Biomedical subjects

W Fan

Publications and source records attributed to W Fan.

At least 55 records · Page 3Linked to original sources

PubMed2000

Prolyl endopeptidase inhibitory activity of fourteen Kampo formulas and inhibitory constituents of Tokaku-joki-to.

Prolyl endopeptidase (PEP, EC 3.4.21.26) is an enzyme playing a role in the metabolism of proline-containing neuropeptides which have been suggested to be involved in learning and memory processes. In screening for PEP inhibitors from fourteen traditional Kampo formulas, we found that Tokaku-joki-to shows a significant inhibitory activity. Examination of the constituents of the Kampo formula resulted in the isolation of two new compounds, cis-3,5,4'-trihydroxystilbene 4'-O-beta-D-(6-O-galloyl)glucopyranoside (10) and 4-(4-hydroxyphenyl)-2-butanone 4'-O-beta-D-(6-O-galloyl-2-O-cinnamoyl)glucopyranoside (16), along with twenty-five known compounds, cinnamic acid (1), protocatechuic acid (2), gallic acid (3), torachrysone 8-O-beta-D-glucoside (4), emodin (5), emodin 8-O-beta-D-glucoside (6), 3,5,4'-trihydroxystilbene 4'-O-beta-D-glucopyranoside (7), 3,5,4'-trihydroxystilbene 4'-O-beta-D-(2-O-galloyl)glucopyranoside (8), 3,5,4'-trihydroxystilbene 4'-O-beta-D-(6-O-galloyl)glucopyranoside (9), 4-(4-hydroxyphenyl)-2-butanone 4'-O-beta-D-glucopyranoside (11), isolindleyin (12), lindleyin (13), 4(4-hydroxyphenyl)-2-butanone 4'-O-beta-D-(2,6-di-O-galloyl)glucopyranoside (14), 4-(4-hydroxyphenyl)-2-butanone 4'-O-beta-D-(2-O-galloyl-6-O-cinnamoyl)glucopyranoside (15), 1-O-galloylglucose (17), 1,2,6-tri-O-galloylglucose (18), gallic acid 4-O-beta-D-(6-O-galloyl)glucopyranoside (19), liquiritigenin (20), liquiritigenin 4'-O-beta-D-glucoside (21), liquiritigenin 7,4'-diglucoside (22), liquiritigenin 4'-O-beta-D-apiofuranosyl-(1-->2)-beta-D-glucopyranoside (23), licuroside (24), (-)-epicatechin (25), (-)-epicatechin 3-O-gallate (26) and (+)-catechin (27). Among these compounds, twelve (7-10, 14-16, 18, 19, 24-26) showed noncompetitive inhibition with an IC50 of 22.9, 3.0, 14.9, 2.8, 10.5, 0.69, 8.2, 0.44, 9.39, 26.5, 28.1 and 0.052 microM, respectively.

Drugs, Chinese Herbal↗

PubMed2000

In vitro evaluation of schedule-dependent interactions between docetaxel and doxorubicin against human breast and ovarian cancer cells.

Docetaxel, a novel member of the taxoid family, has shown greater potency than paclitaxel in the treatment of advanced breast cancer and certain other solid tumors. The promising clinical activity of docetaxel has also promoted considerable interest in combining this drug with other antitumor agents. In this study, we assessed the cytotoxic interaction between docetaxel and doxorubicin administered at various schedules to human breast and ovarian cancer cells. Through a series of in vitro assays including DNA fragmentation analyses, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, and flow cytometric analyses, we found that the antagonistic interaction occurred when tumor cells were exposed to the two drugs simultaneously or exposed to doxorubicin before docetaxel. However, no antagonism was observed when docetaxel was added before doxorubicin. Further analyses demonstrated that doxorubicin could interfere with the cytotoxic effect of docetaxel on both mitotic arrest and apoptotic cell death. In addition, biochemical examinations revealed that docetaxel could induce phosphorylation of both bcl-2 and c-raf-1, but these changes were inhibited when tumor cells were pretreated or simultaneously treated with doxorubicin. These results indicate that the interaction between docetaxel and doxorubicin is highly schedule dependent. Exposure of tumor cells to doxorubicin before docetaxel could result in pronounced antagonism. The optimal schedule for this combination might be sequential exposure to docetaxel followed by doxorubicin.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed2000

[Reconstructing the limb end defects with a free mini-flap, a report of 18 cases].

OBJECTIVE: Some operative procedures were introduced for repairing hand or foot defects using the free mini-flap from the finger or toe. METHODS: Eighteen cases of finger tip or foot wound were treated with the free mini-flap and vessel anastomosis. The size of the flap was 2 cm x 1 cm to 6 cm x 4 cm and the length of the pedicle was 2 to 4 cm. RESULTS: All the eighteen free mini-flaps survived and the results were satisfactory. CONCLUSION: The method of the mini-flap is a good option for covering of thumb, finger or foot defects when a local or an island flap is not available.

Adolescent↗

PubMed1999

Cytokine response gene 6 induces p21 and regulates both cell growth and arrest.

Cytokine response gene #6 (CR6), cloned from interleukin 2-stimulated T lymphocytes, is homologous to GADD45 and MyD118, genes which promote cell cycle arrest and apoptosis. To determine how this gene family could possibly mediate both cell survival/proliferation and cell cycle arrest/death, transfectants were generated so that the genes could be expressed ectopically, independently from their normal inducing agents. In cycling retinoblastoma protein-negative (pRb-) cells, ectopic CR6 expression blocked G2/M transition, but did not prevent G1/S transition so that endoreduplication resulted. By comparison, when CR6, GADD45, and MyD118 genes were expressed ectopically in proliferating pRb+ cells, either G1/S or G2/M transition was effectively blocked, so that there was no endoreduplication. Consistent with these findings, in proliferating pRb-cells, ectopic expression of CR6 promoted the expression of both G1 and G2/M cyclins. By comparison, in pRb+ cells, the expression of G1 cyclins was increased, while expression of the mitotic cyclins was decreased. However, in pRb+ cells, cyclin-dependent kinase activities associated with both G1 and G2/M cyclins were decreased. Moreover, ectopic expression of all three genes resulted in the expression of the CKI, p21, both in pRb- and pRb+ cells. The physiologic induction of CR6 expression by IL2 in quiescent normal human T cells occurs transiently in the first half of G1, coordinately with the expression of p21. Therefore, this gene family regulates G1 and G2, and promotes either cell growth or arrest by a common mechanism.

Carrier Proteins↗

PubMed1999

CR6: A third member in the MyD118 and Gadd45 gene family which functions in negative growth control.

MyD118 and Gadd45 are two related genes which encode for proteins that play important roles in negative growth control, including both growth suppression and apoptosis. A strategy was employed to clone new members of the MyD118 and Gadd45 family of genes. Based on alignment of the deduced amino acid sequences, one cDNA clone was found to encode for the murine homologue of human CR6, originally cloned as an IL-2 immediate-early response gene. The murine and human CR6 proteins were observed to be 97% identical, indicating that CR6 is an evolutionarily conserved protein. Analysis of CR6 expression during hematopoietic cell development associated with growth arrest and apoptotic cell death, upon exposure of hematopoietic cells to a variety of growth arrest and apoptotic stimuli, and in a variety of murine tissues, has revealed that CR6 expression differs significantly from the expression of the related MyD118 and Gadd45 genes. Nevertheless, CR6, like MyD118 and Gadd45, suppressed colony formation of human lung carcinoma H1299 cells. These data suggest that CR6 plays similar, but not identical, roles to MyD118 and Gadd45 in negative control of cell growth.

Amino Acid Sequence↗

PubMed1999

Possible mechanisms of paclitaxel-induced apoptosis.

Paclitaxel, a naturally occurring antimicrotubule agent, has been demonstrated to possess significant cell-killing activity in a variety of tumor cells through induction of apoptosis. It is currently unclear whether this finding suggests a novel mechanism of action for paclitaxel against tumor cells or just represents an end product of the well-known action of paclitaxel on microtubules and cell cycle arrest. Morphologically, a sustained block of mitosis seems to be required for paclitaxel-induced apoptosis because most apoptotic events are observed to occur in cells showing a prior mitotic arrest. However, this morphological correlation alone does not prove that paclitaxel-induced apoptosis is indeed a secondary event resulting from mitotic arrest. Instead, several lines of evidence obtained from recent studies have suggested that apoptotic cell death induced by paclitaxel may occur via a signaling pathway independent of microtubules and G2/M arrest.

Alkaloids↗

PubMed1999

Interaction of NPR1 with basic leucine zipper protein transcription factors that bind sequences required for salicylic acid induction of the PR-1 gene.

The Arabidopsis thaliana NPR1 has been shown to be a key regulator of gene expression during the onset of a plant disease-resistance response known as systemic acquired resistance. The npr1 mutant plants fail to respond to systemic acquired resistance-inducing signals such as salicylic acid (SA), or express SA-induced pathogenesis-related (PR) genes. Using NPR1 as bait in a yeast two-hybrid screen, we identified a subclass of transcription factors in the basic leucine zipper protein family (AHBP-1b and TGA6) and showed that they interact specifically in yeast and in vitro with NPR1. Point mutations that abolish the NPR1 function in A. thaliana also impair the interactions between NPR1 and the transcription factors in the yeast two-hybrid assay. Furthermore, a gel mobility shift assay showed that the purified transcription factor protein, AHBP-1b, binds specifically to an SA-responsive promoter element of the A. thaliana PR-1 gene. These data suggest that NPR1 may regulate PR-1 gene expression by interacting with a subclass of basic leucine zipper protein transcription factors.

Amino Acid Sequence↗

PubMed1999

A MHC-encoded ubiquitin-like protein (FAT10) binds noncovalently to the spindle assembly checkpoint protein MAD2.

Recently a number of nonclass I genes were discovered in the human MHC class I region. One of these, FAT10, encodes a protein consisting of two domains with homology to ubiquitin. FAT10 mRNA is expressed constitutively in some lymphoblastoid lines and dendritic cells and in certain other cells after gamma-interferon induction. FAT10 protein expression is controlled at several levels including transcription, translation, and protein stability. Yeast two-hybrid screening of a human lymphocyte library and immunoprecipitation studies revealed that FAT10 noncovalently associated with MAD2, a protein implicated in a cell-cycle checkpoint for spindle assembly during anaphase. Thus, FAT10 may modulate cell growth during B cell or dendritic cell development and activation.

Amino Acid Sequence↗

PubMed1999

Apoptotic cell death induced by baccatin III, a precursor of paclitaxel, may occur without G(2)/M arrest.

PURPOSE: Paclitaxel has been demonstrated to possess significant cell-killing activity in a variety of tumor cells by induction of apoptosis, but the mechanism by which paclitaxel leads to cell death and its relationship with mitotic arrest is not entirely clear. In this study, baccatin III, a synthetic precursor of paclitaxel, was used to analyze whether paclitaxel-induced apoptosis can be a separate event from microtubule bundling and G(2)/M arrest. METHODS: Several different methods including DNA fragmentation, flow cytometric analyses, TdT-mediated dUTP nick end labeling (TUNEL) and time-lapse video microscopy were used to analyze apoptotic cell death induced by baccatin III and its possible correlation with cell cycle distribution. RESULTS: Our results demonstrated that baccatin III could also cause apoptotic cell death in both BCap37 (a human breast cancer cell line) and KB cells (derived from human epidermoid carcinoma), but had less effect on microtubule bundling and G(2)/M arrest. Furthermore, we demonstrated that most apoptotic events induced by baccatin III were not coupled with G(2)/M arrest. Instead, these apoptotic events occurred predominantly in the cells in other phases of the cell cycle. CONCLUSION: Baccatin III, which contains the core taxane ring, is the fundamental piece of paclitaxel structure. The finding of baccatin III-induced apoptosis independent of cell cycle arrest, on the one hand, implies that the core taxane ring may play a critical role in inducing cell death and, on the other hand, suggests that paclitaxel might induce apoptosis from other phases of the cell cycle by a similar mechanism.

Alkaloids↗

PubMed1999

Active-oxygen scavenging activity of traditional nourishing-tonic herbal medicines and active constituents of Rhodiola sacra.

The active-oxygen scavenging activity of 70 traditional herbal medicines used in China and Japan as nourishing tonics were evaluated by electron spin resonance (ESR) technique, in order to evaluate their effectiveness for anti-aging and to search for new active-oxygen scavengers from natural resources. Most of the 70 herbal medicines showed scavenging activity with various intensities. Areca catechu (methanol extract), Dendrobium plicatile (methanol extract), Juglans regia (water extract), Paeonia lactiflora (methanol extract), Psychotria serpens (water and methanol extracts), Rhodiola sacra (water and methanol extracts) and Uncaria rhynchophylla (water extract) especially showed strong scavenging activity against superoxide anion radical (*O2-), while J. regia (water and methanol extracts), Morus alba (water extract) and Schisandra chinensis (water extract) revealed strong scavenging activity against hydroxyl radical (HO*). In addition, the active-oxygen scavenging activities of 19 compounds isolated from R. sacra were also examined, and hydroquinone (1), caffeic acid (3), protocatechuic acid (6), gallic acid (7), (-)-epigallocatechin 3-O-gallate (8), 3-O-galloylepigallocatechin-(4beta-->8)-epigallocatechin+ ++ 3-O-gallate (10), heterodendrin (17) and gallic acid 4-O-beta-D-glucopyranoside (19) were found to show mild or strong inhibitory activity against superoxide anion radical (*O2-), while 4-hydroxybenzoic acid (2), 3, 4-hydroxycinnamic acid (4), 6-8 and 19 inhibited hydroxyl radical (OH*). These active-oxygen scavengers may contribute, to different extents, to their anti-aging action.

Antioxidants↗

PubMed1999

Integration of NPY, AGRP, and melanocortin signals in the hypothalamic paraventricular nucleus: evidence of a cellular basis for the adipostat.

Energy stores are held relatively constant in many mammals. The circuitry necessary for maintaining energy homeostasis should (1) sense the amount of energy stored in adipose tissue, (2) sense and integrate the multiple opposing signals regarding nutritional state, and (3) provide output regulating energy intake and expenditure to maintain energy homeostasis. We demonstrate that individual neurons within the paraventricular nucleus of the hypothalamus (PVH) are capable of detection and integration of orexigenic (neuropeptide Y [NPY]) and anorexigenic (melanocortin) signals, that NPY and melanocortins are functional antagonists of each other within the PVH in the regulation of feeding behavior, and that melanocortin administration within the PVH regulates both feeding behavior and energy expenditure. These data provide a cellular basis for the adipostat within neurons in the PVH that appear to be jointly regulated by NPY- and melanocortin-responsive neurons.

Agouti-Related Protein↗

PubMed1999

Screening of crude drug extracts for prolyl endopeptidase inhibitory activity.

Prolyl endopeptidase (PEP, EC 3.4.21.26) is an enzyme to play a role in metabolism of proline-containing neuropeptides, such as vasopressin, substance P and thyrotropin-releasing hormone (TRH), which were suggested to be involved with learning and memory processes. Then, specific inhibitor of PEP is expected to have antiamnesic effects, and thus we screened forty-six water- and methanol-extracts from crude drugs selected on the basis of traditional Chinese medicine theory, for Flavobacterium prolyl endopeptidase inhibition. Among them, the water-extracts of Rhodiola sacra (IC50, 0.77 microgram/ml) and the methanol-extracts of Lycopodium clavatum (IC50, 1.3 micrograms/ml), Paeonia lactiflora var. trichocarpa (IC50, 5.7 micrograms/ml), Paeonia veitchii (IC50, 2.4 micrograms/ml) and Rhodiola sacra (IC50, 0.67 microgram/ml) showed strong inhibitory activity. In addition, we also examined the PEP inhibitory activity of eleven compounds from Salvia deserta, and found that in addition to a catechol group alpha-hydroxy-para-quinone group may be related to the PEP inhibition.

China↗

PubMed1999

A preliminary study on the ocular blood flow (OBF) of Hong Kong Chinese.

The Ocular Blood Flow (OBF) Tonometer is an objective instrument to measure the Pulsatile Ocular Blood Flow (POBF), Pulse Amplitude (PA) and Pulse Rate (PR). It also provides the minimum intraocular pressure (IOPmin) simultaneously. This preliminary study applied the OBF Tonometer on 74 Hong Kong Chinese. They had their POBF measured followed by the Goldmann Applanation Tonometer (GAT). The OBF tonometer overestimates the IOP slightly. The average POBF result was smaller (661.2 microliters/min) than the previous studies. The effect of myopia of our sample (mean spherical equivalent -4.27 D) could be one of the contributing factors. Other factors include age and racial difference. Twenty-two subjects had their POBF measured again by the same practitioner and different practitioner for the reproducibility variation assessments. The POBF, PA and IOPmin were found to be repeatable between sessions. The POBF "norm" in Hong Kong Chinese needs to be established from a larger sample size and from a comparison on an age and refraction matched Caucasian group.

Adolescent↗

PubMed1999

Graded and dynamic reflex summation of myelinated and unmyelinated rat aortic baroreceptors.

Unmyelinated (C) and myelinated (A) baroreceptor (BR) axons are present in rat aortic depressor nerve (ADN). With graded ADN electrical activation and anodal conduction blockade, reflex responses in anesthetized rats were assessed as changes in mean arterial pressure (MAP) and heart rate (HR). We tested the hypothesis that C-type BR inputs are effective at low frequencies because they outnumber A-type. Anodal current (Ian) reversibly eliminated all MAP and HR responses to A-selective stimuli. High intensities activated all ADN axons (A+C) and decreased MAP at lower frequencies (<10 Hz) than were effective with A-selective stimulation. I(an) reduced only MAP responses to >10-Hz ADN stimulation. Burst patterns significantly augmented A- but not C-selective reflex responses despite identical numbers of shocks per second. A-selective stimuli failed to evoke significant bradycardia even at 200 Hz. Maximum intensity stimuli plus Ian (C selective) evoked less bradycardia than without I(an) (A+C), indicating supra-additive summation unlike the occlusive summation for MAP responses. However, activation of reduced numbers of C-type BRs with all A-type BRs suggests a strong A to C interaction in reflex bradycardia responses. Surprisingly, Ian block of A-type conduction eliminated all reflex bradycardia at such submaximal intensities despite C conduction and depressor responses. A- and C-type BRs act synergistically, and A-type activity is absolutely required in cardiac but not in depressor pathways. Thus greater numbers do not appear to account for C-type BR efficacy, and critical interactions between these two sensory subtypes appear to occur differentially across cardiac and systemic baroreflex effector pathways.

Animals↗

PubMed1999

Mitotic chromosomal bcl-2. I. Stable expression throughout the cell cycle and association with isolated chromosomes.

Bcl-2 is present in a cytoplasmic distribution in cells that express high levels of this oncoprotein. In contrast, using immunocytochemistry in cells expressing low levels of bcl-2, such as KB human carcinoma cells, we and others have shown that bcl-2 is present on the surface of mitotic chromosomes. However, monoclonal antibodies reactive with an epitope representing amino acids 41-54 of the bcl-2 sequence did not detect bcl-2 in other phases of the cell cycle. This study extended those earlier findings to determine if bcl-2 was expressed as a cyclin or if this pattern was an artifact of immunocytochemistry. Immunofluorescence studies in several other human cell lines showed the same mitotic distribution of bcl-2. Other studies using flow cytometry also showed selective mitotic phase detection of bcl-2. A comparison of available commercial antibodies showed that, in spite of reactivity with denatured bcl-2 on Western blots, clear reactivity with bcl-2 in fixed cells was found only with those reactive with the (a.a. 41-54) epitope. With RNase protection and Western blot analyses, cells synchronized at various stages of the cell cycle showed constant levels of bcl-2 mRNA and protein. Analysis of bcl-2 using Western blots showed a band with the same apparent molecular weight as that seen in comparison with authentic bcl-2 overexpressed in the cytoplasm. The retention of bcl-2 on chromosomes in unfixed, permeabilized preparations was influenced by protease treatment, phosphate, and pH. Studies using isolated chromosomes showed that much of the bcl-2 in these cells was attached to chromosomes in mitosis, had the expected molecular weight, and was phosphorylated in the same manner as that seen in whole-cell extracts. These results show that bcl-2 is not a cyclin and that the bcl-2 localized on chromosomes is the same molecule seen by immunoblotting. These results suggest that the reactive (a.a. 41-54) epitope present in bcl-2 is somehow modified or masked in interphase.

Animals↗

PubMed1999

Mitotic chromosomal bcl-2. II. Localization to interphase nuclei.

We have previously shown, by immunofluorescence of fixed cells, that bcl-2 is found only in mitotic chromosomes in KB cultured human tumor cells expressing low levels of this oncoprotein. However, other studies showed that bcl-2 did not change its levels during the cell cycle when analyzed using Western blots. In this study we analyzed the distribution of bcl-2 during interphase, the point at which it is undetectable by immunofluorescence, using biochemical extraction, immunoprecipitation, and cell fractionation with Western blots. Interestingly, when carefully examined by immunofluorescence in fixed cells, the earliest point in the cell cycle showing bcl-2 localization was early G2, in which bcl-2 could be found within the intact nucleus. In spite of showing no immunofluorescence reaction in fixed interphase cells, immunoprecipitation of gentle detergent extracts showed that bcl-2 from interphase cells reacted readily with the antibody used (#124) after extraction. However, immunoprecipitation using anti-bcl-2 followed by Western blots using anti-Bax showed that, unlike overexpressing cells, this bcl-2 was not complexed with Bax. Classical cell fractionation methods were used to separate nuclei from cytosol and cell membranes. Surprisingly, these experiments clearly showed that essentially all of the bcl-2 in interphase KB cells was present in the nucleus. Therefore, the lack of reaction in fixed cells with anti-bcl-2 antibody reflects either a masking or a conformational change of the reactive epitope in bcl-2 present within the nucleus. By correlation, this change may be related to the phosphorylation of bcl-2 that occurs just before mitosis. The nature of this novel yet highly conserved nuclear form of bcl-2 and the understanding of its function will require further study.

Blotting, Western↗

PubMed1999

Prolyl endopeptidase inhibitors from the underground part of Rhodiola sacra S. H. Fu.

Prolyl endopeptidase (PEP, EC 3.4.21.26) is an enzyme which plays a role in the metabolism of proline-containing neuropeptides, e.g., vasopressin, substance P and thyrotropin-releasing hormone (TRH), which have been suggested to be involved in learning and memory processes. In our systematic screening for PEP inhibitors from traditional Chinese medicines, we found that MeOH extract from the underground part of Rhodiola sacra S. H. Fu shows significant inhibitory activity against PEP from Flavobacterium meningosepticum. Examination of the constituents of the extract resulted in the isolation of nineteen known compounds, identified as hydroquinone (1), 4-hydroxybenzoic acid (2), caffeic acid (3), 4-hydroxycinnamic acid (4), suberic acid (5), protocatechuic acid (6), gallic acid (7), (-)-epigallocatechin 3-O-gallate (8), 2-phenylethyl beta-D-glucopyranoside (9), 3-O-galloylepigallocatechin-(4beta-->8)-epigallocatechin+ ++ 3-O-gallate (10), 2-phenylethyl alpha-L-arabinopyranosyl-(1-->6)-beta-D-glucopyranoside (11), sacranoside A (12), beta-D-glucopyranosyl 4-hydroxybenzoate (13), rhodiocyanoside A (14), rhodiooctanoside (15), sarmentosin (16), heterodendrin (17), arbutin (18) and 4-O-(beta-D-glucopyranosyl)-gallic acid (19). Among these, 1, 2, 5, 8-10, 13, 16, 18 and 19 have been isolated for the first time from R. sacra, among which 5, 9, 10, 13, 16, 18 and 19 have been isolated from Rhodiola plants for the first time. On the PEP inhibition, seven compounds (6-8, 10, 12, 18, 19) showed inhibition with an 1C50 of 27.8, 487, 1.47, 0.437, 348, 391 and 215 microM, respectively. The kinetic study of these inhibitors indicated that they are noncompetitive inhibitors, except for 6 which is a competitive inhibitor.

Drugs, Chinese Herbal↗

PubMed1999

Antagonistic interplay between antimitotic and G1-S arresting agents observed in experimental combination therapy.

Paclitaxel is a naturally occurring antimitotic agent that has been shown to stabilize microtubules, induce mitotic arrest, and ultimately induce apoptotic cell death. The favorable clinical activity of paclitaxel has prompted considerable interest in combining paclitaxel with numerous other antineoplastic agents. Our previous studies have suggested 5-fluorouracil (5-FU), an antineoplastic agent that usually arrests tumor cells at the G1-S phase of the cell cycle, in combination with paclitaxel significantly represses paclitaxel-induced mitotic arrest and apoptosis. In the present study, we have extended this investigation to include several other antimitotic agents (vinblastine, colchicine, and nocodazole) in various combination schedules with the G1-S arresting agents 5-FU and hydroxyurea (HU). We found 5-FU, as well as HU, could significantly interfere with the overall cytotoxicity as compared with treatment with antimitotic agents alone. It appeared that 5-FU or HU severely limited the antimitotic agents' cytotoxic effects on both mitotic arrest and apoptosis. No combination of a G1-S arresting agent with an antimitotic agent in any schedule produced an antitumor effect greater than that of the antimitotic agent alone. In addition, biochemical examination revealed that 5-FU and HU blocked the antimitotic agent-induced increase of p21WAF1/CIP1 protein levels, as well as prevented the hyperphosphorylation of the bcl-2 and c-raf-1 proteins. These findings suggest that careful considerations may be necessary when combining antineoplastic agents that exert their cytotoxic action at different phases of the cell cycle.

Antineoplastic Agents↗