VOLUME 90 NUMBER 4, PART 2

Second-generattan antii:istamrnes

[amine to immediate bronchoconstriction proved by inhaled allergen and adenosine 5’ monophosphate in asthma. Am Rev Respir Dis 1987;136:369-73.

IO. Togias AG, Proud D, Kagey-Sobotka A, Freidhoff L. The in viva and in vitro effect of antihistamines on mast cell mediator release: a potentially important property in the treatment of allergic disease. Ann Allergy 1989;63:467-9.

1 I. Charlesworth EN, Kagey-Sobotka A, Norman PS, Lichtenstein I.. Effect of cetirizine on mast cell mediator release and cellular traffic during the cutaneous late-phase reaction. J ALLERGY CLIN IMMUNOI. 1989;83:905-12.

12. Townley RG, Okada C, Miyagawa H. The effect of cetirizine on eosinophil chemotaxis and release of eosinophil peroxidase

in human allergic subjects [Abstract]. J ~ILI:NC~Y &‘I 11 Ib4.- MUNOL 1991;87:281A.

13. Fade1 R, David B, Herpin-Richard N, Borgnon ~1. Rassemont R, Rihoux JP. In vivo effects of cetirizine on cutaneous re~c- tivity and eosinophil migration induced by platelet acttvating factor (PAF-acether) in man. J ~\LI.ERGY c‘i IX. i2AMI’IIOl

1990;86:314-20. 14. Hennocq E, Vargaftig BB. Accumulation of eoGnophi!\ tn

response to intracutaneous PAF-acether in man [Letrcrl. Lancct 1988;1:1378.

15. Sanjar S, Colditz 1. Pharmacological modulation ol eosinophil accumulation in guinea pig airway. In: Eosinophili in asthma. San Diego: Academic Press 1989:201- 12

Commentary and discussion: Inhibition of mediator release

DETERfVUNWG AN ANTIHISTAMINE’S MAST CELL-STASILIZtNG PROPERTY

Dr. Simons. The central question that we hope to address in this discussion is, “What is the clinical importance of the inhibition of mediator release by antihistamines?” What other key questions does the panel think we should address?

Dr. Kaliner. Another important question is whether H, antihistamines have a mast cell-stabilizing prop- erty. There is no doubt that the primary measurable event that occurs in rhinitis after mast cells are acti- vated is vascular permeability. Because vascular per- meability is one of the major physiologic responses to mast cell degranulation and is largely--albeit not exclusively -mediated by histamine, a good antihis- tamine will control that response.

Dr. Simons. The most potent antihistamines can almost abolish it completely.

Dr. Kaliner. Yes, depending on the dose and all of the other parameters involved. My question is then, how do you interpret the mast cell-stabilizing capac- ity of an agent if the primary event that agent affects is the event that allows you to measure the mast cell mediators?

If you review all of the data on mediator release, it can be confusing. Some studies show reductions in leukotrienes but not in prostaglandin D, or histamine; others show reductions in histamine but not in pros- taglandin D, or leukotrienes; and others show consis- tent reductions in vascular permeability but not ex- clusive or total reductions.

Dr. Simons. The results vary even with the same drug. Depending on dose, duration of treatment, and end-organ studied, levels of mediators may or may not be decreased.

110/40678

Dr. Kaliner. That is right depending on the study. but one thing is still obvious-that all effective anti- histamines clearly block the primary physiologic event. However, after hearing all the data presented, I was less than positive about the mast cell--stabilizing property of these agents. Dr. Naclerio. your data in nasal lavage studies are the best, no question about it. However, even in your studies, when we looked at histamine reductions, we were talking about reduc- tions from 4.5 to 3.5 or 2.5 rig/ml. The data from our laboratory show the same reduction. I am not certain that the difference between 4.5 and 2 .4 ngi ml of nasal washings is a significant change

My point is that we all acknowledge the fenibilities in these measurements. The whole concept of mast cell stabilization is based on them; yet they seem somewhat contradictory from study to study, and they are small changes in a very flexible parameter. Based on that type of evidence, how can we conclude whether terfenadine or any of the H! antihistamines is or is not a mast cell stabilizer’?

Dr. Naclerio. To try to answer your question. I will stay with the data from my laboratory. One of the factors that has always been important in inter- preting these studies is the pattern of response. You also look at the patterns of mediators. Why does one go up and another go down? That is where the strengths are in the lavage studies. In our studies 01 histamine release, I believed the data the first time we did the study. When we repeated the study in another group of subjects, we found the same effects in histamine release. Even though that decrease may be small, it is statistically significant and re- producible.

There is another important question we must an- swer-the question that Dr. Simons already raised: Does the statistical change in the model, which I he- lieve is true. translate into clinical benefit’! I do not

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726 Commentary and discussion J ALLERGY CLIN IMMUNOL OCTOBER 1992

know the answer. Our studies clearly show a reduction in histamine, but I do not know whether a 25% re- duction in histamine release translates into clinical benefit or a physiologic effect.

Dr. Kaliner. Let us divorce the clinical issue from our discussion for the moment. What I am asking about is the mechanism of action. If you have a 25% reduction in histamine in the nasal lavage, couldn’t that just reflect that less fluid is leaked by blood vessels and therefore less volume is coming into the lumen‘? Wouldn’t that give you the same result?

Dr. Naclerio. You have raised a very important question: How are these mediators moved to nasal secretions? I think that if vascular leakage was the entire explanation, then every H, antihistamine should have those effects. In my presentation I summarized our studies of cetirizine, which did not reduce hista- mine. We have also looked at diphenhydramine, loratadine, azelastine, and ketotifen, and they clearly did not decrease histamine release.

Dr. Kaliner. Some of them had an effect on his- tamine release.

Dr. Naclerio. They did not have any significant effect.

Dr. Simons. Cetirizine, which is in fact a very potent H, blocker, did not decrease the release of histamine significantly in the nasal challenge model.

Dr. Naclerio. That is right. The H, antihistaminic effect of terfenadine versus the H, antihistaminic ef- fect of the other antihistamines we have studied was not different. From a theoretic point of view, I believe that fluid movements could be an explanation. That is why the disk model we have just started to use may be more illuminating. With the disk model we can look at both the volume of the secretion and the con- centration of histamine and whether it changes in nasal secretions. Our preliminary results suggest that many fluids contributing to nasal secretions do not contain histamine and tend to dilute the concentration of his- tamine in nasal secretions. Therefore we believe the total amount of histamine recovered is the best re- flection of changes in a mediator.

Dr. Townley. It seems to me that the best way to answer this question would be to determine whether terfenadine or other H, agents inhibit histamine release in nasal mast cells or perhaps basophils in an in vitro system.

Dr. Kaliner. We tried that by challenging nasal turbinates in vitro. There were two problems. First, compared with lung tissue, nasal mucosa has only one tenth the number of mast cells. So your parameters are reduced initially by 90%. Second, you do not obtain much mediator release from nasal mucosa in vitro. A very small amount of histamine escapes from

the turbinate into the supematant in vitro; it is not like in vitro studies with lung tissue, which is extremely thin. Unless blood flow from vascular permeability washes the histamine out, very little histamine is re- leased.

If you look back at the work we did several years ago, you will see that the histamine release is ex- pressed in percent increase above control; it ranged from 15% to 30% above control. With lung tissue, histamine release would be 1000% above control. Thus lung and nasal mucosa are very different tissues.

Dr. Townley. Do we interpret that to mean that the nose is just much more sensitive to histamine than the lungs, and so it takes very little histamine-just a few mast cells-to provide that histamine?

Dr. Kaliner. No, I think actually what those results say is that less histamine is found in the nose, and therefore antihistamines can be more effective in rhi- nitis. Presumably, if the lungs had the same concen- tration of histamine, antihistamines might work much better in patients with asthma.

At any rate, there is much more histamine in the lung than in the nose, and therefore it is much harder to antagonize lung histamine than nasal histamine. I think that difference has a big impact on clinical re- sponses .

Let me ask the panel another question: Based on the information as you know it, would you conclude that one of the actions of terfenadine is to reduce mast

cell mediator release? Dr. Simons. Looking very carefully at the in vitro

evidence from Dr. Townley’s laboratory and Dr. Tas- aka’s group and at the in vivo evidence published by Dr. Naclerio and presented here, I have no doubt that terfenadine has an antiallergic effect. However, the relative contribution of this effect to the overall effi- cacy of terfenadine, which is a good functional an- tagonist, is unknown.

I have no trouble accepting the best of the studies that we reviewed. For the first time we have repro- ducible results in an in vivo human model, and the studies are placebo controlled. Interpretation of the results is much more difficult. Because we agree that the best of these new antihistamines are excellent func- tional antagonists, do we need any additional reason to use them? In other words, because these drugs work very well on the basis of H, blockade, do we need to invoke any further mechanisms, or are we just going through an intellectual exercise? I hasten to add that if it is only an intellectual exercise, it is worthwhile because we are learning much about the mechanisms involved. However, we may not need to invoke this mechanism to explain why the drugs are effective in the treatment of allergic rhinitis and urticaria.

VOLUME 90 NUMBER 4. PART 2

Commentary and discussion 727

CLINICAL SlGIIllFlCANCE OF INHBITION OF HtSTAMfNE RELEASE

Dr. Naclerio. If one does accept the statistical re- duction in histamine release as demonstrated in Dr. Townley’s laboratory and in our laboratory, the im- portant issue that remains is whether that reduction has any clinical significance.

Clinicians want to know whether we can demon- strate in seasonal trials the difference between one H, antihistamine and another. However, to do that we would need better clinical measures, because the large number of subjects that would be needed to show that cetirizine and terfenadine, for example, are not dif- ferent according to symptom diaries almost precludes doing the study.

Dr. Simons. You are right. Terfenadine and astem- izole. the oldest of the “new” H, antagonists, have been most frequently compared “head to head” in the treatment of these two disease entities. Additional “head-to-head” comparative studies of the newer H, antagonists are needed in the treatment of allergic rhinitis and urticaria.

We did a comprehensive “head-to-head” compari- son of five H, antihistamines in a single-dose study and looked only at reduction of histamine-induced wheal-and-flare reactions in the skin. We found that 10 mg of cetirizine and 120 mg of terfenadine were significantly more potent than the other H, antihista- mines we studied.

Dr. Naclerio. Again that study involves a model. The question is, is that difference clinically beneficial to patients? I do not think we can demonstrate a clin- ical difference yet.

Dr. Simons. Certainly it is difficult to compare the allergic rhinitis efficacy studies performed by different investigators in different countries with different H, antagonists in different pollen seasons. However, in- vestigators who have performed many of these studies offer the perspective that these medications are fairly similar in efficacy.

Dr. Townley, what is your position on the clinical importance of the antiallergic effects demonstrated in current studies?

Dr. Townley. The quick answer is that I do not believe that we know. It is clear that these agents are potent antihistamines, but 1 agree that it is difficult to assess the clinical relevance of their antiallergic effects

Dr. Simons. Do you believe the existing literature supports the concept that the antiallergic effect is true in vivo as well as in vitro‘?

Dr. Townley. Yes. The data, I think, are unequiv- ocal. Again. it is the interpretation of the data that is still somewhat uncertain.

@AGONISTS AS MAST CELL STAB-ILIZERS

Dr. Naclerio. I would like to pose a question to Dr. Kaliner. It has been proposed that giving a p- agonist that would block mast cell activatitm in the lung would be harmful because it would ailow pro- liferation of late allergic events, perhaps through re- lease of cytokines. Thus if we had a good mast cell stabilizer that was not an antihistamine. wcruici it he

good or bad for clinical rhinitis? Dr. Kaliner. I believe cromolyn sodium :s an ex-

ample that shows that it would be good clinically. Dr. Naclerio. I agree that cromolyn has efficacy in

patients with allergic rhinitis, but I do not believe that there is good evidence that it is a mast cell stabilizer. It may have other effects that make it clinically useful

Dr. Townley. I believe that the p-agonists are clearly very potent mast cell stabilizers and ?/et seem to be notoriously ineffective in the nose.

Dr. Kaliner. I know of only one study thaf showed efficacy for a P-agonist and that involved fi:noteml.

Given nasally, it was a potent inhibitor of mast cell degranulation. It stopped the symptoms at allergen challenge, but the patients were uncomfortable be- cause of nasal congestion caused by drug-induced va- sodilation. What you are saying is that oral P-agonist& do not have much of an effect in the nose, but that question has not really been studied car&tll::.

Dr. Simons. P-Agonists are another important class of drugs with excellent functional antagomm. and yet, especially for the new long-acting medications in this class, formoterol and salmeterol, much effort is going into defining antiinflammatory effects that ma! or may not be relevant clinically.

Dr. Townley. We have evidence that iormoterol inhibits the late reaction and strikingly inhibits eosin- ophilia in the late reaction in studies done in thr: guinea pig lung.

A HIERARCHY OF MEDiATORS OF THE ALLERGIC REACTION

Dr. Simons. Dr. Kaliner, which of the !,eleasecl mediators have the greatest clinical relevance?

Dr. Kaliner. If you had to outline a hierarchy for the acute allergic reaction, histamine would certainly be at the top based largely on the capacity of antihis- tamines to reduce histamine-mediated symptoms. Bm- dykinin is an extremely important inducer of vascular permeability in every model and also contributes to the acute-phase reaction, as do the eicosanolds, pros- taglandins, and leukotrienes.

Dr. Townley suggested that platelet-activating fat:- tor is important, but my own reading of that literature and my own experiments lead me to disagree. I believe that platelet-activating factor is an extremeI! parent

728 Commentary and discussion J ALLERGY CLIN IMMUNOL OCTOBER 1992

molecule in search of a disease and that rhinitis is not its home. However, there is much debate about that.

Dr. Simons. Are you willing to be quoted on that? Dr. Kaliner. I have been many times over the

years. I have always been skeptical that an agent that has a 20 nsec half-life will have any activity in vivo. It seems to me that most of it is not released from the membrane but is reincorporated back into membrane phospholipid. I have also always been skeptical about the pharmacologic data on platelet-activating factor because it involves up to 1 million times the concen- tration that can ever be recovered in vivo.

Dr. Simons. So for medications such as azelastine and ketotifen, which have broad antimediator effects as well as an antiallergy effect, would you say the antihistaminic effect is much more important than any effect on platelet-activating factor or other mediators?

Dr. Kaliner. Yes. Dr. Townley. I am not quite as ready as Dr. Kaliner

to bury platelet-activating factor. The failure of a platelet-activating factor antagonist to block the late reaction certainly put a few nails in the coffin but not quite enough to close it completely. It is true that it is very difficult to measure platelet-activating factor because of its brief half-life, but it still deserves some attention because of its ability to induce eosinophils in the skin and lung of a variety of species.

Dr. Kaliner. The second part of your question, Dr. Simons, is what mediators are important in the late- phase reaction, and that is where there are many un- knowns. It seems clear that certain mast cell mediators including the inflammatory factors and chemotactic factors initiate the late-phase process. Other molecules that have a long-lasting activity are probably involved, including some of the enzymes as well as mast cell- derived inflammatory cytokines .

What first happens to tissues affected by late-phase reactions is infiltration by neutrophils. Our data in a rat model show that neutrophils are essential for fur- ther propagation of the reaction. By releasing lyso- somes, neutrophils contribute to the sequential acti- vation of the late-phase reaction.

As neutrophils infiltrate tissues, inflammatory cy- tokines are released from mast cells and presumably activate lymphocytes. There are approximately 10 times the number of CDCtype lymphocytes in the mu- cosa as there are mast cells. When these lymphocytes become activated, they generate a number of inflam- matory mediators, much the same as mast cells do.

These events happen 2 to 4 hours after the acute reaction. After 8 to 24 hours, eosinophil migration begins in earnest. At that point an increase also occurs in the number of mononuclear cells and in prolifer- ation of mast cells.

Nobody knows the potential contribution of mac- rophages, which could also be activated. Nobody has studied epithelial cells, which are a potent source of several inflammatory mediators. Endothelial cells have not been examined at all. We know that they are the target for the immediate reaction and presumably have been involved in late reactions as well. Defining the roles of these cells is going to keep many people busy for a decade.

The bottom line is that the mast cell initiates the process. If you inhibit mast cell reactions, you inhibit both the immediate and late reactions.

Dr. Townley. You indicated that mast cells may actually be more important in cytokine production than helper T cells and that mast cells seem to elicit essentially the whole cascade of cytokines. What is the evidence for that?

Dr. Kaliner. If you culture human or mouse mast

cells and compare the quantity of interleukins 3 and 4 produced after activation to the amount produced by an equivalent number of lymphocytes, you find that a mast cell produces much more interleukin4 per cell. As a matter of fact, the only preformed inter- leukin4 found in tissue is in mast cells.

Dr. Naclerio. You stated that histamine is the most important mediator of the immediate allergic reaction. Why do you think you have been unable to measure it in your model?

Dr. Kaliner. I think the answer is that when a decongestant is not used in a nasal lavage model, the histamine released by mediators becomes diluted by the influx of vascular fluid and proteins. We have measured the increase in plasma proteins that come into nasal secretions over the lo-minute period after allergen challenge, and the proteins actually increase 125fold from baseline. Even if all of the histamine were released from all of the mast cells in the nasal mucosa, there would still be only a small amount of histamine. That amount would be diluted enormously by this 125fold increase in plasma protein and fluid that comes into the interstitium.

The issue is not why we cannot measure it, but how anyone can measure it without first reducing vas- cular permeability with a decongestant such as pseu- doephedrine. In a still unpublished study, we chal- lenged patients after pseudoephedrine was adminis- tered, and we observed an increase in histamine levels that we did not see when pseudoephedrine is not used. Our conclusion is that in our model, unless you affect vascular kinetics, there is no reason to anticipate seeing an increase in histamine. However, that does not mean that histamine is not being released. It just means that it is not coming out in a measurable amount in the lavage fluid. I am positive that histamine is released.