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Biomedical subjects

B Silvestrini

Publications and source records attributed to B Silvestrini.

At least 19 recordsLinked to original sources
PubMed1996

Astrocytes synthesize and secrete prostaglandin D synthetase in vitro.

Prostaglandin D synthetase [PGD-S, prostaglandin-H2 D-isomerase, (5Z, 13E)-(15S)-9alpha, 11 alpha-epidioxy-15-hyrdroxyprosta-5,13-dienoate D-isomerase, EC 5,3,99,2], an enzyme that catalyzes the formation of prostaglandin D2, was originally isolated from homogenates of rat brain and spleen and is known to be a membrane-bound enzyme. Subsequent immunohistochemical studies have shown that PGD-S is associated with neurons in the brain of immature rats, whereas in adult rats it is associated with oligodendrocytes. Several recent studies have shown that the beta-trace protein isolated from human cerebrospinal fluid (CSF), the second most abundant protein in human CSF after albumin, is equivalent to PGD-S. In this paper, we report the preparation of a monospecific polyclonal antibody against purified PGD-S isolated from human CSF and the establishment of a specific radioimmunoassay for this protein. Using this radioimmunoassay in conjunction with immunoblot analysis, PGD-S was detected in various biological fluids including serum, aqueous humor, and rete testis fluid. In addition, an antibody prepared against human PGD-S partially cross-reacted with the PGD-S in the rat and ram. Using a monospecific polyclonal antibody prepared against purified rat PGD-S isolated from rat CSF in conjunction with [35S]methionine incorporation and immunoprecipitation techniques, it was shown for the first time that PGD-S is actively synthesized and secreted by astrocytes cultured in vitro, suggesting the astrocyte is the cellular origin of PGD-S in the CSF. The identification of the astrocyte as the cellular origin of this unique enzyme will allow the use of an in vitro system to study its regulation.

Aging↗
PubMed1995

Cerebrin-50, a human cerebrospinal fluid protein whose mRNA is present in multiple tissues but predominantly expressed in the lymphoblastoid cells and the brain.

A full-length cDNA of 2295 bp coding for a human cerebrospinal fluid protein, designated cerebrin-50, has been isolated from a human brain cDNA expression library. Nucleotide sequence analysis of this cDNA revealed that it codes for a 435 amino acid polypeptide which starts with an ATG initiation codon from the 5'-end and ends with a TGA termination codon with a calculated molecular weight of 51,484 daltons. A 10-amino acid peptide of NH2-SGDLETRYWG based on the deduced amino acid sequence of the cerebrin-50 cDNA was synthesized, conjugated to bovine serum albumin and was used to raise a monospecific polyclonal antibody in a rabbit. Immunoblot analysis using this antibody indicated the presence of a 52 kD protein in the human cerebrospinal fluid, brain cytosol, T lymphoblastoid cell-conditioned medium, and human serum; it is also noted that the concentration of this 52 kD protein is several order of magnitude higher in the cerebrospinal fluid than in other biological fluids. The presence of its mRNA in the brain, T lymphoblastoid cells, spleen, liver, and testis was confirmed by sequential use of reverse-transcription and polymerase chain reaction (RT-PCR). Preliminary analysis by quantitative RT-PCR has noted that the expression of the cerebrin-50 mRNA is higher in the lymphoblastoid cells and brain than the spleen, liver, and testis.

Amino Acid Sequence↗
PubMed1994

Facilins, a novel class of biological factors that facilitate the aortic response to dopamine and other biogenic amines.

Biological fluids and tissues extracts were shown to contain biological factors, termed facilins, that facilitate the dopamine-, adrenaline-, and serotonin-mediated aortic contraction at concentrations devoid of any direct effect. Cyproheptadine and phentolamine antagonized the direct contracting effect of biogenic amines, but not the facilitated component of the aortic response thus indicating that the mechanism of action of facilins was unlike that of biogenic amines. Fresh schizophrenics' CSF displayed a stronger facilitating effect than normal CSF on the dopamine-mediated aortic response. This finding, however, was not confirmed with samples kept frozen for prolonged periods of time. Multiple molecular forms of facilins were detected in rabbit serum. Those with a high apparent molecular weight were proteinous and were neither insulin nor other factors known for their contracting effects on the aorta such as epidermal growth factor, transforming growth factor-beta, and platelet-derived growth factor.

Adrenal Glands↗
PubMed1994

Response of alpha 2-macroglobulin messenger ribonucleic acid expression to acute inflammation in the testis is different from the response in the liver and brain.

Recent studies from this laboratory have shown that Sertoli cells derived from 20-day-old rats and cultured in vitro synthesize and secrete a nonspecific protease inhibitor that is structurally and immunologically similar to serum alpha 2-macroglobulin (alpha 2-MG). In contrast to its serum homologue, the testicular alpha 2-MG is not an acute-phase protein in the rat since its protein concentration in the rete-testis fluid does not increase in response to inflammation. In the present study we examined the expression of alpha 2-MG mRNA in the rat testis in comparison to that in the brain and liver following induced inflammation. alpha 2-MG mRNA in the testis did not respond to induced inflammation, whereas its protein concentration in serum and its mRNA level in the brain and liver increased significantly in 20-day-old inflamed rats. In 8-day-old rat testis, where the blood-testis barrier is not yet formed, alpha 2-MG mRNA expression also did not respond to induced inflammation. The mRNA expression of clusterin, another authentic Sertoli cell protein whose secretion appears to be closely related to cell-cell interactions in the seminiferous epithelium, was shown to be unaffected by induced inflammation in the testis, brain, and liver. In view of the unexpected differential expression of alpha 2-MG mRNA to induced inflammation in the testis and liver, we sought to examine whether Sertoli cell alpha 2-MG would respond to FSH and testosterone (T), the major regulators of testicular function. Interestingly, expression of alpha 2-MG and clusterin mRNA in the Sertoli cell was not regulated by FSH, T, or a combination of FSH and T. Since there is an intimate morphological relationship between Sertoli cells and germ cells, we next examined the effect of germ cell-conditioned medium (GCCM) on Sertoli cell alpha 2-MG and clusterin mRNA expression. It was noted that GCCM caused a dose-dependent stimulation of alpha 2-MG and inhibition of clusterin mRNA expression in Sertoli cells, respectively. Therefore, our studies have shown that the regulatory mechanism that modulates the expression of alpha 2-MG mRNA in the rat testis is different from its counterpart in the brain and liver.

Aging↗
PubMed1994

Rat testicular testibumin is identical to sulfated glycoprotein-1 (SGP-1) whose mRNA expression in the testis is age- but not germ cell-dependent.

Using sequential HPLC and capillary electrophoresis (CE), testibumin (CMB-1) has been purified to apparent homogeneity from Sertoli cell-enriched culture medium prepared from 20-day-old rat testes. N-Terminal amino acid sequence analysis of the purified testibumin revealed a partial sequence of NH2-XPVQDPKI. When this partial sequence was compared to existing protein database, it was shown that it is identical to a previously isolated Sertoli cell secretory protein, sulfated glycoprotein I (SGP-1). The fact that testibumin is equivalent to SGP-1 was further confirmed when its full-length cDNA was isolated and sequenced. Studies using quantitative PCR to examine the changes of steady-state mRNA level of testibumin (SGP-1) in the rat testes between 3 and 60 days of age indicated that its mRNA increased rapidly after birth, peaked at 10-20 days, and declined rapidly where the adult testibumin mRNA level was similar to the neonatal rat at 3 days of age. Depletion of germ cells by a single dose of lonidamine, an antispermatogenic drug, did not induce an increase in testibumin (SGP-1) mRNA level indicating its mRNA expression is not dependent on germ cells.

Aging↗
PubMed1993

The use of high-performance electrophoresis chromatography for the micropurification of cerebrospinal fluid proteins in the rat.

Using reversed-phase high-performance liquid chromatography (HPLC) on a Vydac C8 column in conjunction with sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and silver staining, we have identified more than 30 proteins in cerebrospinal fluid collected from adult rats by cannulation of cisterna magna. When these partially purified cerebrospinal fluid proteins were further fractionated by high-performance electrophoresis chromatography (HPEC) on an Applied Biosystems 230A HPEC system using a 10% T SDS-polyacrylamide gel with a phosphate base running buffer system under nonreducing conditions, we have purified more than 10 proteins to apparent homogeneity from a pool of 10 ml of rat cerebrospinal fluid as verified by silver staining and direct N-terminal amino acid sequencing. Two additional series of experiments using rat cerebrospinal fluid over a 12-month period yielded virtually identical results. A major advantage of HPEC over conventional HPLC is that the recovery of protein is almost quantitative and is in the range of 90-95% using as little as 1 microgram of protein. The purified proteins from HPEC are ready for direct protein sequencing following a buffer exchange to remove residual Tris and phosphate without additional manipulation. The potential use of HPEC for micropurification of proteins was discussed.

Amino Acid Sequence↗
PubMed1993

Abnormal glycosylation of alpha 2-macroglobulin, a non-acute-phase protein in patients with autoimmune diseases.

Previous studies from this and other laboratories have shown that abnormal glycosylation of several acute-phase proteins can be detected in various pathological conditions including autoimmune diseases. In the present study, we have investigated if abnormal glycosylation is limited to acute-phase proteins. We used the concanavalin A (Con A) blots in conjunction with the peptide mapping techniques to analyze serum samples and cerebrospinal fluids (CSF) obtained from patients with autoimmune diseases: systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), mixed connective tissue disease (MCTD), scleroderma (SCL), Sjögren's syndrome (SS), and polymyositis (PM); diseases of probable autoimmune origin: hepatopathies (HP); diseases of suspected autoimmune origin: schizophrenia and Alzheimer's disease (AZ); and conditions not related to autoimmunity: pregnancy (PG) and elevation of the carcinoembryonic antigen (CEA), in comparison to normal donors (NHS). We have micropurified two human proteins; alpha 2-macroglobulin, a non-acute-phase protein and beta-chain of haptoglobin, a known acute-phase protein, from serum samples of individual patients with SLE, RA, MCTD, SCL and SS, and from PG and NHS for analysis. The identity of the purified proteins was confirmed by immunoblots using either monospecific polyclonal or monoclonal antibodies, and by direct N-terminal amino acid sequencing. Peptide maps for each of these proteins were generated using Staphylococcus aureus protease V8, a Glu-C endopeptidase. When the peptide fragments of alpha 2-macroglobulin were resolved by SDS-PAGE and visualized using silver staining, no differences were noted between patient samples and controls. However, when they were examined by lectin blots using Con A, the Con A-reactive fragments increased specifically and significantly in samples derived from patients of SLE, SCL, MCTD, and RA. Similarly when the peptide fragments of the beta-chain of haptoglobin were visualized by silver staining, no differences were noted; however, the Con A reactivity of specific fragments increased in SLE, RA, SCL, and SS patients. Analysis of these results indicated that there has been a selective increase in Con A-reactive fragments in both acute-phase and non-acute-phase proteins in autoimmune conditions. Thus, the study of changes in glycosylation patterns in selected serum proteins may be a valuable diagnostic approach to define the pathophysiology of inflammatory and autoimmune disorders.

Acute-Phase Proteins↗
PubMed1993

Changes of immunoreactivity in alpha 1-antitrypsin in patients with autoimmune diseases.

Recent studies from this laboratory have shown that a monoclonal antibody prepared against a specific epitope on alpha 1-antitrypsin is a valuable diagnostic marker for autoimmune conditions. In the present study we have further characterized this monoclonal antibody and reassessed its diagnostic value in screening samples from patients with various autoimmune conditions. alpha 1-Antitrypsin was micropurified from patients with selected autoimmune conditions and from normal donors. The purified alpha 1-antitrypsin isolated from patients with autoimmune conditions and normal donors was deglycosylated using both a mixture of exoglycosidases and endoglycosidase F. The immunoreactivity of the native and deglycosylated alpha 1-antitrypsin was examined using both a monoclonal antibody and a polyclonal antibody in enzyme linked immunosorbent assay (ELISA) and radioimmunoassay (RIA), respectively. It was noted that alpha 1-antitrypsin isolated from patients with autoimmune diseases generated a displacement curve dissimilar to alpha 1-antitrypsin purified from normal donors or alpha 1-antitrypsin from patients with autoimmune diseases subjected to deglycosylation when these samples were examined by ELISA using the monoclonal antibody. However, when the polyclonal antibody was used for these studies, no difference was found between the native and deglycosylated alpha 1-antitrypsin suggesting that the monoclonal antibody recognized an epitope not detectable by the polyclonal antibody.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Micropurification of two human cerebrospinal fluid proteins by high performance electrophoresis chromatography.

Using C8 reversed-phase HPLC in conjunction with sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we have fractionated proteins contained in human CSFs obtained from patients with schizophrenic disorders. When these proteins were electrophoretically blotted onto polyvinylidene difluoride membrane for direct N-terminal amino acid sequencing, several CSF proteins were identified; these included albumin, transferrin, apolipoprotein A-I, beta 2-microglobulin, and prealbumin. We have also identified two structurally related human CSF proteins designated cerebrin 28 (M(r) 28,000) and cerebrin 30 (M(r) 30,000) that have an N-terminal amino acid sequence of NH2-APPAQVSVQPNF and NH2-APEAQVSVQPLFXQ, respectively. Comparison of these sequences with existing database at Protein Identification Resource (R 32.0), GenBank (R 72.0), SWISS-PROT (R 22.0), and EMBL (R 31.0) indicated that they are unique proteins. These proteins were subsequently purified by high performance electrophoresis chromatography (HPEC) using an Applied Biosystems 230A HPEC system. A specific polyclonal antibody was prepared and an ELISA was established for cerebrin 30. It was noted that HPEC is a powerful tool to purify microgram quantities of proteins from human, rabbit, and rat CSFs. Using such a system, we have been able to micropurify as many as 10 proteins simultaneously in a single experiment because the elution of proteins occurred strictly according to their molecular weights. More importantly, we routinely obtained a recovery of > 90%. The potential use of this technology for micropurification of proteins was discussed.

Amino Acid Sequence↗
PubMed1993

Chronic inflammatory response in the rat can be blocked by bindarit.

In the rat, injection of Freund's complete adjuvant was accompanied by a significant increase in concanavalin A (Con A)-reactivity of selected plasma proteins along with an increase in concentrations of selected proteins known as acute phase proteins. We have evaluated the effect of bindarit, (2-[(1-benzyl-indazol-3-yl)methoxy]-2-methyl propionic acid), on the expression of alpha 2-macroglobulin, a known acute-phase protein in the rat. This compound has previously been shown to inhibit heat-induced denaturation of rat serum albumin and to strongly reduce the secondary phase response of adjuvant induced arthritis. Adult rats were induced with chronic inflammation by injection with Freund's complete adjuvant. Bindarit was administered to the chronic inflamed rats as a 0.5% medicated diet. Indomethacin, given by gavage daily at a dose of 1 mg/kg body weight, was used as a reference drug. Qualitative and quantitative changes of Con A-reactive proteins and alpha 2-macroglobulin were examined by lectin- and immuno-blots, and by radioimmunoassay. It was noted that the concentration of alpha 2-macroglobulin increased in rats with adjuvant induced arthritis. The addition of bindarit and indomethacin were able to reduce the concentration of alpha 2-macroglobulin as well as the Con A-reactivity of various proteins to normal level 37 days following treatment. We have also examined the effects of chronic inflammation on the levels of rat clusterin, a testicular and serum glycoprotein related to programmed cell death, tissue regression, and complement cascade reaction; and testibumin, a testicular FSH and testosterone-responsive protein with unknown function. It was noted that chronic inflammation did not induce significant changes in both the clusterin and testibumin concentrations in these experimental groups. The involvement of protein glycosylation and denaturation in the production of new antigenic determinants, their role in the development of chronic inflammatory disease and the potential use of bindarit to investigate the relationship between abnormal glycosylation and autoimmune disease were discussed.

Acute-Phase Proteins↗
PubMed1992

Abnormal glycosylation of hemopexin in arthritic rats can be blocked by bindarit.

Induction of arthritis in rats with Freund's complete adjuvant was accompanied by a distinctive alteration of concanavalin A (Con-A) reactivity in their serum proteins in which the concentrations of selected Con-A reactive proteins were significantly higher when compared to healthy rats. To assess if the observed increase in Con-A reactivity of specific serum proteins reflects an increase in carbohydrate moieties in these proteins in addition to an increase in their protein concentrations, a heme binding serum glycoprotein, hemopexin, also an acute phase reactant, was selected as a marker protein. Hemopexin was purified to apparent homogeneity from pools of serum samples derived from rats with yeast induced inflammation, a monospecific polyclonal antibody was prepared and was used for immunoblot analysis. It was noted that the concentration of hemopexin increased in rats with adjuvant induced arthritis; however, its concentration fell to normal levels after administration with a newly synthesized drug, bindarit, (2-[(1-benzyl-indazol-3-yl)methoxy]-2-methyl propionic acid, C19H20N2O3. Hemopexin was micropurified individually from healthy rats, adjuvant induced arthritic rats, and adjuvant arthritic rats treated with bindarit, cleaved with a Glu-C endopeptidase, Staphylococcus aureus protease V8, and the resultant peptide fragments resolved by SDS-PAGE and examined by silver staining, Coomassie blue staining, and lectin blots using Con-A. It was subsequently noted that hemopexin isolated from adjuvant induced arthritic rats showed a significant increase in Con-A reactivity in selected peptide fragments and that such an increase in glycosylation could be reversed to a pattern similar to healthy rats following treatment with bindarit.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute-Phase Reaction↗
PubMed1992

Identification of multiple biological factors in rabbit serum that modulate dopamine-mediated aortic constriction.

A bioassay was established using freshly prepared rabbit aortic strip to monitor the effects of various biological factors contained in rabbit serum that modulate vasoconstriction. Serotonin was shown to be a major vasoconstriction modulator in rabbit serum. A rapid procedure is described for its isolation from rabbit serum by sequential gel filtration chromatography and reversed-phase HPLC using C4, C18, and C2/C18 columns with an overall cumulative yield of about 20%. It was also noted that rabbit serum contains multiple biological factors that modulate dopamine-mediated aortic contraction other than serotonin. A potent vasoconstriction inhibitor was identified in rabbit serum which appears to be a novel regulator that mediates its effect via both alpha-adrenergic and serotonin receptors.

Animals↗
PubMed1992

A new protein antidenaturant agent, bindarit, reduces secondary phase of adjuvant arthritis in rats.

Bindarit (or 2-[(1-benzyl-indazol-3-yl)methoxy]-2-methyl propionic acid) reduces heat induced denaturation of bovine and rat serum albumin in vitro (EC50 = 8.5 and 65 micrograms/ml, respectively) and inhibits heat induced serum albumin denaturation after in vivo (12.5-25-50 mg/kg po) administration in rats. To assess the relationship between protein denaturation and the development of chronic inflammatory diseases, the drug (0.5 or 0.12% medicated diet) was studied in comparison with indomethacin (1 mg/kg po daily) in rats injected with complete Freund's adjuvant. Bindarit appeared different from aspirin-like drugs, antiinflammatory steroids and immunosuppressants because it does not reduce primary inflammation of arthritic rats and was shown to be completely inactive on cyclo and lipooxygenase activity in vitro and on immune reactions of mice in vivo. Nevertheless, the drug strongly reduced the development of the secondary phase of adjuvant induced arthritis. The most significant effect of bindarit in this phase was a strong inhibition of serum albumin denaturation in arthritic rats. Assessment of both electrophoretic and quantitative changes suggests that the reduction of albumin during inflammation is due, at least in part, to a denaturation of native albumin, which loses its electrophoretic mobility. The involvement of protein denaturation in the production of new antigenic determinants, their pathogenic relevance in the development of adjuvant arthritis and the possibility that protein stabilization by bindarit could be the mechanism of action of the drug are discussed.

Animals↗
PubMed1991

Facilitating effect of amphetamine on the response of rabbit aortic strips to adrenaline, dopamine and serotonin.

Amphetamine increased the response of rabbit aortic strips to adrenaline, dopamine and serotonin at consistently lower doses than those exerting a direct contracting effect. The amphetamine-facilitated contraction had the same shape as that produced by biogenic amines alone, whereas the contraction produced by amphetamine alone was more delayed and flatter. Serotonin and dopamine facilitated each other, but less markedly and with a narrower interval between facilitating and contracting doses than amphetamine. Pargyline exerted no facilitating effect on biogenic amines. Phentolamine and prazosin inhibited the direct response to adrenaline, dopamine and amphetamine, and the amphetamine-facilitated response to adrenaline and dopamine; they were inactive against serotonin alone and combined with a facilitating dose of amphetamine or dopamine. Cyproheptadine inhibited the direct response to serotonin and amphetamine, and the amphetamine-facilitated response to serotonin; it was inactive against dopamine and adrenaline both alone and combined with a facilitating dose of amphetamine or serotonin.

Amphetamine↗
PubMed1991

Pretreatment with bendazac attenuates retinal damage induced by intense light in rats.

Bendazac is a drug which protects proteins from denaturation induced by different agents. It is also effective in protecting rabbits from X-ray-induced cataract. This study deals with the effects of bendazac on the intense light-induced retinal damage in rats. Four groups of animals received orally 0, 50, 100 or 200 mg/kg of bendazac L-lysine salt three times a day for 3 days and once the fourth day, before 1 h exposure to intense-green filtered light. Fourteen days after housing in a dark room, the rats were sacrificed and the retinae were examined by light microscopy. Retinal damage was graded according to a score severity from 0 to 5. The mean score for control animals was 2.23, whereas a dose-related and statistically significant reduction of retinal damage was detected in bendazac treated rats, i.e. 1.72, 1.54 and 1.40. A protective activity in the distribution of the severity score, i.e. a higher incidence of no damaged retinae and a lower frequency in the most severely affected ones, was also observed in treated versus control rats. These results suggest a potential therapeutic value of bendazac in the treatment of those conditions, such as retinitis pigmentosa and senile macular degeneration, in which the light exposure plays a role as a co-factor.

Animals↗
PubMed1991

Alpha 2-macroglobulin is not an acute-phase protein in the rat testis.

Earlier studies from this laboratory have shown that Sertoli cells actively synthesize and secrete a nonspecific protease inhibitor in vitro; N-terminal sequence analysis, subunit structural analysis, and other biological studies revealed that this protein is the homolog of serum alpha 2-macroglobulin. We have now quantified the relative distribution of alpha 2-macroglobulin in the reproductive compartments and their comparison with nonreproductive organs. In serum and all nonreproductive tissues examined, the concentration of alpha 2-macroglobulin progressively decreased with advancing age. However, in both the testis and epididymis, the levels of this protein increased with the age of the animals. Serum alpha 2-macroglobulin levels were consistently higher than those in any other tissues until 60 days when the concentrations of this protein were the highest in the epididymis. The distribution of alpha 2-macroglobulin in various nonreproductive tissues from female rats was similar to that observed for male rats in that its levels tended to decrease with age. However, uterine levels of alpha 2-macroglobulin increased progressively with advancing age, whereas ovarian levels of alpha 2-macroglobulin remained relatively stable with an increase in animal age. As serum alpha 2-macroglobulin is an acute-phase protein in the rat, the response of this protein in the testis to induced inflammation was examined. The concentration of alpha 2-macroglobulin in serum rose about 150-fold after injection of fermented yeast. By contrast, the levels of this protein in rete testis fluid, which is derived exclusively from seminiferous fluid, did not change in response to inflammation. These results suggest that there might be distinctive mechanisms that regulate this protein in the systemic circulation vs. the microenvironment behind the blood-testis barrier in the seminiferous epithelium.

Acute-Phase Proteins↗
PubMed1991

Lonidamine: an overview.

The attention of pharmaco-therapeutic research is shifting from the cell duplication mechanisms to the oncogene expressions and cofactors that are the actual cause of cancer. This trend corresponds to the appearance of a second generation of anticancer agents that are typically represented by tamoxifen and lonidamine. The first is a hormonal agent with endocrine effect that was developed making use of a rationale and methods already available, although in a different context. The second, an energolytic agent, is a more complicated case. It was discovered when neither a solid knowledge of cancer energy systems nor the related pharmacological methods were available. Both had to be developed along with a basic research in which lonidamine was at the same time target and tool. In this way, the indication was obtained that cancer activates a specialized energy system in the repair phase following exposure to hyperthermia, alkylating agents, and radiations. This system confers cancer cells an advantage over the normal ones, but is vulnerable and appears to be lonidamine's target. The presently available data show that lonidamine, when used in appropriate conditions with respect to its mode of action, increases the disease-free interval and survival in some types of tumours.

Antineoplastic Agents↗