FIRAZYR (icatibant)
1
outline
Background on Hereditary Angioedema (HAE) + existing therapies
Rationale for new drug + mechanism of action
Source of lead
Summary of optimization + metabolic stability
Testing funnel
Pharmacokinetics
2
Hereditary angio-edema
angio- vessel
edema- swelling
Acute inflammatory attacks
anywhere
GI tract- hypotension
Larynx- asphyxiation
30% of deaths
Cause: deficiency of functional C1 esterase inhibitor
N Engl J Med 2008;359:1027-36.
3
So what does that long word mean? Well if we break it down to greek we can ease it up to us.
Angio edema vessel swelling
You can see a patient experiencing a mild angioedema attack in his left hand.
Hereditary angioedema is a disease characterized by attacks of inflammation ranging from moderate discomfort to asphyxiation when it occurs in your larynx.
Attacks begin with a tingling sensation. Swelling worsense over the first 24 hours, then gradually subsides over the next few days.
Attacks can occur anywhere in your body and can cause nausea, pain, vomiting, etc.
Can cause severe illness in the GI tract or larynx.
The cause of this disease is a genetic defeciency in a plasma protein known as C1 inhibitor
Now I would like to shift into talking about the biological mechanism of disease.
3
Inflammatory Mechanisms
N Engl J Med 2008;359:1027-36.
Contact activation pathway
Bradykinin-> B2 receptor agonist
Biomaterials. 2009 Apr;30(10):1857-69.
4
This is a figure depicting the inflammatory cascade known as the contact activation pathway. Ill come back to this figure a lot, so get used to seeing it. It is termed the contact activation pathway because factor 12 is activated through contact with a particular type of
Talk about contact inhibition pathway: hagemans factor 12 was originally thought to be activated by contact with a negative tissue surface. More recently opinion has shifted as studies have shown it can be activated by other surfaces. Still, this pathway is called the contact activation pathway because it involves autoactivation of factor 12 after factor 12 comes into contact with a surface.
As you can see, the pathway has elements in common with the blood clotting cascade, and begins with an activation of factor 12 from the blood clotting cascade.
The end result here is the production of the inflammatory peptide bradykinin and activation of the B2 receptor, which is a GPCR and follows typical GPCR mediated signaling, which I will cover in the next slide. Note the presence of C1 inhibitor at many of the steps in this cascade and how vital it is to insuring the cascade does not proceed aberrantly. As I mentioned before, lack of operational C1 Inhibitor is the root cause of HAE. The variety of symptoms are caused directly by the final step in this cascade, which involves bradykinin binding to its receptor and the subsequent signaling events. I would like to start by covering some of the signaling events which bradykinin agnoism of B2 receptor causes.
4
Bradykinin and the B2 receptor
B2 receptor agonism
Gq activation
DAG
Prostacyclins
IP3
Ca++/calmodulin
NO
PKC also
JOURNAL OF CELLULAR PHYSIOLOGY 193:275286 (2002)
http://flipper.diff.org/app/pathways/Bradykinin
5
So we begin with bradykinin mediated agonism of the B2 receptor, which causes a change in conformation of receptor and activation of G alpha q subunit. G alpha q is linked with phospholipase C.
Phospholipase C cleaves PIP3 into IP3 and diacylglycerol.
A variety of responses is caused by this cleavage event, including
Calcium is released from intracellular storage in the sarcoplasmic reticulum by the binding of IP3 to its cognate receptor on the SR membrane.
calcium stimulates Nitric oxide synthase. Which produces nitric oxide that can stimulate vasodilation in adjacent smooth muscle cells.
DAG is a substrate for production of inflammatory prostacyclins these molecules can travel to adjacent smooth muscle cells and produce dilation events.
5
C1 Esterase Inhibitor
N Engl J Med 2008;359:1027-36.
6
Now that we know a little bit about the signaling events leading up to an angio-edema attack, I would like to backtrack a little bit and talk about the natural suppressor of the contact activation pathway, the C1 inhibitor.
6
C1 esterase inhibitor
Serpin-class serine protease inhibitor
-Locks substrate covalently
-inhibition by distortion
Two types of HAE:
1. downregulation of C1INH
2. mutated C1INH (loss of function)
Arch Intern Med. 2001;161:2417-2429
Immunobiol., vol. 199, pp. 358-365 (1998)
purple: trypsin blue: antitrypsin PDB: 1EZX
Nature 405, 586-590 (2000)
7
C1 inhibitor is a member of a class of serine protease inhibitors known as serpins. Named for its original role as discovered in the complement pathway. Serpins such as this one covalently inhibit the protease in question by trapping the active serine residue in the acyl-intermediate state. So one thing to consider, is since the inhibitor acts via suicide, the regulation of its concentration is a bit different than if the inhibition were reverisble.
In hereditary angio- edema, tehre are two types of mutations involving the C1 inhibitor.
Downregulation of a functional protein through the promoter region
Loss of function mutations such as truncations or key residue mutations
Its not a surprise that the first medication for acute angio-edema was a purified C1 inhibitor delivered directly to the circulation intravenously. Ill talk about that soon.
7
Treatment options
8
8
Prophylactic treatment options
Stress reduction
Identification of Triggers
17-Alkylated androgens- general immunosuppresants
jauncdice
Peliosis hepatis
Hepatocellular adenoma
Antifibrinolytics- inhibit activation of factor 12
Tranexamic acid
-aminocaproic acid
Fatigue
cramps
Int. J. lmmunopharmac., Vol. 17, No. I I, pp. 857-863, 1995
N Engl J Med 2008;359:1027-36.
Biochimica et Biophysica Acta, 673 (1981) 75--85
Ann. Rev. Immunol. 1988. 6: 595-628
9
Of course, stress being a primary instigator of an angio-edema attack, doctors will recommend that patients seek to reduce at least some of the stress in their lives. In addition, since many patients seem to have certain external triggers, doctors recommend that patients identify these triggers so that prophylactic treatment can be more effective.
17 alpha alkylated androgens are used as a prophylactic treatment option, they provoke a general immunosuppressive response and lead to increased amount of endogenous C1 inhibitor production by the liver. This therapy is pretty successful in preventing attacks, however it is not without side effects. Here I have listed a few of the side effects that can occur from prolongated use of alpha androgens.
Another class of prophylactic drugs are the anti fibrinolytics. These are both mimics of the amino acid lysine, and they serve to block lysine binding sites on plasminogen, preventing it being cleaved by a its activating protease. Antifibrinolytics are able to inhibit the processing of plasminogen into plasmin which is responsible for producing an amount of factor 12 and also activating the complement cascade which is a non-specific immuno inflammatory response.
These drugs can have side effects such as fatigue and cramps. Neither of these drugs are desirable to use in the long term due to their side effects.
Since this condition manifests as attacks, the focus of most drug development program was to find either a preventative measure or a drug that would instantly relieve an attack at the onset. And now I will transition to talking about treatments for acute attacks.
9
Acute Treatments
N Engl J Med 2008;359:1027-36.
C1 Inhibitor
10
The first idea to treat an attack was to give patients exactly what they were missing, C1 inhibitor. C1 inihibitor can be derived from a vareity of sources.
10
C1 Inhibitor and rationale for new drug
Human derived
Cinryze (2008)
Berinert (2009)
Recombinant
Ruconest (2014)
cheaper
Rationale for new drug
Need immediate relief
Timing (home therapy)
Immunogenicity of full proteins
IV
$$$- producing full protein
N Engl J Med 2008;359:1027-36.
TRANSFUSION 2014;54:2566-2573.
Nature Reviews Drug Discovery 7, 801-802 (October 2008)
http://fc01.deviantart.net/fs70/i/2013/177/a/7/scared_bugs_bunny_by_yetioner-d6asv54.png
11
C1 inhibitor can be derived from a variety of sources. the FDA approved drugs, cinryze and berinert, are isolated from human plasma.
There are also recombinant C1 inhibitors available, such as Rhucin. Interestingly enough, rhucin is produced in the milk of transgenic rabbits.
So C1 inhibitor can be given as an acute prophylaxis, is relatively free of side effects, and works great.
So why even develop a new drug?
One of the main issues is that as icatibant was being developed, most of these C1 inhibitors werent approved for at home treatment, leading to significant symptom onset for most patients before the drug could be administered at a clinic.
Since then, I believe all of these drugs are approved for at home use, resolving this problem.
To note, the recombinant C1 inhibitor has been reported to cause an immunogenic response in a low frequency of patients.
But the biggest issue is that the C1 inhibitor has to be given intravenously, which can be a problem in terms of at home dosing and patient compliance. Improper IV injection can be a problem.
In addition, since the c1 inhibitor inhibits a protease cascade and the resulting receptor mediated signaling cascade, immediate relief is not seen.
Also, money. Less of an issue since the recombinant one, but still producing a whole protein is pretty expensive compared to our final peptidomimetic product.
So from here I would like to talk about a few other treatments approved for acute attacks.
11
Acute Treatments
N Engl J Med 2008;359:1027-36.
Ecallantide(2009)
12
So as Ive highlighted here, ecallantide blocks the final step where kallikrein liberates bradykinin peptide from HMWK. In addition, it can also block further activation of hagemans factor and increased pathway transmission.
12
Ecallantide (Kalbitor)
N Engl J Med 2010;363:523-31.
http://www.kalbitor.com/patient/about-kalbitor/how-kalbitor-works.html
http://www.drugdevelopment-technology.com/projects/dyax-corps-kalbitor/images/1-kalibator.jpg
Small protein inhibitor of plasma kallikrein
Approved in 09
Given in 3 subq shots
Identified by phage display
Small segment of endogenous protein inhibitor of factor 10a
Produced recombinantly in yeast
Significant reduction of symptoms in 4 hours
Problems:
Nurse/doctor required to administer
Still need immediate relief
13
So in 2009 the first subcutaneous treatment for HAE attacks was approved. Known as ecallantide or kalbitor, as it acts as an inhibitor of plasma kallikrein, the protease responsible for liberating bradykinin from HMWK.
This treatment is given in 3 subsequent subq shots, and must be given by a doctor or nurse due to potential immunogenicity.
This drug is a small protein identified by phage display which consists of a small segment of an endogenous serpin.
It is given subcutaneously and thus has a distinct advantage over c1 esterase inhibitor. Symptoms are marked to improve in a couple hours.
So again, this is an improvement. But its not ideal. We are still not seeing immediate relief of symptoms, which is absolutely essential in the case of a laryngeal attack.
13
Acute Treatments
N Engl J Med 2008;359:1027-36.
Icatibant
(2011)
14
And that brings us to our drug for this presentation, Icatibant. Icatibant as a receptor antagonist for the bradykinin B2 receptor at a roughly nanomolar ec50 in vitro. It directly quells the exponential signaling events that occur upon agonism of the B2 receptor by BK.
14
icatibant
Approved in 2011 by FDA
Subcutaneous
At home single use injection
Stable for 2 years at 4C
Peptidomimetic
unnatural amino acids
bioisosteres
conformational restriction
Direct B2 antagonist (nM)
immediate relief
symptoms can reoccur
peptidomimetic strategy proves useful:
high bioavailability- immediate relief
fast clearance- few side effects
No appreciable immune response
Compared to full protein precursors
Maurer M, Aberer W, Bouillet L, Caballero T, Fabien V, et al. (2013) Hereditary Angioedema Attacks Resolve Faster and Are Shorter after Early Icatibant Treatment. PLoS ONE 8(2): e53773.
N Engl J Med 2010;363:532-41.
Vascular Health and Risk Management 2010:6 795802
15
So since icatibant directly agonises the B2 receptor, it should result in the most immediate relief out of any of the drugs ive shown before. And it does, with a response time of blank. Icatibant is a single subcutaneous injection at 30mg. This is easy to do at home, and because of the ease of use, patients can quell the attack at early onset.
so here on the top we see the 9-AA peptide bradykinin, and underneath it the 10 AA icatibant. It is a peptido mimetic, mimicking bradykinin, with a few unnatural modifications that sean will go into more detail on later.
I would like to note that the peptidomimetic response is very useful in terms of an acute treatment not only due to the ease of development using bradykinin as lead.
Ideally, we would like to have a compound that is instantly bidning whos effefcts last until the attack Is over, and is then rapidly cleared to avoid side effects. And we see with a subcutaneous peptidomimetic strategy we achieve all of those points. Peptide is highly soluble and has great bioavailability (need percent here), enters systemic distribution and reaches its target quickly. Its potency is on par with the endogenous ligand bradykinin. Finally, since it is peptidomimetic, it is cleared after one pass throughout the circulature, leading to largely decreased side effets. Also, since it is relatively small, there is less of a risk of immune response than those full protein drugs I mentioned earlier.
15
outline
Background on Hereditary Angiodeema (HAE) + existing therapies
Rationale for new drug + mechanism of action
Source of lead
Summary of optimization + metabolic stability
Testing funnel
Pharmacokinetics
16
Source of lead
History of Bradykinin Antagonists
17
Bradykinin Receptors
Bradykinin is an agonist for the GPCRs bradykinin receptor 1 and 2 (BR1 and BR2), which are responsible for controlling pain and inflammation.
BR1 is inducible, and only present in localized areas at a given time
BR2 is responsible for localized vasodilation and increased vascular edema associated with hereditary angioedema.
BR2 is widespread and constitutive
Takano, Masaoki, and Shogo Matsuyama. "Intracellular and Nuclear Bradykinin B2 Receptors."European Journal of Pharmacology732
(2014): 169-72. Web
Longhurst, Hilary J. Management of Acute Attacks of Hereditary Angioedema: Potential Role of Icatibant.Vascular Health and Risk Management6
(2010): 795802. Print.
http://structbio.vanderbilt.edu/sanders/Research_Julia_Ver_1/temp.html
Golias, Ch et al. The Kinin System - Bradykinin: Biological Effects and Clinical Implications. Multiple Role of the Kinin System - Bradykinin.
Hippokratia11.3 (2007): 124128. Print.
18
18
Bradykinin as a lead
Structure expresses high efficacy in tissue culture
Compound is agonist for these effects, an antagonist is necessary to combat these.
Compound is rapidly degraded in vivo, stability is an issue.
19
Summary of optimization
+ metabolic stability
20
Evaluating Bradykinin Activity
The are no crystal structures to evaluate binding conformation of BR2.
Bradykinin analogs were originally evaluated in tissue cultures, leading to optimization based on efficacy, although with variability depending on tissue type.
Agonism is measure by pD2 , or the negative log of the concentration of compound required to induce 50 % of the maximal response in the tissue
Antagonism is measured by pA2, or the negative log of the concentration of antagonist necessary to double the concentration of agonist in order to induce 50 % of the maximal response
Regoli, D., J. Barabe, and Paula H. Stern.Pharmacology of Bradykinin and Related Kinins. Baltimore: Williams & Wilkins, 1980
21
The Alanine Study
Regoli, D., J. Barabe, and Paula H. Stern.Pharmacology of Bradykinin and Related Kinins. Baltimore: Williams & Wilkins, 1980.
Tissue CultureBradykininAla-1Ala-2Ala-3Ala-4Ala-5Ala-6Ala-7Ala-8Ala-9Cat Ileum (pD2)8.475.276.88.375.175.697.926.71900003020
3.47
8510
794
64.6
18600
3890
115
2190
42700
186000
2.29
1070
468
4.57
166
39800
324
692
3980
4170
22
Positions 5 and 8
Regoli, D., J. Barabe, and Paula H. Stern.Pharmacology of Bradykinin and Related Kinins. Baltimore: Williams & Wilkins, 1980.
Tissue CultureCat Ileum (pD2) Rabbit Jugular Vein (pD2) Dog Carotid Artery (pD2) Bradykinin8.478.468.64Position 5Ala5.695.414.4Tyr6.846.217.55Cha8.068.157.92Position 8Ala>4.374.375.4Tyr(Me)8.518.598.64Cha8.478.157.8223
3.39E-09 3.47E-09 2.29E-09 2.04E-06 3.89E-06 3.98E-05 1.45E-07 6.17E-07 2.82E-08 8.71E-09 7.08E-09 1.20E-08 >4.27E-05 4.27E-05 3.98E-06 3.09E-09 2.57E-09 2.29E-09 3.39E-09 7.08E-09 1.51E-08
23
From Agonist to Antagonist and Increasing Potency
Regoli, Domenico, Suzanne Nsa Allogho, Anna Rizzi, and Fernand Junior Gobeil. "Bradykinin Receptors and Their Antagonists."European Journal of Pharmacology348.1 (1998): 1-10.
Vavrek, Raymond J., and John M. Stewart. "Competitive Antagonists of Bradykinin."Peptides6.2 (1985): 161-64.
Rhaleb, N. E., S. Telemaque, N. Rouissi, S. Dion, D. Jukic, G. Drapeau, and D. Regoli. "Structure-activity Studies of Bradykinin and Related Peptides. B2- Receptor Antagonists."Hypertension17.1 (1991): 107-15.
Dunn, Floyd W., and John M. Stewart. "Analogs of Bradykinin Containing P-2-Thienyl- L-alanine1."Journal of Medicinal Chemistry14.9 (1971): 779-81
CompoundRabbit Jugular VeinDog Carotid ArteryBradykinin (pD2)8.468.64[D-Phe7]-BK (pA2)Residual Agonism6.25[Hyp3, D-Phe7]-BK (pA2)Residual Agonism7D-Arg[Hyp3,D-Phe7]-BK (pA2)8.017.93[Thi5,8,D-Phe7]-BK (pA2)6.76.55[Hyp3,Thi5,8,D-Phe7]-BK (pA2)6.967.01D-Arg[Thi5,8,D-Phe7]-BK (pA2)7.867.86Antagonism!!
24
Activity of analogs compared wit,h bradykinin, based on the dose required to cause a half-maximal isotonic contraction. Dose
(in pg) required to effect a depressor response of 20-25 mm of Hg by the intraaortic (ia) and iv routes. Percentage of an iv dose destroyed
by a single passage through the pulmonary circulation in the rat.
3.47E-09 2.29E-09 1.00E+00 5.62E-07 1.00E+00 1.00E-07 9.77E-09 1.17E-08 2.00E-07 2.82E-07 1.10E-07 9.77E-08 1.38E-08 1.38E-08
24
To Icatibant
Henke, Stephan, Hiristo Anagnostopulos, Gerhard Breipohl, Jochen Knolle, Jens Stechl, Bernward Scholkens, Hans-Wolfram Fehlhaber,
Hermann Gerhards, and Franz Hock. Peptides Having Bradykinin Antagonist Action. Hoechst Aktiengesellschaft., assignee. Patent 5,648,333.
15 July 1997. Print.
CompoundGuinea Pig Pulmonary Artery (pD2) DArg[Hyp3-DTic7]-BK4.92DArg[Hyp3-DTic7-Oic8]-BK8.16DArg[Hyp3-Thi5-DTic7-Oic8]-BK8.27DArg[Hyp3-Thi5-DPhe7-Oic8]-BK7.8525
1.20E-05 6.92E-09 5.37E-09 1.41E-08
25
Metabolic Stability of Bradykinin
Sidorowicz, W., J. Szechinski, P. C. Canizaro, and F. J. Behal. "Cleavage of the ARG-PRO Bond of Bradykinin by a Human Lung Peptidase: Isolation, Characterization, and Inhibition by Several -Lactam Antibiotics." Experimental Biology and Medicine 175.4 (1984): 503-09.
Kaplan, A. P. (2008) Bradykinin Pathways and Allergic Disease, in Allergy and Allergic Diseases, Volume 1, Second Edition (eds A. B. Kay, A. P. Kaplan, J. Bousquet and P. G. Holt), Wiley-Blackwell, Oxford, UK. doi:10.1002/9781444300918.ch20
Regoli, Domenico, Suzanne Nsa Allogho, Anna Rizzi, and Fernand Junior Gobeil. "Bradykinin Receptors and Their Antagonists."European Journal of Pharmacology348.1 (1998): 1-10.
Bradykinin destruction during
first pass through vasculature: 98%
26
Icatibant Antagonist Stability
T1/2=1.2-1.5 hrs
Ghazi, Aasia, and J Andrew Grant. Hereditary Angioedema: Epidemiology, Management, and Role of Icatibant.Biologics: Targets & Therapy7
(2013): 103113.PMC. Web. 17 Apr. 2015.
Kaplan, A. P. (2008) Bradykinin Pathways and Allergic Disease, in Allergy and Allergic Diseases, Volume 1, Second Edition (eds A. B. Kay, A. P.
Kaplan, J. Bousquet and P. G. Holt), Wiley-Blackwell,
Oxford, UK. doi:10.1002/9781444300918.ch20
27
27
Bradykinin and Icatibant
28
outline
Background on Hereditary Angioedema (HAE) + existing therapies
Rationale for new drug + mechanism of action
Source of lead
Summary of optimization + metabolic stability
Testing funnel
Pharmacokinetics
29
Testing funnel
Biochemical and animal models of inflammation
30
In Vitro Studies
Receptor binding
Downstream effects
Bradykinin induced releases of:
EDRF
Guanylate cyclase activation
32P quantification
Ca2+
indo-1 Fluorescent probe
Prostacyclin
radioimmunoassay
31
Hock, et al.Br. J. Pharmacol. 1991,102, 769
Radioligand competition binding assays in:
Guinea-pig ileum (crude membrane)
Rat uterus
Guinea-pig pulmonary artery
Rabbit aorta
Receptor binding: essentially a scintillation proximity assay using tritiated compound. In brief, crude membranes from guinea-pig ileum were incubated for 1 h at 25C in a buffer solution with [3H]-BK (1 nmolI 1-) and various concentrations of the test compounds (11 concentrations; range: 1 x 10' to 1 x 10-10moll-'). Non specific binding was determined in the presence of 1 ,umol 1-unlabelled BK.
Bradykinin induced releases: all cell based assays. Calcium and EDRF (endothelium derived relaxing factor) in endothelial pig aorta. EDRF itself is not stable, but is known to active guanylate cyclase. So cyclic GMP is a good indicator of this activation. Of note, in the 90s it was pretty well accepted that EDRF was the free radical NO (nitric oxide). Prostacyclin from endothelial bovine aorta. About EDFRs, now it is accepted that there are a few and this is a class of factors that include NO, endothelin-1, prostaglandin H2, and endothelium derived hyperpolarizing factor.
31
32
Inhibitory effect based on Ca2+ release
Summary of radioligand binding results
Hock, et al.Br. J. Pharmacol. 1991,102, 769
Hoe 140s IC50 is 3x lower in each model
Both compounds exhibit agonist activity at low concentrations (20 M)
Hoe 140 shown to block downstream effect of increased free Ca2+
As Sean discussed earlier, there was a BK antagonist developed in the mid 80s that the Hoest company was using for their lead. In a pair of papers published in 1991, they reported their best antagonist, Hoe 140, as compared to this 80s antagonist in all of the testing.
And as Sumit showed us, BK binding to the receptor activates a g protein, and then activates phospholipase C invokes an enhanced production of inositol tris (1,4,5) phosphate increases free calcium.
32
EDRF release
prostacyclin release
33
Hock, et al.Br. J. Pharmacol. 1991,102, 769
Endothelium- derived relaxing factor: measured as the formation of cyclic GMP. This was a plate assay coupled with the calcium probe shown previously. Briefly, 24 well plates of pig aorta endothelial cells were incubated with drugs and superoxide dismutase (SOD is an anti-inflammatory agent, the enzyme reacts with reactive oxygen species) and then dosed with BK to induce inflammation. Supernatant was removed at certain time points and the cGMP assay was performed. The left graph shows the time points comparing control (vehicle) to differing concentrations of Hoe140. Starting with 0.1 nM going to 1 uM.
PGF assay: RIGHT. C is baseline. Log scale X axis is concentrations of BK. Open circles are Hoe140, open triangles are 1980s BK antagonist. Solid circles are BK alone. *= significance to baseline. They had a radioimmunoassay for the hydrolysis product of PGI2, = 6-keto PGF 1 alpha. So it is actually kind of the same assay as EDRF release, except in bovine aorta.
33
In vivo studies
Comparison of BK antagonists
Duration of action
Anesthetized rats
BK induced bronchoconstriction
anesthetized guinea-pig
Standard Inflammation Model
Carrageenin-induced rat paw oedema
34
Wirth, et al. Br. J. Pharmacol. 1991, 102, 774
Winter, et al. Exp. Biol. Med. 1962, 111, 544
Goal of these studies was to test the efficacy of Hoe140 in vivo in models where BK served as an agonist For this purpose, models of BK-induced hypotensive reaction and BK-induced bronchoconstriction were used to demonstrate its high potency and long duration of action.
Comparison: 1985 class of antagonists synthesized by Vavrek & Stewart 1985, D-Arg-[Hyp2, Thi5,8, D-Phe7] BK for the blood pressure experiments
Carrageenin: class of natural products from seaweds, linear sulfated polysacharides.
34
Duration of action
Effect on BK-induced bronchoconstriction
35
Wirth, et al. Br. J. Pharmacol. 1991, 102, 774
Right: systemic blood pressure was measured in anesthetized rats; BK was given as an intra-arterial bolus injection 3 times. Subcutaneous administration of the antagonist (Hoe140). Control was vehicle. Shown in the figure is time versus %change in mean arterial pressure (MAP). Solid columns are Hoe140, open= vehicle. You see an immediate change in MAP at 1 hour, with it not quite returning at 5 hours.
Left: Effect of bronchoconstriction in guinea-pigs shows dose dependence of the antagonist. RL (top)= pulmonary resistance. Cdyn (bottom)=dynamic compliance. BK-induced bronchoconstriction normally does two things compared to baseline, it increases RL 42-58% and decreases Cdyn 33-39%. This graph shows the effect of increasing concentrations of Hoe140. Open columns are before the i.v infusion of Hoe140, solid columns are after.
35
36
Wirth, et al. Br. J. Pharmacol. 1991, 102, 774
So, this study is exactly what you think it is. They initiate inflammation in a rats paw, drain it and measure the volume.
Note about Patent life: two patents are most easily tied to this drug, the original 1995 one from Hoescht Aktiengesellscaft. Initially they report the same in vitro type radioligand binding data from the 1991 papers. There is a 2014 paper from Shire, which includes more updated experiments like knockout mice, and null mice in addition to the radioligand binding assay. Because if it aint broke, dont fix it.
36
Pharmacokinetics
+ final product formulation
37
Icatibant
38
PropertiesLog P-2.3# Hydrogen Acceptors23# Hydrogen Donors15MW1304.522 DaIcatibant (DB06196) [online]. Available from URL:
http://www.drugbank.ca/drugs/DB06196#admet
Not going to be orally bioavailable, but who cares! As you can see, this is a huge drug and breaks the rule of 5 in multiple ways. First of all, it is a very hydrophilic decapeptide. The logP is -2.3, so incredibly soluble, but membrane permeability is a huge issue! Other faults in the rule of 5 are its many hydrogen bond donors and acceptors, and huge molecular weight. The polarity is also a big problem for permeability. When this drug was tested using the Caco-2 cell layer assay, it was found to not be cell permeable. However, the target is in the blood so it really didnt matter. There might be some reabsorption in the kidneys by OATP1, (it was found to not be an inhibitor).
If they had cared about permeability, they should have increased LogP, decreased ionizable groups, decreased size, and decreased polarity.
38
Icatibant acetate (active) + NaCl, glacial acetic acid, NaOH, H20
Available as a 3mL injection (10 mg/ml) subcutaneously
Requires no mixing or measuring
39
Formulation
Firazyr (Icatibant Injection) Prescribing information [online]. Available from URL:
http://pi.shirecontent.com/PI/PDFs/Firazyr_USA_ENG.pdf
PharmacokineticS
Metabolism/ elimination
Proteolytic enzymes (2 inactive metabolites)
CYPs: non-substrate, non-inhibitor
Elimination: urine (
Top Related