Cardiovascular pharmacology satellite conference

TIPS - February I982

IInterferon

45

The pmblemsof cloning IFN DNA have twen overcomein variousways and thishas Shakespearewmfe of a‘tide in the affairs of shown that. for instance, humaa IMn is men’ and 1think wrily at times of the way c&d for in at leasteight genes”.This tindthat enth~i~m for work on interferon has ing is con&lent with evidence that IFNO gone upanddownoverfheyearssince 1957 purified from human lymphobl~toid cells when the first paper by isaacsand Linden. contains at least five different mokcular mann appeared’. This described how a species*. In addition to cloning DNA into chicken cell exposedto an inuctivafedvirus ~c~~c~iu coil if has been possible to pmduceda protein that renderedother cells have the DNA trauslafed and so human resistantto infeetion with viruses.and thus inferfemn is now being pmduced in batmediated virus interference. ferial cultures in considerableamounts’. ft Thereafter there was a premature hope will therefon:aof he long before there will that this interferon could be usedas a treat- be no pracfical limitation to the amountsof ment for virus infectrons.This was dashed ices available, at least for research becauseIhe subsfauccpmved to be ‘species purposes.and producing IF% from verfespecific’, and did not protect human cells brate cells will be an outdatedtechnology. and was also difficult to pmduce. Tben The purification of interfemns can be there was enthusi~m for treating patients greatly helped by using monoclonal anti. with inferfemn inducers. parficularly bodies producedby :be mouse hybridoma ~ubl~s~~ RKAs such as poly l:C, ~ehn~~~. Such i?ntibodycan be atfached produced synthefically, or viral dsRNA to columnsonto which the interferonis then extracted horn infected culturesof penicilabsorbed. h can ‘w: eluted from a washed lium. This hope founded on fhe relatively column in a greatly purifed sfafe. W:: have small amountsof interferoninducedand fhe rccentfy used such interferon prepared by toxicity of the inducers. D. Se&r of LMB. Cambridge, fmm maThen K. Cantell in Helsinki developed teriai suppliedby K. Cantell. This purified methodsof making substantialamountsof material containedno deteetabk contamininterferon from vi~~fect~ human ant except for a fraee of mouse immunoleucocyfes. This made if possible to proglobulin and was given under double duce sufficient amounts of interferon to bii~co~tjons to vofunfeersat the U.K. give a realistic and effective humandose of Clinical Research Cenue’. These subjects IV-10 units, and to show that virus ~rn~~n~ of headache, malaise, and infections such as herpes msfer in ofber generalized symptoms. and the imm~om~mi~d patient or developedfever, fachycardia, a rise in cirrhinovinfs-inducedcolds in normal volunculating neufrophils and a fall in lymph+ teers could be modified by exogenous cytes. which lasted for some hours. This infertemn. Following fhe suggesrionsof I. confums conclusionsdrawn f?om erperiGresserof Paris, H. Stranderin Stockholm merits in animzds, that interfemns have a also obtained preliminary evidence that a wide range of effects. and it may well br tumour, the osfeogenicsarcoma, could be &af the general symptoms experienced influenced by using interferon as an adjuafter a virus infection of the respiratory vant therapy after the patient had received tract, such as influenza. are actually the conventional initial therapy by surgery or radiation*. There have been considerableadvances in fun~rnen~i k~wl~ge about inferfemns. Setting aside our new understanding of how if is induced, and the biochemical In the idyllic setting of Canada’s Ban@ mechanisms by which it inhibifs virus National Park the second biiinnudl carreplication. pamcutarly by inhibif~ngfrans diov&scuhxp~~acol~;gy satellite conferlation of the virus messengerRNA, inter- ence was held on 21 and 22 August at the Banff Cenfre. Aml~ngstthe more than 70 feronhasnow beenpurifiedto ho~ge~i~. h turns ouf fhaf there are many different purticipans, our Canadian hostswelcomed inferfemns. Firstly. the inferferons vary ovemeas colleagues fmm France (G. from specieslo species and secondly,each Cms), New Zealand (W. Lubbe) and species produces at least three types of Nigeria (G. ~n~~ulu~hi). Sessionswere interferon (EN), namely IFNtv from virus held in the morningsand afier dinner in the available for infected leusocyfes. lFN(3 from inducer- evenings, leaving a~fe~s treated fibmblasts, and IFNr from informal discussion, hiking md sighfimmunologically stimulatedlymphocytes. seeing.

re*ulf of the pre.senceof inlerferon in the circulation; the concentrafions found in such dissses are nof veer)different fmm fhosefound in the circulation of the voluu1~333when they were feeling so uncomforfable. There i\ evidence that YLlc& WIIK: of these effects are mediafed by prosfaglandins and mare wmk must be done on thi, subject It is also fascinatingto ~onfcmpl.ae tbe wide range of biological effecs thdf interferons have; the pyrogenic reqmuz has been mentioned. but they alu, inducz d rather slow inflammafor? respon’le, increasethe expressionof tissuefrdnsplplirrifation antigens on cells, increase natural killer cell activity. modif) delayed hypetsensifivity. tissue rejection and serological responses, present ceff division. and soon. Even if we diquver ho% to use interferons in the treatmentof certain virus infections and cancm there ma) hi still other therapeutic po!+sibilitiesto biconsidered. 13 -\ f Tk’RELl

Reading Iii

Cardiovascularpharmacology satelliteconference The firhi se&on organizculb? P. E. Dresel (Dalhouaie fJniverrti> . Halib\ 1 ~wenut somenew anti~~~thrni~ drug\ fp his openingcommentsDr Drebefnotedthat the real cgmcernIn) with the treatment of’ post-infarctionarrhythmias. and reminded us that we still don’t Lnow the mechanism of action of’ currendy avail;lble anti~hythmi~s, All we really know is that they occasionaliywork. and he mised the quesfion as IO whether Ihe mechanismof their action must necessarilvbc confin& tu both re-entry and aut~~at~~ity. Should we not

,.,,L.LrC, !b.ard,.*, VW.. NJ lllhr h!: >2&WIt*xut tic ‘C

examine their effects upon cardiac muscle and not just Purkinje tibers? It is known that some drugs have more efficacy on ‘sick’ than on healthy Pnrkinje cells. whilr: others have an opposite pattern. Would newer agentswhich ~fe~nti~ly exetl their antiatrhythmic action on the ventticuhtr myocardium be equally good antiatrhyth mics? Following this brief introdttction. J. Byme (Mead Jam, Co., Evansville. htdian~~)repnrted on the development of enc&Gde in a variety ofin I&O andin vivu preparations. This new drug appeats to exhibit effiacious antiatrhythmic activity as gauged by a number of currently acceptedcriteria. For example, eneainide decreasesV,,: of the Purkinje actian potential while slowing conduction in both myocardial ant! Pnrkinje fibers. Intetestingly, rtudks nave shown that encainide prolongsthe ac*;attpotentialmom in infarc ted than in normal muscle. Moreover, it increases the ventricular fibrillation thteshoid in anesthetizeddogs, and does not appear to affect ‘slow channef’ action potentials.The Odemethylated metabolite appearsto be two to four times mote potent than the parent compound. Results to date. indiiate that encainide does not display specif~ity. but exhibits a spectrumof antinrrhythmic activity similar to that of quinidine. A compar%on of the antiarrhythmic effects of disobutamide (DB) and dis~y~mide (DP) was presented by R. Novotney (G. D. Searle Br Co., Chiiago). Both agentsdisplay antiarrhythmicactivity againsta variety of arrhythmic laboratory modeIs (e.g. coronary artery ligation, cat~~a~~ and oua~i~i~~ed artbythmias,Harris technique.etc. ). However. resultsobtainedusingtmntr~!embrane a&m potentialand voltage clamp studies, indicate that DB unlike DP, but similar to tetrodotoxin.if specific as a sodium &annel blocker. Unfortunately, histological lesions have been observed following chronic animal toxicity studies, so that furtherclinical investigationhas ceasedfor the tbne being. The specitiiity of clotilium to increase cardoacrefractorinesswithout stowing the conductionof normal impulses,0; altering the hrnction of the rapid sodium channel, was diiussed by M. Steinberg (Eli Litly Co., Indianapolis). It was hypheked that these effects should be beneficial in the

treatment of reentry atrhyth&s, but unforttntateIy , the drug can also indncecardiac arrhythmias, espe&By in h$otharn+ anesthetizedanimals where its effects on mfraetorinesSare also markedly potentiated. (Is there a good antiarrttythmicthat is not anfiythmogenic in suffiiently high doses?).Clofitium sehzetivelyprolongsthe

action potential duration and refractory period of the heatt, but apparently ha no effect upon normal automaticity.It displays great specificity for the my~a~ium. and a prolongedduration of action (f I .=9 days). However, it is ineffective againstmany of the usual arrhythmogeniclaboratory models having enhancedautotnaticity as their ele~ff~hysio~~eal basis. The tinal presentationin this sessionwas given by W. Wastila (Burroughs Wellcome, Co.. Research Triangle, NC) who spoke on m&mine. an analog of &~idi~ whiih, however, is not an adtonergic nenronal blocker. Like bretyt ium, meobentine increases ventricular fibrillation thresholdand preventsarrhyth miss during and after coronaryligationboth in vitro andin v& , aswell as thoseresulting from ouabain. The drug also selectively accumulatesin the myocardiumand not in other muscle masses,nor is this affected by ~~os~~~torny. It hasa biological half-Me of between 5 and 12 h. in discussion it was notedthat it would be helpfui if we could be more definitive in differentiating between the atuiarrhythmicand antifih rillatory actionof a drug. ’ David Severson(Unive~i~ of Calgary, Calgary) chaired the second session on cyclic nucleotidesand cardiac metabolism. Speaking on ‘Factors Influencing the Release of Cyclii Nucleotides from PerfusedHearts’, W. R. Ingebretsennotedthat the administrationIofisopmterenolnot only resultsin the accumulationof CAMP in tbe tissue, but also induces the tzlease ol cAMP into the effluent. The dre&nktced release of CAMP occursin the sametime frame as

the release of glycerol and glycerol phos phate. Proknecid (0.01-l mkt) inhibits CAMP efflux without affecting tissuecontent of cychc nu~le~jdes. The acuity of choiineqic agentsto stimulate the release of cGMP is blocked by atmpine. Pm benecid, but not atmpine, is able to inhibit sodium ni~pmssi~induced release of cGMP. N~ally, anoxia produces an increasein CAMP release, but in the presence of diabetesthis does not occur. These results(and others) suggestthat the cyclic nucleotide release process is not tightly linked to changesin tissuecontent, and that the regulationof this processis complex. 3. Heller-Brown (University of California, San Diego) speakingon the ‘Biochem ical Effectsof the Pa~ympa~etic Nervous Systemof the Atrium’, notedthat in murine atria micromolar concentrations of carbachol and other muscarinic agonistspro duce a rapid non-competitiveblockade of ~at~~lamin5stimu~t~ cAMP accumw I&on. Cholinesteraseinhibits as well as K’ depolarizatmn-induced ACh release, also attenuate isopmterenol-stimulated CAMP formation. Studies on ACh release fmm murine atria suggestthat this may be a presynaptic site of an~gonism between sympatheticand parasympatheticstimula tion. Muscarinic receptorsin the atrium can be competitivelyantagonizedby verapamil suggesting a ~lat~onship between these receptorsand Gas’ channels. f. Diamond (University of British Cot umbia. Vancouver) then spoke on the ‘Possible Role of Cyclic GNP in Cardiac Con~~tility’, and presentedevidencesuggestingthat the negative inotropicactionof

ACh is cot necessarily linked to cGMP. For example. no negative inntropic action occurs Following sodium nitropruuside, hut marked increases in cGMP occur.

oxygen. even though coron3rq flow dew% I PC. 3nJ mducrt anahlg6~1~ clcctr+ not change. Thcv results were imcrprcteJ ph)wtiopcdl dltrrautm\ m c;lmnc m?tF as suggesting that intracellular mechani\m> t3rdium. cspcciltll! In the prc\cncc ot’ 111ido\i\.

either ACh or carbachol sufficient to elo

may be pla)ing prominent roles in Ihe arrhythmogenic and mtiarrhythmic actions, and suggests th3t a relative hypnxia

vate cGMP. do not produce a negative ino-

accomp;l?ies

qs

tropic action. Diamond suggested that the nitroprussidc data must be interpreted cautiously in light of the recent report of

mirs.

Moreover.

very

low

concentrations

of

Lincoln and Keely (1980). thrt large increases in cGMP produced by nitroprusside are not accompanied by activation of cGMP-dependent protein kinase, whereas the smaller increases produced by ACh are accompanied by activation of the kinase. Thus, many recent studies fail to suppoti 3 causal relationship between increases in tig sue cGMP levels and negative inotmpy. The Rnal paper in !his session was presented by Dr Seversc;n who spoke on the ‘Regulation oFLipolysis in Cardi3c Tissue’. From studies using Langendorff heart prep. arations he noted that the appearance of glycerol in the effluent is a good indication of the mobilization of Free Fatty acids in the hear? by the hydrolysis of endogenous triglycerides. Mobilization of free fatty acids is enhanced by such things as glucagon. catecholamines, dibutyrylcAMP, calcium, ouabain, theophylline and work. while inhibition of lipolysis occurs Following an increase in free fatty acids, ketone bodies. hypoxia and nieotinic acid. At present, there is no evidence for the activation of any cardiac lipase activity by acovalent moditication such as CAMP-dependent protein kinase phosphorylation. The possibility that the principal mechanism For cardiac lipase regulation may involve allosteric effecters was discussed. The thii session, organized by R. Tanz (Oregon Health Sciences University, Ponland), was on DN~S and Myocardial Metabolism, with arrhythmogenesis being emphasized. In the opening paper the chairman proposed a ‘Hypothesis For the Role of CAMP in Ouabaibinduced Arrhythmogenesis’. Evirience was pre sented showing that very soon after the addition of a ‘toxic’ concentration of oua bain to guinea-pig Langendotff heart prep arations, ventricular CAMP levels show a

transienttwcFfold increase,which return to normal levels just before the occurrence of occasional arrhythmic phenomena. Later, with the development of tachyarrhythmias and then asystole, ventricular ATP levels decline SigniFicantly and the release of lactic dehydrogenase and cnatine kinase into the coronary eFtluent increases dramatically. Pretreatment with pmpranolol negates these effects. Moreover, pmliminqj results suggest that pretreatment with lidocaine increirws the ability of the ouabain-treated heart to utilize more

ouahdin-induced

arrh+

C’IIIT \ugpc\tcd th,n rhrw f3clorr

m3y be major precipitant5 oi mdlignant ventricular arrhythmia\. More recent tind-

have shown that LPC \rimulate\ of aden>latc c)cla\e Independent W. Lubbe (Univenity of AuckBnd. catcuhol;lminc\. enhancer Ca” Ir;ln\prn New Zealand) next reported \ome result\ acrtn\ 1*ol31ed m,cs)tc+. and maintain\ From his studies on ‘Cyclic Adenine Sic tranrmemhrdne po(cntlal\ dcpendcnt cleotides and Arrhythmia\ in Acute exclusively 6,n the 40% ;nu.ud current (I..) Myocardiol Is&mid. using cornnary dc\pite concomnarn acidnu.. anery ligated isolated perfused r3t he;lrt\. The tinal paper in thi* q\clon war.prcThese studies were aimed at attempting to rented by K. R. Courtnq (P3lo Alro Me&correlate biochemicrl alterations with c;lrl\ cal Rewe;lrch Foundauon. California) am changes in vulnerability to ventricular tih ‘Interval Dependent Drug Acturn in Dcprcrillation. The rapid reduction of wntricuhr wed Myocardium’. l!Gng p3pill.n) muwle* fibrillation threshold (reflected by spoa From guineil-pig\. the myoc3rdial dcpret3neously occurring ventricular tachycardia \%mt action of %\en antiarrh)rhmlc drug\ 3nd tibrillation) is accompanied hy high ua\ studied u\ing Ihc up~ohc vclocn! of energy phosphate depletion and cyclic the sction potenoal 3, a mc3.urc of AMP accumulation in ischcmic myccar- m>cxlardial e%citilhiln> $Vhen up\trokc dium. Interventions that increase tl+ur kelocity wa\ further dcprcs.4 b> \ub\otuIcyclic AMP (e.g. phosphodiesterase inhihi. ing Tyrode’\ 4ution ~nhou~ bicarbondtr tion with theophylline or I-mcth) I in rhe prtfusate, theu drug ctfccts ucrc .3-isobutyl xanthine) increase vulneral~ility significand) potcmi.ncd 3% indlc&J b) Me%uremenr\ were to tibrillation without altering high energy longer hslf-times. phosphate depletion. Pretreirtment of hsans made on the time tI> recobq of the with pmpranolol or amiod;irone reduce upstroke tetocity follou mg cqn.~sure to cyclic AMP in ischemic tissue and dirnuu~h drug. Lipophilic drug \tmcture\ h;l\ine small mol. wIs (belou Xl) all &pi+ vulnerability to fibrillation without increas ing high energy phosphat%,s.In this model short half-time* (to rcco\ce 1. anlf thus of acute myocardial ischemia. alreration\ in confe: upon *uch Jru:\ Ihe tqdbrlir\ lo .md klrhcr vulnerability to ventricular fibrillation in Moth aslectivcl! c\trdr!\role\ the early phase of ischemia are coupled to high frcqurncy 3rrhlthmldr. The fourth and tinal ***ion chaired b) J changes in the tissue content ofcsclic .A%lP !4cNsill (Uni\ersir) ot Btiirh Columbis. rather than to changes in tissue high eneg phosphates. Thus, it appears that. d> in Vancomer) ~3s 3 p*pourri (II r,jpic\ of currcnf imcrc*I. T. Irl BroJ! (Xlichipdn ouabain-induced toxicity. the initid ISt Ldnbin$ bIcJ off a rth increase in tissue CAMP may play a role in Srate l_5\rr\it>. 3 bnef rc\ica oi propouJ mcchdnitm\ initiating an arrhythmogenic state. uhereb\ c3rJi3c pl)ccrsiJc> mcrcau crjnThe third paper presented by P. B. Corr traclilc &r tissue in viva. and increases rulrfold within IO min of coronary occlusion. Th;lr noted that lithium. uhich inhibir3 thr &ium pump, c3use\ a po4vr inotroplc accumul3ted metahllitcs in ischemrd may respon\r even though it Jock noI :nhibir cause arrhythmias is supported by the t&ts luch btoreo\er. (N3. +K‘ &ATPdse. that, (1) supert$sion negates ischemically Jiverse subst3nce~ aa vctdmtdinc. moneninduced electmphysiolopical altewtions: ain. batr3chotouin and gny3notoGn nnl (2) application of venous blood draining only incrc3se the sodium manslent. bur al*o ischemic regions in vivy results m electmphysiological changes analogous to produce arrhythmias. tie tbm suggebled that the r&son c3rdi3c glycosides produce those seen during ischemi.rfi, ~ivo. and (3) no increase inskeletal muscle Force. resides isehemia in rive is more arrhyth~og~ni~ than hypoxia with maintained flo\h Long-’ in the fact rhat thev I3ck sn effective NaiCa chain acyl camitine is structurally similar to exchange mechan&,m. .Also presenteJ ~3s

48 the status of the endogenous liprnd for (Na’ t K* )-ATPase. An upd21e on calmodulin was fresented

CAMP

rises, and subsequently, tension than !Subshlnces other increases. catecholammes. such as histamine, also

where they exen a primary effect upon he;u! rale. and in the left ventricle where an

by F. Vincenzi (University of Washington, Seattle). who noted that iI is an inlI’2cehht acidic protein with .1 mol. wt of about

increase cunliac cAMP lttveis. The thought

predominant in the lcfi atrium. and if you look hard enough, some are also

?O,ooO that possesses four calciunv binding

‘was expressed that perhaps cAMP is a triggering event Ibr certain types of arrhythmias as well as fornhe enhancement of con-

sites, and therefore acts as a calcium recep Ior. Apparently, it is invol\,ed in the reguls tion of a variety of celluar responses, and can bind a number of different drugs. For example. the antipsychoric tritluoperazine may act by being an amicahnodulin agent. as do the amiarrhyrhmics quinidine and propranolol. as well as the local anesthetic dibucaine. These recent observations will hopefully assist in the development of more specific annicalmodulin drugs in the future. G. Drummond (University of Calgary, Calgary) then talked on ‘Cyclic Nlc cleotides and the Heart’. He indicated that we still don’1 know how an increase in CAMP by epinephrine leads 10 enhanced myocardial contractile force. The activa tion of protein kinase occurs shortly after

tractile force. The final presentation on cardiac histamine receptors was given by J. McNcill. As noted b> Drummond. the addition of histamine to the media perfusing isolated guineapig hearts results in the augmentation of both force and rate which appears to be related to enhanced cAMP. if, in addition, theophylline is added 10 the perfusate. the dose-response curve i:, displaced to the left. And conversely, pt lor exposure 10 HZ amagonists (e.g. cimetidine) shirts it 10 rhe right in a manner suggesting competiGve antagonism. However, there are distinct differences with respect tothc prevalence of HI and HY receptors in different portions of the guinea-pig heart. Thus, HI receptors seem to be predominanl in the right atrium

The Pharmacologiical Society of Canada: newslettbr Included below. as the maim contribution to’ this newsletter,. is a profile of ihe Department of Pharmacology of the University of Ottawa prepared by Drs Stan Kalsner and! Radhey Singhal. Hopefully, this will be thej fust of several Canadian departments to be: profded in the future.. The long-range aspir-/ ation is to familiarize TIPS readers with! the Canadian pharmacological scene and! the names and faces behind the desks and! benches.

a charter vested control in a representative Board cf Governors. Bilingual since its inception, like the surrounding community it serves, the University of Ottawa today is Canada’s oldest and largest officially bilingual University. with most of its progams offered in both English and French.

increase in force is manifest. HI receptors are

located in the lefi venlricle. McNeil1 then noted Ihat apparently HI rcccptor stimulation increases CAMP Icvcls. bul stimulation of HI receptors does not. There are also wide differences with respect 10 species. Thus. rat heart muscle contains neither HI nor l-L. The pathophysiological mle of histamine receptors is still unresolved. The relatively small sizeofthegroup was conducive for. and led to. considerable dis cussion which is really the object of conferences such as this. Its succrss was to a large measure due to the superb organization of Dr McNeil1 and his outstanding staff. The nexl cardio! 2scuhr pharmacology satellite conference will be held in 1983. RALPH TANZ

The Department of Pharmacology was set up in 1948, 3 years after the establishment of the Medical School. In keeping with the bicultural nature of the City, the eight full-time staff have brought a varied ethnic background to the Department. Fortunately, almost everyone speaks English wnh a ‘smattering’ of French, but it is not unusual to overhear a vigorous conversa tion in progress in the hallway in an unidentified foreign tongue.

Pm&Z the Departmentof Pharmacology at the University of

Ottawa Pimred here in Fig. 1 are the full-time. memlbers of the Department of Pharmacol-! ogy of the University of Ottawa, a modest, sized group at the Medical School in this; bilingual Canadian city. The beginnings of the University of’ Ottawa go back lo 1848. Originally called;

the College of Bytown. iI is located near the f Ottawa River and Rideau Canal in the capL I tal of Canada, one of the xnostscenic andi beautifulcities in North America. but with a distinct European flavour. The Canal, ~.]ich borders the University of Ottawa. campus, was fmt built for militar?, defense ! pqoses. but now serves the pleasurabler functions of beiig the longest skating rink in the world and in summer. a channel for small boats. In its present form and structure. the University of Ot?awa dates from I%5 when

Fig. I. The membm of dw Lvptrtmmt at Ottmw. L. to R. (seated):P. D. Hrditta, I. Ma.wkiew’~~zKwilecki. 2. Merali. L. to R. (sranding):S. Kahter, M. MocConuill.R. L. Sin&d, D. A. V. Peters.