Characterisation of bioenergetic pathways and related regulators by multiple assays in human tumour cells.

BACKGROUND: Alterations in cellular metabolism are considered as hallmarks of cancers, however, to recognize these alterations and understand their mechanisms appropriate techniques are required. Our hypothesis was to determine whether dominant bioenergetic mechanism may be estimated by comparing the substrate utilisation with different methods to detect the labelled carbon incorporation and their application in tumour cells. METHODS: To define the bioenergetic pathways different metabolic tests were applied: (a) measuring CO2 production from [1-(14)C]-glucose and [1-(14)C]-acetate; (b) studying the effect of glucose and acetate on adenylate energy charge; (c) analysing glycolytic and TCA cycle metabolites and the number of incorporated (13)C atoms after [U-(13)C]-glucose/[2-(13)C]-acetate labelling. Based on [1-(14)C]-substrate oxidation two selected cell lines out of seven were analysed in details, in which the highest difference was detected at their substrate utilization. To elucidate the relevance of metabolic characterisation the expression of certain regulatory factors, bioenergetic enzymes, mammalian target of rapamycin (mTOR) complexes (C1/C2) and related targets as important elements at the crossroad of cellular signalling network were also investigated. RESULTS: Both [U-(13)C]-glucose and [1-(14)C]-substrate labelling indicated high glycolytic capacity of tumour cells. However, the ratio of certain (13)C-labelled metabolites showed detailed metabolic differences in the two selected cell lines in further characterisation. The detected differences of GAPDH, ß-F1-ATP-ase expression and adenylate energy charge in HT-1080 and ZR-75.1 tumour cells also confirmed the altered metabolism. Moreover, the highly limited labelling of citrate by [2-(13)C]-acetate-representing a novel functional test in malignant cells-confirmed the defect of TCA cycle of HT-1080 in contrast to ZR-75.1 cells. Noteworthy, the impaired TCA cycle in HT-1080 cells were associated with high mTORC1 activity, negligible protein level and activity of mTORC2, high expression of interleukin-1ß, interleukin-6 and heme oxygenase-1 which may contribute to the compensatory mechanism of TCA deficiency. CONCLUSIONS: The applied methods of energy substrate utilisation and other measurements represent simple assay system using (13)C-acetate and glucose to recognize dominant bioenergetic pathways in tumour cells. These may offer a possibility to characterise metabolic subtypes of human tumours and provide guidelines to find biomarkers for prediction and development of new metabolism related targets in personalized therapy.

Characteristics of cystic breast disease with special regard to breast cancer development.

BACKGROUND: This prospective study compares the characteristics of cystic disease of the breast (CDB) of patients who developed breast cancer (BCa) during the follow-up (1.25-4 years) period with those who did not. MATERIALS AND METHODS: K+, Na+, albumin, dehydroepiandrosterone (DHA), DHA-sulphate, oestrone, oestradiol, testosterone and progesterone levels were determined in breast cyst fluid (BCF). Patients presented data about their menstrual status, reproductive history, lactation period, date of first and the number of BCF aspirations, gynaecological interventions, use of oral contraceptives, family history of cancer, smoking habits and coffee consumption. The BCa incidence of patients was compared with the expected number of BCas in an age-matched group of 5143 women. RESULTS: Out of 147 patients 6 developed BCa. The standardized incidence rate was 6.29. There were significant differences in testosterone, oestrone and progesterone levels and also reproductive history of patients who developed BCa compared with patients without BCa. CONCLUSION: The above markers outline a subgroup of patients with the highest BCa risk.

Bursting strength and collagen content changes in esophageal anastomoses. Comparative experimental study in dogs.

BACKGROUND AND AIMS: The Biofragmentable Anastomosis Ring (BAR) is a mechanical device composed of absorbable material, performs an inverted anastomosis by atraumatic compression. It is a safe method recommended for bowel ends covered by serosal layer. The feasibility for esophageal surgery hasn't been proved. In our study we compared the healing of transsected intrathoracic esophagus anastomosed with BAR versus Stapler on an experimental dog model by evaluating the bursting strength (BS) and collagen (Hydroxyproline (HP)) concentration and content in different sites and defined time points after surgery (4th, 7th, 14th, 28th day). MATERIAL AND METHODS: Forty-six mongrel dogs were randomly separated into two groups (28 BAR, 18 Stapler) and parameters of surgical repair evaluated after sacrification on certain days. RESULTS: Although we had a total of 4 leaks (14,3%) only in the BAR group (2 were clinically relevant and fatal), there was no significant difference in the leakage rate between the two groups. BS was significantly higher in the BAR group during the first week with values almost equal from the 14th day, in both groups. BS measurement reflects anastomotic strength only during the first two postoperative weeks. After this rupture is often produced outside the anastomotic line. HP concentration showed no significant differences from samples taken next to and far from the anastomotic line. The HP rate showed a significant reduction in the 4th and 7th day subgroups, and values close to the reference from the 14th day with both techniques. CONCLUSIONS: BS and HP changes proved a normal tissue repair with no significant difference, showing that both methods are suitable to perform anastomoses on the intrathoracic esophagus in mongrel dogs. The higher mechanical stability of the BAR anastomosis could suggest the feasibility of the method in conditions where higher mechanical strength is requested.

Influence of hormonal status of patients with cystic disease on the composition of cyst fluid and breast cancer risk.

The relationship between the composition of breast cyst fluid (BCF), the menstrual status and in addition some endocrine events in the history of patients (n = 131) with gross cystic breast disease was investigated. The dehydroepiandrosterone (DHA) levels in type II (K+/Na+ < 1) cysts of the follicular group were significantly higher compared to the type II cysts of the luteal or postmenopausal groups. For testosterone a significant difference existed between the type I (K+/Na+ > or = 1) follicular and type I postmenopausal groups. Estrone levels were significantly higher in type I BCF of patients in the luteal phase compared to both the follicular and postmenopausal type I cysts. Progesterone levels were lowest in the postmenopausal subgroups (both in type I and II cyst). Significant correlations were found between the number of pregnancies and the levels of DHA-sulfate and also progesterone in BCF. DHA levels were correlated with the period of lactation. The K+/Na+ ratios were the lowest in women who lactated for the longest period. The estrone was lowest in BCF of current oral contraceptive (o.c.) users while the estradiol was lowest in patients who had never used o.c. A history of previous o.c. use was associated with a significantly high mean DHA level. A significantly higher DHA and lower testosterone level were demonstrated in BCF of patients who had some previous gynecological interventions. The composition of BCF and the "life of cysts" and thus the rate of breast cancer risk may depend on hormonal status during the menstrual cycles or postmenopause and also on endocrine history of patients.

5-ethyl-2'-deoxyuridine, a modulator of both antitumour action and pharmacokinetics of 5-fluorouracil.

The aim of the present studies was to elucidate the effects and optimal modulatory conditions of 5-ethyl-2'-deoxyuridine (EtdUrd) on the antitumour efficacy, pharmacokinetics and catabolism of 5-fluorouracil (5-FU) on Colon-26 and Colon-38 murine tumours. HPLC and GC-MS techniques were used to measure the concentrations of 5-FU, dihydro-5-fluorouracil, EtdUrd, 5-ethyluracil and uridine in the plasma and that of 5-FU and 5-fluoro-2'-deoxyuridine monophosphate (FdUMP) in the tumours. It was shown that EtdUrd, given 1 h before 5-FU, selectively enhanced the antitumour action of 5-FU, without significantly increasing its toxic side-effects, thus resulting in an approximately three times higher therapeutic index. Pharmacokinetic studies revealed that 1 h after 400 mg/kg EtdUrd administration - i.e. at the time of 5-FU treatment - the plasma concentration of EtdUrd was 269 microM, and that of 5-ethyluracil, as the major metabolite of EtdUrd, was 421 microM. It is of interest that EtdUrd pretreatment did not change the maximal plasma concentration of 5-FU; however, the half-life of the terminal elimination increased from 114.5 min to 171.2 min and thus the mean residence time of 5-FU rose significantly (P < 0.05). After the combined treatment, the maximal concentration of dihydro-5-fluorouracil in the plasma decreased from 61.06 microM to 29.70 microM (P < 0.01). The intratumoral concentrations of 5-FU were 34%-158% higher 6-96 h after the combined treatment than after the single 5-FU treatment. EtdUrd also caused a moderate increase in the intratumoral level of FdUMP. It is noteworthy, that EtdUrd increased the endogenous uridine concentration in the plasma from 18 microM to a maximum of 249 microM, and the level remained high for longer than 6 h. The present studies indicate that EtdUrd enhances the therapeutic index of 5-FU by reducing the catabolism, prolonging the plasma and intratumoral concentrations of 5-FU and, at the same time, offering protection to normal organs by increasing the endogenous uridine level.

[Principles of drug therapy of malignant tumors: determinants of the efficacy of cytostatic agents]. / A rosszindulatú daganatok gyógyszeres terápiájának irányelvei: a citosztatikumok hatékonyságának feltételei.

Successful therapy with the anticancer cytostatic agents are remarkably dependent on the appropriate dosage and schedules of the treatment. To this end however it is highly important to have insights into the tumorbiological, pharmacokinetics, pharmacobiochemical and molecular-biological factors which underline the efficacy of the cytostatic agents. The present communication intend to provide a comprehensive survey for the medical doctors attending cancer patients concerning the mode of action of those anti tumour agents traditionally classified as alkylating agents, antimetabolites, topoisomerase inhibitors, and antimitotic agents. It will be emphasized that the currently available agents show no specific action on the malignant cells because they are inhibitors of cell proliferation and target certain molecular mechanism implicated in the cell cycle. The basis for the relatively selective antitumour action are rather different for the various cytostatic agents, nevertheless the differential activation and also the repair capacity in the tumour and in the normal organs offer an explanation for the agents acting directly or indirectly on DNA. Knowledge of the mode of action of the cytostatic agents offer not only a better understanding for their therapeutic failure but gives guidelines for those tumorbiological features which are necessary for their efficacy. In the last years substantial data have accumulated which indicate the possibility to improve the therapeutic action by modulating the pharmacokinetic or the pharmacobiochemical determinants of the antitumour drug action.

Modulation of 5-fluorouracil by 5-ethyl-2'-deoxyuridine on cell lines expressing different dihydropyrimidine dehydrogenase activities.

The purpose of the present study was to clarify the significance of the inhibition of dihydropyrimidine dehydrogenase (DPD) in the modulation of 5-fluorouracil (5-FU) action by 5-ethyl-2'-deoxyuridine (EUdR). Four human cell lines, which differed in their susceptibility to 5-FU and in their DPD activity, were selected as biological objects. Several other enzymes of pyrimidine metabolism, i.e. thymidylate synthase (TS), thymidine kinase (TK) and pyrimidine nucleoside phosphorylase (PNP), which might be involved in the 5-FU action were also studied to elucidate their potential role in the modulation of 5-FU cytotoxicity. Two out of the four cell lines, i.e. COLO1 and SW620, showed low (57 and 28 pmol/min/mg protein) and the other two cell lines, i.e. CAL51 and CAL33, showed high (235 and 184 pmol/min/mg protein) DPD activity, respectively. In our study, contrary to our expectation, no correlation between the DPD and TS activity of the cell lines and their 5-FU sensitivity could be observed. EUdR alone was cytotoxic only on CAL33 cells in a concentration below 1 mM (IC50=194 microM) which might be due to the high TK activity (857 pmol/min/mg protein) measured in this cell line, favoring the formation of the phosphorylated nucleotides EdUMP and EdUTP indispensable for the inhibition of TS and DNA polymerase, respectively. Surprisingly, although EUdR by metabolizing to EUra was able to reduce the high activity of DPD in CAL33 and CAL51 cells by 47 and 55%, respectively, no potentiation of the 5-FU action occurred on these cell lines. On the contrary, enhancement of the 5-FU cytotoxicity was demonstrated on COLO1 and SW620 cells with low DPD activity. Our findings suggest that the 5-FU modulatory action of EUdR may be directed on other molecular targets than DPD as well, i.e. the augmentation of TS inhibition by EdUMP as demonstrated on SW620 cells might be one of these mechanisms.

Putative role of dihydropyrimidine dehydrogenase in the toxic side effect of 5-fluorouracil in colorectal cancer patients.

Dihydropyrimidine dehydrogenase (DPD) is the first and rate limiting enzyme in the catabolism of 5-fluorouracil (5-FU). It has been reported from various laboratories that the plasma concentration of 5-FU was influenced by DPD activities in various normal human organs (e.g. liver or lymphocytes). Since the congenital deficiency in DPD caused severe, in some cases lethal, FU-related toxicity, it was decided to collect data about the DPD activity in colorectal cancer patients in order to investigate the possible correlation between the enzyme activity and appearance of the side effects of 5-FU. Assuming that DPD activity in lymphocytes represents the 5-FU catabolic capacity of the organism, DPD activity was determined in the lymphocytes of 48 patients with colorectal cancer after surgery during the therapeutic course with 5-FU and folinic acid. On the basis of the enzyme activity, patients were divided into three categories: low (DPD <5.03 pmol/min/10(6) lymphocytes); medium (DPD = 5.04-13.25 pmol/min/10(6) lymphocytes), and high (DPD > 13.26 pmol/min/10(6) lymphocytes) activity groups. By evaluating the toxic side effects during the 5-FU + folinic acid treatment, the following results were obtained. In the low DPD activity group, 9 of 11 patients had 5-FU-related side effects (mucositis, diarrhea, myelotoxicity, angina pectoris, hypertension). In 3 patients, no change of the therapy was needed, in 3 patients symptoms could be reversed by dose reduction of 5-FU while in 3 patients interruption of 5-FU therapy was needed. In the medium DPD activity group, mild toxicity (diarrhea, transitory hypertension) occurred in 5 of 29 and in the high activity group (diarrhea) in 1 of 8 patients, respectively. In these last two groups, no dose reduction of 5-FU was necessary. The present study furnished further evidence for the possible correlation between the 5-FU side effects and DPD function. Consequently, it is recommended to measure DPD activity prior to 5-FU based chemotherapy, which might be helpful in avoiding drug-related toxicity by adjusting the dose of 5-FU individually.

Pharmacokinetics of panomifene in healthy volunteers at phase I/a study.

Panomifene (PAN) /E/-1,2,-diphenyl-1-[4-[2-(2-hydroxyethylamino)-ethoxy]-phenyl]-3, 3,3-trifluoropropene is a new original Hungarian compound and is a tamoxifen (TMX) analog. In the phase I/a study presented here the human tolerance, pharmacokinetics and endocrine effects of a single oral dose of panomifene were evaluated in healthy, post-menopausal, female volunteers. As to the dose escalation, pharmacokinetic studies were carried out at doses of 24, 48 and 96 mg in two volunteers, and 120 mg in one volunteer. To find a suitable dose or dose range, for further evaluation of the drug detailed pharmacokinetics were performed at a selected dose level (24 mg) in 10 volunteers. The pharmacokinetic study showed considerable interindividual variability of the parameters, and only a medium correlation between dose and AUC (r=0.876). At the selected dose level (24 mg p.o.) the peak concentration of the plasma was 67.7 +/- 17.4 ng/ml (Cmax(meas)), the time to peak was 3.6 +/- 1.8 h (t[max(meas)]). The mean of the terminal half-life was 70.0 +/- 23.1 h (t(1/2beta)). The area under the plasma concentration time curve (AUC) calculated by the kinetic equation (AUCcalc) was 4814 +/- 1172 and by the trapezoidal rule (AUCtrap) was 4612 +/- 1357 (ng/ml) h.

[Determination of dihydropyrimidine dehydrogenase in the prediction of toxic side effects of 5-fluorouracil]. / A dihidropirimidin-dehidrogenáz meghatározás jelentösége az 5-fluorouracil toxikus mellékhatásainak elörejelzésében.

In the chemotherapy of colorectal cancers the most frequently given drug is 5-fluorouracil, which in certain cases reduces or delays the appearance of the local recurrence or metastasis. It is well known that the patient's response to 5-fluorouracil is very different concerning both, effects and side effects. More than 80% of the infused drug is catabolised in the first 20 minutes after the treatment. The first and rate limiting enzyme of the catabolism is dihydropyrimidine dehydrogenase, which has the highest activity in the liver and lymphocytes. The activity of this enzyme shows correlation with the blood level of 5-fluorouracil. The deficiency of this enzyme caused severe, in some cases lethal toxicity, its congenital deficiency is responsible for familial pyrimidinaemia. Authors intended to collect data about the dihydropyrimidine dehydrogenase activity of colorectal cancer patients, in order to screen enzyme deficiency or very low enzyme activity, which might be in connection with the appearance of severe side effects, moreover to determine the optimal dose of 5-fluorouracil before the treatment. Dihydropyrimidine dehydrogenase activity was determined in the lymphocytes of 48 colorectal cancer patients, treated by 5-fluorouracil, at the beginning of each cytostatic cycle. The enzyme activity of the patients was between 1.2 and 24.4 pmol/min/10(6) lymphocyte. The value of the enzyme activity fluctuated in a range, characteristic for the individual patients and this value was not modified by the 5-fluorouracil treatment. Dividing the patients in two groups, low (lower than 5 pmol/min 10(6) lymphocyte) and high (higher than 15 pmol/min 10(6) lymphocyte) dihydropirimidine dehydrogenase activity, we found that decrease in the white blood cell number and appearance of the side effects occurred with much higher frequency in the low activity group which resulted in the reduction of the dose or in more serious cases interruption of the treatment. Authors conclude that the determination of the dihydropyrimidine dehydrogenase activity in the lymphocytes is a valuable method in the prediction of the toxic side effects of 5-fluorouracil, in the screening of the congenital enzyme deficiency and in the individualization of the 5-fluorouracil dosage.

[Steroid hormone receptors in squamous cell carcinoma of the head and neck]. / Szteroidhormon-receptorok vizsgálata fej-nyak laphámrákban szenvedó betegeken.

In this study we have tried to find new prognostic markers to extend the therapeutical modalities for patients with head and neck squamous cell carcinoma. During evolution the development of the pharyngolaryngeal region differs in males and females, therefore this region can be considered as one of the target organs for sex steroids. Some of the tumours, originating from this area, contain hormone receptors that theoretically makes them susceptible for hormone therapy. Therefore the real concentration of steroid receptors is of great clinical importance. We determined the estradiol, progesterone and testosterone receptor content using biochemical method in tumour tissue of 33 male patients with head and neck squamous cell carcinoma. The receptors in the macroscopically intact mucosa in 15 of all tumour cases were also measured. The patients were followed for 18-24 month after operation and postoperative irradiation performed according to the protocol of the Head and Neck Surgery department. There were 26/33 (79%) estradiol receptor positive, 14/33 (42%) progesterone receptor positive and 18/30 (60%) testosterone receptor positive cases among the tumour samples. The healthy mucosa samples were positive in 6/15 (40%), 2/15 (13%) and 3/15 (20%) of cases, respectively. The differences in proportion of positive status between tumour and healthy mucosa was statistically significant. We established that during the control period the highest rate of the tumour-free survival was in the estradiol receptor positive, progesterone receptor negative group. Consequently the steroid receptor status of head and neck squamous cell carcinomas might help in assessing the prognosis of survival, and in possible choice for endocrine treatment, in order to complete the complex tumour therapy.

Modulation of 5-fluorouracil in mice using uridine diphosphoglucose.

Uridine diphosphoglucose (UDPG) is a precursor of uridine that can be used as a rescuing agent from 5-fluorouracil (5FU) toxicity. Four doses of UDPG (2000 mg/kg i.p. or p.o. at 2, 6, 24, and 30 h after 5FU bolus) allowed the escalation of a weekly bolus of 5FU from 100 mg/kg (5FU100) to 150 mg/kg (5FU150) in healthy and tumor-bearing BALB/c, C57/BI, and CD8F1 (BALB/c x DBA/8) mice. 5FU150 without rescuing agents is not tolerated by the animals. When followed by UDPG, on the contrary, it is possible to increase the dose of 5FU even when it is modulated by leucovorin. Toxicity was the same for 5FU100 and 5FU150 + UDPG, and the nadir values (expressed as a percentage of pretreatment values) were 83 and 85% for weight, 45 and 45% for hematocrit, and 45 and 61% for leukocytes, respectively. Platelets were not affected by treatment. A protective effect was also shown for the gastrointestinal tract. The enzymes thymidine kinase, maltase, and sucrase were measured in the intestinal mucosa at different times after 5FU treatment with or without UDPG rescue. Even if the nadir values in enzyme activities were similar in mice receiving or not receiving UDPG, the pattern of recovery showed that cell repopulation was more rapid in the group treated with UDPG. 5FU150 + UDPG had enhanced antitumor activity against CD8F1 mammary carcinoma and against the resistant tumor Colon 26 (tumor doubling time 1.9 days for controls, 8.5 days for 5FU100, 13.7 days for 5FU150 + UDPG, and 15.9 days for 5FU150 + leucovorin + UDPG). We demonstrated that UDPG administered at 2, 24, and 30 h after 5FU100 does not reduce the antitumor activity of 5FU in two sensitive tumors (Colon 38 and Colon 26-10). In conclusion, UDPG is a promising rescuing agent for 5FU; it reduces the toxic side effects and increases the therapeutic index.

Effect of ethyldeoxyuridine on 5-fluorouracil-induced neutropenia.

It is supposed that the toxic effect of 5-fluorouracil (5FU) on tumour cells may be increased by pretreatment with ethyldeoxyuridine (EDU). We studied the effect of this combination on neutrophil count in mice. Our present studies demonstrated that the neutropenia induced by 5 x 20 mg/kg 5FU became more severe when each dose of 5FU was preceded by 200 mg/kg EDU.

Determination of panomifene in human plasma by high-performance liquid chromatography.

An ion-pair HPLC method was developed for the determination of the antiestrogenic drug panomifene, (E)-1,2-diphenyl-1-(4-[2-(2-hydroxyethylamino)ethoxy]phenyl)-3,3,3 - trifluoropropene, in human plasma. Tamoxifen, 20 ng in 1 ml of plasma, was used as an internal standard. The compounds were isolated from plasma by liquid-solid extraction. Fluorescence detection was achieved by on-line photochemical conversion of the compounds into highly fluorescent phenanthrene derivatives. The sensitivity of the method was 1 ng/ml. The within-day and between-day precision, linearity, extraction recovery and stability of panomifene in plasma and in deproteinized plasma were determined for validation of the method. The method is suitable for measuring plasma levels of panomifene and tamoxifen and for pharmacokinetic studies.

[Hormonal and biochemical characterization of breast cyst fluid in gross cystic mastopathy]. / A nagycisztás mastopathia cisztafolyadékának hormonális és biokémiai jellemzése.

A total of 93 breast cyst fluids (BCF) obtained by needle aspiration of women suffering from gross cystic mastopathy was hormonally investigated. The mean age of the patients was 45 years (range 27-65). Estradiol (E2), progesterone (PROG), testosterone (TE), prolactin (PROL), estriol (E3), dehydroisoandrosterone and its sulfate (DHA, DHA-S) levels were investigated in the BCF and in the respective sera. Tumour marker beta-HCG and CA 15-3 as well as cations (K+, Na+) were determined, too. E2, E3, PROG, TE, PROL, DHA, DHA-S and K+ showed significant accumulation in the BCF compared to the serum values. The K+/Na+ ratio proved to be a useful tool to divide cysts into type I (> or = 1), type II (< 1 but > or = 0.1) and type III (< 0.1) subgroups. In case of type I BCF, higher E2, DHA, DHA-S and PROL levels could be detected, while PROG and TE contents proved to be the highest in type II cysts. These findings indicate that the type I BCF is a marker for "active" GCD of the breast and suggest that it may be associated with increased breast cancer risk. It is suggested therefore when macrocysts are aspirated, sex steroids, steroid hormone precursors and cations in the BCF should be examined routinely, and women with type I cysts should be controlled carefully.

Influence of uridine treatment in mice on the protection of gastrointestinal toxicity caused by 5-fluorouracil.

C57B1/6 male normal mice were treated with 5-FU (200 mg/kg i.p.) alone or in combination with a bolus injection of uridine (2 x 3500 mg/kg i.p.) in order to study the potential rescue effect of uridine on 5-FU-induced gastrointestinal toxicity. 5-FU alone inhibited the activity of different enzymes (thymidine-kinase, alkaline-phosphatase, sucrase and maltase) which were selected as the early biochemical markers for the injured small intestinal mucosa. The nadir of the enzyme activities was between 24-96 hrs after 5-FU administration, and the complete regeneration took a week. In the combination of 5-FU plus uridine bolus injection the seriousness of gastrointestinal damage caused by 5-FU was significantly (p < 0.05) milder and the recovery time was shorter by 2 days. Comparing the rescue effect of two dose schedules of uridine, both high dose (2 x 3500 mg/kg) or repeated lower doses of uridine (7 x 800 mg/kg) resulted in a similar protection from the gastrointestinal side effect of 5-FU.

Biochemical consequences of 5-fluorouracil gastrointestinal toxicity in rats; effect of high-dose uridine.

Selective protection of the normal host tissues from the toxic effects of anticancer agents would allow the use of higher, probably more effective, doses of the drugs. It has been demonstrated that delayed high-dose uridine administration after 5-fluorouracil decreases the extent of myelosuppression and causes faster regeneration of the bone marrow. We studied the biochemical consequences of the gastrointestinal toxicity caused by 5-fluorouracil and the potential of high-dose uridine treatment to influence these adverse effects. 5-Fluorouracil caused dose-related decreases in the biochemical parameters (thymidine kinase, sucrase, maltase, alkaline phosphatase) selected as early markers of the impaired metabolic activity of the intestinal mucosa. The nadir of the biochemical changes was reached between 24 h and 72 h after 5-fluorouracil treatment, and complete regeneration of the mucosa took 6-7 days. Delayed high-dose uridine administration failed to mitigate the severity of the gastrointestinal damage that ensued after 5-fluorouracil treatment, but caused significantly earlier regeneration of the mucosa.

[Clinical pathology of gastric mucosal dysplasia]. / A gyomornyálkahártya dysplasiájának klinikai patológiája.

The stomach cancer develops on dysplastic lesions of gastric mucosa. It can be found in every precancerous condition, as chronic gastritis, gastric adenoma, giant rugal hypertrophy, chronic peptic ulcer, gastric stump after partial resection, pernicious anaemia. So, this dysplastic change is not a specific lesion. Different classifications are known for grading of gastric dysplasia. Authors evaluated them compared with each other. The signs of dysplasia were studied in 306 gastric aimed biopsy specimens from 233 patients between 1979-1990. In this material severe dysplasia occurred in 20.6%. It means a frequency of 0.84% regarding all gastric endoscopies in the same period of time. The endoscopic investigation revealed a protruded lesion in 18.5% and excavated one in 45.9%. What is very important, local change could not be detected in 35.6%. Follow-up study could be performed in 49 patients in a period of 1-7 years. In this group cancer developed in five patients. By the other hand, 22 gastric carcinomas were proved amongst 233 patients. The authors' recommendation is to follow-up the patients bearing gastric dysplasia at least during 10 years after the diagnosis.

Alkaline phosphatase activity in human and rat liver tumors.

Enzyme activity measurements of alkaline phosphatase in surgically removed human liver tumors showed elevated level of the enzyme in 6 focal nodular hyperplasias, reduction in 8 primary hepatocellular carcinomas, and no change in the 4 adenoma samples. The activity represented liver type of alkaline phosphatase nearly in all cases because it could be inhibited by L-homoarginine more extensively than by L- phenylalanine. Studies on polyacrylamide gel electrophoresis indicated the presence of a variant type isoenzyme only in one focal nodular hyperplasia and in two hepatocellular carcinomas, one of which showed a fibrolamellar structure whereas the other was associated to cirrhosis. The importance of the elevated amount of connective tissue in the tumor, resulting in an isoenzyme shift of alkaline phosphatase, received substantial support upon comparing chemically induced rat liver tumors with and without cirrhosis.

Isolation of viable intestinal epithelial cells and their use for in vitro toxicity studies.

The application of the collagenase portal vein perfusion technique for the isolation of intestinal cells resulted in the preparation of highly viable enterocytes. Cell viability was found to be greater than 90% as tested by LDH release and Trypan blue exclusion techniques. According to the results of marker enzyme determinations, collected cells were mostly of matured villus type, characterized by high disaccharidase and very low thymidine kinase activity. In vitro treatment of the isolated cells with the anticancer agent cis-diamminedichloroplatinum (II) caused decrease of the metabolic processes, i.e. glucose oxidation and protein synthesis, demonstrating that beyond the production of DNA-crosslinks other mechanisms may play a role in the cytotoxic effect of the drug. It should be stressed, however, that prolonged incubation of the cell suspension over 30 min at physiological temperature may itself lead to gradual decrease of the viability and to disturbance of the metabolic activity of the cells.