To treat or not to treat? An evidence-based practice guide for the management of endometrial polyps.

Endometrial polyps are a common finding, with a prevalence of about 40%, and are usually diagnosed incidentally as most are asymptomatic. Symptomatic polyps usually present with abnormal uterine bleeding and/or sub-fertility. About 25% of polyps resolve spontaneously if managed conservatively. The usual management of endometrial polyps, symptomatic or asymptomatic, is polypectomy, performed primarily to exclude malignancy within the polyp despite the overall risk of malignancy being low (about 3%). The main risk factors for malignancy are menopause and abnormal uterine bleeding, with hypertension, obesity, diabetes mellitus, and tamoxifen use thought to play a lesser role. Transvaginal ultrasonography is the primary diagnostic tool for endometrial polyps although visualization by hysteroscopy is the gold standard for diagnosis. There is no proven preventative or medical treatment, with complete polyp removal under hysteroscopic guidance the recommended surgical treatment. Some women may decline surgical endometrial polyp management due to the small inherent risks. Conservative management is an option for asymptomatic premenopausal and postmenopausal women, whilst polypectomy is recommended for all women with abnormal uterine bleeding. Management should be individualized and made in consultation with the patient.

Improvement in pelvic pain with botulinum toxin type A - Single vs. repeat injections.

The aim of this prospective study was to report the outcomes of pain and vaginal pressures of successive botulinum toxin type A injections for women with objective pelvic floor muscle overactivity and a two-year history of pelvic pain. Between 2005 and 2008, 37 women underwent injection of 100 IU of botulinum toxin type A into the puborectalis and pubococcygeous muscles with dysmenorrhoea, dyspareunia, dyschesia, and non-menstrual pelvic pain assessed using a visual analogue scale (VAS), and vaginal pressure measured by vaginal manometry, at 0, 4, 12 and 26 weeks from each injection. 26 women (70%) had one injection of botulinum toxin type A and 11 (30%) had 2 or more injections. The second injection was performed at the earliest at 26 weeks after the first, with subsequent injections having a median time to re-injection of 33.4 weeks (range 9.4-122.7 weeks). Single and repeated injections both demonstrated a statistically significant reduction in dyspareunia by VAS scores from 54 to 30 in the single injection group and from 51 to 23 in the multiple injection group (p = .001), non-menstrual pelvic pain VAS from 37 to 25 (p = .04), as well as vaginal pressures; 40 versus 34 cm H(2)O (p = .02). No statistically significant difference in dysmenorrhoea or dyschesia was observed for either group from their baseline scores. Multiple injections of botulinum toxin type A in women with pelvic floor muscle overactivity provide significant relief from dyspareunia and non-menstrual pelvic pain. The upper limit between re-injection is not yet determined, nor is the maximum number of treatments. Clinical outcomes for single and subsequent injection of botulinum toxin type A for recurrent pelvic pain are equivalent. Women who have had benefit from a single injection of botulinum toxin type A can be reassured that if symptoms reoccur, repeated injections can be expected to be equally efficacious.

Metabolic effects of thiopurine derivatives against human CCRF-CEM leukaemia cells.

UNLABELLED: BACKGROUND and aims. To compare the metabolic effects induced by the anticancer drugs, 6-mercaptopurine (6-MP), 6-thioguanine (6-TG) and 6-methylmercaptopurine riboside (MMPR), which may inhibit the de novo biosynthesis of purine nucleotides or be mis-incorporated into DNA or RNA. METHODS: Leukaemia cells were grown in culture, exposed to a thiopurine and cell extracts were analyzed for NTPs, dNTPs, drug metabolites and P-Rib-PP. RESULTS: In leukaemia cells, 6-MP was converted to MPR-MP, thio-XMP, thio-GMP, thio-GDP and thio-GTP. Metabolites of 6-TG included thio-XMP, thio-GMP, thio-GDP and thio-GTP, while MMPR-MP was the only major metabolite of MMPR, MMPR (25 microM, 4 h) induced a 16-fold increase in P-Rib-PP and 6-MP (25 microM, 4 h) induced a delayed 5.2-fold increase. MPR-MP, thio-GMP and MMPR-MP are inhibitors of amido phosphoribosyltransferase from leukaemia cells with Ki values of 114 +/- 7.10 microM, 6.20 +/- 2.10 microM and 3.09 +/- 0.30 microM, respectively. CONCLUSION: The nucleoside-5'-monophosphate derivatives of the 3 thiopurines inhibit amido phosphoribosyltransferase in growing leukaemia cells but there is also an initial inhibition of the further conversion of IMP in the pathway. In growing cells, MMPR acts solely as an inhibitor of de novo purine biosynthesis while 6-TG and to a lesser extent, 6-MP, are converted to significant concentrations of di- and tri-phosphate derivatives which may have other mechanisms of cytotoxicity.

Effects of cytosine arabinoside on human leukemia cells.

Cytosine arabinoside (Ara-C) is used to treat leukemias, with complete remission induced by combination chemotherapy in approximately 70% of cases of acute myelogenous leukemia (AML). Ara-CTP acts as a competitive inhibitor of DNA polymerase and may also be incorporated into DNA. Accumulation of deoxyribonucleoside triphosphates (dNTPs) induced by Ara-C may indicate disruption of DNA synthesis in susceptible leukemia cells. A procedure has been developed for the quantification of Ara-CTP and dNTPs from small samples of leukaemia cells from patients (4 x 10(7) cells) activated with concanavalin A (10 micrograms/ml, 48 hr) and grown in the presence of [32P]orthophosphate (1.1 microM, 9 x 10(6) Ci/mol, 16 hr). The susceptibilities to Ara-C of the human leukemia cell lines CCRF-CEM (IC50 = 6.30 nM), CCRF-HSB-2 (IC50 = 10.4 nM) and MOLT-4 (IC50 = 10.0 nM) may be correlated with their abilities to accumulate high concentrations of Ara-CTP (> 1000 amol/cell) with increases of between 1.3- and 3.4-fold in dATP, dGTP and dTTP for the four cell lines, while dCTP decreased between 0.23- and 0.78-fold. By contrast, an Ara-C-resistant derivative of HL-60 cells (IC50 = 400 nM) accumulated only low concentrations of Ara-CTP (71 amol/cell) without significant changes in dNTPs. High concentrations of Ara-CTP in leukemia cells induce accumulations of dATP, dGTP and dTTP due to inhibition of DNA synthesis, and depletion of dCTP. This imbalance in the pools of the four dNTPs could lead to genetic miscoding and cell death.

Cytotoxic effects of inhibitors of de novo pyrimidine biosynthesis upon Plasmodium falciparum.

The malarial parasite Plasmodium falciparum can only synthesize pyrimidine nucleotides via the de novo pathway which is therefore a suitable target for development of antimalarial drugs. New assay procedures have been developed using high-pressure liquid chromatography (HPLC) which enable concurrent measurement of pyrimidine intermediates in malaria. Synchronized parasites growing in erythrocytes were pulse-labeled with [14C]bicarbonate at 6-h intervals around the 48-h asexual life cycle. Analysis of malarial extracts by HPLC showed tht incorporation of [14C]bicarbonate into pyrimidine nucleotides was maximal during the transition from trophozoites to schizonts. The reaction, N-carbamyl-L-aspartate-->L-dihydroorotate (CA-asp-->DHO) catalyzed by malarial dihydroorotase is inhibited by L-6-thiodihydroorotate (TDHO) in vitro (Ki = 6.5 microM), and TDHO, as the free acid or methyl ester, induces a major accumulation of CA-asp in malaria. Atovaquone, a naphthoquinone, is a moderate inhibitor of dihydroorotate dehydrogenase in vitro (Ki = 27 microM) but induces major accumulations of CA-asp and DHO. Pyrazofurin induces accumulation of orotate and orotidine in malaria, consistent with inhibition of orotidine 5'-monophosphate (OMP) decarboxylase with subsequent dephosphorylation of the OMP accumulated. Although TDHO, atovaquone, and pyrazofurin arrest the growth of P. falciparum, only moderate decreases in UTP, CTP, and dTTP were observed. 5-Fluoroorotate also arrests the growth of P. falciparum with major accumulations of 5-fluorouridine mono-, di-, and triphosphates and the most significant inhibition of de novo biosynthesis of pyrimidine nucleotides.

Antifolates induce inhibition of amido phosphoribosyltransferase in leukemia cells.

The pathway for de novo biosynthesis of purine nucleotides contains two one-carbon transfer reactions catalyzed by glycinamide ribotide (GAR) and 5-aminoimidazole-4-carboxamide ribotide (AICAR) transformylases in which N10-formyltetrahydrofolate is the one-carbon donor. We have found that the antifolates methotrexate (MTX) and piritrexim (PTX) completely block the de novo purine pathway in mouse L1210 leukemia cells growing in culture but with only minor accumulations of GAR and AICAR to less than 5% of the polyphosphate derivatives of N-formylglycinamide ribotide (FGAR) which accumulate when the pathway is blocked completely by azaserine. This azaserine-induced accumulation of FGAR polyphosphates is completely abolished by MTX, indicating that inhibition of the pathway is at or before GAR transformylase (reaction 3; Lyons, S. D., and Christopherson, R. I. (1991) Biochem. Int. 24, 187-197). Three h after the addition of MTX (0.1 microM), cellular 5-phosphoribosyl-1-pyrophosphate has accumulated 3.4-fold while 6-methyl-mercaptopurine riboside (25 microM) induces a 6.3-fold accumulation. These data suggest that amido phosphoribosyltransferase catalyzing reaction 1 of the pathway is the primary site of inhibition. In support of this conclusion, we have found that dihydrofolate-Glu5, which accumulates in MTX-treated cells, is a noncompetitive inhibitor of amido phosphoribosyltransferase with a dissociation constant of 3.41 +/- 0.08 microM for interaction with the enzyme-glutamine complex in vitro. Folate-Glu5, MTX-Glu5, PTX, dihydrotriazine benzenesulfonyl fluoride, and AICAR also inhibit amido phosphoribosyltransferase.

Antifolates induce primary inhibition of the de novo purine pathway prior to 5-aminoimidazole-4-carboxamide ribotide transformylase in leukemia cells.

Polyglutamated dihydrofolate, accumulated as a result of potent inhibition of dihydrofolate reductase (DHFR), has been postulated to directly inhibit the purine pathway at 5-aminoimidazole-4-carboxamide ribotide (AICAR) transformylase (reaction 9) in leukemia cells exposed to methotrexate (MTX). We have observed that 25 microM MTX or piritrexim, a "non-classical" antifolate, induce several-fold accumulations of AICAR and N-succino-AICAR to a combined cellular concentration of 89 microM in mouse L1210 leukemia cells after 2 h. By contrast, complete inhibition of reaction 4 by 25 microM azaserine results in accumulation of N-formyl-glycinamide ribotide (FGAR) polyphosphates to a combined cellular concentration of greater than 10 mM. MTX prevented azaserine-induced accumulation of FGAR polyphosphates. Hence, these antifolates induce primary inhibition of the de novo purine pathway at, or prior to, glycinamide ribotide transformylase (reaction 3).

Cytotoxic effects of dihydroorotase inhibitors upon human CCRF-CEM leukemia.

6-L-Thiodihydroorotate (TDHO) and 2-oxo-1,2,3,6-tetrahydropyrimidine-4,6-dicarboxylate (HDDP) are potent inhibitors of mammalian dihydroorotase in vitro (R. I. Christopherson, K. J. Schmalzl, E. Szabados, R. J. Goodridge, M. C. Harsanyi, M. E. Sant, E. M. Algar, J. E. Anderson, A. Armstrong, S. C. Sharma, W. A. Bubb, and S. D. Lyons, Biochemistry, 28: 463-470, 1989). Using human CCRF-CEM leukemia cells growing in culture, TDHO and HDDP as the free acids have 50% inhibitory concentration (IC50) values of 32 microM and greater than 1000 microM, respectively, whereas for TDHO methyl ester, the IC50 value is 25 microM, and for HDDP dimethyl ester, the IC50 value is 21 microM. These IC50 values were not affected by addition of dihydroorotate, uridine, or deoxycytidine to the culture medium. TDHO methyl ester (25 microM) had only slight inhibitory effects upon the dihydroorotase reaction of de novo pyrimidine biosynthesis in growing leukemia cells, cells arrested in G2 + M phases of the cell cycle. At 250 microM TDHO methyl ester, analysis of cell extracts by high-performance liquid chromatography showed that after 4 h carbamyl aspartate had accumulated from undetectable levels to 760 microM, whereas UTP decreased from 580 to 110 microM and CTP from 350 to 86 microM, indicating inhibition of dihydroorotase in growing leukemia cells. IMP accumulated from 63 to 350 microM, total guanylates increased while adenylates decreased, and the adenylate energy charge decreased from 0.91 to 0.69 after 4 h. The cellular concentration of 5-phosphoribosyl 1-pyrophosphate increased from 180 to 290 microM due to sparing from pyrimidine nucleotide biosynthesis resulting in complementary stimulation of the de novo purine pathway. HDDP dimethyl ester at concentrations of up to 250 microM had no discernable effect upon pyrimidine or purine nucleotide biosynthesis. At 25 microM HDDP-dimethyl ester, cells arrested in G2 + M phases initially, with accumulation of cells in G1/G0 at later times. These data suggest that the primary mechanisms of growth inhibition for TDHO and HDDP involve inhibition of cell cycle progression from late G2 or M phase to G1 phase and that blockade of the pyrimidine pathway by TDHO is a secondary effect found at higher concentrations.

Effects of brequinar and ciprofloxacin on de novo nucleotide biosynthesis in mouse L1210 leukemia.

Exposure of mouse L1210 leukemia cells to 25 microM brequinar for 4 h results in large accumulations of N-carbamyl-L-aspartate and L-dihydroorotate to cellular concentrations of 8.5 mM and 0.8 mM, respectively, while UTP and CTP decrease to 4% of their initial levels; incorporation of [14C]bicarbonate into nucleic acids (DNA and RNA) was decreased to 47%. These data provide direct evidence for inhibition of DHO dehydrogenase by brequinar in growing cells. Exposure of leukemia cells to 200 microM ciprofloxacin for 4 h did not affect de novo pyrimidine nucleotide biosynthesis or the incorporation of [14C]bicarbonate into nucleic acids but resulted in a general decrease in nucleoside triphosphates, with concomitant accumulation of nucleoside mono- and diphosphates (the adenylate energy charge decreased from 0.89 to 0.69), consistent with inhibition of the electron transport chain or uncoupling of oxidative phosphorylation.

Cytotoxic mechanisms of glutamine antagonists in mouse L1210 leukemia.

The glutamine antagonists, acivicin (NSC 163501), azaserine (NSC 742), and 6-diazo-5-oxo-L-norleucine (DON) (NSC 7365), are potent inhibitors of many glutamine-dependent amidotransferases in vitro. Experiments performed with mouse L1210 leukemia growing in culture show that each antagonist has different sites of inhibition in nucleotide biosynthesis. Acivicin is a potent inhibitor of CTP and GMP synthetases and partially inhibits N-formylglycineamidine ribotide (FGAM) synthetase of purine biosynthesis. DON inhibits FGAM synthetase, CTP synthetase, and glucosamine-6-phosphate isomerase. Azaserine inhibits FGAM synthetase and glucosamine-6-phosphate isomerase. Large accumulations of FGAR and its di- and triphosphate derivatives were observed for all three antagonists which could interfere with the biosynthesis of nucleic acids, providing another mechanism of cytotoxicity. Acivicin, azaserine, and DON are not potent inhibitors of carbamyl phosphate synthetase II (glutamine-hydrolyzing) and amidophosphoribosyltransferase in leukemia cells growing in culture although there are reports of such inhibitions in vitro. Blockade of de novo purine biosynthesis by these three antagonists results in a "complementary stimulation" of de novo pyrimidine biosynthesis.

Chromatographic analysis of purine precursors in mouse L1210 leukemia.

A number of antagonists of nucleotide metabolism with anti-cancer activity affect the de novo purine pathway. To determine the biochemical mechanisms of cytotoxicity of these drugs, assay procedures have been developed for measurement of the levels of intermediates proximal to IMP in the pathway for de novo purine biosynthesis in mouse L1210 leukemia cells. Purine precursors have been synthesized in vitro from [14C]glycine using enzymes from chicken liver. These 14C-labeled intermediates have been used as marker compounds to define retention times for metabolites of leukemia cells separated by HPLC and the chromatographic mobilities of these intermediates after two-dimensional thin-layer chromatography. These new chromatographic procedures have been used in combination to determine the steady-state concentrations for purine precursors in mouse L1210 leukemia cells in the exponential phase of growth: N-formylglycineamide ribotide (16 microM); N-formylglycineamidine ribotide (4.7 microM); 5-aminoimidazole ribotide (4.0 microM); 4-carboxy-5-aminoimidazole ribotide (0.46 microM); N-succino-5-aminoimidazole-4-carboxamide ribotide (11 microM); 5-aminoimidazole-4-carboxamide ribotide (16 microM); 5-formamidoimidazole-4-carboxamide ribotide (2.7 microM); and IMP (57 microM). The metabolic effects of tiazofurin (25 microM) upon mouse L1210 leukemia cells growing in culture define a "metabolic crossover point" at the reaction catalyzed by IMP dehydrogenase (EC 1.1.1.205) which confirms previous reports of inhibition of this enzyme.

Dual effects of pyrazofurin and 3-deazauridine upon pyrimidine and purine biosynthesis in mouse L1210 leukemia.

Pyrazofurin (NSC 143095) as the monophosphate derivative is a potent inhibitor of orotidine 5'-monophosphate (OMP) decarboxylase of the pyrimidine pathway and has been proposed to inhibit 5-aminoimidazole-4-carboxamide ribotide (AICAR) transformylase (EC 2.1.2.3) of the purine pathway (J. F. Worzalla, and M. J. Sweeney, Pyrazofurin inhibition of purine biosynthesis via 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranosyl 5'-monophosphate formyltransferase. Cancer Res., 40: 1482-1485, 1980). Measurement of levels of pyrimidine and purine intermediates in cultured mouse L1210 leukemia cells has shown that 25 microM pyrazofurin induces an 8-fold accumulation of OMP and large accumulations of intermediates proximal to the blockade with abrupt decreases in uridine and cytidine nucleotides. Considerable increases in the cellular concentrations of N-succino-AICAR (SAICAR), AICAR, 5-formamidoimidazole-4-carboxamide ribotide (FAICAR), IMP, XMP, and GMP at later times indicate that AICAR transformylase is not significantly inhibited in cultured cells; rather the purine pathway and the GMP branch are stimulated. However, addition of 25 microM 3-deazauridine (NSC 126849) to leukemia cells did result in inhibition of AICAR transformylase: AICAR and SAICAR accumulated, IMP disappeared and there was a large accumulation of guanosine nucleotides. Blockade of pyrimidine biosynthesis by derivatives of pyrazofurin or 3-deazauridine spares 5-phosphoribosyl-1-pyrophosphate and L-glutamine, elevated concentrations of which may stimulate initial reactions of purine biosynthesis and the reaction XMP----GMP.

Comparison of 1% sodium tetradecyl sulfate to a thrombogenic sclerosant cocktail for endoscopic sclerotherapy.

This preliminary report suggests superior efficacy of a mixture of Sodium Tetradecyl Sulfate, thrombin and cefazolin (ST-Thr-Cef) over 1% Sodium Tetradecyl Sulfate (ST) in the early control of variceal bleeding. The overall management of patients treated with ST-Thr-Cef was superior as indicated by fewer hospital days and lesser transfusion requirements. Though trends were suggestive, we were unable to demonstrate a survival advantage for patients treated with ST-Thr-Cef. There was a low number of complication in both treatment groups and they were easily managed.

Effects of acivicin and dichloroallyl lawsone upon pyrimidine biosynthesis in mouse L1210 leukemia cells.

Acivicin (NSC 163501) and dichloroallyl lawsone (NSC 126771) are potent inhibitors of nucleotide biosynthesis with consequent anti-cancer activity against certain experimental tumors. To determine in detail the metabolic events induced by each inhibitor, we have devised a new two-dimensional chromatographic procedure for measurement of the concentrations of all pyrimidine intermediates and some purine nucleotides from 100 microliter of an extract of cells grown in the presence of [14C]bicarbonate. Addition of acivicin (25 microM) to mouse L1210 leukemia cells causes severe depletion in the cellular levels of CTP and GTP, accumulation of uridine nucleotides, and abrupt but transient increases in the concentrations of the early intermediates of both the pyrimidine and purine pathways. Addition of dichloroallyl lawsone (25 microM) results in a rapid depletion of uridine and cytidine nucleotides; carbamyl aspartate and dihydroorotate accumulate to high levels in an equilibrium ratio of 20.5:1, and orotate, orotidine, and UMP increase transiently before decreasing to levels approaching their original steady states. The predominant inhibitory effects of acivicin are upon the reactions UTP----CTP and XMP----GMP, but there is also an initial transient activation of both the pyrimidine and purine pathways by acivicin. The data obtained with dichloroallyl lawsone are consistent with inhibition of the conversion of UMP----UDP initially followed by potent inhibition of dihydroorotate----orotate.

Regulation of hamster carbamoyl-phosphate synthase II by 5-phospho-alpha-D-ribosyl 1-diphosphate and uridine 5'-triphosphate.

In mammals, carbamoyl phosphate for utilization in pyrimidine biosynthesis is synthesized by a glutamine-dependent carbamoyl-phosphate synthase II which is subject to regulation by 5-phospho-alpha-D-ribosyl 1-diphosphate (PRib-PP), a positive effector, and MgUTP, a negative effector [Mori, M., Ishida, H. and Tatibana, M. (1975) Biochemistry 14, 2622-2630]. We have found that Lineweaver-Burk plots of carbamoyl phosphate synthase activity versus 1/[MgATP] are described by a velocity equation which is a ratio of quadratic polynomials, consistent with a positive homotropic interaction between two catalytic sites for the binding of MgATP (Ks = 16.6 +/- 3.1 mM, interaction factor a = 0.00538 +/- 0.00245). The activating effect of PRib-PP upon carbamoyl-phosphate synthase is consistent with PRib-PP binding at an allosteric site (Ka = 31.4 +/- 6.4 microM) and promoting the binding of a first molecule of MgATP as substrate (interaction factor l = 0.0437 +/- 0.0063). Thus MgATP and PRib-PP bind to the E X MgATP complex with respective dissociation constants of a X Ks = 0.089 mM and l X Ka = 1.4 microM while MgATP binds to the E X PRib-PP complex with a dissociation constant of l X Ks = 0.73 mM. Data for the inhibitory effect of MgUTP upon carbamoyl-phosphate synthase indicate that MgUTP competes with MgATP for binding at the catalytic site (Ki = 0.203 +/- 0.016 mM). A computer model has recently been developed which enables quantitative stimulation of the time-dependent effects of blockade of the pyrimidine pathway by a tight-binding enzyme inhibitor [Duggleby, R.G. and Christopherson, R.I. (1984) Eur. J. Biochem. 143, 221-226]. The velocity equation derived in the present paper provides a quantitative basis for predicting changes in the flux through the de novo pyrimidine pathway in growing cells.