icatibant .pdf

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

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

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

C1 ESTERASE INHIBITOR

N Engl J Med 2008;359:1027-36.

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

TREATMENT OPTIONS

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

ACUTE TREATMENTS

N Engl J Med 2008;359:1027-36.

C1 Inhibitor

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

ACUTE TREATMENTS

N Engl J Med 2008;359:1027-36.

Ecallantide (2009)

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

ACUTE TREATMENTS

N Engl J Med 2008;359:1027-36.

Icatibant (2011)

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

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

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

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

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 Culture

Bradykinin Ala-1 Ala-2 Ala-3 Ala-4 Ala-5 Ala-6 Ala-7 Ala-8 Ala-9

Cat Ileum (pD2)

8.47 5.27 6.8 8.37 5.17 5.69 7.92 6.7

POSITIONS 5 AND 8

Regoli, D., J. Barabe, and Paula H. Stern.Pharmacology of Bradykinin and Related Kinins. Baltimore: Williams & Wilkins, 1980.

Tissue Culture Cat Ileum (pD2)

Rabbit Jugular Vein (pD2)

Dog Carotid Artery (pD2)

Bradykinin 8.47 8.46 8.64

Position 5 Ala 5.69 5.41 4.4

Tyr 6.84 6.21 7.55

Cha 8.06 8.15 7.92

Position 8 Ala >4.37 4.37 5.4

Tyr(Me) 8.51 8.59 8.64

Cha 8.47 8.15 7.82 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

Compound Rabbit Jugular Vein Dog Carotid Artery

Bradykinin (pD2) 8.46 8.64 [D-Phe7]-BK (pA2) Residual Agonism 6.25 [Hyp3, D-Phe7]-BK (pA2) Residual Agonism 7

D-Arg[Hyp3,D-Phe7]-BK (pA2) 8.01 7.93

[Thi5,8,D-Phe7]-BK (pA2) 6.7 6.55

[Hyp3,Thi5,8,D-Phe7]-BK (pA2) 6.96 7.01

D-Arg[Thi5,8,D-Phe7]-BK (pA2) 7.86 7.86

Antagonism!!

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.

Compound Guinea Pig Pulmonary Artery (pD2) DArg[Hyp3-DTic7]-BK 4.92 DArg[Hyp3-DTic7-Oic8]-BK 8.16 DArg[Hyp3-Thi5-DTic7-Oic8]-BK 8.27 DArg[Hyp3-Thi5-DPhe7-Oic8]-BK 7.85

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

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

IN VITRO STUDIES

R E C E P T O R B I N D I N G D O W N S T R E A M E F F E C T S

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

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+

E D R F R E L E A S E P R O S TA C Y C L I N R E L E A S E

33 Hock, et al.Br. J. Pharmacol. 1991,102, 769

IN VIVO STUDIES C O M P A R I S O N O F B K A N T A G O N I S T S

Duration of action Anesthetized rats

BK induced bronchoconstriction

anesthetized guinea-pig

S T A N D A R D I N F L A M M A T I O N M O D E L

Carrageenin-induced rat paw oedema

34 Wirth, et al. Br. J. Pharmacol. 1991, 102, 774 Winter, et al. Exp. Biol. Med. 1962, 111, 544

D U R A T I O N O F A C T I O N E F F E C T O N B K - I N D U C E D B R O N C H O C O N S T R I C T I O N

35 Wirth, et al. Br. J. Pharmacol. 1991, 102, 774

36 Wirth, et al. Br. J. Pharmacol. 1991, 102, 774

ICATIBANT

38

Properties

Log P -2.3

# Hydrogen Acceptors 23

# Hydrogen Donors 15

MW 1304.522 Da

Icatibant (DB06196) [online]. Available from URL: http://www.drugbank.ca/drugs/DB06196#admet

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

M E T A B O L I S M / E L I M I N A T I O N

Proteolytic enzymes (2 inactive metabolites)

CYPs: non-substrate, non-inhibitor

Elimination: urine (

Guinea pig ileum Rat uterus Guinea pig

pulmonary artery Rabbit aorta BK1-receptor (Regoli &

Barabe, 1980)

pig aorta endothelial cells

Bovine aorta endothelial cells

42

ANIMAL TISSUES- AN EXPLANATION

43

FURTHER WORK IN DRUG DISCOVERY WITH BK ANTAGONISTS

Huang, et al. J. Med. Chem. 2010, 53, 53835399

INDO-1 FLUORESCENCE PROBE

44 Grynkiewicz, G. et al. J. Biol. Chem. 1985, 260 (6), 3440-3450.

CLINICAL TRIALS- PHASE III

F A S T - 1 A N D F A S T - 2

Single dose not significantly faster than placebo

More rapid symptom relief than oral (tranexamic acid)

No rescue therapy required of icatibant patients

45 Deeks, E.D. Drugs, 2010, 70 (1), 73-81

SMALL MOLECULE B2 ANTAGONIST DEVELOPMENT

Oral drug usually desired

Clinical trials

Easier dosing

Could be prophylactic measures if patient has prior knowledge of oncoming trigger

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