them from senescence.…”

Impact of local anesthetics on epigenetics in cancer

“…The reversal of cancer-associated epigenetic dysregulations represents one possible antineoplastic strategy. Various demethylating molecules were characterized at the preclinical level (as exemplified by curcumin, (−)-epigallocatechin-3gallate, N-phthalyl-tryptophan and zebularine) (90)(91)(92)(93)(94), and two agents (5-azacytidine and decitabine) have been approved by the FDA and EMA to treat patients with myelodysplastic syndrome or acute myeloid leukemia. These agents inhibit DNMT and hence reduce the global DNA methylation level in cancer cells.…”