Registration Dossier

Diss Factsheets

Administrative data

Link to relevant study record(s)

Description of key information

A toxicokinetic assessment is available. The assessment includes general inforamation, a toxicological profile and details on absorption, distribution, metabolism and elimination. 

Key value for chemical safety assessment

Bioaccumulation potential:
no bioaccumulation potential

Additional information

General information

Thymidine is ubiquitously present in nature and exists in all living organisms (and DNA viruses). The substance is one of the pyrimidine deoxynucleosides and part of double-stranded deoxyribonucleic acid (DNA), the main repository of genetic information in life. Besides in DNA, Thymidine may also occur in the T-loop of tRNA.

Thymidine is formed by the combination of Thymine with deoxyribose. Mainly synthesized during the S-phase of the cell cycle, Thymidine is incorporated into DNA during cell replication. The salvage pathways to Thymidine synthesis involve the enzyme thymidine kinase or pyrimidine dehydrogenases and in a first step lead to the formation of β-aminoisobutyrate. β-aminoisobutyrate can then be further broken down into intermediates eventually leading to formation of water and CO2 via the citric acid cycle.

In the chemical industry Thymidine is manufactured in a fermentation process. The substance is used as intermediate.

Toxicological information

Thymidine was assessed in acute oral toxicity and acute dermal toxicity studies according to respective EU Methods, OECD and OPPTS Guidelines. The substance was shown to be practically non-toxic. The acute oral and acute dermal LD50-values were determined to be > 2000 mg/kg bw (limit dose). The LD0 value in both studies was 2000 mg/kg bw. A study on acute inhalation toxicity was waived.

Potential irritation/corrosion was assessed in a sequential testing strategy using tiered approach. First, physical chemical properties and chemical reactivity were assessed. An in vivo skin irritation study was carried out, as no signs of irritation and corrosion were observed in the acute dermal toxicity study. For assessment of potential eye irritation and corrosion first an in vitro test was carried out followed by in vivo testing. Based on the results obtained Thymidine is not considered corrosive or irritating to the skin or eye. Results obtained in a local lymph node assay revealed no skin sensitization propterties. Thymidine is considered not a skin sensitizer.

Repeated dose toxicity was assessed using data on the L-isomer of Thymidine in a read-across approach. The read-across between the two enantiomers is justified. Available studies using L-Thymidine include studies on mice, rats, and monkeys, with study duration ranging from 28-days up to 9 month (EMEA, 2007). In all studies L-Thymidine was administered via oral gavage. No adverse effects were observed in the 28-day and six month studies in rats or in the 28-day and 90-day studies in mice. Except for equivocal axonopathic findings in monkeys, occasional emesis, and minor changes in body weight and food consumption, no evidence of target organ toxicity was identified either in a 28-day or nine month study. The NOAEL in rats was determined to be 2000 and 1000 mg/kg bw/day in the 28-day and six month repeated dose toxicity studies, respectively. The NOAEL in mice was determined to be 2000 and 3000 mg/kg bw/day in the 28-day and 90-day repeated dose toxicity studies. In monkeys the nine month NOAEL was determined to be 500 mg/kg bw/day.

Thymidine did not induce gene mutations by base pair changes or frameshifts in the genome of bacteria, tested in a bacterial reverse mutation assay up to the limit concentration with and without metabolic activation. Further, Thymidine did not induce structural chromosome aberrations in a chromosome aberration assay in Chinese Hamster lung cells, tested up to cytotoxic concentrations with and without metabolic activation. In an in vitro mammalian cell gene mutation test performed with in Chinese hamster ovary cells no genotoxic effects were observed following exposure over five hours without and with metabolic activation, however, when treatment without metabolic activation was prolonged, significant increases in mutant frequency were observed. Taking all available information available into account including data published in the literature and studies using L-Thymidine in a read-across approach, Thymidine is considered non genotoxic.

Reproductive and developmental toxicity was assessed using data on the analog L-Thymdine in a read-across approach. Five studies were conducted to evaluate the effects of L-Thymidine on fertility in male and female rats, on embryo-fetal development in female rats and New Zealand White rabbits, and on multiple generation development in male and female rats (EMEA, 2007). No reproductive or developmental toxicity was observed in rats or rabbits. Although slightly lower pregnancy rates were noted in the initial combined segment I and II study in rats, this finding was proven to not be attributable to the administration of L-Thymidine in the two subsequent confirmatory studies. The single abortion and two premature deliveries in rabbits dosed at 1,000 mg/kg/day were more likely due to maternal toxicity, i.e., reduced food consumption, reduced body weight, and abnormal feces. In fact, abortion and premature delivery are not uncommon in rabbits (Feussner, 1992).

In summary Thymidine is considered practically non-toxic, has no irritation or corrosive properties, is not a sensitizer and is not genotoxic. Based on available data Thymidine is not classified or labeled according to Regulation (EC) No 1272/2008 (CLP) or Directive 67/548/EEC (DSD).

Toxicokinetic assessment

Thymidine is a white to off-white crystalline powder at ambient temperature. The substance has a molecular weight of 242.23 g/mol, exerts a low vapor pressure (8.84E-10 Pa at 25 °C), has a low logPow (< 0.3 at 25 °C) and is very soluble in water (43870 mg/L at 20 °C). A pKa of 9.28 was determined. Particle size distribution analysis revealed a range of D10, D90 [63 μm, 500 μm] with 37.6 % < 100 um and 1.5 % < 45 um.

Based on the particle size, Thymidine is more likely to settle in the nasopharyngeal region than in the tracheobronchial or pulmonary regions. Due to the low logPow direct absorption across the respiratory tract epithelium is not very likely. Instead, the high water solubility points towards retention within mucus. Thus, absorption via respiratory tract is expected to be low.

Upon exposure to the skin the substance may dissolve into the surface moisture of the skin. However, Thymidine may be too hydrophilic to cross the lipid rich environment of the stratum corneum. Thus, absorption via the skin is expected to be low.

Highest rates of absorption are expected via the oral route. Upon ingestion Thymidine is likely to dissolve in the gastrointestinal (GI) fluids. As a weak acid, with a pKa of 9.28, the main site of absorption most likely is the small intestine. Maximum plasma concentrations are expected within the first few hours following exposure.

Several studies on absorption using the structural analog L-Thymidine were carried out (EMEA, 2007). Orally administered doses of 10 mg/kg bw resulted in bioavailability of 60% in rats, 59% in monkeys and 38% in woodchucks, maximum plasma concentrations reached within 1 to 3 h.

Studies conducted with fed and fasted female rats suggested that food had no effect on the absorption of L-Thymidine (10 mg/kg dose). L-Thymidine exposure was less than dose-proportional, especially at doses greater than 1000 mg/kg bw. As shown in an in vitro Caco-2 study, L-Thymidine has moderate permeability properties. Various in vitro studies demonstrated that L-Thymidine does not interact with P-gp and MRP. A lack of interaction between L-Thymidine and transporters was also shown in humans. Finally, there were no sex-related differences in the pharmacokinetics profile of L-Thymidine.

Following uptake and becoming bioavailable Thymidine is likely to distribute throughout the whole body water, tissues and brain. Takeda, et al. (1984) assessed the distribution of tritiated Thymidine in various tissues of rat 24 hours after oral administration of tritiated thymidine. Highest levels were found in the small intestine and the spleen. Lower levels were observed in the liver, lung, testis, kidney, brain and muscle. Following i.v. injection in mice, tritiated thymidine was apparently rapidly absorbed by all cells, leaving the blood stream within a few minutes and found in all tissues, with highest levels in kidney and liver (Hughes et al., 1958). In human pharmacokinetic studies with tritiated Thymidine plasma levels following i.v. injection reached uniform mixing throughout the total body water within several minutes (Rubini et al, 1960; Straus et al., 1977).

Oral administration of [14C]-L-Thymidine (10 mg/kg bw) to male rats also resulted in whole body distribution, with the highest concentrations observed in small and large intestines, urinary bladder, kidneys, prostate gland, mesenteric lymph nodes, stomach, and pancreas. There was limited penetration to the central nervous system, moderate crossing the placenta, and substantial secretion into rat milk as evidenced by a milk/plasma AUC ratio of 2.3. L-Thymidine was weakly bound to plasma protein (from 3.3 to 7.5 % in rat, monkey and human) and was independent of L-Thymidine concentration over the range evaluated (0.4 to 40 μg/ml). L-Thymidine partitioned into the erythrocytes of rats, monkeys and humans independently of its concentration (range 32 to 43 %).

Park and Mitra (1992) assessed the metabolism of Thymidine in the GI tract. Before becoming bioavailable Thymidine was rapidly metabolized into nucleobase and sugar in the upper tract. No metabolites appeared in the colon. Notably, a free 3'-OH group seemed required for the metabolism (catabolism) of thymidine analogues in the rat intestine mainly by pyrimidine nucleoside phosphorylase.

Bioavailable Thymidine may be rapidly incorporated into the DNA of proliferating cells. Following i.v. injection of tritiated Thymidine in humans up to 90 % was lost from the blood within the first minutes (Rubini et al., 1960). About one-half of the plasma tritiated thymidine is catabolized to tritiated water, while the rest was presumably incorporated into DNA or converted to degradation products.

The salvage pathway to Thymidine synthesis involves the enzyme thymidine kinase and in a first step leads to the formation of β-aminoisobutyrate. Ultimately degradation leads to the formation water and CO2 via the citric acid cycle. Fink et al. (1955) incubated Thymidine with rat liver and identified a number of metabolites via the formation of β-aminoisobutyric acid, including p-aminoisobutyric acid, 5-hydroxymethyluracil, alanine and glucose, as well as 5-hydroxymethyluracil, urea, dihydrothymine, P-ureidoisobutyric acid, uracil-5-carboxylic acid and riboside (5-methyluridine). All of the metabolites are practically non-toxic.

Notably, while the mono-, di- and triphosphate metabolites of L-Thymidine have been observed in vitro, they have not been seen in plasma, urine or faeces in vivo. Following a single oral dose of radiolabelled L-Thymidine to rats, the parent compound was the predominant radioactive component excreted in both males and females. One minor and unidentified metabolite was observed in females. This metabolite represented up to 3.9% of the radioactivity present in plasma and no more than 7.3% of radioactivity in urine (representing no more than 1.1% of the administered radioactivity). As less than 0.80% of the total administered dose was eliminated in the bile, these were considered to be minor metabolites. No metabolites were observed in plasma or urine of monkeys or woodchucks receiving radiolabelled L-Thymidine.

Based on its molecular weight, water solubility and pKa Thymidine and its metabolites are likely to be excreted via the urine. Zaharko et al. (1979) identified kidney clearance of intact Thymidine the primary route of removal at high plasma concentrations and noted that metabolism, which plays a major role in the clearance of Thymidine at micromolar concentrations in plasma, appears to be saturated at millimolar concentrations of Thymidine. CO2 produced during metabolism may be exhaled.

After a single oral administration of [14C]-L-Thymidine to rats (10 mg/kg) radiocarbon was eliminated in both urine and faeces in approximately equivalent amounts (40-50 %) over 168 hr; overall recovery of radiocarbon was > 91 %. The comparison of results after both routes of administration suggested that urine was the primary route of excretion, and that radioactivity recovered in the faeces after oral administration corresponded to unabsorbed substance.

In monkeys, drug-related radioactivity was primarily eliminated in the urine following intravenous administration of [3H]-L-Thymidine (74 % of the dose as L-Thymidine). After oral administration, 36.6 % of the administered compound was eliminated in the urine.

L-Thymidine was eliminated at a moderate to rapid rate (t1/2 2-8 hr) in mice, rats, and woodchucks, but more slowly in monkeys (t1/2 7.5-18 hr) and humans (t1/2 41.1 hr). No gender differences were observed in elimination.

L-Thymidine did not inhibit in vitro human CYP450 enzymes including CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4. In addition, L-Thymidine did not induce rat CYP1A2, CYP2B, 3A or 4A in vivo. The occurrence interactions with other compounds mediated via cytochrome P450 therefore seems unlikely.

Summary

Thymidine is ubiquitously present in nature and exists in all living organisms. The substance is one of the pyrimidine deoxynucleosides and part of double-stranded DNA. Degradation of Thymidine ultimately leads to the formation of CO2 and water. The parent compound and its metabolites are practically non-toxic.

The main route of Thymidine exposure is via the oral pathway. Following oral uptake the substance is likely to solubilise in GI-fluids and is absorbed via the small intestine, if not metabolised in the GI-tract. Bioavailable Thymidine distributes throughout the body water, cells and tissues. Thymidine may then be incorporated into DNA by replicating cells or degraded via salvage pathways. Metabolism is mainly via thymidine kinase leading to the formation of β-aminoisobutyraten and ultimately resulting in the formation of water and CO2. Degradation by Cytochrome P450 metabolism seems unlikely. The substance is not activated and does not bioaccumulate. Elimination of the parent compound and metabolites is most likely via the kidneys and urine.

Categories Display