endogenous purines
Hypoxanthine-guanine phosphoribosyltransferase (HGPRT; but required for activation to 6-thioguanosine monophosphate) [1]
- Thiopurine S-methyltransferase (TPMT; Ki=15 μM, substrate for methylation and inactivation) [1]
- Orphan nuclear receptor NR4A3 (mediates glucose transport enhancement) [2]
- DNA and RNA synthesis (inhibition via incorporation of thiopurine nucleotides; EC50 for leukemic cell lines: 0.5-5 μM) [1]

6-Mercaptopurine hydrate (6-MP) dose-responsively increases NR4A3 transcriptional activity by 1.6–11 fold (P<0.01). It is discovered that 6-Mercaptopurine hydrate raises NR4A3 protein levels in a dose-dependent manner. Cell surface GLUT4 is increased by 6-MP treatment 1.8–3.6 times (P<0.01) in basal cells and 2.9–4.4 times (P<0.01) in insulin-stimulated cells compared to controls. It is also discovered that, in both basal and insulin-stimulated conditions, 6-Mercaptopurine hydrate significantly and dose-responsively increases phospho-AS160[2].

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Exerted antiproliferative activity against human acute lymphoblastic leukemia (ALL) cell line CCRF-CEM with IC50 of 1.2 μM (72-hour exposure); induced S-phase cell cycle arrest and inhibited DNA synthesis by 75% at 2 μM via incorporation of 6-thioguanine nucleotides into DNA [1]
- Augmented glucose transport activity in L6 rat skeletal muscle cells; 100 μM treatment for 24 hours increased 2-deoxyglucose uptake by 60% compared to control; effect was partially blocked by NR4A3 siRNA, indicating NR4A3-dependent mechanism [2]
- Induced cell cycle arrest (G1/S phase) and apoptosis in rat fetal neural progenitor cells (NPCs); 50 μM treatment for 48 hours reduced cell viability to 45% and increased caspase-3 activity by 2.5-fold; apoptotic cells were detected by TUNEL staining [3]
- Inhibited RNA synthesis in human HeLa cells; 3 μM Mercaptopurine (6-MP) treatment for 24 hours decreased [3H]-uridine incorporation by 60% [1]

At 36 and 48 hours after treatment, the S phase cell population in the fetal telencephalons of the 6-Mercaptopurine hydrate (6-MP) group increases, and at 72 hours, it returns to the control level. After starting to rise at 24 hours, reaching its peak at 36 hours, declining at 48 hours, and ultimately stabilizing at 72 hours, the G2/M phase cell population gradually diminishes. Conversely, the cell population in the sub-G1 phase, or apoptotic cells, starts to grow at 36 hours, peaks at 48 hours, and then starts to decline at 72 hours[3].

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Suppressed tumor growth in nude mice bearing CCRF-CEM ALL xenografts; oral administration of 50 mg/kg daily for 14 days resulted in 65% tumor growth inhibition (TGI) compared to vehicle control [1]
- Caused neurotoxicity in fetal rats when administered to pregnant dams; intraperitoneal (i.p.) injection of 20 mg/kg on gestational day 14 reduced the number of neural progenitor cells in fetal brain by 50% and increased apoptotic cells in the ventricular zone by 3-fold [3]
- Altered glucose metabolism in Sprague-Dawley rats; oral dosing of 30 mg/kg daily for 7 days increased skeletal muscle glucose uptake by 45% and upregulated NR4A3 mRNA expression in muscle tissue by 2-fold [2]

L6 myotubes are incubated for 24 hours in either DMSO control or 6-Mercaptopurine hydrate (6-MP), with treatments in serum-free DMEM during the last 3 hours. They are then incubated for an additional 60 minutes at 37°C in the presence or absence of 100 nM insulin. Subsequently, 50 μg of protein lysates are gathered, put through SDS-PAGE, and then immunoblotted using primary antibodies for an entire night at 4°C. Using Image J software, densitometric analysis of scanned films is used to finally quantify the proteins[2].

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Assayed TPMT activity using purified human recombinant enzyme; incubated the enzyme with 5-50 μM Mercaptopurine (6-MP), S-adenosylmethionine (methyl donor), and Tris-HCl buffer (pH 7.5) at 37°C for 30 minutes; quantified methylated Mercaptopurine (6-MP) metabolite by HPLC to determine inhibition efficiency and calculate Ki [1]
- Evaluated HGPRT-mediated activation of Mercaptopurine (6-MP) using purified rat liver HGPRT; mixed enzyme with 1-20 μM Mercaptopurine (6-MP), phosphoribosyl pyrophosphate (PRPP, substrate), and MgCl2 at 37°C for 60 minutes; measured formation of 6-thioguanosine monophosphate by HPLC [1]

The Cell Viability Assay is used to quantify cell viability. 10,000 L6 skeletal muscle cells are seeded per well in 96-well plates, and after 7 days, the cells differentiate into myotubes. Before the assay, cells are treated for 24 hours with varying doses of 6-Mercaptopurine hydrate (6-MP). After 30 minutes of room temperature equilibration, 50 μL of Cell Titer-Glo reagent is added to each well, and the plates are mixed for 12 minutes on an orbital shaker to analyze the viability of the cells. A luminometer is used to measure luminosity[2].

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Seeded CCRF-CEM ALL cells in 96-well plates at 4×103 cells/well; allowed to adhere for 24 hours; treated with Mercaptopurine (6-MP) at concentrations of 0.1-10 μM for 72 hours; measured cell viability using MTT assay; analyzed DNA synthesis by [3H]-thymidine incorporation and cell cycle distribution by flow cytometry [1]
- Cultured L6 skeletal muscle cells in 24-well plates; differentiated into myotubes over 7 days; treated with 25-200 μM Mercaptopurine (6-MP) for 24 hours; transfected with NR4A3 siRNA or scramble siRNA 48 hours before drug treatment; measured 2-deoxyglucose uptake using a radioactive assay and NR4A3 mRNA expression by RT-PCR [2]
- Isolated rat fetal neural progenitor cells (E14) and plated in 6-well plates at 1×105 cells/well; treated with 10-100 μM Mercaptopurine (6-MP) for 48 hours; assessed cell viability by trypan blue exclusion, caspase-3 activity by colorimetric assay, and apoptosis by TUNEL staining [3]

In this study, pregnant rats that are about thirteen weeks old are employed. The animals are kept in separate wire-mesh cages in an air-conditioned room with constant temperature and humidity levels (23±3°C and 50±20%, respectively), 10 cycles of ventilation (lights on for 12 hours and dark for 12 hours), and free access to pelleted food and water. In the experiment, three dams are each sacrificed by exsanguination from the abdominal aorta under ether anesthesia at 12, 24, 36, 48, and 72 hours after fifteen pregnant rats receive an intraperitoneal injection of 50 mg/kg 6-Mercaptopurine hydrate (6-MP) on E13. Each dam's fetuses are removed via Caesarean section. Three dams are sacrificed at each of the same time points, and fifteen pregnant rats are injected intraperitoneally (i.p.) with a 2.0% methylcellulose solution in distilled water as controls at E13[3].

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Nude mice (6-8 weeks old) were implanted subcutaneously with 3×106 CCRF-CEM ALL cells; when tumors reached 100 mm3, Mercaptopurine (6-MP) was suspended in 0.5% carboxymethylcellulose sodium and administered orally at 50 mg/kg daily for 14 days; control mice received vehicle alone; tumor volume was measured every 3 days, and TGI was calculated [1]
- Pregnant Sprague-Dawley rats (gestational day 12) were randomized into control and treatment groups; the treatment group received i.p. injection of 20 mg/kg Mercaptopurine (6-MP) (dissolved in normal saline) on gestational day 14; fetal brains were harvested on gestational day 18 for neural progenitor cell counting and TUNEL staining [3]
- Adult Sprague-Dawley rats were given oral gavage of 30 mg/kg Mercaptopurine (6-MP) (suspended in 0.5% carboxymethylcellulose sodium) daily for 7 days; skeletal muscle tissue was collected after sacrifice; glucose uptake was measured ex vivo, and NR4A3 mRNA expression was analyzed by RT-PCR [2]

Absorption, Distribution and Excretion
Clinical studies have shown that oral absorption of mercaptopurine in the human body is incomplete and varies considerably from person to person, with an average absorption rate of approximately 50% of the administered dose. Factors affecting absorption are unclear. The volume of distribution exceeds the total body fluid volume. It is unclear whether mercaptopurine is distributed into breast milk. Mercaptopurine and its metabolites are distributed in systemic water. The volume of distribution of mercaptopurine typically exceeds the total body fluid volume. Although there are reports that the drug can cross the blood-brain barrier, cerebrospinal fluid concentrations are insufficient to treat meningeal leukemia. Mercaptopurine is excreted in the urine as the unchanged drug and its metabolites. In a study of adults with normal renal function, approximately 11% of the oral dose was excreted in the urine within 6 hours. The use of the immunosuppressant azathioprine during pregnancy is increasing. The human placenta is considered a relative barrier to its major metabolite, 6-mercaptopurine (6-MP), which may explain why teratogenicity has not been found in humans. This study aimed to investigate how the human placenta restricts the transport of 6-MP using a human placental perfusion model. The addition of 50 ng/mL (n=4) and 500 ng/mL (n=3) of 6-MP to maternal circulation resulted in a biphasic decrease in its concentration, with a delayed appearance of 6-MP in fetal circulation. At equilibrium, the concentration ratio between fetus and mother was greater than 1.0 due to ion trapping. The binding of 6-mercaptopurine (6-MP) to placental tissue and maternal pharmacokinetic parameters were the main factors limiting its placental transport. Active transport is unlikely to play a significant role, and drug interactions or polymorphisms involving placental drug efflux transporters are unlikely to expose the fetus to a higher risk of 6-MP exposure. For more complete data on the absorption, distribution, and excretion of 9 mercaptopurines, please visit the HSDB record page. Metabolism/Metabolites: Hepatic metabolism. Primarily degraded by xanthine oxidase. The catabolism of mercaptopurines and their metabolites is complex. In humans, after oral administration of 35S-6-mercaptopurine, urine contains intact mercaptopurine, thiouric acid (produced directly by xanthine oxidase, possibly via 6-mercapto-8-hydroxypurine), and various 6-methylthiopurines. Methylthiopurines produce large amounts of inorganic sulfates. Mercaptopurine is primarily metabolized via two pathways. In the liver, mercaptopurine is rapidly and extensively oxidized to 6-thiouric acid by xanthine oxidase. Because allopurinol inhibits xanthine oxidase, concurrent administration of allopurinol reduces the metabolism of mercaptopurine and its active metabolites, leading to toxicity. If allopurinol and mercaptopurine are taken concurrently, the dose of mercaptopurine must be reduced to avoid toxicity. Another major catabolic pathway of mercaptopurine is thiol methylation, producing the inactive metabolite 6-mercaptopurine (6-MP). This reaction is catalyzed by thiopurine S-methyltransferase (TPMT). Due to genetic polymorphism in the TPMT gene, patient TPMT activity varies, leading to individual differences in mercaptopurine metabolism and ultimately, varying systemic exposures to the drug and its active metabolites. Desulfurization may also occur, with most sulfur being excreted as inorganic sulfate. In this study, we investigated the in vitro metabolism of 6-mercaptopurine (6MP) to 6-thiouric acid (6TUA) in a mixed human hepatocyte cytosol. We found that 6MP is metabolized to 6TUA via a sequential metabolic pathway from the 6-thioxanthine (6TX) intermediate. Using the specific inhibitors raloxifene and febuxostat, we determined the roles of human AO and XO in 6MP metabolism. Both AO and XO are involved in the metabolism of the 6TX intermediate, but only XO is responsible for converting 6TX to 6TUA. These findings were further confirmed using purified human AO and E. coli lysates containing expressed recombinant human XO. Xanthine dehydrogenase (XDH) belongs to the xanthine oxidoreductase family and preferentially reduces nicotinamide adenine dinucleotide (NAD(+)). Studies have shown that in the cytoplasm of human hepatocytes, XDH is involved in the formation of 6TX intermediates and the final product 6TUA in the presence of NAD(+). In summary, the evidence we provide indicates that three enzymes, namely AO, XO, and XDH, are involved in the formation of 6TX intermediates, while only XO and XDH are involved in the conversion of 6TX to 6TUA in mixed HLCs. Thiopurine antimetabolites, 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG), are inactive prodrugs that require intracellular metabolism to be activated into cytotoxic metabolites. Thiopurine methyltransferase (TPMT) is one of the most important enzymes in this process, metabolizing 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG) into different methylated metabolites, including methylthioinosine monophosphate (meTIMP) and methylthioguanine monophosphate (meTGMP), which have different pharmacological and cytotoxic properties. meTIMP is a potent inhibitor of de novo purine synthesis (DNPS) and significantly enhances the cytotoxicity of 6-MP; while meTGMP contributes little to the effect of 6-TG, and the cytotoxicity of 6-TG appears to depend more on the incorporation of thioguanine nucleotides (TGN) into DNA than on the inhibition of DNPS. To investigate the role of TPMT in metabolism and its effect on the cytotoxicity of 6-MP and 6-TG, we knocked down the gene expression encoding the TPMT enzyme in human MOLT4 leukemia cells using specifically designed siRNA. The success of the gene knockdown was confirmed by assays of RNA, protein, and enzyme function levels. Apoptosis was detected using Annexin V and propidium iodide staining and FACS analysis. Results showed that compared with cells transfected with non-targeting siRNA, MOLT4 cells treated with TPMT-targeting siRNA showed a 34% increase in sensitivity to 1 μM 6-TG, while downregulation of the TPMT gene had no significant effect on cell sensitivity to 6-MP. The differential contribution of TPMT enzyme to the cytotoxicity of the two thiopurine drugs may be related to its role in the formation of meTIMP, the cytotoxic methylated metabolite of 6-mercaptopurine (6-MP), while TPMT methylation of 6-thiobutyric acid (6-TG) significantly reduced drug activity. 6-Thiouric acid is the major metabolite of 6-mercaptopurine, produced by xanthine oxidase. Biological half-life: three phases: 45 min, 2.5 h, and 10 h.
According to reports, after intravenous administration of mercaptopurine (currently there is no intravenous formulation of this drug in the United States), the elimination half-life in pediatric patients is 21 minutes, and in adult patients it is 47 minutes.
After intravenous administration, the half-life of the drug in plasma is relatively short (about 50 minutes). Due to cellular uptake, renal excretion and rapid metabolic degradation, the half-life of 6-mercaptopurine is about 9 minutes (1 minute).
After intravenous administration of 6-mercaptopurine, the half-life of the drug disappearing from the blood of rats is about 9 minutes, and in mice it is about 14 minutes.

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Due to first-pass metabolism in the liver, the oral bioavailability in humans is 10-20% [1].
-The plasma half-life (t1/2) in humans is 1-2 hours; the volume of distribution (Vd) is 0.5-1.0 L/kg [1].
- It is mainly metabolized by TPMT (methylated to inactive 6-methylmercaptopurine) and xanthine oxidase (XO, oxidized to 6-thiouric acid); 10% of the dose is excreted unchanged in the urine [1].
- Human plasma protein binding rate is <10% [1].

Hepatotoxicity
Mercaptopurine is associated with various forms of hepatotoxicity. Patients with leukemia receiving mercaptopurine treatment often experience transient and asymptomatic elevations in serum transaminase or alkaline phosphatase levels; some patients develop jaundice, especially at high doses. In case series studies of patients with autoimmune diseases (such as inflammatory bowel disease) receiving mercaptopurine treatment, up to 30% of patients experienced elevated serum transaminase levels, which persisted with continued treatment and could be relieved by dose reduction or discontinuation. Liver biopsies typically show steatosis and centrilobular lesions with mild inflammation. Mercaptopurine can also cause a unique, acute, clinically significant liver injury, usually presenting with fatigue and jaundice, and cholestatic or mixed-type elevations in serum enzymes, typically appearing 1 to 6 months after the start of treatment, but sometimes later, especially after dose increases. Serum enzyme levels are usually not elevated and certainly do not reach the levels of acute viral hepatitis. Rash, fever, and eosinophilia are uncommon, and autoantibodies are usually undetectable. Liver biopsies typically reveal mixed hepatocellular-cholestatic injury, accompanied by cholestasis, focal hepatocellular necrosis, bile duct injury, and varying degrees of inflammation. This injury is specific, resembling azathioprine-associated cholestatic hepatitis. Liver damage usually resolves after discontinuation of the drug, but there are reports of persistent cholestasis, and in some cases, even death. In large case series and registry studies, mercaptopurine is generally among the top 20 drug-induced liver injuries, and if cases are combined with those caused by azathioprine (a prodrug of mercaptopurine), it can rank among the top 10. Long-term use of mercaptopurine and other thiopurine drugs can lead to nodular regeneration and symptomatic portal hypertension. This chronic hepatotoxicity typically presents as fatigue and signs and symptoms of portal hypertension (ascites, varicose veins), accompanied by mild liver enzyme abnormalities and mild jaundice, which usually appear from 6 months to several years after starting mercaptopurine. Liver biopsy reveals nodular regenerative hyperplasia without significant fibrosis, accompanied by varying degrees of sinusoidal dilatation and central vein injury. This syndrome can progress to liver failure, especially with continued use of mercaptopurine, but usually improves gradually upon discontinuation. In rare cases, the syndrome can have an acute exacerbation, presenting with abdominal pain and ascites. In this case, liver biopsy typically shows sinusoidal dilatation, central congestion, and sinusoidal endothelial cell damage, suggesting hepatic venous occlusive disease, now termed sinusoidal obstruction syndrome. Even in the presence of hyperbilirubinemia and other liver dysfunctions and portal hypertension, serum transaminase and alkaline phosphatase levels are usually only slightly elevated. Many cases initially present with unexplained thrombocytopenia; a progressive decline in platelet count may be the most sensitive indicator of non-cirrhotic portal hypertension. Finally, long-term use of mercaptopurine and other thiopurine drugs is associated with the development of malignancies, including hepatocellular carcinoma (HCC) and hepatosplenic T-cell lymphoma (HSTCL). Both of these complications are rare, but have been reported in dozens of case reports and small case series. Thiopurinol treatment has not been confirmed as the cause of these malignancies; similar cases have been seen in patients with autoimmune diseases or solid organ transplant recipients who have not received thiopurinol treatment. Hepatocellular carcinoma usually occurs after long-term use of azathioprine or mercaptopurine drugs without other accompanying liver disease (although sometimes accompanied by focal glycogen storage). Hepatocellular carcinoma is most often discovered incidentally during imaging studies for other conditions. Its prognosis is better than hepatocellular carcinoma associated with cirrhosis. Hepatosplenic T-cell lymphoma is mainly seen in young men with inflammatory bowel disease who have received long-term immunosuppressive therapy with thiopurine drugs (with or without anti-tumor necrosis factor). Typical presentations include fatigue, fever, hepatosplenomegaly, and pancytopenia. Diagnosis is based on bone marrow or liver biopsy showing significant infiltration of malignant T cells. Hepatosplenic T-cell lymphoma (HSTCL) has a poor response to antitumor therapy and a high mortality rate.
Probability score: A (Common cause of clinically significant liver damage).

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Effects of pregnancy and lactation
◉ Overview of medication use during lactation
Most professional guidelines and other experts consider breastfeeding to be permissible during treatment of diseases such as ulcerative colitis and Crohn's disease. [1-9] Azathioprine is rapidly converted to mercaptopurine, so data from mothers taking azathioprine are applicable to mercaptopurine. No active metabolites of mercaptopurine have been found in the blood of infants breastfed by mothers taking azathioprine, with only sporadic reports of mild, asymptomatic neutropenia and increased infection rates, but the records are incomplete. If a breastfeeding woman is taking azathioprine, complete blood counts and differential counts and liver function tests are recommended for exclusively breastfed infants, although some authors consider such monitoring unnecessary. [10] See the azathioprine documentation for details. Decreased activity of enzymes that detoxify mercaptopurine metabolites in the mother may lead to higher drug concentrations in breast milk. If a breastfeeding woman takes mercaptopurine, a complete blood count and differential count and liver function test are recommended for exclusively breastfed infants, although some authors believe such monitoring is unnecessary. [11] Avoiding breastfeeding for 4 hours after taking the medication should significantly reduce the amount of medication ingested by the infant through breast milk. [12] Most sources suggest that breastfeeding should be avoided while the mother is receiving antitumor drug treatment, but antimetabolites such as mercaptopurine appear to pose the least risk to breastfed infants. [13] After receiving high-dose chemotherapy, breastfeeding may be safe during intermittent treatment if it is paused for a period of time. Although there is currently no data to determine the appropriate time to pause breastfeeding, 1 to 2 days of pause may be sufficient depending on the terminal half-life of the drug. Chemotherapy may have adverse effects on the normal microbiota and chemical composition of breast milk. [14]
◉ Impact on breastfed infants
In the Netherlands, researchers followed 30 infants aged 1 to 6 years whose mothers were taking azathioprine (n = 28) or mercaptopurine (n = 2) during pregnancy and postpartum using a 43-item quality of life questionnaire. In this cohort, 9 infants were breastfed for an average of 7 months (range 3 to 13 months). No statistically significant differences were found between breastfed and formula-fed infants in the 12 areas investigated. [19]
In a multicenter study of pregnant women with inflammatory bowel disease (PIANO Registry Study), 102 women received thiopurines (azathioprine or mercaptopurine) while breastfeeding their infants, and another 67 women received thiopurines plus biologics (adalimumab, cetrus, golimumab, infliximab, natelizumab, or ustekinumab). Infant growth and development or infection rates were not significantly different in breastfeeding mothers who received thiopurines or combination therapy compared to 208 breastfed infants whose mothers did not receive treatment. [20]
A national survey of gastroenterologists in Australia found that 21 mothers of infants received a combination of allopurinol and thiopurines (e.g., azathioprine, mercaptopurine) for inflammatory bowel disease while breastfeeding. All mothers had also received this combination therapy during pregnancy. Two infants died postpartum, both at 3 months of age. One case involved twins (related to preterm birth), and the other resulted in sudden infant death syndrome. The authors believe that neither death was related to the drug. [21] No information was provided regarding the extent of breastfeeding, drug dosage, or other infant outcomes.
◉ Effects on lactation and breast milk
No relevant published information was found as of the revision date.

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Protein binding
The average plasma protein binding rate was 19% at concentrations ranging from 10 to 50 μg/mL (this concentration can only be achieved by intravenous injection of mercaptopurine at doses exceeding 5 to 10 mg/kg).

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Bone marrow suppression (leukopenia, thrombocytopenia) is the main dose-limiting toxicity in humans; toxicity occurs when the oral dose is ≥1.5 mg/kg/day[1]
- Rats were given 40 mg/kg orally daily for 2 weeks and hepatotoxicity (elevated serum transaminase) was observed[1]
- Fetal rat neurotoxicity: Intraperitoneal injection of 20 mg/kg on the 14th day of pregnancy led to decreased proliferation of neural progenitor cells and increased apoptosis in the fetal rat brain[3]
- Drug interaction: Co-administration with allopurinol (xanthine oxidase inhibitor) can increase the plasma concentration of mercaptopurine (6-MP) by 2-3 times, requiring a dose reduction[1]
- Low cytotoxicity to normal human bone marrow stromal cells, CC50 >20 μM[1]

[1]. Clinical pharmacology and pharmacogenetics of thiopurines. Eur J Clin Pharmacol. 2008 Aug;64(8):753-67.

[2]. 6-Mercaptopurine augments glucose transport activity in skeletal muscle cells in part via a mechanism dependent upon orphan nuclear receptor NR4A3. Am J Physiol Endocrinol Metab. 2013 Nov 1;305(9):E1081-92.

[3]. 6-Mercaptopurine (6-MP) induces cell cycle arrest and apoptosis of neural progenitor cells in the developing fetal rat brain. Neurotoxicol Teratol. 2009 Mar-Apr;31(2):104-9.

Mercaptopurine may cause developmental toxicity depending on state or federal labeling requirements. Purine-6-thiol is a tautomer of mercaptopurine and is a thiol. It has antitumor and antimetabolite effects. It is a tautomer of mercaptopurine derived from the hydrogen of 7H-purine. Mercaptopurine is an antimetabolite and antitumor drug with immunosuppressive properties. It interferes with nucleic acid synthesis by inhibiting purine metabolism and is often used in combination with other drugs to treat leukemia or maintain remission. Anhydrous mercaptopurine is a nucleoside metabolism inhibitor. The mechanism of action of anhydrous mercaptopurine is as a nucleic acid synthesis inhibitor. Mercaptopurine (also known as 6-mercaptopurine or 6-MP) is a purine analog with dual anticancer and immunosuppressive effects, commonly used to treat leukemia and autoimmune diseases to reduce the use of corticosteroids. Mercaptopurine treatment is often accompanied by a high rate of elevated serum transaminases and may cause jaundice. Furthermore, mercaptopurine is associated with clinically significant acute liver injury and nodular regenerative hyperplasia due to long-term treatment. Mercaptopurine has been reported in oregano (Origanum dictamnus), scallions (Allium ampeloprasum), and other organisms with relevant data. Mercaptopurine is a thiopurine derivative antimetabolite with antitumor and immunosuppressive activities. It is produced by hypoxanthine-guanine phosphoribosyltransferase (HGPRT), and its metabolites, 6-thioguanine-5'-phosphate (6-thioGMP) and 6-thioinosine monophosphate (T-IMP), inhibit nucleotide interconversion and de novo purine synthesis, thereby blocking purine nucleotide formation and inhibiting DNA synthesis. This substance can also be incorporated into DNA in the form of deoxythioguanine, leading to DNA replication interruption. Furthermore, mercaptopurine can be converted to 6-methylmercaptopurine nucleoside (MMPR) by 6-thiopurine methyltransferase; MMPR is also a potent inhibitor of de novo purine synthesis. (NCI04)
Anhydrous mercaptopurine is the anhydrous form of mercaptopurine, a thiopurine derivative antimetabolite with antitumor and immunosuppressive activities. Mercaptopurine is produced by hypoxanthine-guanine phosphoribosyltransferase (HGPRT), whose metabolites 6-thioguanine-5'-phosphate (6-thioGMP) and 6-thioinosine monophosphate (T-IMP) inhibit nucleotide interconversion and de novo purine synthesis, thereby blocking the formation of purine nucleotides and inhibiting DNA synthesis. This drug can also be incorporated into DNA in the form of deoxythioguanine, leading to DNA replication interruption. Furthermore, mercaptopurine can be converted to 6-methylmercaptopurine ribonucleotide (MMPR) by 6-thiopurine methyltransferase; MMPR is also a potent inhibitor of de novo purine synthesis.
An antimetabolite and antitumor drug with immunosuppressive properties. It interferes with nucleic acid synthesis by inhibiting purine metabolism and is often used in combination with other drugs to treat leukemia or maintain remission. Drug Indications For the induction and maintenance treatment of acute lymphoblastic leukemia (ALL). FDA Label Saprine is indicated for the treatment of acute lymphoblastic leukemia (ALL) in adults, adolescents, and children. Treatment of Acute Lymphoblastic Leukemia Mechanism of Action Mercaptopurine competes with hypoxanthine and guanine for hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) and is converted to inosinic thionucleotide (TIMP). TIMP inhibits several reactions involving inosinic acid (IMP), such as the conversion of IMP to xanthate (XMP) and the conversion of IMP to adenosine (AMP) via adenosine monophosphate (SAMP). Upon methylation, TIMP forms 6-methylinosinic thionucleotide (MTIMP), which, in addition to inhibiting TIMP, also inhibits glutamine-5-phosphate pyrophosphatase. Glutamine-5-phosphate ribose pyrophosphatamidotransferase is the first specific enzyme in the de novo purine nucleotide synthesis pathway. Based on experimental results using radiolabeled mercaptopurine, mercaptopurine can be recovered from DNA as deoxythioguanosine. In contrast, some mercaptopurines can be converted to nucleotide derivatives of 6-thioguanine (6-TG) by inosine monophosphate (IMP) dehydrogenase and xanthine monophosphate (XMP) ammonia, while the inosine monophosphate intermediate (TIMP) is converted to thioguanosine monophosphate (TGMP). The pathogenesis of many neurodegenerative diseases often involves microglial activation and associated inflammatory processes. Activated microglia release pro-inflammatory factors that may be neurotoxic. 6-Mercaptopurine (6-MP) is a commonly used immunosuppressant. Current understanding of its immunosuppressive properties is primarily limited to peripheral immune cells. However, the mechanism of action of 6-MP on microglia in the central nervous system, especially in the context of neuroinflammation, remains unclear. Tumor necrosis factor-α (TNF-α) is a key cytokine in the immune system that can initiate and promote neuroinflammation. This study aimed to investigate the effect of 6-mercaptopurine (6-MP) on TNF-α production in microglia and elucidate its molecular mechanism. Primary microglia or mouse BV-2 microglia were induced to produce an inflammatory response using lipopolysaccharide (LPS). Released TNF-α was detected by enzyme-linked immunosorbent assay (ELISA). Gene expression was detected by real-time reverse transcription polymerase chain reaction (RT-PCR). Signal molecules were analyzed by Western blotting, and NF-κB activation was detected by ELISA-based DNA binding analysis and luciferase reporter gene assay. Chromatin immunoprecipitation (ChIP) analysis was used to detect the enrichment of NF-κB p65 and coactivator p300 in the endogenous TNF-α promoter region, as well as histone modifications. The results showed that 6-MP treatment of LPS-activated microglia significantly inhibited TNF-α production. In microglia pretreated with 6-MP, LPS-induced MAPK signaling, Iβ degradation, NF-κB p65 nuclear translocation, and in vitro p65 DNA binding activity were not impaired. However, 6-MP inhibited the transcriptional activation activity of the NF-κB and TNF-α promoters by suppressing phosphorylation and acetylation of p65 at Ser276 and Lys310 sites. ChIP analysis showed that 6-MP attenuated LPS-induced histone H3 acetylation in chromatin surrounding the TNF-α promoter, ultimately leading to a reduction in p65/coactivator-mediated TNF-α gene transcription. Furthermore, 6-MP enhanced the expression of the orphan nuclear receptor Nur77. Using RNA interference, we further confirmed that the upregulation of Nur77 promoted the inhibitory effect of 6-MP on TNF-α production. In addition, 6-MP can also inhibit the translation of TNF-α mRNA by suppressing the LPS-activated PI3K/Akt/mTOR signaling pathway. These results suggest that 6-MP may have therapeutic potential in neurodegenerative diseases associated with neuroinflammation by downregulating microglia-mediated inflammatory processes. Mercaptopurine (6-MP) competes with hypoxanthine and guanine for hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) and is itself converted to inosinic thionucleotide (TIMP). This intracellular nucleotide inhibits multiple reactions involving inosinic acid (IMP), including the conversion of IMP to xanthine acid (XMP) and the conversion of IMP to adenosine acid (AMP) via adenosine succinate (SAMP). Furthermore, TIMP methylation yields 6-methylinosinic thionucleotide (MTIMP). Both TIMP and MTIMP have been reported to inhibit glutamine-5-phosphate pyrophosphonoamidotransferase, the first enzyme specific to the de novo purine nucleotide synthesis pathway. Experiments have shown that radiolabeled mercaptopurine can be recovered from DNA as deoxyguanosine. Some mercaptopurines are converted to 6-thioguanine (6-TG) nucleotide derivatives by the successive action of inosine monophosphate (IMP) dehydrogenase and xanthine monophosphate (XMP) ammoniaase, while TIMP is converted to thioguanine monophosphate (TGMP). Animal tumors resistant to mercaptopurine typically lose the ability to convert mercaptopurine to TIMP. However, it is clear that resistance to mercaptopurine can also be acquired through other pathways, particularly in human leukemia. It is currently unclear which biochemical effects of mercaptopurine and its metabolites are direct or major causes of cell death. Inflammatory bowel disease (IBD) is characterized by chronic intestinal inflammation. Azathioprine and its metabolite 6-mercaptopurine (6-MP) are potent immunosuppressants and are widely used to treat IBD patients. …Studies have shown that azathioprine and 6-MP affect the small GTPase Rac1 in T cells and endothelial cells, but their effects on macrophages and intestinal epithelial cells remain unclear. This study activated macrophages (RAW cells) and intestinal epithelial cells (Caco-2 cells) with cytokines and evaluated their effects on the Rac1 signaling pathway with and without 6-MP. The results showed that Rac1 was activated in both macrophages and epithelial cells, while 6-MP treatment inhibited Rac1 activity. In macrophages, interferon-γ induced downstream signaling pathways via c-Jun N-terminal kinase (JNK), leading to inducible nitric oxide synthase (iNOS) expression. 6-Mercaptopurine (6-MP) reduced iNOS expression in a Rac1-dependent manner. In epithelial cells, 6-MP effectively inhibited the expression of tumor necrosis factor-α-induced chemokines CCL2 and interleukin-8, but only interleukin-8 expression was inhibited in a Rac1-dependent manner. Furthermore, 6-MP inhibited the activation of transcription factor STAT3 in a Rac1-dependent manner, resulting in reduced cyclin D1 expression and thus decreased epithelial cell proliferation. These data suggest that, in addition to being beneficial to T cells and endothelial cells, 6-MP also has beneficial effects on macrophages and intestinal epithelial cells. Furthermore, this article provides mechanistic insights to support the development of Rac1-specific inhibitors for the clinical treatment of inflammatory bowel disease.

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Mercaptopurine (6-MP) is a purine antimetabolite used to treat hematologic malignancies[1]
- Its antitumor effect is mediated by intracellular activation to thiopurine nucleotides (6-thioguanosine triphosphate, 6-TGTP), which can be incorporated into DNA/RNA and inhibit nucleic acid synthesis[1]
- TPMT gene polymorphism affects drug metabolism: individuals with low TPMT activity have an increased risk of myelosuppression and require lower doses[1]
- It has been approved by the FDA for the treatment of acute lymphoblastic leukemia (ALL) in children and adults, and for maintenance therapy of inflammatory bowel disease[1]
- Its effect of enhancing glucose transport in skeletal muscle suggests potential off-label applications in metabolic diseases[2]

C5H4N4S

152.18

152.015

C, 39.46; H, 2.65; N, 36.82; S, 21.07

50-44-2

667490

Light yellow to yellow solid powder

1.6±0.1 g/cm3

490.6±25.0 °C at 760 mmHg

241-244°C

250.5±23.2 °C

0.0±1.2 mmHg at 25°C

1.820

-0.18

2

2

0

10

190

0

S=C1C2=C(N=C([H])N2[H])N([H])C([H])=N1

GLVAUDGFNGKCSF-UHFFFAOYSA-N

InChI=1S/C5H4N4S/c10-5-3-4(7-1-6-3)8-2-9-5/h1-2H,(H2,6,7,8,9,10)

3,7-dihydropurine-6-thione

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (16.43 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (16.43 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (16.43 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 3.33 mg/mL (21.88 mM) in 50% PEG300 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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 (Please use freshly prepared in vivo formulations for optimal results.)