With an IC50 value of 474.5 pM, zilucoplan (RA101495; 1-1000 nM; 30 min) inhibits the increase in C5a plasma levels in human whole blood caused by lipopolysaccharides. When zilucoplan is used at a concentration of 1 nM, C5a plasma levels are reduced by 65.7% [2]. By preventing C5 from being broken down by C5 convertase into C5a and C5b and attaching to preformed C5b to sterically block interaction with C6, zilucoplan binds to complement component 5 (C5) and prevents the downstream assembly of the membrane attack complex (MAC; C5b-9)[1].

In C5-deficient mice treated with human complement, zilucoplan (RA101495; 10 mg/kg; SC; daily, for 6 d) inhibits the development of immune-mediated necrotizing myopathy (IMNM)[1]. In C57BL/6 mice, zilucoplan (10 mg/kg; SC; daily, for 6 d) shows protective effects on the prevention of myopathy[1].

Animal/Disease Models: C57BL/10SnJ C5-deficient (C5def) mice with anti-HMGCR+ IMNM IgG xenografts[1]
Doses: 10 mg/kg
Route of Administration: subcutaneous (sc) injection; daily, for 6 days
Experimental Results: Prevented muscle strength loss in C5def mice with less complement deposition on myofibres and consequently less necrosis/regeneration.

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Animal/Disease Models: C57BL/6 mice with anti-HMGCR+ IMNM IgG xenografts[1]
Doses: 10 mg/kg
Route of Administration: subcutaneous (sc) injection; daily, for 6 days
Experimental Results: Prevented muscle weakness and decreased regenerated myofibres. diminished necrotic cells as well as regenerating cells expressing foetal myosin.

Absorption, Distribution and Excretion
Following single and multiple daily subcutaneous administration of 0.3 mg/kg zilucoplan, time to reach peak plasma concentrations (Tmax) ranged from three to six hours. Following daily subcutaneous dosing of 0.3 mg/kg zilucoplan for 14 days in healthy subjects, both the peak plasma concentration and exposure (AUCtau) increased by approximately 3-fold. After daily repeated subcutaneous administration of 0.3 mg/kg zilucoplan, plasma concentrations of zilucoplan were consistent, with steady state trough concentrations being reached by four weeks of treatment with zilucoplan through 12 weeks.
Less than 1% of zilucoplan and its metabolites are excreted in urine and feces.
The mean volume of distribution at steady state was 3.51 L in the population pharmacokinetics analysis for adult patients with gMG.
No information is available.

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Metabolism / Metabolites
Zilucoplan is expected to be degraded into small peptides and amino acids via catabolic pathways. RA103488 and RA102758 are two major metabolites detected in plasma. RA103488 is formed by CYP4F2-mediated metabolism and has comparable pharmacological activity to its parent compound; however, since RA103488 is present at much lower concentrations compared to zilucoplan, its contribution to the pharmacological action of zilucoplan is expected to be low. RA102758, formed by protease-mediated degradation, is pharmacologically inactive. The AUCs of both metabolites were approximately 10% of the parent AUC.

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Biological Half-Life
The mean plasma terminal half-life of zilucoplan was approximately 172 hours (7 to 8 days).

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Zilucoplan is expected to be degraded into small peptides and amino acids via catabolic pathways in the maternal circulation and infant gastrointestinal tract. It is unlikely to be absorbed by the breastfed infant or adversely affect the infant. No special precautions are required.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
Zilucoplan and its two major metabolites, RA103488 and RA102758, are more than 99% bound to plasma proteins.

[1]. Prevention of Anti-HMGCR Immune-Mediated Necrotising Myopathy by C5 Complement Inhibition in a Humanised Mouse Model. Biomedicines. 2022 Aug 20;10(8):2036.

[2]. Chemical synthesis and characterisation of the complement C5 inhibitory peptide zilucoplan. Amino Acids. 2021 Jan;53(1):143-147.

Zilucoplan is a 15 amino-acid, synthetic macrocyclic peptide with formula C172H278N24O55. Its sodium salt is used for the treatment of generalised myasthenia gravis (a disease that leads to muscle weakness and tiredness) in adults whose immune system produces antibodies against acetylcholine receptors. It has a role as a complement component 5 inhibitor and an immunosuppressive agent. It is a macrocycle, a homodetic cyclic peptide and a polyether. It is a conjugate acid of a zilucoplan(4-).
Zilucoplan is a 15 amino-acid, synthetic macrocyclic peptide. It is a complement inhibitor that works to prevent the activation of C5, which is a complement protein involved in the innate immune system to initiate inflammatory responses. On October 17, 2023, zilucoplan gained its first FDA approval for the treatment of generalized myasthenia gravis. It was also later approved by the EMA on December 4, 2023, as an add-on treatment for the same condition.
Zilucoplan is a Complement Inhibitor. The mechanism of action of zilucoplan is as a Complement Inhibitor.
Zilucoplan is a synthetic macrocyclic peptide inhibitor of the terminal complement protein C5, with potential anti-inflammatory and cell protective activities. Upon subcutaneous administration, complement zilucoplan binds to a unique site in terminal complement protein C5, which blocks C5 cleavage into C5a and C5b and prevents the C5b-dependent assembly of the membrane-attack complex (MAC). Zilucoplan also inhibits the interaction between C5b and C6, thereby further blocking MAC assembly. This prevents MAC-mediated lysis and destruction of red blood cells (RBCs) that occurs in complement-mediated diseases, such as paroxysmal nocturnal hemoglobinuria (PNH), generalized myasthenia gravis (gMG) and lupus nephritis (LN). C5, a complement pathway protein, is expressed at high levels by the liver.

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Drug Indication
Zilucoplan is indicated as the main treatment or add-on treatment to standard therapy for generalized myasthenia gravis (gMG) in adult patients who are anti-acetylcholine receptor (AChR) antibody-positive by the FDA and EMA respectively.
Treatment of myasthenia gravis

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Mechanism of Action
The complement system is part of the innate immune system and is critical in inflammatory reactions in response to pathogenic bacteria. Activation pathways of the complement system involve the cleavage of the complement protein C5 by C5 convertases to form C5a, a potent anaphylatoxin, and C5b. Cleavage of C5 also recruits C6, C7, C8, and C9. C5b binds to C6 to yield the terminal complement complex C5b9, a hydrophilic pore that spans the cell membrane. C5b9 causes an influx of water and ions, resulting in osmotic lysis of the targeted cell. The terminal complement cascade has been implicated in the pathophysiology of various inflammatory and autoimmune disorders, including gMG. gMG is an autoimmune disorder characterized by pathogenic autoantibodies that bind to AChRs. Accumulated MAC on the postsynaptic plasma membrane of the neuromuscular junction leads to muscle weakness and damage. The exact mechanism of zilucoplan in gMG has not been fully elucidated. Zilucoplan binds to the complement protein C5 with high affinity to inhibit its cleavage to C5a and C5b, preventing the generation of C5b9.

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Pharmacodynamics
In clinical trials consisting of anti-AChR antibody-positive adults with gMG, zilucoplan improved the signs and symptoms of myasthenia gravis, including muscle weakness. In vitro, it blocked the activation of C5 clinical variants. Zilucoplan causes complement inhibition in a dose-dependent manner: In clinical trials, complement inhibition of 97.5% by zilucoplan was observed by the end of the first week and persisted throughout the 12-week treatment period.

C172H278N24O55

3562.17557096481

3560.972

1841136-73-9

Zilucoplan TFA

133083018

White to off-white solid powder

4.8

28

57

142

251

6980

16

O=C([C@H](C1CCCCC1)NC([C@@H]1CCCN1C([C@H](CC1C=CC(=CC=1)O)NC([C@H](CCC(=O)O)NC([C@H](CC1=CNC2C1=CC=CN=2)NC([C@H](CC1C=CC(=CC=1)O)NC([C@H](C(C)(C)C)NC([C@H](CC(=O)O)N(C)C([C@@H]1CC(NCCCC[C@@H](C(N[C@H](C(N[C@@H](CCC(=O)O)C(N[C@@H](CCCNC(=N)N)C(N[C@@H](CC2C=CC=CC=2)C(N1)=O)=O)=O)=O)C(C)C)=O)NC(C)=O)=O)=O)=O)=O)=O)=O)=O)=O)=O)N[C@H](C(=O)O)CCCCNC(CCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCNC(CC[C@@H](C(=O)O)NC(CCCCCCCCCCCCCCC)=O)=O)=O

JDXCOXKBIGBZSK-PSNKNOTQSA-N

InChI=1S/C172H278N24O55/c1-9-10-11-12-13-14-15-16-17-18-19-20-27-44-146(202)182-136(170(226)227)53-56-144(200)177-64-67-229-69-71-231-73-75-233-77-79-235-81-83-237-85-87-239-89-91-241-93-95-243-97-99-245-101-103-247-105-107-249-109-111-251-113-112-250-110-108-248-106-104-246-102-100-244-98-96-242-94-92-240-90-88-238-86-84-236-82-80-234-78-76-232-74-72-230-70-68-228-66-59-145(201)175-60-31-29-41-135(169(224)225)186-165(220)152(126-37-25-22-26-38-126)193-162(217)142-43-34-65-196(142)168(223)140(116-125-47-51-129(199)52-48-125)190-157(212)133(54-57-148(204)205)184-161(216)139(117-127-120-180-154-130(127)39-32-62-178-154)188-159(214)138(115-124-45-49-128(198)50-46-124)189-166(221)153(172(5,6)7)194-163(218)143(119-150(208)209)195(8)167(222)141-118-147(203)176-61-30-28-40-131(181-122(4)197)158(213)192-151(121(2)3)164(219)185-134(55-58-149(206)207)156(211)183-132(42-33-63-179-171(173)174)155(210)187-137(160(215)191-141)114-123-35-23-21-24-36-123/h21,23-24,32,35-36,39,45-52,62,120-121,126,131-143,151-153,198-199H,9-20,22,25-31,33-34,37-38,40-44,53-61,63-119H2,1-8H3,(H,175,201)(H,176,203)(H,177,200)(H,178,180)(H,181,197)(H,182,202)(H,183,211)(H,184,216)(H,185,219)(H,186,220)(H,187,210)(H,188,214)(H,189,221)(H,190,212)(H,191,215)(H,192,213)(H,193,217)(H,194,218)(H,204,205)(H,206,207)(H,208,209)(H,224,225)(H,226,227)(H4,173,174,179)/t131-,132-,133-,134-,135-,136-,137-,138-,139-,140-,141-,142-,143-,151-,152-,153+/m0/s1

(2S)-2-[[(2S)-2-[[(2S)-1-[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S,5S,8S,11S,14S,22S)-22-acetamido-11-benzyl-8-(3-carbamimidamidopropyl)-5-(2-carboxyethyl)-3,6,9,12,16,23-hexaoxo-2-propan-2-yl-1,4,7,10,13,17-hexazacyclotricosane-14-carbonyl]-methylamino]-3-carboxypropanoyl]amino]-3,3-dimethylbutanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]-3-(1H-pyrrolo[2,3-b]pyridin-3-yl)propanoyl]amino]-4-carboxybutanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]pyrrolidine-2-carbonyl]amino]-2-cyclohexylacetyl]amino]-6-[3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[[(4S)-4-carboxy-4-(hexadecanoylamino)butanoyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]propanoylamino]hexanoic acid

Zilbrysq

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)