Formation and anion-exchange of crystalline echinocandin...

Details Formation and anion-exchange of crystalline echinocandin... Formation and anion-exchange of crystalline echinocandin...

2001-08-30

2003-07-08

Low, Christopher S. F. (Department: 1653)

Chemistry: natural resins or derivatives; peptides or proteins;

Peptides of 3 to 100 amino acid residues

Cyclic peptides

C530S344000, C530S345000, C530S300000, C514S011400, C514S009100, C514S002600, C435S071100

Reexamination Certificate

active

06590073

ABSTRACT:

FIELD OF THE INVENTION
The present invention relates to a process for formation and anion-exchange of crystalline salts of an echinocandin nucleus, in particular, salts of an Echinocandin B nucleus.
BACKGROUND OF THE INVENTION
Echinocandin cyclopeptides are natural antifungal products. Included in the Echinocandin cyclopeptide family are natural products such as Echinocandin B (ECB), Echinocandin C, Aculeacin A&ggr;, Mulundocandin, Sporiofungin A, Pneumocandin A
0
, WF11899A, and Pneumocandin B
0
. These are typically produced by culturing various microorganisms. For example, Echinocandin B is produced from the fermentation of the fungus,
Aspergillus Nidulans.
In the search for more active materials, the natural products have been modified in a variety of ways. One of the most common has been replacement of the N-acyl side chain on the natural product to produce a semi-synthetic derivative. For example, U.S. Pat. Nos. 4,293,489; 4,320,052; 5,166,135; and 5,541,160; and EP 359529; 448353; 447186; 462531; and 561639 describe a variety of N-acyl derivatized Echinocandin compounds with varying degrees of antifungal activity.
The N-acyl derivatives are produced by deacylating the natural product followed by reacylation with a different acyl group. Deacylation is typically achieved by means of an enzyme (e.g., deacylase enzyme). The deacylase enzyme may be obtained from the microorganism
Actinoplanes utahensis
or Pseudomonas species (see i.e., U.S. Pat. Nos. 4,293,482 and 4,304,716; and EP 460,882). The deacylated compound is typically referred to as the nucleus of the corresponding natural product (e.g., the deacylated product of Echinocandin B is referred to as the Echinocandin B nucleus (ECBN)). Unfortunately, both the acylated and unacylated products are difficult to purify due to their limited solubility and amorphous state. In addition, the free amino compound (e.g., ECBN) is generally unstable and subject to ring opening.
It is well-known in the art that crystalline materials in general are easier to purify than their amorphous counterparts. Hence, it is desirable to produce cyclopeptide compounds in their crystalline state to obtain optimal purity. Since the potency of the final pharmaceutical product is dependent upon the purity of the intermediates used to make the final product, improvements in purity at any stage of the manufacturing process is highly desirable. Ideally, the contaminants are removed at the earliest stage possible in the manufacturing process. Hence, there is a need for a process that simplifies and improves the purification of cyclopeptide compounds containing a free amino group prior to subsequent attachment of an amino substituent.
BRIEF SUMMARY OF THE INVENTION
The present invention provides a method for forming a crystalline echinocandin nucleus salt from its mixed broth or partially purified process streams by the steps of (i) concentrating a solution comprising an echinocandin nucleus or amorphous salt thereof, an aldehyde impurity and a solvent by means of a nanofiltration process to form a concentrate; (ii) adding an aldehyde derivatizing agent; (iii) adjusting the pH to a value less than 4.0 (preferably between about 2.0 and about 3.0); (iv) adding an acid or metal salt; and (v) cooling the concentrate to crystallize an echinocandin nucleus salt having an anion corresponding to the anion of the acid or metal salt added in step (iv). A seed crystal may optionally be added to initiate crystallization.
In another embodiment of the present invention, a process for exchanging the anion of an Echinocandin ammonium salt (including simple derivatives thereof) is provided as well as various forms of crystalline echinocandin nucleus salts.
DEFINITIONS
“Echinocandin compounds” refers to compounds having the following general structure including any simple derivatives thereof:
wherein R is a hydrogen or —C(O)R′ where R′ is an alkyl group, an alkenyl group, an alkynyl group, an aryl group, or heteroaryl group; R
1
is —H or —OH; R
2
is —H, —NH
2
or —CH
3
; R
3
is —H, —CH
3
, —CH
2
CONH, or —CH
2
CH
2
NH
2
; R
4
is —H or —OH; R
5
is —OH, —OSO
3
H, or —OPO
2
HR
a
, where R
a
is hydroxy, C
1
-C
6
alkyl, C
1
-C
6
alkoxy, phenyl, phenoxy, p-halophenyl, p-halophenoxy, p-nitrophenyl, p-nitrophenoxy, benzyl, benzyloxy, p-halobenzyl, p-halobenzyloxy, p-nitrobenzyl, or p-nitrobenzyloxy; R
6
is —H, —OH, or —OSO
3
H; R
7
is —H or —CH
3
; and pharmaceutically acceptable salts, esters, hydrates or solvates thereof. Also included within the meaning of echinocandin are the various enantomeric forms of structure I illustrated above even though specific chiral centers are depicted. “Echinocandin nucleus” refers to the deacylated Echinocandin compound where R is a hydrogen. “ECBN” refers to the Echinocandin B nucleus where R1, R4 and R5 are hydroxyl groups, R2, R3, and R7 are methyl groups; and R and R6 are hydrogens.
“Alkyl” refers to a hydrocarbon radical of the general formula C
n
H
2n+1
, containing from 1 to 30 carbon atoms unless otherwise indicated. The alkane radical may be straight (e.g. methyl, ethyl, propyl, butyl, etc.), branched (e.g., isopropyl, isobutyl, tertiary butyl, neopentyl, etc.), cyclic (e.g., cyclopropyl, cyclobutyl, cyclopentyl, methylcyclopentyl, cyclohexyl, etc.), or multi-cyclic (e.g., bicyclo[2.2.1]heptane, spiro[2.2]pentane, etc.). The alkane radical may be substituted or unsubstituted. Similarly, the alkyl portion of an alkoxy group or alkanoate have the same definition as above.
“Alkenyl” refers to an acyclic hydrocarbon containing at least one carbon carbon double bond. The alkene radical may be straight, branched, cyclic, or multi-cyclic. The alkene radical may be substituted or unsubstituted.
“Alkynyl” refers to an acyclic hydrocarbon containing at least one carbon carbon triple bond. The alkyne radical may be straight, or branched. The alkyne radical may be substituted or unsubstituted.
“Aryl” refers to aromatic moieties having single (e.g., phenyl) or fused ring systems (e.g., napthalene, anthracene, phenanthrene, etc.). The aryl groups may be substituted or unsubstituted.
“Heteroaryl” refers to aromatic moieties containing at least one heteroatom within the aromatic ring system (e.g., pyrrole, pyridine, indole, thiophene, furan, benzofuran, imidazole, pyrimidine, purine, benzimidazole, quinoline, etc.). The aromatic moiety may consist of a single or fused ring system. The heteroaryl groups may be substituted or unsubstituted.
Within the field of organic chemistry and particularly within the field of organic biochemistry, it is widely understood that significant substitution of compounds is tolerated or even useful. In the present invention, for example, the term alkyl group allows for substituents which is a classic alkyl, such as methyl, ethyl, propyl, hexyl, isooctyl, dodecyl, stearyl, etc. The term specifically envisions and allows for substitutions on alkyls which are common in the art, such as hydroxy, halogen, alkoxy, carbonyl, keto, ester, carbamato, etc., as well as including the unsubstituted alkyl moiety. However, the substituents should be selected so as to not adversely affect the pharmacological characteristics of the compound or adversely interfere with the use of the medicament. Suitable substituents for any of the groups defined above include alkyl, alkenyl, alkynyl, aryl, halo, hydroxy, alkoxy, aryloxy, mercapto, alkylthio, arylthio, mono- and di-alkyl amino, quaternary ammonium salts, aminoalkoxy, hydroxyalkylamino, aminoalkylthio, carbamyl, carbonyl, carboxy, glycolyl, glycyl, hydrazino, guanyl, and combinations thereof.
“Solvate” means an aggregate that comprises one or more molecules of the solute, such as Compound I, with one or more molecules of a solvent, such as water, ethanol, and the like.
“Suitable solvent” refers to any solvent, or mixture of solvents, inert to the ongoing reaction that sufficiently solubilizes the reactants to afford a medium within which to effect the desired anion exchange or salt formation.
“Mixed broth” refers to a conversion mixture whe

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