Acta Clinica Belgica International Journal of Clinical and Laboratory Medicine
ISSN: 1784-3286 (Print) 2295-3337 (Online) Journal homepage: http://www.tandfonline.com/loi/yacb20
Pharmacological Properties Of Drugs Inhibiting Platelet Aggregation G. de Gaetano To cite this article: G. de Gaetano (1975) Pharmacological Properties Of Drugs Inhibiting Platelet Aggregation, Acta Clinica Belgica, 30:3, 184-189, DOI: 10.1080/17843286.1975.11716995 To link to this article: http://dx.doi.org/10.1080/17843286.1975.11716995
Published online: 21 May 2016.
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Date: 11 August 2017, At: 05:55
184
PHARMACOLOGICAL PROPERTIES OF DRUGS INHIBITING PLATELET AGGREGATION
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G. de GAETANO*
The influence of drugs on platelet function can be tested in vitro and in vivo systems both in man and in ex perimental animals. In some in vitro systems, either native or anticoagulated whole blood can be used, whereas for other tests the preparation of platelet-rich plasma is required; in addition , platelet function can also be studied after separating the cells from plas ma proteins by one of many different techniques (" was hed " platelets) (1 2). In vivo studies of drugs influencing platelet fun ction include testingJn vitro the platelet fun ction in blood taken after administration of the drug and the use of certain in vivo techniques such as the bleeding time test (duration , pattern and amount of blood loss), the determination of platelet survival and the accumulation of radioactive platelets in certain diseased organs (6, 7, 14, 21, 28); in experimental animals it is possible to evaluate the effect on platelets of both aggregating substances and inhibitory compounds (23). Some drugs may inhibit one of these numerous test systems developed for stud ying platelet function but not others, making it difficult to establish the significance of such inhibition for clinical use of these compounds. For example, aspirin inhibits platelet release reaction (19 , 36) and prolongs bleeding time (14, 21), but does not modify the retention of platelets in a glass bead
" Laboratory for Hae mostasis and Th ro mbosis Research, Istituto di Ricerche Farmacologiche «Mario Negri», Via Eritrea, 62, 201 57 Milano, Italy. Acta C/inica Belgica, 30, 3 (1 975)
column nor the shortened platelet survival in certain pathological conditions (6). In contrast, dipyridamole is quite ineffective in in vitro systems and does not prolong bleeding time, but increases the shortened platelet survival (6). Let us add that drugs effective in some animal models are devoid of antiplatelet activity in others (23). No single test can therefore reveal all aspects of platelet fun ction and its pharmacological inhibition. In in vitro systems the binding of drugs to plas ma proteins has to be considered : f.i. su1finpyrazone is very active as an inhibitor of the release reaction of washed platelets, but it is almost without effect on platelet-rich plas ma (17 , 35, 36). Another problem which should be carefully considered is the disposition of dru gs affecting platelet fun ction . After ingestion , dru gs are subjected to many processes including absorption, protein binding, distribution in tissues, biotransform ation and excretion. Comparison of the effects of a dru g in vitro and in vivo requires quantitative determination of the drug in the blood; a lack of effect in vivo may be simply due to too low an active concentration of the drug in the blood. This in turn may result from poor gastrointestinal absorption , high protein binding, extensive biotransformation into inactive metabolites or rapid excretion. Conversely, a dru g inactive in vitro may be active in vivo because it requires biotransform ation into active metabolites or acts indirectly, for instance, by releasing physiological compounds which are not avail able in vitro. This problem is furth er
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PHARMACOLOGY OF ANTI-AGGREGA TION AGENTS
complicated when different animal species are used, since drug disposition may be quan titatively and often qualitatively different (39). The main reactions of platelets which can be studied in vitro are the following (16, 29): I. Adhes ion to collagen and to glass (12); 2. Shape change in res ponse to ADP (2); 3. Development of procoagul ant acti vity (platelet factor 3 ava ilability (12); 4. Change in over-all membrane electric charge (5); 5. Reversible aggregation (3); 6. Adherence to polymerizi ng fibrin an d fibrin retraction (4, 18); 7. Release reaction and irreversible aggregation (3); 8. Uptake and release of serotonin , independent of the ad hesion-aggregation-release · reaction (20). Many compounds inhibit these reactions in vitro, but only a few are suitable for administration to man (I 6, 17, 29, 31) (table 1). Table I In vitro inhibitors of platele t fun ction Group I - Adenos ine and a na logues - Anti-a drenerg ic su bstances - Anti-se ro tonin dru gs - Su lph-scyd ryl gro up inhibitors - Degrad a tion products of some proteins - Pro stag landin s E 1 and 0 1 - Methy l xa ntines - Dipyrid a mo le and Conge ners Group 2 -· Non-steroida l anti- infl am m a tory d rugs - Anti-depressa nt dru gs
Clinically useful compounds inhibiting in vitro platelet function can be divided in two major groups : the first mainly inhibits the reactions numbered l-6 above and the second mainl y inhibits reaction 7. It is not clear whether drugs which inhibit the uptake of serotonin and/or induce its release (reaction no. 8) have any in -
185 nuence on hae mostasis , although cyproheptadine has been shown to inhibit thrombosis of injured vessels in th e hamster check pouch and the accumulation of platelets in renal transplants (22). Among th e dru gs belonging to the first group , dipyrid amole and a number of pyrimid opyrimidine compounds have particul arl y been investigated. These drugs also inhibit the release reaction, but this effect may be due to th eir action on ADP, rath er th an on th e re lease reacti on itself. Indeed, wi th low co ncentrations of releaseinduci ng stimuli , th e small amount of AD P which is released has an enh anci ng effect on further release. Elimination of this ADP effect ca uses a significant reduction in th e extent of release when stimuli are used at low concentrations (17). The concentrations of these dru gs th at inhi bit platelet fun ction in vitro are several times higher th an th ose which ca n be achieved safely in vivo in man ; nevertheless, th ese drugs may be clinicall y effective as antithromboti c age nts since, bes ides platelets, they may also modify oth er relevant parameters such as blood pressure and blood now velocit y. The most potent age nts of the first group of compou nds are Prostaglandin 0 2 and Prostaglandin E1 (I , 8, 15, 26). PGE 1 has little va lue for in vivo administratio n since it is cleared very rapidly from the circu latio n; this property however is advantageous for preparing platelet concentrates free of aggregates for transfusion purposes (25). No information is yet ava ilable on the in vivo effect of PGD 2• A common mechanism of action of the drugs belonging to the first group is their capacity to in crease the levels of platelet cyclic-AMP (table II). Cyclic-AMP inhibits platelet aggregation both when added in vitro and after its intravenous infusion to man (I, 13, 24). Increasing knowledge has led to th e development of a un ifying concept th at substances which stimulate platelets to aggregate tend to induce a decrease in platelet cyclic-A MP, whereas the reverse is tru e for substances which inhibit platelet aggregation . The intracellular level of cyclic-A MP depends on the activity of at least two enzy mes, firstl y adenyl Acta Clinica Belgica. 30. 3 (1975)
186
PHARMA COLOGY OF ANTI-AGGREGATION AGENTS
Table II ADENYL CYCLASE
ATP~j
AMP
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methyl xantines dipyrida11ole
+
PHOSPHODIESTERASES
cyclase which catalyses the conversion of ATP to cyclic-AMP and phosphodiesterase, which inactivates cyclic-AMP to AMP. The presence of both enzymatic activities in platelets has been reported (24). P'rostaglandin E 1 for example increases platelet cyclic-AMP concentration by stimulating adenyl cyclase, whereas caffeine elevates the cyclic-AMP levels by inhibiting phosphodiesterase. The inhibitory effect of PGE, is strongly potentiated by caffeine (or by other similarly acting drugs). The mode of action of cyclic-AMP in platelet has not yet been established; it is conceivable however that it is linked to the availability of membranebound Ca++ in the cytosol of the platelet. In this regard , very recent observations that ionophores induce several platelet reactions by making available intracellular Ca++ are of the greatest interest (ll). . Among the drugs belonging to the second group, i.e. inhibiting the release reaction, nonsteroidal anti-inflammatory agents have received much attention, particularly because they are readily available, inexpensive and relatively nontoxic. Aspirin is certainly the drug most extensively investigated for its effects on platelet funct-ion, haemostasis and thrombosis . Doses of antiinflammatory drugs much lower than those generally used in human therapeutics are highly effective in inhibiting some of the platelet reactions; this property should minimize the occurActa C!inica Belgica, 30. 3 (19 75)
renee of side effects, particularly at the gastrointestinal level. Aspirin and related drugs do not increase the levels of platelet cyclic-AMP (1 ); they inhibit the platelet synthesis of one or more cyclic endoperoxide intermediates which appear during the biosynthesis of PGE 2 and PGF2a from arachidonate (table Ill) Smith and Willis (27), first reported that the formation and release of prostaglandins E2 and F2a which occurs in thrombin-stimulated platelets is inhibited by indomethacin and aspirin, whereas sodium salicylate has only a weak effect. Phenylbutazone has activity .intermediate between aspirin and sodium salicylate, whereas acetaminophen (paracetamol) is inactive. To aremarkable degree, these results parallel the relative ability of the drugs to suppress the second phase or irreversible platelet aggregation and the accompanying release reaction. In addition, Kocsis et at. (9) have shown that after oral administration of aspirin or indomethacin, the duration of the inhibitory effects on aggregation and release parallels their effects on platelet prostaglandin production. Subsequently it has been demonstrated that during the biosynthesis of prostaglandins in the platelets one or two endoperoxide intermediates transiently accumulate in the cell and trigger irreversible aggregation and the release reaction (32, 33). These intermediates have been called: "Labile AggregationStimulating Substance" (LASS). Aspirin and
187
PHARMACOLOGY OF ANTI-AGGREGATION AGENTS
Table Ill
ARACHIDONIC ACID
T YA I
I I I I I I
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~
111
PLATELET ENZYMES
.
t
ASPIRIN
_..._
IRREVERSIBLE A6GRE6ATIDN and
RELEASE REACTION
possibly other anti-inflammatory drugs block the enzymatic conversion of arachidonic acid (the precursor for biosynthesis of PGE 2 and PGFta) to this pro-aggregating material and to PGE 2; the latter sensitizes the platelets to the aggregating effects of LASS (33). It has also been observed that arachidonic acid is able to prevent the inhibitory effects of aspirin on platelet aggregation (10) and that an acetylenic analog of arachidonic acid (TY A) acts like aspirin on platelets since it competes with arachidonic acid towards prostaglandin synthetases (34). In conclusion, all the compounds that influence platelet function seem to work through two main enzymatic mechanisms : the first one involves the formation of increased levels of platelet cyclic-AMP and the second one the inhibition of the formation of a pro-aggregating factor during the biosynthesis of prostaglandins. It is unknown, at the present time, whether these two proposed mechanisms are independent or interrelated in some way . One would welcome the development of more potent and' more selective agents, active as stimulants of platelet adenyl cyclase or as inhibitors of phosphodiesterases or of prostaglandin synthetases. ;
Possibly , association of one or more of these drugs could have much more benefici al effects on thromboembolic complications than those obtained until now with the drugs presentl y available. KEY WORDS Blood Platelets- Platelet Aggregation - Platelet Agg regation Inh ibitors pirin - Dipyridamole - Cyclic-A MP - Pros taglandins.
SUMMARY The pharmacological properties of drugs inh ibiting platelet fun ction have been reviewed. Some of the problems arising in ex perimentation of potent iall y useful substances have also been discussed. Animal species, anticoagulants, plas ma proteins and drug disposition are the main factors innuencing platelet fun ction tests. Clinically useful drugs inhibit platelet fun ction by either increasing the levels of platelet cyclic AMP (PGE " di py ridamole, pyrimido-py rimidine compounds) or inhibiting the platelet biosy nthesis of one or more intermediates appearing during the formatio n of PGE2 and PGF2a from arac hidonate (aspirin , indometh ac in). Possibl y the two mechanisms are interrelated in some way. Knowl edge of the bioc hemical pathways underlyi ng platelet fun ction, should result in the deve lopment of more potent and more selective agents, potentiall y useful as antithrombotic dru gs. Acta C/inica Belgica, 30, 3 (1 975)
188
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SAMENV ATIING de funcDe farmacologische eigenschappen van farm aca die ken. tie van de bloedplaatjes inhi beren worde n bespro teren men experi het bij en voordo zich die Enkele problemen eveneens van potentieel bruikb are geneesmiddelen worde n a-proteiplasm tia, agulan antico , n ~te i~rsoo D gediscuteerd .. testen van nen en stofwt ssellng ZtJn de hoofdfactore n die de de bloedplaatjes functi e beinvloeden. l aat. Klin isch bruikb are farm aca verhinderen de bloedp AMP in isch cycl n va ntratie conce de zowel oor d . cue Jesfun le, pyrim idobl~~l a.atJeS te verh ogen (PGE 1, dipyridamo these in de pynmtdm~ stoffe n) als de bloedplaatjes biosyn produkten ediaire interm ere meerd of een n bloedplaatjes va t arac higedurende de vorming van PG E2 en PGF 2• vanui mogeis Het ne). aci eth indom ne, (aspiri n ~?naa t te belette manier IIJk dat de twee mecha nisme n op de een of andere gekoppeld zijn. Een betere biochemische kennis van de bloedplaatjes selecti ve functie zou de ontwikkeling van krachtige en meer mbotische genees middelen, potentieel bruikb aar als antitro farmaca, in de hand werken . RESUME iq ues L'aute ur a discut e certain es pro pri etc~s pharm acolog ction plafon Ia de teur inhibi s ament medic aux princip des se posent quettaire et les proble mes le plus communs qui s anidans !'etud e de ces substa nces. Les differe ntes espece Jes yes, emplo ts agulan antico nts differe les es, males etudie nts icame med des me olis proteines plas matiqu es et le metab des tests sont des facteurs tres importants dans !'evalu ation yes en emplo nts icame med Les re. pou r Ia fon ction plaque ttai nt en clinique et qui inhibent Ia fon ction plaque ttaire agisse (PGE ttaire plaque ue 1, cycliq MP d'A augme ntant le niveau :se pladipyri damole et ses derives) ou en inhibant Ia synthi apparaisquettaire de certai ns produits intermediaires qui partir de sent pendant Ia formation de PGE 2 et PGF2• a pas n'est II ne). ethaci indom ne, (aspiri e doniqu l'acide arachi correla en scient action d' exclu que ces deux mecanismes s qui mique biochi voies des e issanc conna La eux. tion entre Ia mise au reglent Ia fon ction plaque ttaire devrai t abouti r a plus tiques rombo antith ent iellem potent nces substa point de puissa ntes et plus specifiques. REFE RENCES M., IREI. BALL , G. , BR ERETON , G.G. , FULWOOD, glandin prosta of Effect P.S, YATE and LAND, D.M. as piE I alone and in combination with theophylline or plaon and ation aggreg t platele ced n-indu rin on collage yclic telet nucleo tides including adenosi ne 3':5'-c monophosphate. Biochem. J .. 120. 709, 1970. in shape 2. BORN , G.V.R.- Observation s on the chang e pof blood platelets brought abo ut by adenosine diphos 1970. 487, 209. .}, (Lone/ l. hate. J. Physio the blood 3. DAY , H.J . and HOLMSEN, H.- Concepts of 197 1. 3, 4, .. awl J-/aem Ser. n. reactio platelet release Acta Clinica Belgica, 10, J (19 75)
PHAR MACOLOG Y OF ANTI -A GGREGATION AGEN
TS
Y4. DE GAETANO, G. , BOTTECCH IA, D. and VERM ce presen the in ts se-clo reptila of tion Retrac LEN, J. ionhes ad t platele the ting of age nts indu ci ng or inh ibi agg regation reaction . Throm bas. RC's .. 2. 71, 1973. s of 5. GROTTUM , K.A . - Electrophoretic investigation , blood platelets. In : J. Caen eel ., Platele t Aggregation 1971. , 11 199-2 pp. Paris, Masso n & Cie., s of pla6. HARKER , L.A . and SLICHTER. S.J. - Studie etic prosth with ts patien in s kinetic gen fibrino telet and heart valves . New Engl. J. M NI.. 283. 1302. 1970. bleedi ng 7. HARK ER, L.A . and SLICHTER, S.J . - The t fun cplatele of ation evalu tor time as a screening test tion. New Engl. J. Med .. 287. 155. 1972 . struc ture 8. KLO EZE, J. - Relatio nship between chemi cal Bioand platele t aggregation ac ti vity of prostaglandins. 1969. 285. 187. mst.}, (A Acta s. chim. hiophy M.J. , 9. KOCS IS, J.J , HERN ANDOY ICH, J .. SILV ER, inh iof on Durati C. AN, RM INGE and J.B. , SMITH regaagg and ation form n bition of platele t prostaglandi f(lantion by ingested as pirin or indometh ac in . Prosta 1973. dins. 3, 141 , E, F. 10. LEONARDI. R.G. , ALEXANDER, B. and WHIT release t platele of bition inhi pirin as of - Preve ntion greaction by the fatty ac id precursor of platele t prosta 1973. 327, J, .. Res bos. landins. Throm effects of II. MASSINI , P. and LUSCHER, E.F - Some ts . Complatele blood on s cation valent di for ores ionoph s. pariso n wi th the effects of th rombin . /Jiochim. biophy Acta (A rnst.}, 372. 109, 1974. - Platele t 12. MANNUCCI , P.M . and GORIN! , S. eds. PleFunct ion and Thrombosis. A Revie w of Meth ods, . 1972 , York New Press, num NUCC I 13. MANNUCCI, P.M., PARETI , F.l. and MAN stratio ~ admini enous intrav L. - Platele t fun ctions after clin. in man of cyclic-AM P and related drugs. J. Lab. 1974. , Med.. 84, 828 EN 14. MIELKE, C. I-1 . Jr. , RAMOS, J.C. and BRITT blee: te templa : agent atelet antipl an as n Aspiri A.F.H... 59. ding time as a monitor of therapy. Am. J . c/in. Path 1973. 236, Stimu la15. MILLS , D.C.B. and MACFARLANE, D.E.g lanprosta by e cyclas nylate ade t platele n huma of tion 1974. 1, 40 5. .. din D2. Thrombos. Res - Factors 16. MUSTARD, J.F. and PACK HAM, M.A. and release esion, adh ction: fun t platele ncing influe 1970. 97, 22. agg regatio n. Ph armacol. Rev., inhi 17. MUSTARD, J.F. and PACKHAM , M.A. - Drugs 51, 31 22, acal., Pharm m. Bioche bi ting platelet fun ction. 1973. ART, 18. NIEWI AROW SK I, S., REGOECZ I, E., STEW t G.J. , SENY I, A.F. and MUSTARD, J.F. - Platele 51, , Invest. clin. J. . rin fib ng erizi polym with interac tion 685, 1972. n pla19. O' BRIEN , J.R. - Effects of salicylates on huma telets. Lance t, I. 779, 1968. storage of 20. PLETSCHER, A. - Metabolism , transfe r and Pharm aJ. Brit. ts. platele blood in 5-hydroxy trypta mine cal.. 32, I, 1968 .
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PHARMACOLOGY OF ANTI-AGGREGATION AGENTS 21. PRAGA , C., MALISA RDI , P , POLLINI , C. COR TELLARO , M. and MARS, G. - Bleeding time and antiaggregatin g drugs: a cont rol led study in elderly patients . Thrombos. Res .. 3. 13, 1973 . 22. RATTAZZ I. L. , HAIMOV , M.N., GLABMAN, S., PAPA TEST AS , A., GALERNT, LM . and BURROWS, L. - Role of th e platelet in th e obliterati ve vasc ular tra nsplant rejection phenomenon. Swx. Fomm .. 21. 241 , 1970. 23. REYERS- DEG LIIN!\OCENTI, 1. , POGGI , A. and DE GAETANO , G. - Platelet aggregati on and hae molys is induced in rats by intrave nous infusion of ADP. Effect of potentiall y antithrombotic dru gs. Sca/1(/. J . Ha('lnaro /., 13, 33 1, 1974. 24. SA LZMAN , E.W.- Cyclic AMP and platelet function . New Engl. J. M ed .. 286. 358, 1972. 25 . SHIO , H. and RAMW ELL, P.W.- Prostag landin E1 in pl atelet harvestin g: an in vitro stud y. Science, I 75. 536, 1972. 26. SMITH , J.B , SILVER , M.J ., INGE RMAN , C.M . and KOCS IS, J.J . - Pros tag landin D2 inhibits the aggregati on of human platelets. Thrombos. Res .. 5. 29 1, 1974. 27. SMITH , J.B. and WILLIS, A.L. - Aspi rin selecti vely inhibits prostag landin producti on in human platelets. Na rure New Bioi., 23 1. 235 , 197 1.
189 28 . VERMYLEN, J.- Acra clin. Be/g.. th is issue, 1975. 29. VERMYLEN , J., DE GAETANO , G. and VERSTRAETE, M.- Platelets and thrombosis . In : L. Pa ller ed., Recent Advances in Thrombosis, Churchill Livingstone, London , pp. 11 3-1 50, 1973 . 30. WEINER , M.- Absorption, binding, metabolism, inactivati on and excretion. Canad. med. Ass. J. . 108. 447, 1973. 31. WEISS , H.J . - The pharm aco logy of platelet inh ibition. In: Spaet T.H . eel. , Progress in Hemostasis and Th rombosis, Grune & Stratton Inc., New York, pp. 199-23 1, 1972 . 32. WILLIS, A.L. - Isolation of a chemical trigger for thrombosis. Prostaglandins, 5, I, 1974. 33. WILLIS , A.L. - An enzy matic mechanism for the antithrombotic and antihemostatic ac ti ons of as pirin. Science, 183, 325 , 1974 . 34. WILLIS, A.L. , KUHN , D.C. and WEISS, H.J . - Acetylenic analog of arachidonate th at acts like as pirin on platelets. Science, /83, 327 , 1974. 35. ZUCKER, M.B.,- Perso nal com munication, 1974 . 36. ZUCKER, M.B. and PETERSON , J.- Effect of acetylsalicylic acid, other nonsteroidal anti-innammatory agents, and dipyridamole on human blood platelets. J. Lab. clin. Med., 76, 66, 1970.
Acta C/inica Belgica. 30, 3 (19 75)
ISSN: 1784-3286 (Print) 2295-3337 (Online) Journal homepage: http://www.tandfonline.com/loi/yacb20
Pharmacological Properties Of Drugs Inhibiting Platelet Aggregation G. de Gaetano To cite this article: G. de Gaetano (1975) Pharmacological Properties Of Drugs Inhibiting Platelet Aggregation, Acta Clinica Belgica, 30:3, 184-189, DOI: 10.1080/17843286.1975.11716995 To link to this article: http://dx.doi.org/10.1080/17843286.1975.11716995
Published online: 21 May 2016.
Submit your article to this journal
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=yacb20 Download by: [Australian Catholic University]
Date: 11 August 2017, At: 05:55
184
PHARMACOLOGICAL PROPERTIES OF DRUGS INHIBITING PLATELET AGGREGATION
Downloaded by [Australian Catholic University] at 05:55 11 August 2017
G. de GAETANO*
The influence of drugs on platelet function can be tested in vitro and in vivo systems both in man and in ex perimental animals. In some in vitro systems, either native or anticoagulated whole blood can be used, whereas for other tests the preparation of platelet-rich plasma is required; in addition , platelet function can also be studied after separating the cells from plas ma proteins by one of many different techniques (" was hed " platelets) (1 2). In vivo studies of drugs influencing platelet fun ction include testingJn vitro the platelet fun ction in blood taken after administration of the drug and the use of certain in vivo techniques such as the bleeding time test (duration , pattern and amount of blood loss), the determination of platelet survival and the accumulation of radioactive platelets in certain diseased organs (6, 7, 14, 21, 28); in experimental animals it is possible to evaluate the effect on platelets of both aggregating substances and inhibitory compounds (23). Some drugs may inhibit one of these numerous test systems developed for stud ying platelet function but not others, making it difficult to establish the significance of such inhibition for clinical use of these compounds. For example, aspirin inhibits platelet release reaction (19 , 36) and prolongs bleeding time (14, 21), but does not modify the retention of platelets in a glass bead
" Laboratory for Hae mostasis and Th ro mbosis Research, Istituto di Ricerche Farmacologiche «Mario Negri», Via Eritrea, 62, 201 57 Milano, Italy. Acta C/inica Belgica, 30, 3 (1 975)
column nor the shortened platelet survival in certain pathological conditions (6). In contrast, dipyridamole is quite ineffective in in vitro systems and does not prolong bleeding time, but increases the shortened platelet survival (6). Let us add that drugs effective in some animal models are devoid of antiplatelet activity in others (23). No single test can therefore reveal all aspects of platelet fun ction and its pharmacological inhibition. In in vitro systems the binding of drugs to plas ma proteins has to be considered : f.i. su1finpyrazone is very active as an inhibitor of the release reaction of washed platelets, but it is almost without effect on platelet-rich plas ma (17 , 35, 36). Another problem which should be carefully considered is the disposition of dru gs affecting platelet fun ction . After ingestion , dru gs are subjected to many processes including absorption, protein binding, distribution in tissues, biotransform ation and excretion. Comparison of the effects of a dru g in vitro and in vivo requires quantitative determination of the drug in the blood; a lack of effect in vivo may be simply due to too low an active concentration of the drug in the blood. This in turn may result from poor gastrointestinal absorption , high protein binding, extensive biotransformation into inactive metabolites or rapid excretion. Conversely, a dru g inactive in vitro may be active in vivo because it requires biotransform ation into active metabolites or acts indirectly, for instance, by releasing physiological compounds which are not avail able in vitro. This problem is furth er
Downloaded by [Australian Catholic University] at 05:55 11 August 2017
PHARMACOLOGY OF ANTI-AGGREGA TION AGENTS
complicated when different animal species are used, since drug disposition may be quan titatively and often qualitatively different (39). The main reactions of platelets which can be studied in vitro are the following (16, 29): I. Adhes ion to collagen and to glass (12); 2. Shape change in res ponse to ADP (2); 3. Development of procoagul ant acti vity (platelet factor 3 ava ilability (12); 4. Change in over-all membrane electric charge (5); 5. Reversible aggregation (3); 6. Adherence to polymerizi ng fibrin an d fibrin retraction (4, 18); 7. Release reaction and irreversible aggregation (3); 8. Uptake and release of serotonin , independent of the ad hesion-aggregation-release · reaction (20). Many compounds inhibit these reactions in vitro, but only a few are suitable for administration to man (I 6, 17, 29, 31) (table 1). Table I In vitro inhibitors of platele t fun ction Group I - Adenos ine and a na logues - Anti-a drenerg ic su bstances - Anti-se ro tonin dru gs - Su lph-scyd ryl gro up inhibitors - Degrad a tion products of some proteins - Pro stag landin s E 1 and 0 1 - Methy l xa ntines - Dipyrid a mo le and Conge ners Group 2 -· Non-steroida l anti- infl am m a tory d rugs - Anti-depressa nt dru gs
Clinically useful compounds inhibiting in vitro platelet function can be divided in two major groups : the first mainly inhibits the reactions numbered l-6 above and the second mainl y inhibits reaction 7. It is not clear whether drugs which inhibit the uptake of serotonin and/or induce its release (reaction no. 8) have any in -
185 nuence on hae mostasis , although cyproheptadine has been shown to inhibit thrombosis of injured vessels in th e hamster check pouch and the accumulation of platelets in renal transplants (22). Among th e dru gs belonging to the first group , dipyrid amole and a number of pyrimid opyrimidine compounds have particul arl y been investigated. These drugs also inhibit the release reaction, but this effect may be due to th eir action on ADP, rath er th an on th e re lease reacti on itself. Indeed, wi th low co ncentrations of releaseinduci ng stimuli , th e small amount of AD P which is released has an enh anci ng effect on further release. Elimination of this ADP effect ca uses a significant reduction in th e extent of release when stimuli are used at low concentrations (17). The concentrations of these dru gs th at inhi bit platelet fun ction in vitro are several times higher th an th ose which ca n be achieved safely in vivo in man ; nevertheless, th ese drugs may be clinicall y effective as antithromboti c age nts since, bes ides platelets, they may also modify oth er relevant parameters such as blood pressure and blood now velocit y. The most potent age nts of the first group of compou nds are Prostaglandin 0 2 and Prostaglandin E1 (I , 8, 15, 26). PGE 1 has little va lue for in vivo administratio n since it is cleared very rapidly from the circu latio n; this property however is advantageous for preparing platelet concentrates free of aggregates for transfusion purposes (25). No information is yet ava ilable on the in vivo effect of PGD 2• A common mechanism of action of the drugs belonging to the first group is their capacity to in crease the levels of platelet cyclic-AMP (table II). Cyclic-AMP inhibits platelet aggregation both when added in vitro and after its intravenous infusion to man (I, 13, 24). Increasing knowledge has led to th e development of a un ifying concept th at substances which stimulate platelets to aggregate tend to induce a decrease in platelet cyclic-A MP, whereas the reverse is tru e for substances which inhibit platelet aggregation . The intracellular level of cyclic-A MP depends on the activity of at least two enzy mes, firstl y adenyl Acta Clinica Belgica. 30. 3 (1975)
186
PHARMA COLOGY OF ANTI-AGGREGATION AGENTS
Table II ADENYL CYCLASE
ATP~j
AMP
Downloaded by [Australian Catholic University] at 05:55 11 August 2017
methyl xantines dipyrida11ole
+
PHOSPHODIESTERASES
cyclase which catalyses the conversion of ATP to cyclic-AMP and phosphodiesterase, which inactivates cyclic-AMP to AMP. The presence of both enzymatic activities in platelets has been reported (24). P'rostaglandin E 1 for example increases platelet cyclic-AMP concentration by stimulating adenyl cyclase, whereas caffeine elevates the cyclic-AMP levels by inhibiting phosphodiesterase. The inhibitory effect of PGE, is strongly potentiated by caffeine (or by other similarly acting drugs). The mode of action of cyclic-AMP in platelet has not yet been established; it is conceivable however that it is linked to the availability of membranebound Ca++ in the cytosol of the platelet. In this regard , very recent observations that ionophores induce several platelet reactions by making available intracellular Ca++ are of the greatest interest (ll). . Among the drugs belonging to the second group, i.e. inhibiting the release reaction, nonsteroidal anti-inflammatory agents have received much attention, particularly because they are readily available, inexpensive and relatively nontoxic. Aspirin is certainly the drug most extensively investigated for its effects on platelet funct-ion, haemostasis and thrombosis . Doses of antiinflammatory drugs much lower than those generally used in human therapeutics are highly effective in inhibiting some of the platelet reactions; this property should minimize the occurActa C!inica Belgica, 30. 3 (19 75)
renee of side effects, particularly at the gastrointestinal level. Aspirin and related drugs do not increase the levels of platelet cyclic-AMP (1 ); they inhibit the platelet synthesis of one or more cyclic endoperoxide intermediates which appear during the biosynthesis of PGE 2 and PGF2a from arachidonate (table Ill) Smith and Willis (27), first reported that the formation and release of prostaglandins E2 and F2a which occurs in thrombin-stimulated platelets is inhibited by indomethacin and aspirin, whereas sodium salicylate has only a weak effect. Phenylbutazone has activity .intermediate between aspirin and sodium salicylate, whereas acetaminophen (paracetamol) is inactive. To aremarkable degree, these results parallel the relative ability of the drugs to suppress the second phase or irreversible platelet aggregation and the accompanying release reaction. In addition, Kocsis et at. (9) have shown that after oral administration of aspirin or indomethacin, the duration of the inhibitory effects on aggregation and release parallels their effects on platelet prostaglandin production. Subsequently it has been demonstrated that during the biosynthesis of prostaglandins in the platelets one or two endoperoxide intermediates transiently accumulate in the cell and trigger irreversible aggregation and the release reaction (32, 33). These intermediates have been called: "Labile AggregationStimulating Substance" (LASS). Aspirin and
187
PHARMACOLOGY OF ANTI-AGGREGATION AGENTS
Table Ill
ARACHIDONIC ACID
T YA I
I I I I I I
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~
111
PLATELET ENZYMES
.
t
ASPIRIN
_..._
IRREVERSIBLE A6GRE6ATIDN and
RELEASE REACTION
possibly other anti-inflammatory drugs block the enzymatic conversion of arachidonic acid (the precursor for biosynthesis of PGE 2 and PGFta) to this pro-aggregating material and to PGE 2; the latter sensitizes the platelets to the aggregating effects of LASS (33). It has also been observed that arachidonic acid is able to prevent the inhibitory effects of aspirin on platelet aggregation (10) and that an acetylenic analog of arachidonic acid (TY A) acts like aspirin on platelets since it competes with arachidonic acid towards prostaglandin synthetases (34). In conclusion, all the compounds that influence platelet function seem to work through two main enzymatic mechanisms : the first one involves the formation of increased levels of platelet cyclic-AMP and the second one the inhibition of the formation of a pro-aggregating factor during the biosynthesis of prostaglandins. It is unknown, at the present time, whether these two proposed mechanisms are independent or interrelated in some way . One would welcome the development of more potent and' more selective agents, active as stimulants of platelet adenyl cyclase or as inhibitors of phosphodiesterases or of prostaglandin synthetases. ;
Possibly , association of one or more of these drugs could have much more benefici al effects on thromboembolic complications than those obtained until now with the drugs presentl y available. KEY WORDS Blood Platelets- Platelet Aggregation - Platelet Agg regation Inh ibitors pirin - Dipyridamole - Cyclic-A MP - Pros taglandins.
SUMMARY The pharmacological properties of drugs inh ibiting platelet fun ction have been reviewed. Some of the problems arising in ex perimentation of potent iall y useful substances have also been discussed. Animal species, anticoagulants, plas ma proteins and drug disposition are the main factors innuencing platelet fun ction tests. Clinically useful drugs inhibit platelet fun ction by either increasing the levels of platelet cyclic AMP (PGE " di py ridamole, pyrimido-py rimidine compounds) or inhibiting the platelet biosy nthesis of one or more intermediates appearing during the formatio n of PGE2 and PGF2a from arac hidonate (aspirin , indometh ac in). Possibl y the two mechanisms are interrelated in some way. Knowl edge of the bioc hemical pathways underlyi ng platelet fun ction, should result in the deve lopment of more potent and more selective agents, potentiall y useful as antithrombotic dru gs. Acta C/inica Belgica, 30, 3 (1 975)
188
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SAMENV ATIING de funcDe farmacologische eigenschappen van farm aca die ken. tie van de bloedplaatjes inhi beren worde n bespro teren men experi het bij en voordo zich die Enkele problemen eveneens van potentieel bruikb are geneesmiddelen worde n a-proteiplasm tia, agulan antico , n ~te i~rsoo D gediscuteerd .. testen van nen en stofwt ssellng ZtJn de hoofdfactore n die de de bloedplaatjes functi e beinvloeden. l aat. Klin isch bruikb are farm aca verhinderen de bloedp AMP in isch cycl n va ntratie conce de zowel oor d . cue Jesfun le, pyrim idobl~~l a.atJeS te verh ogen (PGE 1, dipyridamo these in de pynmtdm~ stoffe n) als de bloedplaatjes biosyn produkten ediaire interm ere meerd of een n bloedplaatjes va t arac higedurende de vorming van PG E2 en PGF 2• vanui mogeis Het ne). aci eth indom ne, (aspiri n ~?naa t te belette manier IIJk dat de twee mecha nisme n op de een of andere gekoppeld zijn. Een betere biochemische kennis van de bloedplaatjes selecti ve functie zou de ontwikkeling van krachtige en meer mbotische genees middelen, potentieel bruikb aar als antitro farmaca, in de hand werken . RESUME iq ues L'aute ur a discut e certain es pro pri etc~s pharm acolog ction plafon Ia de teur inhibi s ament medic aux princip des se posent quettaire et les proble mes le plus communs qui s anidans !'etud e de ces substa nces. Les differe ntes espece Jes yes, emplo ts agulan antico nts differe les es, males etudie nts icame med des me olis proteines plas matiqu es et le metab des tests sont des facteurs tres importants dans !'evalu ation yes en emplo nts icame med Les re. pou r Ia fon ction plaque ttai nt en clinique et qui inhibent Ia fon ction plaque ttaire agisse (PGE ttaire plaque ue 1, cycliq MP d'A augme ntant le niveau :se pladipyri damole et ses derives) ou en inhibant Ia synthi apparaisquettaire de certai ns produits intermediaires qui partir de sent pendant Ia formation de PGE 2 et PGF2• a pas n'est II ne). ethaci indom ne, (aspiri e doniqu l'acide arachi correla en scient action d' exclu que ces deux mecanismes s qui mique biochi voies des e issanc conna La eux. tion entre Ia mise au reglent Ia fon ction plaque ttaire devrai t abouti r a plus tiques rombo antith ent iellem potent nces substa point de puissa ntes et plus specifiques. REFE RENCES M., IREI. BALL , G. , BR ERETON , G.G. , FULWOOD, glandin prosta of Effect P.S, YATE and LAND, D.M. as piE I alone and in combination with theophylline or plaon and ation aggreg t platele ced n-indu rin on collage yclic telet nucleo tides including adenosi ne 3':5'-c monophosphate. Biochem. J .. 120. 709, 1970. in shape 2. BORN , G.V.R.- Observation s on the chang e pof blood platelets brought abo ut by adenosine diphos 1970. 487, 209. .}, (Lone/ l. hate. J. Physio the blood 3. DAY , H.J . and HOLMSEN, H.- Concepts of 197 1. 3, 4, .. awl J-/aem Ser. n. reactio platelet release Acta Clinica Belgica, 10, J (19 75)
PHAR MACOLOG Y OF ANTI -A GGREGATION AGEN
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Y4. DE GAETANO, G. , BOTTECCH IA, D. and VERM ce presen the in ts se-clo reptila of tion Retrac LEN, J. ionhes ad t platele the ting of age nts indu ci ng or inh ibi agg regation reaction . Throm bas. RC's .. 2. 71, 1973. s of 5. GROTTUM , K.A . - Electrophoretic investigation , blood platelets. In : J. Caen eel ., Platele t Aggregation 1971. , 11 199-2 pp. Paris, Masso n & Cie., s of pla6. HARKER , L.A . and SLICHTER. S.J. - Studie etic prosth with ts patien in s kinetic gen fibrino telet and heart valves . New Engl. J. M NI.. 283. 1302. 1970. bleedi ng 7. HARK ER, L.A . and SLICHTER, S.J . - The t fun cplatele of ation evalu tor time as a screening test tion. New Engl. J. Med .. 287. 155. 1972 . struc ture 8. KLO EZE, J. - Relatio nship between chemi cal Bioand platele t aggregation ac ti vity of prostaglandins. 1969. 285. 187. mst.}, (A Acta s. chim. hiophy M.J. , 9. KOCS IS, J.J , HERN ANDOY ICH, J .. SILV ER, inh iof on Durati C. AN, RM INGE and J.B. , SMITH regaagg and ation form n bition of platele t prostaglandi f(lantion by ingested as pirin or indometh ac in . Prosta 1973. dins. 3, 141 , E, F. 10. LEONARDI. R.G. , ALEXANDER, B. and WHIT release t platele of bition inhi pirin as of - Preve ntion greaction by the fatty ac id precursor of platele t prosta 1973. 327, J, .. Res bos. landins. Throm effects of II. MASSINI , P. and LUSCHER, E.F - Some ts . Complatele blood on s cation valent di for ores ionoph s. pariso n wi th the effects of th rombin . /Jiochim. biophy Acta (A rnst.}, 372. 109, 1974. - Platele t 12. MANNUCCI , P.M . and GORIN! , S. eds. PleFunct ion and Thrombosis. A Revie w of Meth ods, . 1972 , York New Press, num NUCC I 13. MANNUCCI, P.M., PARETI , F.l. and MAN stratio ~ admini enous intrav L. - Platele t fun ctions after clin. in man of cyclic-AM P and related drugs. J. Lab. 1974. , Med.. 84, 828 EN 14. MIELKE, C. I-1 . Jr. , RAMOS, J.C. and BRITT blee: te templa : agent atelet antipl an as n Aspiri A.F.H... 59. ding time as a monitor of therapy. Am. J . c/in. Path 1973. 236, Stimu la15. MILLS , D.C.B. and MACFARLANE, D.E.g lanprosta by e cyclas nylate ade t platele n huma of tion 1974. 1, 40 5. .. din D2. Thrombos. Res - Factors 16. MUSTARD, J.F. and PACK HAM, M.A. and release esion, adh ction: fun t platele ncing influe 1970. 97, 22. agg regatio n. Ph armacol. Rev., inhi 17. MUSTARD, J.F. and PACKHAM , M.A. - Drugs 51, 31 22, acal., Pharm m. Bioche bi ting platelet fun ction. 1973. ART, 18. NIEWI AROW SK I, S., REGOECZ I, E., STEW t G.J. , SENY I, A.F. and MUSTARD, J.F. - Platele 51, , Invest. clin. J. . rin fib ng erizi polym with interac tion 685, 1972. n pla19. O' BRIEN , J.R. - Effects of salicylates on huma telets. Lance t, I. 779, 1968. storage of 20. PLETSCHER, A. - Metabolism , transfe r and Pharm aJ. Brit. ts. platele blood in 5-hydroxy trypta mine cal.. 32, I, 1968 .
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PHARMACOLOGY OF ANTI-AGGREGATION AGENTS 21. PRAGA , C., MALISA RDI , P , POLLINI , C. COR TELLARO , M. and MARS, G. - Bleeding time and antiaggregatin g drugs: a cont rol led study in elderly patients . Thrombos. Res .. 3. 13, 1973 . 22. RATTAZZ I. L. , HAIMOV , M.N., GLABMAN, S., PAPA TEST AS , A., GALERNT, LM . and BURROWS, L. - Role of th e platelet in th e obliterati ve vasc ular tra nsplant rejection phenomenon. Swx. Fomm .. 21. 241 , 1970. 23. REYERS- DEG LIIN!\OCENTI, 1. , POGGI , A. and DE GAETANO , G. - Platelet aggregati on and hae molys is induced in rats by intrave nous infusion of ADP. Effect of potentiall y antithrombotic dru gs. Sca/1(/. J . Ha('lnaro /., 13, 33 1, 1974. 24. SA LZMAN , E.W.- Cyclic AMP and platelet function . New Engl. J. M ed .. 286. 358, 1972. 25 . SHIO , H. and RAMW ELL, P.W.- Prostag landin E1 in pl atelet harvestin g: an in vitro stud y. Science, I 75. 536, 1972. 26. SMITH , J.B , SILVER , M.J ., INGE RMAN , C.M . and KOCS IS, J.J . - Pros tag landin D2 inhibits the aggregati on of human platelets. Thrombos. Res .. 5. 29 1, 1974. 27. SMITH , J.B. and WILLIS, A.L. - Aspi rin selecti vely inhibits prostag landin producti on in human platelets. Na rure New Bioi., 23 1. 235 , 197 1.
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Acta C/inica Belgica. 30, 3 (19 75)
