Title: Pulmonary insulin crystals
United States Patent: 6,310,038
Inventors: Havelund; Svend (Bagsvaerd, DK)
Assignee: Novo Nordisk A/S (Bagsvaerd, DK)
Appl. No.: 045038
Filed: March 20, 1998
Foreign Application Priority Data: Mar 20, 1997[DK]
The present invention relates to zinc free insulin crystals having a
diameter below 10 .mu.m and to therapeutic powder formulations suitable
for pulmonary administration containing such insulin crystals. The
crystals of the present invention exhibit a better stability profile than
powders of essentially the same composition prepared by spray drying,
freeze-drying, vacuum drying and open drying. The therapeutic powder
formulations elucidate better flowing properties than corresponding
amorphous powder formulations.
The zinc free insulin crystals of the invention are advantageously
provided in a crystal structure belonging to the cubic crystal system,
preferably in the octadecahedral or dodecahedral crystal forms, since
these crystal forms result in readily soluble product having excellent
The diameter of the insulin crystals is advantageously kept in the range
of 0.2 to 5 .mu.m, preferably in the range of 0.2 to 2 .mu.m, more
preferably in the range of 0.5 and 1 .mu.m, to ensure high bioavailability
and suitable profile of action, see PCT application No. WO 95/24183 and
PCT application No. WO 96/32149.
In a preferred embodiment the insulin used is selected from the group
consisting of human insulin, bovine insulin or porcine insulin, preferably
In another preferred embodiment the insulin used is selected from the
group consisting of rapid-acting insulins, preferably des(B30) human
insulin, AspB28 human insulin or LysB28 ProB29
In another preferred embodiment the insulin used is an insulin derivative,
preferably selected from the group consisting of B29-N.epsilon.
-myristoyl-des(B30) human insulin, B29-N.epsilon.
-palmitoyl-des(B30) human insulin, B29-N.epsilon. -myristoyl
human insulin, B29-N.epsilon. -palmitoyl human insulin,
B28-N.epsilon. -myristoyl LySB28 ProB29 human insulin,
B28-N.epsilon. -palmitoyl LysB28 ProB29 human insulin,
B30-N.epsilon. -myristoyl-ThrB29 LysB30 human insulin,
B30-N.epsilon. -palmitoyl-ThrB29 LysB30 human insulin,
B29-N.epsilon. -(N-palmitoyl-.gamma.-glutamyl)-des(B30) human
insulin, B29-N.epsilon. -(N-lithocholyl-.gamma.-glutamil)-des(B30)
human insulin, B29-N.epsilon. -(.omega.-carboxyheptadecanoyl)des(B30)
human insulin and B29-N.epsilon. -(.omega.-carboxyheptadecanoyl)
human insulin, more preferably LysB29 (N-.epsilon. acylated) des(B30)
The insulin derivatives have a protracted onset of action and may thus
compensate the very rapid increase in plasma insulin normally associated
with pulmonary delivery. By carefully selecting the type of insulin, the
present invention enables adjustment of the timing and to obtain the
desired biological response within a defined time span.
In order to avoid irritation of the lungs and to eliminate immunological
reactions, the employed insulin is preferably insulin which has been
purified by chromatography, such as MC insulin (Novo), Single Peak insulin
(E. Lilly) and RI insulin (Nordisk).
In a preferred embodiment the zinc free insulin crystals according to the
invention further comprise a stabilizing amount of a phenolic compound,
preferably m-cresol or phenol, or a mixture of these compounds.
The present invention is furthermore concerned with a therapeutic powder
formulation suitable for pulmonary administration comprising the zinc free
crystals described above.
In a preferred embodiment this therapeutic powder formulation further
comprises an enhancer which enhances the absorption of insulin in the
lower respiratory tract.
The enhancer is advantageously a surfactant, preferably selected from the
group consisting of salts of fatty acids, bile salts or phospholipids,
more preferably a bile salt.
Preferred fatty acids salts are salts of C.sub.10-14 fatty acids, such as
sodium caprate, sodium laurate and sodium myristate.
Lysophosphatidylcholine is a preferred phospholipid.
Preferred bile salts are salts of ursodeoxycholate, taurocholate,
glycocholate and taurodihydrofusidate. Still more preferred are powder
formulations according to the invention wherein the enhancer is a salt of
taurocholate, preferably sodium taurocholate.
The molar ratio of insulin to enhancer in the powder formulation of the
present invention is preferably 9:1 to 1:9, more preferably between 5:1 to
1:5, and still more preferably between 3:1 to 1:3.
The powder formulations of the present invention may optionally be
combined with a carrier or excipient generally accepted as suitable for
pulmonary administration. The purpose of adding a carrier or excipient may
be as a bulking agent, stabilizing agent or an agent improving the flowing
Suitable carrier agents include 1) carbohydrates, e.g. monosaccharides
such as fructose, galactose, glucose, sorbose, and the like; 2)
disaccharides, such as lactose, trehalose and the like; 3)
polysaccharides, such as raffinose, maltodextrins, dextrans, and the like;
4) alditols, such as mannitol, xylitol, and the like; 5) inorganic salts,
such as sodium chloride, and the like; 6) organic salts, such as sodium
citrate, sodium ascorbate, and the like. A preferred group of carriers
includes trehalose, raffinose, mannitol, sorbitol, xylitol, inositol,
sucrose, sodium chloride and sodium citrate.
The crystals of the present invention are advantageously produced
according to the following procedure:
a) providing a solution of insulin having a pH between 7.0 and 9.5;
b) mixing said solution with a solution of a salt of an alkali metal or an
ammonium salt; and
c) recovering the formed crystals.
The salt of an alkali metal or ammonium is preferably selected from the
group consisting of the hydrochloride or acetate of sodium, potassium,
lithium or ammonia, or mixtures thereof, more preferably sodium acetate.
In order to suppress the solubility of the crystals formed, the solution
of insulin and/or the solution of a salt of an alkali metal or an ammonium
salt preferably comprises a water miscible organic solvent in an amount
which corresponds to 5 to 25% (v/v) in the solution obtained after mixing.
The water miscible organic solvent is preferably selected from the group
consisting of ethanol, methanol, acetone and 2-propanol, more preferably
A very uniform distribution of crystal sizes and crystals of the same
crystallographic form are obtained when the two solutions are mixed within
a period of less than 2 hours, preferably less than 1 hour, more
preferably less than 15 minutes, still more preferably less than 5
The crystallization process by which uniformly sized, small, zinc free
crystals is obtained directly, without the use of milling, micronizing,
sieving and other dust generating steps, is much to be preferred from the
present state of the art in the manufacture of insulin powders for
The concentration of insulin after mixing is preferably between 0.5% and
10%, more preferably between 0.5% and 5%, still more preferably between
0.5% and 2%.
The concentration of salt after mixing is preferably between 0.2 M and 2
M, more preferably about 1 M.
The method according to the present invention may further comprise a
washing step, in which the crystals obtained are washed with a solution
comprising auxiliary substances to be included in the final dry powder,
preferably an enhancer and/or a carbohydrate, and optionally comprising
5-25% of an alcohol, preferably ethanol, 5-50 mM of a preservative
preferably phenol, and 0.1-2 M of a salt such as sodium acetate.
Claim 1 of 18 Claims
What is claimed is:
1. Zinc free insulin crystals having a diameter below 10 .mu.m, wherein
said insulin is selected from the group consisting of human insulin and
LysB29 (N-.epsilon.-acylated) des(B30) human insulin.
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