Halogenated Pyrimidine Analogs Research Articles

Incorporation of 5-Bromo-2′-deoxyuridine into DNA and Proliferative Behavior of Cerebellar Neuroblasts: All That Glitters Is Not Gold

The synthetic halogenated pyrimidine analog, 5-bromo-2′-deoxyuridine (BrdU), is a marker of DNA synthesis. This exogenous nucleoside has generated important insights into the cellular mechanisms of the central nervous system development in a variety of animals including insects, birds, and mammals. Despite this, the detrimental effects of the incorporation of BrdU into DNA on proliferation and viability of different types of cells has been frequently neglected. This review will summarize and present the effects of a pulse of BrdU, at doses ranging from 25 to 300 µg/g, or repeated injections. The latter, following the method of the progressively delayed labeling comprehensive procedure. The prenatal and perinatal development of the cerebellum are studied. These current data have implications for the interpretation of the results obtained by this marker as an index of the generation, migration, and settled pattern of neurons in the developing central nervous system. Caution should be exercised when interpreting the results obtained using BrdU. This is particularly important when high or repeated doses of this agent are injected. I hope that this review sheds light on the effects of this toxic maker. It may be used as a reference for toxicologists and neurobiologists given the broad use of 5-bromo-2′-deoxyuridine to label dividing cells.

Enhancement of IUdR Radiosensitization by Low-Energy Photons Results from Increased and Persistent DNA Damage.

Low-energy X-rays induce Auger cascades by photoelectric absorption in iodine present in the DNA of cells labeled with 5-iodo-2’-deoxyuridine (IUdR). This photoactivation therapy results in enhanced cellular sensitivity to radiation which reaches its maximum with 50 keV photons. Synchrotron core facilities are the only way to generate such monochromatic beams. However, these structures are not adapted for the routine treatment of patients. In this study, we generated two beams emitting photon energy means of 42 and 50 keV respectively, from a conventional 225 kV X-ray source. Viability assays performed after pre-exposure to 10 μM of IUdR for 48h suggest that complex lethal damage is generated after low energy photons irradiation compared to 137Cs irradiation (662KeV). To further decipher the molecular mechanisms leading to IUdR-mediated radiosensitization, we analyzed the content of DNA damage-induced foci in two glioblastoma cell lines and showed that the decrease in survival under these conditions was correlated with an increase in the content of DNA damage-induced foci in cell lines. Moreover, the follow-up of repair kinetics of the induced double-strand breaks showed the maximum delay in cells labeled with IUdR and exposed to X-ray irradiation. Thus, there appears to be a direct relationship between the reduction of radiation survival parameters and the production of DNA damage with impaired repair of these breaks. These results further support the clinical potential use of a halogenated pyrimidine analog combined with low-energy X-ray therapy.

Effects of prenatal exposure to 5-bromo-2′-deoxyuridine on reproductive function in male mouse offspring

The effect of the halogenated pyrimidine analog 5-bromo-2′-deoxyuridine (BrdU), on reproductive functions of male mouse offspring treated prenatally was studied. BrdU was administered intraperitoneally to pregnant ICR mice at 100 mg/kg/d on days 8 through 13 of gestation, and at 100 or 200 mg/kg/d on days 14 through 18 of gestation. Dams were allowed to deliver naturally. Male offspring were aged for 10 weeks and then cohabited with untreated female mice for assessment of reproductive performance. Histopathologic examination of the testes and pituitary, sperm analysis, and determination of plasma testosterone concentrations of offspring at 12 weeks of age were performed. In the subsequent study, pregnant ICR mice were treated with 200 mg BrdU/kg on day 10,13, or 15 of gestation. The embryos or fetuses were obtained from mothers from 6 to 48 h after treatment, and pyknotic cells in the ventricular zone of the telencephalon were counted. There was a significant decrease in body weight gain of offspring in all of the BrdU-treated groups. A marked decrease in copulation rate was noted in the male offspring of dams treated on days 8 through 13 of gestation, whereas no significant decreases in copulation and fertility rates were found in the male offspring of dams treated on days 14 through 18 of gestation. Neither histopathologic examination of testes nor sperm analyses revealed adverse effects of this compound, whereas cysts in the pars distalis of the pituitary were observed in the male offspring treated on days 8 through 13 of gestation. Dilatation of the lateral ventricles was also observed in male offspring at 12 weeks of age in the group treated on days 8 through 13 of gestation. The incidence of pyknotic cells in the ventricular zone of embryos was markedly increased 24 h after treatment on day 10 of gestation. These results indicate that the impaired fertility of the male offspring of dams treated with BrdU on days 8 through 13 of gestation may be due to the effects of BrdU exposure on central nervous system function that result in loss of libido rather than to the direct effects of this compound on the male reproductive organs.

Radiotherapy of glioblastoma

Glioblastoma cells appear to be inherently radioresistant and to present a significant fraction of hypoxic cells. The most significant prognostic factors to compare results achieved in several series of patients are the age, performance status and quality of surgical resection. Several randomized trials have provided evidence supporting the efficacy of radiation therapy in the treatment of glioblastoma. Prescription of a 60-Gy dose delivered according to a conventional dose-fractionation scheme (single daily fractions of 1.7 to 2 Gy five times per week) in a target volume with a 2-3 cm margin of tissue surrounding the perimeter of the contrast enhancing lesion on computerized tomography and magnetic resonance imaging is derived from observations made in several retrospective and prospective studies. Evidence of improvement in survival was observed neither in patients receiving hyperfractioned and accelerated radiotherapy, nor in patients for whom radiation sensitizers such as nitroimidazole compounds or halogenated pyrimidine analogs were associated to radiation therapy. The addition of nitrosourea to radiotherapy increases the 2-year survival rate by about 10%. Combination of full-dose external beam radiotherapy and brachytherapy or radiosurgery boost in selected patients with glioblastoma leads to an increase in the median survival, while external beam radiation alone in patients with similar prognosis does not.

Radiosensitization with carotid intra‐arterial bromodeoxyuridine ± 5‐fluorouracil biomodulation for malignant gliomas

Bromodeoxyuridine (BUdR), a nonhypoxic radiosensitizing drug, is a halogenated pyrimidine analog that is incorporated into the DNA of dividing cells in a competitive process with thymidine. BUdR sensitizes cells to radiation therapy. 5-Fluorouracil (5-FU) inhibits the endogenous synthesis of thymidine, resulting in increased incorporation of the BUdR. Neurons and glial cells have a very low mitotic rate; they will not incorporate BUdR and will not be sensitized. BUdR and 5-FU are best delivered intra-arterially (IA) because of their regional advantage. We infused BUdR +/- 5-FU over 8 1/2 weeks, before and during 59.4-Gy focal conformal external beam radiation therapy, through a permanently implanted pump with a catheter placed retrograde through the external carotid artery to the carotid bifurcation. Sixty-two patients with grades III or IV glioma were entered into one of two trials, with 23 patients receiving BUdR alone and 39 patients receiving BUdR + 5-FU. The maximum tolerated dose (MTD) of BUdR alone was 400 mg/m2/d for 8 1/2 weeks. The Kaplan-Meier median survival (KMS) was 20 months. In the BUdR + 5-FU trial, the MTD of BUdR was also 400 mg/m2/d and 5-FU was 5 mg/m2/d with a KMS of 17 months. The KMS of all 62 patients in both trials 1 and 2 was 18 months. Pathologic grading used both the original World Health Organization (WHO) and 1993 modified WHO systems. The KMS of grade IV patients was 13.8 months (48 patients) with the original system and 17 months (58 patients) with the modified system.(ABSTRACT TRUNCATED AT 250 WORDS)

The management of malignant gliomas with radiation therapy: Therapeutic results and research strategies

I t is estimated that in 1991, 16,700 new cases of primary central nervous system tumors will be diagnosed in the United States, and that 11,500 patients will succumb to their disease.’ Malignant gliomas comprise 33% to 45% of primary brain tumors,2’3 and of these, nearly 85% are glioblastoma multiforme.2’4 The 1980 patterns of care survey for brain tumor patients conducted by the Joint Section on Central Nervous System Tumors of the American Association of Neurological Surgeons and Congress of Neurological Surgeons and the American College of Surgeons Commission on Cancer found that the 5-year survival rate for patients with anaplastic astrocytomas was 18%, while it was only 5% for those with glioblastoma multiforme.* With few exceptions, there has been little improvement in the outcome of patients with malignant gliomas over the past decade. Prospective clinical studies have shown both the efficacy and the shortcomings of conventional radiation therapy in the treatment of malignant gliomas.5,6 The response of these lesions to standard radiotherapeutic techniques is limited by their striking inherent radioresistance’ and the radiosensitivity of the surrounding normal brain tissue.’ In addition to pursuing more effective chemotherapy programs, research strategies have focused on methods designed to enhance the effectiveness of external irradiation using hypoxic cell radiosensitizers, halogenated pyrimidine analogs, and altered fractionation schemes, or to selectively augment the tumor dose using interstitial brachytherapy. Single-institution and cooperative group studies using these approaches are now in progress. This review examines

Intra-arterial 5-bromo-2-deoxyuridine (BUdR) radiosensitization with external beam radiation in rhesus monkeys: toxicity study.

A primate toxicity study was performed to test the hypothesis that BUdR does not increase the likelihood of unilateral or bilateral central nervous system damage secondary to radiation therapy. BUdR, a halogenated pyrimidine analog is incorporated into DNA of dividing cells and sensitizes them to radiation. It is best given unilaterally, intra-arterially by continuous infusion because of its regional advantage (Rd) estimated to be between 11 and 16. Six rhesus monkeys were implanted with a Model 400 Infusaid pump perfusing the right internal carotid artery. Three of the six monkeys received intra-arterial (IA) BUdR infusion plus whole brain external beam radiation (6,000 R over 6 weeks) and three received radiation alone. The two BUdR treated animals completing radiation developed symmetric bilateral high signal aberrations on MRI in the frontal, parietal, and occipital centrum semiovale and corona radiata at nine months. At autopsy, confluent microinfarcts were found to correspond topographically to the MRI abnormalities. In the radiation alone group, two animals had normal MRI and autopsy while the third animal had bilateral MRI high signal aberrations develop sequentially with corresponding microinfarcts at autopsy. These changes were greater in severity than those seen in the BUdR treated animals. We support previous evidence that there is differential intraspecies sensitivity to radiation. We find that BUdR produces no unilateral potentiation of radiation toxicity.

Enhancement of X ray induced DNA damage by pre-treatment with halogenated pyrimidine analogs

The use of the halogenated pyrimidine analogs, including bromodeoxyuridine (BrdUrd) and iododeoxyuridine (IdUrd), as potential clinical radiosensitizers is an area of renewed clinical research. Cellular radiation effects of these analogs using clinically achievable steady state plasma levels (10 −7−10 −5 M) were measured in vitro using exponentially growing Chinese hamster V79 cells. Radiation response was determined by clonogenic survival and by the production of DNA single strand (SSB) and double strand breaks (DSB) using filter elution techniques. Replacement of thymidine in DNA by BrdUrd or IdUrd was also determined. Drug exposure of BrdUrd or IdUrd for two population doublins (17 hr) prior to irradiation resulted in a progressive reduction of n̄ and D 0 compared to untreated controls over the drug dose range of 10 −7-10 −5 M. The percent thymidine replacement increased from 1% at 10 −7 M to 5% at 10 −6 M to 23% at 10−5 M. Using a 17 hr exposure at 10 −5 of BrdUrd of IdUrd, the production of SSB and DSB was increased by ≥2× and ≥1.5× respectively. No differences in the kinetics of repair of SSB and DSB were found following 3 hr of post-irradiation repair. We conclude that in this in vitro model, there appears to be a direct relationship between percent thymidine replacement, reduction of radiation survival parameters (n, D 0), and the production of DNA strand breaks in the clinically achievable dose range of these halogenated pyrimidine analogs. These filter elution assays may be adapted to in vivo studies and possibly may allow for monitoring of radiation-induced DNA damage and repair in clinical tumor specimens treated with these radiosensitizers.

The use of halogenated thymidine analogs as clinical radiosensitizers: Rationale, current status, and future prospects: Non-hypoxic cell sensitizers

The halogenated pyrimidine analogs, bromodeoxyuridine (BUdR) and iododeoxyuridine (IUdR) have been recognized as potential clinical radiosensitizers for over two decades. In vivo and in vitro experimental studies document that radiosensitization is directly dependent on the amount of thymidine replacement in DNA by these analogs. Early clinical studies in Japan using selective intra-arterial infusions of BUdR and conventional fractionated radiation suggested improved survival in patients with primary brain tumors, although there were significant catheter-related complications. Based on recent in vivo and clinical pharmacology studies on continuous intravenous infusions of these drugs, clinical trials are underway evaluating the potential of radiosensitization in high grade gliomas and other poorly radioresponsive tumors using the technically safer intravenous route of administration. In this paper, we review the basic strategy for the use of these analogs, the ongoing clinical trials and the potential areas for future experimental and clinical studies.

The effects of pretreatment of synchronized CHO cells with 5-BUdR and TdR on chromosomal damage and cell death.

Chinese hamster ovary (CHO) cell populations were exposed independently to thymidine (TdR) or 5-bromodeoxyuridine (5-BUdR) for either one cell generation (approximately 18 h) or two (36 h) and subsequently synchronized in mitosis either by chemical arrest with colcemid or by mechanical shaking. Upon subsequent X-irradiation at various stages in the cell cycle, the synchronized populations were analyzed for cellular lethality and chromosomal damage. This investigation confirmed that each radiation response was "age-dependent" and essentially similar for both methods of synchrony. Cells irradiated in mitosis were found to be most radio-sensitive while cells irradiated during the DNA synthesis stage were most radio-resistant. A comparative analysis revealed that the two responses of radiation were more pronounced for cells labelled with 5-BUdR than they were for TdR-labelled cells. A further enhancement of radiation responses was noted for cells that were grown in the presence of the halogenated pyrimidine analog for two cell generations, thus re-affirming that the nucleus is a primary target for radiation damage. The comprehensive conclusion sought with regard to the degree of correlation between chromosomal damage and loss of cellular reproduction integrity awaits such information as the molecular organization of mammalian chromosomes and the mechanisms by which aberrations are produced within these chromosomes.

Preferential Radiosensitization of Mouse Sarcoma Relative to Normal Skin by Chronic Intra-Arterial Infusion of Halogenated Pyrimidine Analogs<xref ref-type="fn" rid="FN2">2</xref>

Preferential Radiosensitization of Mouse Sarcoma Relative to Normal Skin by Chronic Intra-Arterial Infusion of Halogenated Pyrimidine Analogs<xref ref-type="fn" rid="FN2">2</xref>