Insulin receptor (IR). Insulin lispro binds to and activates the insulin receptor (IR), a receptor tyrosine kinase. Binding of insulin to the extracellular alpha-subunits of IR induces conformational changes that activate the intracellular tyrosine kinase domain, leading to autophosphorylation and subsequent phosphorylation of downstream substrates (IRS proteins). This initiates signaling cascades including PI3K-Akt and MAPK pathways, promoting glucose uptake, glycogen synthesis, lipogenesis, and protein synthesis, while inhibiting gluconeogenesis and lipolysis.

Insulin lispro binds to the insulin receptor with similar affinity to human insulin in cell-free binding assays. IR binding affinity is typically measured using competition binding assays with radiolabeled [¹2⁵I]-insulin. Membranes prepared from cells overexpressing human insulin receptor (e.g., CHO-IR) or purified soluble IR ectodomain are incubated with various concentrations of unlabeled insulin lispro and a fixed concentration of [¹2⁵I]-insulin. Bound radioactivity is separated by filtration, and IC50/Ki values are calculated from displacement curves. Insulin lispro shows comparable receptor binding characteristics to regular human insulin.

Insulin lispro promotes glucose uptake and metabolism in cultured cells, such as 3T3-L1 adipocytes or L6 myotubes. In glucose uptake assays, cells are serum-starved and then treated with insulin lispro (0.01-100 nM) for 15-30 minutes. 2-deoxy-[3H]-glucose or fluorescent 2-NBDG is added, and uptake is measured. The EC50 for glucose uptake stimulation is typically in the sub-nanomolar range. Insulin lispro also activates insulin receptor autophosphorylation and downstream Akt phosphorylation as assessed by Western blotting. In INS-1 beta-cells, insulin does not directly potentiate insulin secretion (negative feedback), but the analog may have altered feedback properties.

Insulin receptor binding affinity is measured using a competition radioligand binding assay format. Human insulin receptor (isoform B) is immobilized on a solid support (e.g., 96-well plate or scintillation proximity assay (SPA) beads). Wells are incubated with 0.1-1 nM [¹2⁵I]-labeled human insulin (tracer) and increasing concentrations of unlabeled insulin lispro (0.001-1000 nM) for 2-4 hours at room temperature. Unbound tracer is removed by washing, and bound radioactivity is measured using a gamma counter or scintillation counter. Specific binding is calculated by subtracting nonspecific binding (excess unlabeled insulin, 10 uM). IC50 values are determined by nonlinear regression, and Ki values are calculated using the Cheng-Prusoff equation.

Glucose uptake assays are performed using 3T3-L1 adipocytes differentiated from preadipocytes. Differentiated adipocytes are serum-starved for 2-4 hours in low-glucose DMEM. Cells are washed with KRPH buffer (20 mM HEPES, pH 7.4) and incubated with insulin lispro (0.01-100 nM) for 20 minutes at 37degC. 2-deoxy-D-[3H]-glucose (0.5 uCi/well, 50 uM) is then added for 10 minutes. Uptake is stopped with ice-cold PBS containing 20 uM cytochalasin B. Cells are lysed, and radioactivity is measured by scintillation counting. Nonspecific uptake is determined in the presence of 20 uM cytochalasin B. EC50 values are calculated from dose-response curves.

In vivo insulin activity is assessed using the hypoglycemic clamp technique in rodents or large animals. For euglycemic clamp studies, conscious, catheterized rats or dogs are infused intravenously with insulin lispro at a constant rate (e.g., 1-10 mU/kg/min), while glucose is infused at a variable rate to maintain euglycemia (blood glucose ~100 mg/dL). The glucose infusion rate (GIR) required to maintain euglycemia reflects insulin sensitivity. Insulin lispro has a faster onset and shorter duration of action compared to regular human insulin, making it suitable for prandial glucose control. In diabetic animal models (STZ-induced diabetes), subcutaneous insulin lispro (0.1-1 U/kg) rapidly lowers blood glucose within 30 minutes.

Insulin lispro is rapidly absorbed following subcutaneous injection, achieving peak blood levels in 30-70 minutes compared to 60-120 minutes for regular human insulin. The absolute bioavailability after subcutaneous injection ranges from 55% to 77% with doses between 0.1 to 0.2 unit/kg. Absorption is faster from the abdomen compared to arm or thigh, but total exposure is similar across injection sites. The time-to-peak (Tmax) is approximately 60 minutes. Insulin lispro has a shorter duration of action (3-5 hours) compared to regular human insulin (5-8 hours). It is metabolized primarily by insulin-degrading enzyme in the liver and kidney. The half-life of elimination is approximately 1 hour. Renal impairment may affect clearance, but relative absorption remains faster than human insulin.

Adverse effects of insulin lispro include hypoglycemia (most common), hypokalemia, injection site reactions (lipodystrophy, pruritus, rash), and weight gain. Hypoglycemia is the most serious adverse event and can manifest as sweating, palpitations, confusion, seizures, or coma. Allergic reactions to insulin lispro are rare but can include anaphylaxis. Compared to regular human insulin, insulin lispro is associated with less nocturnal hypoglycemia due to its shorter duration of action. In clinical trials, the incidence of severe hypoglycemia is similar to or lower than with regular human insulin. Insulin lispro should not be used during episodes of hypoglycemia.

[1]. Insulin lispro: a review of its use in the management of diabetes mellitus. Drugs. 2007;67(3):407-34.

Insulin is modified so that amino acid 28 of its B chain is replaced with lysine, and amino acid 29 is replaced with lysine. It is used to control blood glucose levels in patients with type 2 diabetes.
See also: Lispro insulin (note moved to).

---

Insulin lispro was the first rapid-acting insulin analog and is marketed under the brand name Humalog® (Eli Lilly and Company). It was approved by the FDA in 1996. It is used in conjunction with intermediate- or long-acting insulin (e.g., insulin glargine, insulin detemir, isophane NPH insulin) for basal-bolus therapy in Type 1 diabetes. In Type 2 diabetes, it may be used without a longer-acting insulin when given with sulfonylureas. Insulin lispro is also available as a premixed formulation with insulin lispro protamine (Humalog Mix 75/25, Humalog Mix 50/50). Unlike human insulin, insulin lispro can be administered immediately before a meal, offering greater dosing convenience and flexibility.

C158H234N40O42S2.C99H155N25O35S4

5813.61785999995

5811.691

133107-64-9

16132438

White to off-white solid powder

84

89

185

405

13000

52

[H]/N=C(/NCCC[C@@H](C(NCC(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N1[C@H](C(N[C@H](C(=O)O)[C@H](O)C)=O)CCC1)=O)CCCCN)=O)[C@H](O)C)=O)CC1=CC=C(O)C=C1)=O)CC1=CC=CC=C1)=O)CC1=CC=CC=C1)=O)=O)NC([C@@H](NC(CNC([C@@H](NC([C@H](C(C)C)NC([C@@H](NC([C@@H](NC([C@@H](NC([C@@H](NC([C@@H](NC([C@H](C(C)C)NC([C@@H](NC([C@@H](NC([C@@H](NC(CNC([C@@H](NC([C@@H](NC([C@@H](NC([C@@H](NC([C@@H](NC([C@H](C(C)C)NC([C@H](CC1=CC=CC=C1)N)=O)=O)CC(=O)N)=O)CCC(=O)N)=O)CC1N=CNC=1)=O)CC(C)C)=O)CS)=O)=O)CO)=O)CC1N=CNC=1)=O)CC(C)C)=O)=O)CCC(=O)O)=O)C)=O)CC(C)C)=O)CC1=CC=C(O)C=C1)=O)CC(C)C)=O)=O)CS)=O)=O)CCC(=O)O)=O)\N.CC[C@@H]([C@@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(N[C@H](C(=O)O)CC(=O)N)=O)CS)=O)CC1=CC=C(O)C=C1)=O)CC(=O)N)=O)CCC(=O)O)=O)CC(C)C)=O)CCC(=O)N)=O)CC1=CC=C(O)C=C1)=O)CC(C)C)=O)CO)=O)CS)=O)NC([C@@H](NC([C@H]([C@H](O)C)NC([C@@H](NC([C@@H](NC([C@@H](NC([C@@H](NC([C@H](C(C)C)NC([C@H]([C@H](CC)C)NC(CN)=O)=O)=O)CCC(=O)O)=O)CCC(=O)N)=O)CS)=O)CS)=O)=O)CO)=O)C

WNRQPCUGRUFHED-DETKDSODSA-N

InChI=1S/C158H234N40O42S2.C99H155N25O35S4/c1-79(2)57-104(181-131(213)86(15)173-136(218)102(50-53-125(211)212)179-152(234)127(84(11)12)194-148(230)107(60-82(7)8)184-145(227)113(67-95-70-166-78-172-95)189-150(232)115(74-199)176-123(208)73-170-134(216)116(75-241)191-140(222)105(58-80(3)4)182-144(226)112(66-94-69-165-77-171-94)188-138(220)101(48-51-119(161)204)178-146(228)114(68-120(162)205)190-153(235)126(83(9)10)193-132(214)98(160)61-89-31-21-18-22-32-89)139(221)185-110(64-92-40-44-96(202)45-41-92)142(224)183-106(59-81(5)6)147(229)195-128(85(13)14)154(236)192-117(76-242)135(217)169-71-121(206)174-100(49-52-124(209)210)137(219)177-99(38-29-55-167-158(163)164)133(215)168-72-122(207)175-108(62-90-33-23-19-24-34-90)141(223)186-109(63-91-35-25-20-26-36-91)143(225)187-111(65-93-42-46-97(203)47-43-93)149(231)196-129(87(16)200)155(237)180-103(37-27-28-54-159)156(238)198-56-30-39-118(198)151(233)197-130(88(17)201)157(239)240;1-12-46(9)77(121-73(134)36-100)97(156)122-76(45(7)8)95(154)108-56(25-29-75(137)138)80(139)105-54(23-27-70(102)131)83(142)117-66(40-161)93(152)119-68(42-163)94(153)124-79(48(11)127)98(157)116-64(38-126)90(149)123-78(47(10)13-2)96(155)120-67(41-162)92(151)115-63(37-125)89(148)110-58(31-44(5)6)85(144)111-59(32-49-14-18-51(128)19-15-49)86(145)106-53(22-26-69(101)130)81(140)109-57(30-43(3)4)84(143)107-55(24-28-74(135)136)82(141)113-61(34-71(103)132)88(147)112-60(33-50-16-20-52(129)21-17-50)87(146)118-65(39-160)91(150)114-62(99(158)159)35-72(104)133/h18-26,31-36,40-47,69-70,77-88,98-118,126-130,199-203,241-242H,27-30,37-39,48-68,71-76,159-160H2,1-17H3,(H2,161,204)(H2,162,205)(H,165,171)(H,166,172)(H,168,215)(H,169,217)(H,170,216)(H,173,218)(H,174,206)(H,175,207)(H,176,208)(H,177,219)(H,178,228)(H,179,234)(H,180,237)(H,181,213)(H,182,226)(H,183,224)(H,184,227)(H,185,221)(H,186,223)(H,187,225)(H,188,220)(H,189,232)(H,190,235)(H,191,222)(H,192,236)(H,193,214)(H,194,230)(H,195,229)(H,196,231)(H,197,233)(H,209,210)(H,211,212)(H,239,240)(H4,163,164,167);14-21,43-48,53-68,76-79,125-129,160-163H,12-13,22-42,100H2,1-11H3,(H2,101,130)(H2,102,131)(H2,103,132)(H2,104,133)(H,105,139)(H,106,145)(H,107,143)(H,108,154)(H,109,140)(H,110,148)(H,111,144)(H,112,147)(H,113,141)(H,114,150)(H,115,151)(H,116,157)(H,117,142)(H,118,146)(H,119,152)(H,120,155)(H,121,134)(H,122,156)(H,123,149)(H,124,153)(H,135,136)(H,137,138)(H,158,159)/t86-,87+,88+,98-,99-,100-,101-,102-,103-,104-,105-,106-,107-,108-,109-,110-,111-,112-,113-,114-,115-,116-,117-,118-,126-,127-,128-,129-,130-;46-,47-,48+,53-,54-,55-,56-,57-,58-,59-,60-,61-,62-,63-,64-,65-,66-,67-,68-,76-,77-,78-,79-/m00/s1

(4S)-4-[[(2S)-2-[[(2S)-5-amino-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2R)-2-[[(2S,3S)-2-[[(2S)-2-[[(2S,3R)-2-[[(2R)-2-[[(2R)-2-[[(2S)-5-amino-2-[[(2S)-2-[[(2S)-2-[[(2S,3S)-2-[(2-aminoacetyl)amino]-3-methylpentanoyl]amino]-3-methylbutanoyl]amino]-4-carboxybutanoyl]amino]-5-oxopentanoyl]amino]-3-sulfanylpropanoyl]amino]-3-sulfanylpropanoyl]amino]-3-hydroxybutanoyl]amino]-3-hydroxypropanoyl]amino]-3-methylpentanoyl]amino]-3-sulfanylpropanoyl]amino]-3-hydroxypropanoyl]amino]-4-methylpentanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]-5-oxopentanoyl]amino]-4-methylpentanoyl]amino]-5-[[(2S)-4-amino-1-[[(2S)-1-[[(2R)-1-[[(1S)-3-amino-1-carboxy-3-oxopropyl]amino]-1-oxo-3-sulfanylpropan-2-yl]amino]-3-(4-hydroxyphenyl)-1-oxopropan-2-yl]amino]-1,4-dioxobutan-2-yl]amino]-5-oxopentanoic acid;(4S)-4-[[2-[[(2R)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[2-[[(2R)-2-[[(2S)-2-[[(2S)-2-[[(2S)-5-amino-2-[[(2S)-4-amino-2-[[(2S)-2-[[(2S)-2-amino-3-phenylpropanoyl]amino]-3-methylbutanoyl]amino]-4-oxobutanoyl]amino]-5-oxopentanoyl]amino]-3-(1H-imidazol-5-yl)propanoyl]amino]-4-methylpentanoyl]amino]-3-sulfanylpropanoyl]amino]acetyl]amino]-3-hydroxypropanoyl]amino]-3-(1H-imidazol-5-yl)propanoyl]amino]-4-methylpentanoyl]amino]-3-methylbutanoyl]amino]-4-carboxybutanoyl]amino]propanoyl]amino]-4-methylpentanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]-4-methylpentanoyl]amino]-3-methylbutanoyl]amino]-3-sulfanylpropanoyl]amino]acetyl]amino]-5-[[(2S)-1-[[2-[[(2S)-1-[[(2S)-1-[[(2S)-1-[[(2S,3R)-1-[[(2S)-6-amino-1-[(2S)-2-[[(1S,2R)-1-carboxy-2-hydroxypropyl]carbamoyl]pyrrolidin-1-yl]-1-oxohexan-2-yl]amino]-3-hydroxy-1-oxobutan-2-yl]amino]-3-(4-hydroxyphenyl)-1-oxopropan-2-yl]amino]-1-oxo-3-phenylpropan-2-yl]amino]-1-oxo-3-phenylpropan-2-yl]amino]-2-oxoethyl]amino]-5-carbamimidamido-1-oxopentan-2-yl]amino]-5-oxopentanoic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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.

---

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).

View More

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)

---

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).

View More

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

---

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.

---

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