Orotic acid analogues

C20 Chelbova, K. V., Golovinsky, E. V. and Hadjiolov, A. A. The action of some orotic acid analogues on the in vitro incorporation of [i C]-orotate into pyrimidine nucleotides. Biochem. Pharmacol., 19, 2785-2789 (1970)... 

C21 Chelbova-Khadzhiolova, K. V. Effect of some orotic acid analogues on the incorporation of [ Hj-uridine and pH]-thymidine into RNA and DNA of cultured mouse embryo fibroblasts. Dokl. Bulg. Akad. Nauk, 25, 533-536 (1972)... 

G23 Grigoryan, L. A. Pyrimidine derivatives. XLIII. Synthesis of some orotic acid analogues. Arm. Khim. Zh., 28,564-571 (1975) (Russ.)... 

Maleuric acid (A -carbamoylmaleamic acid, XVIII), when injected into mice bearing Ehrlich ascites tumours, can produce cytoplasmic abnormalities in all phases of mitosis. This acid also inhibits the incorporation of tritiated thymidine into DNA, and prevents the progression of premitotic cells into mitosis [217]. This substance, which is an open-chain analogue of orotic acid, may possibly be an antimetabolic of this pyrimidine or related compounds. 

Ethyl 2-ethylthio-4-chloro-5-pyrimidinecarboxylate (XXIIa), as well as the corresponding4-hydroxy-(XXIIb) and 4-amino-(XXIIIa) derivatives, possess-anti-cytogenic activity on Neurospora crassa [223, 224]. Compounds (XXIIIa, b and c) were found to inhibit the conversion of orotic acid to the uridine nucleotides [202]. Ethyl 2-methylthio-4-(halo-substituted anilino)-5-pyrimidinecarboxylates (XXIV), particularly the o-bromo- and the o-chloro- derivatives, substantially inhibit the growth of five experimental mouse tumours (Krebs-2 ascites carcinoma, Ehrlich carcinoma clone 2, leukaemia L-1210, carcinoma 755 and lymphocytic neoplasm P-288) [225]. Compounds of this type are usually prepared by the base catalysed condensation of ethoxymethylenemalonic esters or related derivatives with urea, thiourea, guanidine, or substituted amidine-type analogues [212, 225-237]. 

Orotic acid readily forms dimers even when irradiated in liquid medium [582, 583]. 5-Bromouracil (5-BrU) in DNA is dehalogenated, rather than forming cyclobutane-type dimers. Such DNA derivatives are more sensitive to ultraviolet irradiation than normal DNAs [584-594], Irradiation of 5-bromo-uracil and derivatives in aqueous medium produces 5,5 -diuracil [590, 591]. However, derivatives such as 3-sbutyl-5-bromo-6-methyluracil have been reported to yield cyclobutane dimers either by irradiation of frozen aqueous solutions, or by catalysis with free radical initiators, such as aluminium chloride, ferric chloride, peroxides or azonitriles [595]. 5-Hydroxymethyluracil is reported to dimerize very slowly in frozen water at 2537 A [596]. The fundamental research in the photochemistry of the nucleic acids, the monomeric bases, and their analogues has stimulated new experiments in certain micro-organisms and approaches in such diverse fields as template coding and genetic recombination [597-616]. 

The first reaction is catalysed by orotate phosphoribosyltransferase (orotidine 5 -phosphate pyrophosphate phosphoribosyltransferase, EC 2.4.2.10) which is readily reversible. The equilibrium constant for the forward reaction [109] is about 0.1. The reaction is specific for orotate (the enzyme usually does not accept uracil) and some synthetic analogues of orotic acid (Chapter 6). Orotate phosphoribosyltransferase activity was found in many animal tissues [110] and there are several phosphoribosyl-transferases of broad specifity which are distinct from the enzyme involved in the orotate pathway [111-113]. 

Although allopurinol and oxipurinol are potent inhibitors of UMP synthesis [120,131] through the inhibition of orotidylic acid decarboxylase (oxipurinol with a 2,4-diketo pyrimidine ring is capable of acting as an analogue of orotic acid, and 1-ribosyl-oxipurinol 5 -phosphate [132] is a... 

The first known drug affecting the orotate pathway was 6-azauridine [245,246], This analogue is phosphorylated to 6-azauridine 5 -monophosphate which acts as a competitive inhibitor of orotidylic acid decarboxylase [247], Therapeutic use of 6-azauridine [248,249] is occasionally complicated by a pronounced crystalluria. Owing to the block of orotidylic acid decarboxylase, large amounts of orotidine and orotic acid are excreted in urine. After the infusion of 6-azauridine the excretion of orotic acid precedes orotidine and the former disappears more rapidly from the urine. Psoriatic patients on azaribine (triacetylated form of 6-azauridine given orally) excreted 0.2-1.3 g of orotic acid and orotidine per day [250]. 

There are several synthetic derivatives of orotic acid and pyrimidine analogues which, after their conversion, interfere with the activity of orotidylic acid decarboxylase [263,264]. i ile 6-azacytidine 5 -phosphate is only one tenth as active as 6-azauridine 5 -phosphate [265], 5-hydroxyuridine 5 -phosphate [266] and aminouridine 5 -phosphate [267] are potent inhibitors of orotidylic acid decarboxylase. The inhibitory action of allopurinol and of its metabolites on pyrimidine synthesis de novo [268] was mentioned in Chapter 3. 

Fluoro-orotic acid undergoes in the liver the same conversion as orotic acid [269,270]. The 5-fluoro analogue serves as a substrate for orotate phosphoribosyltransferase [270] and the anomalous nucleoside 5 -phosphate so produced inhibits orotidylic acid decarboxylase. A number of... 

However, 5-fluoro-orotic and orotic acids are utilized differentially for the synthesis of cytoplasmic liver RNA. 5-Fluoro-orotate is incorporated preferentially into a fraction of non-ribosomal RNA which has several properties in common with messenger RNA [271]. Analysis of microsomal RNA showed Uttle or no incorporation of 5-fluoro-orotic acid into either 18 S or 28 S ribosomal RNA. The analogue is rapidly incorporated into 45 S ribosomal precursor RNA but its subsequent processing into mature 18 S and 28 S RNA is inhibited [272]. The analogue also greatly inhibits the incorporation of orotic acid into ribosomal RNA but has little effect on its incorporation into messenger RNA [273]. 

Azaorotic (oxonic) acid represents another analogue of orotic add with cytostatic activity [274]. The drug inhibits metabolic transformation and incorporation of orotic acid in the liver and kidney [275]. Similarly to... 

G13 Golovinsky, E., Kaneti, J., Yukhnovsky, I. and Genchev, D. A study of some orotic acid derivatives and analogues by... 

Huckel s method of molecular orbitals. II. Analogues of the orotic acid in the reaction with phosphoribosyl-pyrophosphate. J. Theor. Biol., 26, 29-32 (1970)... 

Kelley and Beardmore, 1970 Beardmore, Cashman and Kelley, 1972). Similar values are seen with oxipurinol therapy. This represents a 7-10 fold increase over pretreatment values for orotic acid (<2.0 mg/24 hr) and orotidine (6.7 mg/24 hr) but is much less than the quantity excreted by patients with hereditary orotic aciduria who may excrete up to 1500 mg/24 hr. This pattern of excretion of pyrimidines is analogous to that observed during therapy with the pyrimidine analogue, 6-azauridine (Fallon, et al., 1961). 

Fluorine has the closest atomic dimensions to the hydrogen atom, and pyrimidine analogues where hydrogen is substituted by fluorine are powerful antimetabolites. 5-fluoro-orotic acid (Fig. 12), for instance, prevents the conversion of orotic acid to orotidylic acid (Fig. 2) and the methylation of deoxyuridylic acid to form thymi-dylic acid (Fig. 9). 

Many potent pyrimidine antagonists have also been obtained by isosteric replacement in the heterocyclic ring. Oxonic acid (Fig. 12) an analogue of orotic acid, prevents the conversion of the latter to orotidylic acid (Fig. 2). A later stage of pyrimidine biosynthesis, the decarboxylation of orotidylic acid to form uridylic acid (Fig. 2) is strongly inhibited by 6-azauracil (Fig. 12 this chemical should be correctly termed 4-azauracil). The riboside of 6-azainacil acts on the same pathways as the base, but its inhibition of the orotidylate carboxylase enzyme is some 20 times more potent... 

A pyrimidine phosphoribosyltransferase activity with a broader specificity than the yeast enzyme has been demonstrated in animal tissues. Highly purified preparations from calf thymus (15) and beef erythrocytes (16) accepted orotate and 5-fluorouracil as substrates. Uracil phosphoribosyltransferase activity has also been demonstrated in extracts from mouse leukemia cells. Fluorouracil is a better substrate for this enzyme than uracil at pH 7.5, possibly because the acid dissociation constant for the analogue (pif 8.15) is higher than that of uracil (pK, 9.45) (17). This reasoning would suggest that the anionic form of the substrate might be the species required by the enzyme. This enzyme has been implicated in the... 

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