Patent Grant
9828341
This invention relates generally to compounds that are intermediates for the preparation of dendrimer compounds, the use of these compounds for preparing dendrimers and processes for preparing the compounds.
Interest in dendritic compounds has grown in recent times, and dendrimers have found application in a wide range of areas, such as pharmaceuticals, drug delivery, gene delivery, sensor technologies (cation photodetection and fluorescence signal quenching), in the synthesis of monodisperse metallic nanoparticles, in environmental remediation, as blood substitutes usually in the oxygen-carrying sense (oxygen therapeutics) and plasma substitutes (volume expenders).
Dendritic structures tend to be complex. However, for some applications, such as pharmaceuticals, it is desirable to employ compounds that are discrete, well-characterised, single entities. This can be an issue in the dendrimer space, so it is desirable to use intermediates and processes which produce such single entities. There is therefore a need for simple dendritic cores that can be used to prepare dendritic compounds for use in certain applications.
Newkome et al. (Macromolecules, 1993, 26, 2394-2396) have described the synthesis of a dodecaacid dendritic core, starting from the tetraacid compound (structure shown in Scheme 1, below) and coupling this with a branched amine. This produced a compound with twelve terminal acids (attachment points), which was further elaborated (through five generations) to produce dendrimers having up to 972 terminal acids.
Newkome et al. (J. Org. Chem. 2002, 67, 3957-3960) have also reported the synthesis of a tetraamine (amine terminated) dendritic core, starting from the tetraacid compound. The tetraamine core was further elaborated by reaction with excess acrylonitrile to generate an octanitrile as an oily product.
Hukkamäki et al. (Hukkämaki, J.; Pakkanen, T. T. Journal of Molecular Catalysis A: Chemical 2001, 174, 205-21,) have also described the synthesis of the same amine terminated dendritic core. The tetraamine core was further elaborated by reaction with acrylonitrile to produce a (two generation) dendrimer having eight terminal amine groups.
For the synthesis of simple, well-defined dendritic molecules as single compounds, it would be desirable to employ a simple, stable dendritic core as an intermediate which could provide a route to the synthesis of, for example, dendrimers having pharmaceutical uses.
It is therefore an object of the present invention to provide intermediate compounds for the preparation of dendritic compounds, or to at least provide a useful choice.
In a first aspect, the invention provides a compound of formula (I)
wherein:
Y is O;
B is O;
R1 and R2 are absent; and
either A, E, D and X are all CH2; or A, D and X are all CH2 and E is (CH2CH2O)t#CH2 wherein # indicates a point of attachment of E to its adjacent carbonyl group;
t is an integer from 1 to 10;
or wherein:
Y is C;
R1 and R2 are both H; and
A, E, B and D are CH2 and X is O;
or wherein:
Y is C;
A is (CH2)u
R1 and R2 are both H;
B, X, D and E are all absent; and
u is an integer from 1 to 10;
or wherein:
Y is C;
X is O;
B is (CH2)p;
A, E and D are all CH2; and
R1 is H, NHZ or C1-6alkyl and R2 is a radical of formula (i) or a radical of formula (ii)
Z is H, acyl, C(O)(CH2)wN(H)G, *CH3*C(O)— where *C denotes 13C or 14C, 5-TAMRA (4-carboxytetramethylrhodamine), Fluorescein (resorcinolphthalein), Alexa Fluor 350 (7-amino-4-methyl-6-sulfocoumarin-3-acetic acid), BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) or Alkyne MegaStokes dye 608 (1-{3-{[4-(2-cyclooctyn-1-ylmethyl)benzoyl]amino}propyl}-4-{2-[4-(dimethylamino)phenyl]ethenyl}pyridinium hexafluorophosphate);
w is an integer from 1 to 11;
G is H, acyl, Boc (t-butoxycarbonyl), Troc (2,2,2-trichloroethyloxycarbonyl), Fmoc (9-fluorenylmethoxycarbonyl), Cbz (benzyloxycarbonyl),*CH3*CO— where *C denotes 13C or 14C, 5-TAMRA (4-carboxytetramethylrhodamine), Fluorescein (resorcinolphthalein), Alexa Fluor 350 (7-amino-4-methyl-6-sulfocoumarin-3-acetic acid), BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) or Alkyne MegaStokes dye 608 (1-{3-{[4-(2-cyclooctyn-1-ylmethyl)benzoyl]amino}propyl}-4-{2-[4-(dimethylamino)phenyl]ethenyl}pyridinium hexafluorophosphate);
or wherein:
Y is C;
X is O;
B is (CH2)p;
A, E and D are all CH2; and
R1 and R2, both the same, are a radical of formula (i) or a radical of formula (ii)
R3 is a radical of formula (iii) or a radical of formula (iv)
each T is independently selected from the group consisting of (CH2CH2O)xCH2CH2 and CH2;
each x is independently an integer from 1 to 12;
m is an integer from 1 to 11, provided that when T is (CH2CH2O)xCH2CH2 then m is 1;
n is an integer from 1 to 11, provided that when T is (CH2CH2O)xCH2CH2 then n is 1;
p is an integer from 1 to 5;
R5 is H,
and
M is sodium or ammonium.
Preferably each T is CH2.
Alternatively preferably at least one T is CH2.
Alternatively preferably at least one T is (CH2CH2O)xCH2CH2.
More preferably each T is CH2 and m is an integer from 3 to 10, e.g. an integer from 4 to 9, e.g. an integer from 5 to 8, e.g. an integer from 6 to 7. Most preferably m is 7. Preferably each T is CH2 and n is an integer from 3 to 10, e.g. an integer from 4 to 9, e.g. an integer from 5 to 8, e.g. an integer from 6 to 7. Most preferably n is 7.
Alternatively preferably at least one T is CH2 and m is an integer from 3 to 10, e.g. an integer from 4 to 9, e.g. an integer from 5 to 8, e.g. an integer from 6 to 7. Most preferably m is 7. Preferably at least one T is CH2 and n is an integer from 3 to 10, e.g. an integer from 4 to 9, e.g. an integer from 5 to 8, e.g. an integer from 6 to 7. Most preferably n is 7.
Alternatively preferably at least one T is (CH2CH2O)xCH2CH2 and x is an integer from 2 to 10, e.g. an integer from 2 to 9, e.g. an integer from 2 to 8, e.g. an integer from 2 to 7, e.g. an integer from 2 to 6, e.g. an integer from 2 to 5, e.g. an integer from 2 to 4. Most preferably x is 3.
Preferably Y is O. Alternatively, it is preferred that Y is C.
Preferably R3 is a radical of formula (iii)
Alternatively it is preferred that R3 is a radical of formula (iv)
It is further preferred that, in R3, one T is (CH2CH2O)xCH2CH2 and one T is CH2.
It is further preferred that, in R3, (T)m is (CH2CH2O)xCH2CH2 and (T)n is (CH2)n. Alternatively it is preferred that, in R3, (T)n is (CH2CH2O)xCH2CH2 and (T)m is (CH2)m.
Alternatively preferably R3 is a radical of formula (iv) wherein each T is (CH2CH2O)xCH2CH2, wherein each x in each radical of formula (iv) is independently selected.
Preferably R1 and R2 are both a radical of formula (i)
Alternatively it is preferred that R1 and R2 are both a radical of formula (ii)
It is further preferred that, in R1, one T is (CH2CH2O)xCH2CH2 and one T is CH2 and, in R2, one T is (CH2CH2O)xCH2CH2 and one T is CH2.
It is further preferred that, in both R1 and R2, (T)m is (CH2CH2O)xCH2CH2 and (T)n is (CH2)n. Alternatively it is preferred that, in both R1 and R2, (T)n is (CH2CH2O)xCH2CH2 and (T)m is (CH2)m. Alternatively it is preferred that, in both R1 and R2, each T is (CH2CH2O)xCH2CH2, wherein each x in each radical of formula (ii) is independently selected.
Alternatively it is preferred that R1 is H or C1-6alkyl, e.g. CH3 or CH2CH3. Alternatively it is preferred that R1 is NH2. Alternatively it is preferred that R1 is NHZ, more preferably where Z is C(O)(CH2)wN(H)G, e.g. where G is Troc (2,2,2-trichloroethyloxycarbonyl), Fmoc (fluorenylmethyloxycarbonyl) or Cbz (benzyloxycarbonyl). Preferably w is 7.
Preferably R5 is selected from the group consisting of
Preferably R5 is
Alternatively preferably R5 is H. Alternatively preferably R5 is
Preferably Y is O; B is O; R1 and R2 are absent; and either A, E, D and X are all CH2 or A, D and X are all CH2 and E is (CH2CH2O)tCH2; and t is an integer from 1 to 10, preferably an integer from 1 to 2.
Alternatively it is preferred that Y is C; R1 and R2 are both H; A, E, B and D are CH2 and X is O.
Alternatively it is preferred that Y is C; A is (CH2)u; R1 and R2 are both H; B, X, D and E are all absent; and u is an integer from 1 to 10.
Alternatively it is preferred that:
Y is C; X is O; A, E and D are all CH2; B is (CH2)p;
R1 is H, NHZ or C1-6alkyl;
R2 is a radical of formula (i) or a radical of formula (ii)
and
Z is H, acyl, C(O)(CH2)wN(H)G, *CH3*CO— where *C denotes 13C or 14C, 5-TAMRA (4-carboxytetramethylrhodamine), Fluorescein (resorcinolphthalein), Alexa Fluor 350 (7-amino-4-methyl-6-sulfocoumarin-3-acetic acid), BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) or Alkyne MegaStokes dye 608 (1-{3-{[4-(2-cyclooctyn-1-ylmethyl)benzoyl]amino}propyl}-4-{2-[4-(dimethylamino)phenyl]ethenyl}pyridinium hexafluorophosphate);
w is an integer from 1 to 11;
G is H, acyl, (t-butoxycarbonyl), Troc (2,2,2-trichloroethyloxycarbonyl), Fmoc (9-fluorenylmethoxycarbonyl), Cbz (carboxybenzyl),* *CH3*CO— where *C denotes 13C or 14C, 5-TAMRA (4-carboxytetramethylrhodamine), Fluorescein (resorcinolphthalein), Alexa Fluor 350 (7-amino-4-methyl-6-sulfocoumarin-3-acetic acid), BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) or Alkyne MegaStokes dye 608 (1-{3-{[4-(2-cyclooctyn-1-ylmethyl)benzoyl]amino}propyl}-4-{2-[4-(dimethylamino)phenyl]ethenyl}pyridinium hexafluorophosphate).
Preferably R2 is a radical of formula (ii)
and R1 is H, NHZ or C1-6alkyl. It is further preferred that, in R2, one T is (CH2CH2O)xCH2CH2 and one T is CH2.
It is further preferred that, in R2, (T)m is (CH2CH2O)xCH2CH2 and (T)n is (CH2)n. Alternatively it is preferred that, in R2, (T)n is (CH2CH2O)xCH2CH2 and (T)m is (CH2)m. Alternatively it is preferred that, in R2, each T is (CH2CH2O)xCH2CH2, wherein each x in each radical of formula (ii) is independently selected.
Alternatively it is preferred that Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 and R2, both the same, are a radical of formula (i) or a radical of formula (ii)
Preferably, Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 and R2, both the same, are a radical of formula (i)
R3 is a radical of formula (iii)
and p is 1.
Alternatively preferably Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 and R2, both the same, are a radical of formula (ii)
R3 is a radical of formula (iv)
and p is 1.
Preferably Y is C; R1 and R2 are both H; A, E, B and D are CH2; X is O; and R3 is a radical of formula (iii)
Alternatively preferably Y is C; R1 and R2 are both H; A, E, B and D are CH2; X is O; and R3 is a radical of formula (iv)
Preferably Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 is H; R2 is a radical of formula (i)
R3 is a radical of formula (iii)
and p is 1.
Alternatively preferably Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 is H; R2 is a radical of formula (ii)
R3 is a radical of formula (iv)
and p is 1.
Alternatively preferably Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 is H; R2 is a radical of formula (ii)
R3 is a radical of formula (iii)
and p is 1.
Alternatively preferably Y is C; X is O; A, E and D are all CH2; B is (CH2)p; R1 is H; R2 is a radical of formula (i)
and R3 is a radical of formula (iv)
and p is 1.
Preferably Y is C; A is (CH2)u; R1 and R2 are both H; B, X, D and E are all absent; u is an integer from 1 to 10; and R3 is a radical of formula (iii)
Alternatively preferably Y is C; A is (CH2)u; R1 and R2 are both H; B, X, D and E are all absent; u is an integer from 1 to 10; and R3 is a radical of formula (iv)
Preferably Y is O; B is O; R1 and R2 are absent; and either A, E, D and X are all CH2 or A, D and X are all CH2 and E is (CH2CH2O)tCH2; t is an integer from 1 to 10, preferably an integer from 1 to 2; and R3 is a radical of formula (iii)
Alternatively preferably Y is O; B is O; R1 and R2 are absent; and either A, E, D and X are all CH2 or A, D and X are all CH2 and E is (CH2CH2O)tCH2; t is an integer from 1 to 2; and R3 is a radical of formula (iv)
Preferably p is 1.
Preferably t is an integer from 1 to 2.
Preferably M is sodium.
Preferably the compound of formula (I) is selected from the group consisting of:
In another aspect, the invention provides a compound selected from the group consisting of:
In another aspect, the invention provides the use of a compound of formula (I) as defined above for preparing a dendritic compound.
In still another aspect, the invention provides a method of preparing a dendritic compound, including the step of contacting a compound of formula (I) with a glycoside compound that contains a free amino group.
It will be appreciated that any of the sub-scopes disclosed herein, e.g. with respect to A, B, D, E, G, X, R1, R2, R3, R4, R5, n, m, p t, u, w, x, T, Y and Z may be combined with any of the other sub-scopes disclosed herein to produce further sub-scopes.
Definitions
The term “C1-C6alkyl” means any saturated hydrocarbon radical having up to 6 carbon atoms and is intended to include both straight- and branched-chain alkyl groups. Examples of alkyl groups include: methyl group, ethyl group, n-propyl group, iso-propyl group, n-butyl group, iso-butyl group, sec-butyl group, t-butyl group, n-pentyl group, 1,1-dimethylpropyl group, 1,2-dimethylpropyl group, 2,2-dimethylpropyl group, 1-ethylpropyl group, 2-ethylpropyl group, n-hexyl group and 1-methyl-2-ethylpropyl group.
The term “acyl” means C(═O)R′ group, where R′ is a C1-C30alkyl group, where C1-C30alkyl means any saturated hydrocarbon radical having up to 30 carbon atoms, and is intended to include straight chain alkyl groups. Examples include acetyl group.
Compounds of the Invention
The compounds of the invention are simple, easy to use “star burst PET-PEG” dendritic cores.
They are useful for the preparation of dendrimers, thereby providing a facile route to well-defined cluster molecules, e.g. pharmaceutical cluster molecules, as single compounds. The compounds of the invention are two-, three- or four-directional short-armed and long-armed dendritic cores. Some compounds of the invention are “pre-activated” (e.g. those where R5 is a succinimidyl group) so that they can be used, without coupling reagents, for synthesising dendritic compounds. The dendritic core “arms” can be elongated in a controlled manner with suitable linkers, such as 6-8-carbon linkers or PEG linkers (one or two generations), making the “arms” shorter or longer as desired. The linkers are flexible and the tetrahedral model of the core is stable. Compounds of the invention that are succinimidyl esters are, advantageously, crystalline compounds. They are therefore stable and can be easily stored. Conveniently, the compounds of the invention can be purified by flash chromatography. Because of their simplicity of design, the compounds provide a novel approach to reaching larger sized dendrimers, but employing fewer “arms” (2-4 “arms” instead of 32-64), thus simplifying the dendrimer product. Dendritic cores with long carbon or PEG chains can be considered to be mimics of carbohydrate chains, but they avoid the costs and synthetic challenges associated with preparing linear oligosaccharides.
The invention relates to:
and
R1 and R2 are both H; A, E, B and D are CH2 and X is O.
and
Synthesis of the Compounds of the Invention
The compounds of the invention may be prepared by a variety of different methods. The following are representative non-limiting general methods for synthesising compounds of the invention.
Those skilled in the art will realise that Schemes 2 to 7 below show compounds of the invention where R5 is succinimidyl group. However, compounds of the invention where R5 is other than succinimidyl can be prepared analogously. For example, compounds where R5 is H can be prepared by treatment of the OBn precursors (e.g. Schemes 2-7, below) with palladium on carbon or palladium hydroxide on carbon or platinum on carbon catalysts in solvents such as aqueous THF, methanol, ethanol, ethyl acetate, stirred under a hydrogen atmosphere at ambient temperature and pressure or at 5-50 psi, preferably at 5-25 psi.
“Tetrameric” dendritic core compounds of the invention (where R1 and R2 are both a radical of formula (i) or a radical of formula (ii)) are prepared from compound 1, which is synthesised in three steps from pentaerythritol (Scheme 1).
Compound 1 is then converted to compounds of formula (I) via reaction with a suitable amino-substituted carboxylic acid, such as 3-aminopropanoic acid, 4-aminobutanoic acid, 5-aminopentanoic acid, 6-aminohexanoic acid, 7-aminoheptanoic acid, 8-aminooctanoic acid, 9-aminononanoic acid, 10-aminodecanoic acid, 11-aminoundecanoic acid or 12-aminododecanoic acid (Scheme 2).
“Dimeric” dendritic core compounds of the invention where Y is C and R1 and R2 are both H are prepared from 3,3′-(propane-1,3-diylbis(oxy)dipropanoic acid, using a suitable amino-substituted carboxylic acid, such as 3-aminopropanoic acid, 4-aminobutanoic acid, 5-aminopentanoic acid, 6-aminohexanoic acid, 7-aminoheptanoic acid, 8-aminooctanoic acid, 9-aminononanoic acid, 10-aminodecanoic acid, 11-aminoundecanoic acid or 12-aminododecanoic acid (Scheme 3).
“Dimeric” dendritic core compounds of the invention where Y is C; A is (CH2)u; and R1 and R2 are both H are prepared from a suitable diacid, as shown in Scheme 4.
“Dimeric” dendritic core compounds of the invention where Y is O and E is (CH2CH2O)tCH2 are prepared as shown in Scheme 5. Suitable starting materials include, for example, 3,6,9-trioxaundecanedioic acid, 3,6,9,12-tetraoxatetradecanedioic acid or 3,6,9,12,15-pentaoxaheptadecanedioic acid.
“Trimeric” dendritic core compounds of the invention where Y is C and R1 is NHZ are prepared from 2-amino-2-hydroxymethyl-propane-1,3-diol, as shown in Scheme 6. Those skilled in the art will appreciate that trimeric dendritic core compounds where R1 is NH2 can be used to further elaborate the substitution at the R1 position. For example, such compounds can be linked to radio- or fluorescent labels or the amino functionality can be converted to other functional groups.
“Trimeric” dendritic core compounds of the invention where Y is C and R1 is H, CH3 or CH2CH3 are prepared from 2-hydroxymethyl-propane-1,3-diol, 1,1,1-tris(hydroxymethyl)ethane or 1,1,1-tris(hydroxymethyl)propane, respectively (Scheme 7).
Use of the Compounds of the Invention for Preparing Dendrimers
The compounds of the invention are useful for the preparation of dendritic compounds. Conveniently, the compounds of the invention allow for facile synthesis of dendrimers.
Advantageously, with some compounds of the invention there is no need to use coupling reagents and then to remove an excess of reagents from the dendritic products. The coupling procedure is simple and requires a suitable solvent (DMF, DMSO, water, for example), a small amount of base, e.g. triethylamine, and any glycoside with a free amino group (at least about 2 equivalents of glycoside, e.g. about 2.2 equivalents of glycoside are used for coupling with dimeric compounds of the invention, at least about 3 equivalents of glycoside, e.g. about 3.3 equivalents of glycoside are used for coupling with trimeric compounds of the invention and at least about 4 equivalents of glycoside e.g. about 4.4 equivalents of glycoside are used for coupling with tetrameric compounds of the invention).
Using this method, it is possible to attach a variety of carbohydrate fragments (e.g. heparan sulfates, sulfated monosaccharides, oligosaccharides, amino acids, radioligands, imaging agents, fluorescent probes, antibiotics, cytostatic drugs (chemotherapy), veterinary pharmaceuticals, proton and pH sensors, metal ions and ferrocene) with various linkers to the dendritic cores. By way of example, Scheme 8 shows how a “star burst PET-PEG” dendritic cluster glycomimetic of heparan sulfate can be synthesised from a dendritic core of the present invention. This heparan sulfate mimetic is useful as an inhibitor of Alzheimer's β-secretase.
Those skilled in the art will appreciated that, using the tetrameric compounds of the invention, it is possible to achieve dendrimers that are similar size to 16-mer and 32-mer ball-clusters, but which have fewer copies of glycosides attached. Typically, when the use of 16-, 32- and 64-mer dendritic cores is reported, the dendrimer products are described as mixtures with only partial capping of attachment points due to electrostatic repulsion and steric crowding effects. Using the compounds of the invention, with fewer linkers, the glycosides are well separated in space so the capping leads to fully substituted dendrimers. This is advantageous, for example, in the field of pharmaceuticals where it is desirable to produce discrete, well-characterised chemical compounds, rather than mixtures.
NMR Nuclear magnetic resonance
HRMS High resolution mass spectrometry
DCM Dichloromethane
DMF N,N-Dimethylformamide
EtOH Ethanol
MeOH Methanol
THF Tetrahydrofuran
EtOAc Ethyl acetate
RT Room temperature
TAMRA 4-Carboxytetramethylrhodamine N-succinimidyl ester
BODIPY 4,4-Difluoro-4-bora-3a,4a-diaza-s-indacene
EDC 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride
NHS N-Hydroxysuccinimide
The following examples further illustrate the invention. It is to be appreciated that the invention is not limited to the examples.
Preparation of 1
Tetranitrile precursor (Ref. 1, Hukkamaki, J.; Pakkanen, T. T. Journal of Molecular Catalysis A: Chemical 2001, 174, 205-211) is prepared via Michael-type addition of acrylonitrile to pentaerythritol. Acidic hydrolysis of tetranitrile (Ref. 2, Newcombe, G. R.; Mishra, A; Moorfield, C. N. J. Org. Chem. 2002, 67, 3957-3960) furnishes the tetraacid. Tetraacid (1.0 g, 2.35 mmol) is dissolved in dry DMF (15 mL). N-Hydroxysuccinimide (1.62 g, 14.14 mmol) and 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 2.71 g, 14.14 mmol) are added to the reaction mixture at room temperature and stirring continued for 24 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is purified by flash chromatography on silica gel eluting with EtOAc followed by EtOAc:MeOH, 19:1→9:1→7:1→4:1 to give the tetra-succinimidyl ester (1, 1.2 g, 1.48 mmol, 63%). Rf=0.25 (Ethyl Acetate:MeOH, 9:1). 13C-NMR (125 MHz, DMSO-D6) δ 170.7, 170.1, 68.7, 65.5, 44.9, 31.5, 25.7. HRMS calcd for C33H40N4O20Na (M+Na)+ m/z 835.2134. found 835.2128.
Preparation of 2
Aminooctanoic acid is treated with benzyl alcohol in the presence of thionyl chloride (Ref. 3, Patel, R. P; Price, S. J. Org Chem. 1965, 30 (10), 3575-3576) to give a tetra benzyl ester (2 g, 8.12 mmol). This and tetra-succinimidyl ester (1, 1.1 g, 1.35 mmol) are dissolved in a mixture of dry THF (55 mL) and dry DMF (3 mL) and treated with triethylamine (1.5 mL, 10.83 mmol). After stirring for 24 hrs the mixture is diluted with ethyl acetate and washed with water twice, dried over magnesium sulfate and concentrated. The residue is dissolved in hot EtOAc, the crystals are filtered off and discharged. The mother liquor is concentrated and the residue is purified by flash chromatography on silica gel eluting with Chloroform: EtOAc: MeOH, 5:2:0.5 to afford the tetra-benzyl ester (2, 1.7 g, 1.26 mmol, 93%). Rf=0.3 (Chloroform:Ethyl Acetate: MeOH, 5:2:1). 13C-NMR (125 MHz, CDCl3) δ 173.5, 171.2, 136.1, 128.5, 128.2, 128.1, 69.1, 67.4, 66.1, 45.3, 39.5, 36.9, 34.2, 29.6, 29.4, 28.9, 26.7, 26.6, 24.8. HRMS calcd for C77H112N4O16Na (M+Na)+ m/z 1371.7971. found 1371.7977.
Preparation of 3
Tetra benzyl ester (2, 0.595 g, 441 μmol) is dissolved in dry THF (16 mL). Water (4 mL) and glacial acetic acid (5 drops) are added. The reaction mixture is treated with palladium hydroxide on carbon (20% Pd, 1 g) and stirred for 3 hours under hydrogen at ambient temperature and pressure. The catalyst is filtered off and washed with 50% aqueous EtOH. The solution is concentrated to dryness to give a “long-armed” tetraacid (3, 0.42 g, 429 μmol, 97%). The product is used in the next step without further purification. Rf=0.0 (base line, Chloroform:Ethyl Acetate: MeOH, 5:2:1). 13C-NMR (125 MHz, DMSO-D6) δ 174.5, 169.9, 68.8, 67.3, 45.0, 39.0, 38.4, 36.1, 33.8, 29.1, 28.5, 38.4, 26.3, 25.2, 24.5. HRMS calcd for C49H87N4O16 (M−H)− m/z 987.6117. found 987.6110.
Preparation of 4
“Long-armed” tetraacid (3, 424 mg, 429 μmol) is dissolved in dry DMF (7 mL). N-Hydroxysuccinimide (296 mg, 2.57 mmol) and 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 493 mg, 2.57 mmol) are added to the reaction mixture at room temperature and stirring continued for 24 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is purified by flash chromatography on silica gel eluting with Chloroform:Ethyl Acetate:MeOH, 5:2:0.5→5:4:1 to give the “long-armed” tetra-succinimidyl ester (4, 415 mg, 301 μmol, 70.3%). Rf=0.25 (DCM:MeOH, 9:1). 13C-NMR (125 MHz, CDCl3) δ 171.5, 169.4, 168.6, 68.9, 67.4, 45.3, 39.4, 36.7, 30.8, 29.5, 29.4, 28.6, 28.5, 26.6, 25.5, 25.4, 24.4. HRMS calcd for C65H100N8O24Na (M+Na)+ m/z 1399.6748. found 1399.6737.
Preparation of 5
H2N(PEG)3CH2CH2COOH (or PEG aminoacid) (1.0 g, 4.52 mmol) is dissolved in benzyl alcohol (30 mL, 287 mmol) and cooled to 0° C. Thionyl chloride (6 mL, 82.2 mmol) is added slowly dropwise. The reaction mixture is stirred at 0° C. for 15 min followed by heating at 100° C. for 5 hours. Then this is diluted with diethyl ether and the oily residue is collected and purified by flash chromatography on silica gel eluting with Dichloromethane: MeOH, 9:1→1:1 to afford the benzyl ester (5, 1.2 g, 3.9 mmol, 85%). Rf=0.15 (Dichloromethane: MeOH, 9:1). 13C-NMR (125 MHz, CDCl3) δ 171.6, 135.8, 128.5, 128.2, 128.1, 70.2, 70.14, 70.13, 69.9, 66.7, 66.4, 66.3, 50.0, 39.7, 35.0. HRMS calcd for C1-6H26NO5 (M+H)+ m/z 312.1811. found 312.1806.
Preparation of 6
The benzyl ester 5 (65 mg, 210 μmol) and tetra-succinimidyl ester (4, 58 mg, 42.1 μmol) are dissolved in dry DMF (2 mL) and treated with triethylamine (47 μL, 336 μmol). After stirring for 24 hrs the mixture is diluted with ethyl acetate and washed with water twice, dried over magnesium sulfate and concentrated. The residue is dissolved in hot EtOAc, the crystals are filtered off and discharged. The mother liquor is concentrated and the residue is purified by flash chromatography on silica gel eluting with Chloroform:EtOAc:MeOH, 5:2:0.5 to afford the tetra-benzyl ester (6, 71 mg, 32.8 μmol, 78%). Rf=0.3 (Chloroform:Ethyl Acetate: MeOH, 5:2:1). 13C-NMR (125 MHz, CDCl3) δ 173.2, 171.3, 135.8, 128.5, 128.2, 128.1, 70.5, 704, 70.2, 69.9, 69.2, 67.5, 66.5, 66.3, 45.3, 39.4, 39.1, 36.9, 36.5, 35.1, 29.6, 29.1, 29.0, 26.7, 25.5.
Preparation of 7
Tetra benzyl ester (6, 16 mg, 7.21 μmol) is dissolved in dry THF (4 mL). Water (1 mL) and glacial acetic acid (2 drops) are added. The reaction mixture is treated with palladium hydroxide on carbon (20% Pd, 20 mg) and stirred for 3 hours under hydrogen at ambient temperature and pressure. The catalyst is filtered off and washed with 50% aqueous EtOH. The solution is concentrated to dryness to give a “long-armed” PEG tetraacid (7, 13 mg, 7.21 μmol, 97%). The product is used in the next step without further purification. Rf=0.0 (base line, Chloroform:Ethyl Acetate: MeOH, 5:2:1). 13C-NMR (125 MHz, MeOD) δ 176.7, 174.2, 71.4, 71.3, 71.2, 70.6, 68.8, 67.8, 62.8, 48.5, 40.6, 40.3, 37.8, 37.1, 35.8, 30.4, 30.2, 30.1, 30.0, 27.9, 26.9.
Preparation of 8
“Long-armed” PEG tetraacid (7, 13 mg, 7.21 μmol) is dissolved in dry DMF (1 mL). N-Hydroxysuccinimide (5.1 mg, 43.2 μmol), DIPEA (7.6 μL, 43.2 μmol) and 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 8.3 mg, 43.2 μmol) are added to the reaction mixture at room temperature and stirring continued for 24 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is purified by flash chromatography on silica gel eluting with Chloroform:Ethyl Acetate: MeOH, 5:2:0.5→5:4:1 to give the “long-armed” PEG tetra-succinimidyl ester (8, 15 mg, 6.85 μmol, 94%). Rf=0.25 (DCM:MeOH, 9:1).
Preparation of 9
Dodecanedioic acid (1.0 g, 4.34 mmol) is dissolved in dry DMF (10 mL). N-Hydroxysuccinimide (1.51 g, 13.0 mmol, 3 eq.) and 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 2.55 g, 13.0 mmol, 3 eq.) and N,N-diisopropylethylamine (1.53 mL, 8.68 mmol) are added to the reaction mixture at room temperature and stirring continued for 24 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is re-crystalized from hot EtOAc to give the di-succinimidyl ester (9, 1.84 g, 4.2 mmol, 98%). 13C-NMR (125 MHz, DMSO-D6) δ 170.2, 168.9, 30.2, 28.7, 28.4, 27.9, 25.4, 24.2. HRMS calcd for C20H28N2O8Na (M+Na)+ m/z 447.1743. found 447.1743.
Preparation of 10
Di-succinimidyl ester (9, 500 mg, 1.18 mmol) and an amino benzyl ester (881 mg, 3.53 mmol, 3 eq.) are dissolved in dry DMF (8 mL) and treated with triethylamine (0.66 mL, 4.71 mmol, 4 eq.). After stirring for 24 hrs the mixture is concentrated. The residue is dissolved in hot MeOH, a few drop of chloroform added, the crystals are filtered off and dried to afford the di-benzyl ester (10, 0.75 g, 1.1 mmol, 92%). 13C-NMR (125 MHz, MeOD) δ 176.2, 175.5, 137.4, 129.8, 129.4, 129.4, 67.5, 40.6, 37.6, 35.5, 30.7, 30.6, 30.5, 30.2, 30.1, 27.9, 27.2, 26.1. HRMS calcd for C42H64N2O6Na (M+Na)+ m/z 715.4662. found 715.4664.
Preparation of 14
3,6,9-Trioxaundecanedioic acid (500 mg, 2.25 mmol) is dissolved in dry DMF (10 mL). N-Hydroxysuccinimide (785 mg, 6.75 mmol, 3 eq.), 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 1.32 g, 6.75 mmol, 3 eq.) and N,N-diisopropylethylamine (2.38 mL, 13.5 mmol) followed by benzyl 8-aminooctanate (1.68 g, 6.75 mmol, 3 eq.) are added to the reaction mixture at room temperature and stirring continued for 4 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is dissolved in hot EtOAc, the crystals are filtered off and discharged. The mother liquor is concentrated and the residue is purified by flash chromatography on silica gel eluting with Chloroform: EtOAc: MeOH, 5:2:0.3 to afford the di-benzyl ester (14, 1.1 g, 1.61 mmol, 71%). 13C-NMR (125 MHz, CDCl3) δ 173.4, 169.9, 136.1, 128.4, 128.3, 128.1, 127.8, 127.7, 65.9, 63.9, 60.3, 39.5, 36.8, 31.9, 30.9, 29.3, 28.8, 26.6, 25.3, 24.7, 24.4, 21.2, 20.9. HRMS calcd for C38H56N2O9Na (M+Na)+ m/z 707.3884. found 707.3876.
Preparation of 15
Di-benzyl ester (14, 204 mg, 297 μmol) is dissolved in dry THF (8 mL). Water (2 mL) and glacial acetic acid (3 drops) are added. The reaction mixture is treated with palladium hydroxide on carbon (20% Pd, 0.5 g) and stirred for 3 hours under hydrogen at ambient temperature and pressure. The catalyst is filtered off and washed with 50% aqueous EtOH. The solution is concentrated to dryness to give a “long-armed” diacid (15, 150 mg, 297 μmol, 99.8%). The product is used in the next step without further purification. Rf=0.0 (base line, Chloroform:Ethyl Acetate: MeOH, 5:2:1). 13C-NMR (125 MHz, MeOD) δ 178.1, 175.3, 172.9, 72.3, 71.7, 71.6, 40.4, 35.5, 30.9, 30.6, 30.5, 28.3, 26.5. HRMS calcd for C24H44N2O9Na (M+Na)+ m/z 527.2945. found 527.2943.
Preparation of 16
“Long-armed” diacid (15, 150 mg, 297 μmol) is dissolved in dry DMF (4 mL). N-Hydroxysuccinimide (104 mg, 892 μmol), 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC, 174 mg, 892 μmol) and N,N-diisopropylethylamine (0.157 mL, 892 μmol) are added to the reaction mixture at room temperature and stirring continued for 24 hrs. The mixture is diluted with DCM and washed with water, then with diluted HCl and water, dried over magnesium sulfate and concentrated. The residue is purified by flash chromatography on silica gel eluting with Chloroform:Ethyl Acetate: MeOH, 5:2:0.5→5:4:1 to give the “long-armed” di-succinimidyl ester (16, 188 mg, 269 μmol, 90%). Rf=0.25 (DCM:MeOH, 9:1). 13C-NMR (125 MHz, CDCl3) δ 170.9, 169.9, 169.5, 168.5, 70.7, 70.6, 70.5, 70.4, 70.0, 38.7, 31.4, 30.7, 29.4, 28.6, 28.5, 28.4, 26.5, 25.5, 24.4. HRMS calcd for C32H50N4O13Na (M+Na)+ m/z 721.3272. found 721.3259.
Although the invention has been described by way of example, it should be appreciated that variations and modifications may be made without departing from the scope of the invention. Furthermore, where known equivalents exist to specific features, such equivalents are incorporated as if specifically referred to in the specification.
The invention relates to compounds that are useful for the preparation of dendrimer compounds, the use of these compounds for preparing dendrimers and processes for preparing the compounds.
| Number | Date | Country | Kind |
|---|---|---|---|
| 603908 | Nov 2012 | NZ | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/NZ2013/000215 | 11/28/2013 | WO | 00 |
| Publishing Document | Publishing Date | Country | Kind |
|---|---|---|---|
| WO2014/084743 | 6/5/2014 | WO | A |
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| 5104815 | Garner et al. | Apr 1992 | A |
| 6030640 | Shimizu et al. | Feb 2000 | A |
| 20030097019 | Newkome et al. | May 2003 | A1 |
| 20050008571 | Newkome et al. | Jan 2005 | A1 |
| 20100272660 | Malle | Oct 2010 | A1 |
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| 0891875 | Oct 2003 | EP |
| H11152256 | Jun 1999 | JP |
| WO9525736 | Sep 1995 | WO |
| WO0220469 | Mar 2002 | WO |
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| Number | Date | Country | |
|---|---|---|---|
| 20150291522 A1 | Oct 2015 | US |
