IE65159B1 - Solid phase synthesis of peptide alcohols - Google Patents
Solid phase synthesis of peptide alcoholsInfo
- Publication number
- IE65159B1 IE65159B1 IE244391A IE244391A IE65159B1 IE 65159 B1 IE65159 B1 IE 65159B1 IE 244391 A IE244391 A IE 244391A IE 244391 A IE244391 A IE 244391A IE 65159 B1 IE65159 B1 IE 65159B1
- Authority
- IE
- Ireland
- Prior art keywords
- group
- residue
- resin
- acetalised
- peptide
- Prior art date
Links
- 108090000765 processed proteins & peptides Proteins 0.000 title claims description 53
- 238000010532 solid phase synthesis reaction Methods 0.000 title claims description 4
- 150000001298 alcohols Chemical class 0.000 title description 6
- 229920005989 resin Polymers 0.000 claims description 38
- 239000011347 resin Substances 0.000 claims description 38
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 22
- 150000001875 compounds Chemical class 0.000 claims description 20
- 238000000034 method Methods 0.000 claims description 19
- 125000006239 protecting group Chemical group 0.000 claims description 15
- 229920000642 polymer Polymers 0.000 claims description 12
- 125000003158 alcohol group Chemical group 0.000 claims description 11
- -1 formyl-phenyl group Chemical group 0.000 claims description 11
- 238000004519 manufacturing process Methods 0.000 claims description 11
- 125000004036 acetal group Chemical group 0.000 claims description 8
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 8
- 239000004793 Polystyrene Substances 0.000 claims description 7
- 229920002223 polystyrene Polymers 0.000 claims description 7
- 229920003002 synthetic resin Polymers 0.000 claims description 7
- DHKHKXVYLBGOIT-UHFFFAOYSA-N acetaldehyde Diethyl Acetal Natural products CCOC(C)OCC DHKHKXVYLBGOIT-UHFFFAOYSA-N 0.000 claims description 6
- 210000004899 c-terminal region Anatomy 0.000 claims description 6
- 239000000057 synthetic resin Substances 0.000 claims description 6
- 229910052739 hydrogen Inorganic materials 0.000 claims description 5
- 239000001257 hydrogen Substances 0.000 claims description 5
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 5
- 125000003396 thiol group Chemical group [H]S* 0.000 claims description 5
- 230000002378 acidificating effect Effects 0.000 claims description 4
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 3
- 108010016076 Octreotide Proteins 0.000 claims description 2
- 238000005903 acid hydrolysis reaction Methods 0.000 claims description 2
- 125000004435 hydrogen atom Chemical group [H]* 0.000 claims description 2
- 125000002777 acetyl group Chemical class [H]C([H])([H])C(*)=O 0.000 claims 1
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 27
- 239000000203 mixture Substances 0.000 description 21
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Natural products OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 20
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 14
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 12
- 150000001413 amino acids Chemical group 0.000 description 11
- 102000004196 processed proteins & peptides Human genes 0.000 description 11
- 150000003254 radicals Chemical class 0.000 description 11
- 235000001014 amino acid Nutrition 0.000 description 10
- 238000003776 cleavage reaction Methods 0.000 description 8
- 230000007017 scission Effects 0.000 description 8
- DTQVDTLACAAQTR-UHFFFAOYSA-N trifluoroacetic acid Substances OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 8
- 230000015572 biosynthetic process Effects 0.000 description 7
- 238000006243 chemical reaction Methods 0.000 description 7
- 238000003786 synthesis reaction Methods 0.000 description 7
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 6
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 description 6
- 239000002253 acid Substances 0.000 description 6
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 6
- 229920001184 polypeptide Polymers 0.000 description 6
- 235000000346 sugar Nutrition 0.000 description 6
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 6
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 5
- 125000003277 amino group Chemical group 0.000 description 5
- 239000000706 filtrate Substances 0.000 description 5
- NPZTUJOABDZTLV-UHFFFAOYSA-N hydroxybenzotriazole Substances O=C1C=CC=C2NNN=C12 NPZTUJOABDZTLV-UHFFFAOYSA-N 0.000 description 5
- 125000006850 spacer group Chemical group 0.000 description 5
- YEDUAINPPJYDJZ-UHFFFAOYSA-N 2-hydroxybenzothiazole Chemical compound C1=CC=C2SC(O)=NC2=C1 YEDUAINPPJYDJZ-UHFFFAOYSA-N 0.000 description 4
- QOSSAOTZNIDXMA-UHFFFAOYSA-N Dicylcohexylcarbodiimide Chemical compound C1CCCCC1N=C=NC1CCCCC1 QOSSAOTZNIDXMA-UHFFFAOYSA-N 0.000 description 4
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 4
- NQRYJNQNLNOLGT-UHFFFAOYSA-N Piperidine Chemical compound C1CCNCC1 NQRYJNQNLNOLGT-UHFFFAOYSA-N 0.000 description 4
- 150000001241 acetals Chemical class 0.000 description 4
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 4
- 239000007790 solid phase Substances 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- 150000001414 amino alcohols Chemical group 0.000 description 3
- 150000002148 esters Chemical class 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- 230000003647 oxidation Effects 0.000 description 3
- 238000007254 oxidation reaction Methods 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- MUVQIIBPDFTEKM-QWWZWVQMSA-N (2r,3r)-2-aminobutane-1,3-diol Chemical group C[C@@H](O)[C@H](N)CO MUVQIIBPDFTEKM-QWWZWVQMSA-N 0.000 description 2
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 description 2
- OYNIIKHNXNPSAG-UHFFFAOYSA-N 2-(4-formylphenoxy)acetic acid Chemical compound OC(=O)COC1=CC=C(C=O)C=C1 OYNIIKHNXNPSAG-UHFFFAOYSA-N 0.000 description 2
- QTBSBXVTEAMEQO-UHFFFAOYSA-M Acetate Chemical compound CC([O-])=O QTBSBXVTEAMEQO-UHFFFAOYSA-M 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- NQTADLQHYWFPDB-UHFFFAOYSA-N N-Hydroxysuccinimide Chemical compound ON1C(=O)CCC1=O NQTADLQHYWFPDB-UHFFFAOYSA-N 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 2
- 229920002125 Sokalan® Polymers 0.000 description 2
- 238000006359 acetalization reaction Methods 0.000 description 2
- 125000003368 amide group Chemical group 0.000 description 2
- 229960002684 aminocaproic acid Drugs 0.000 description 2
- IJOOHPMOJXWVHK-UHFFFAOYSA-N chlorotrimethylsilane Chemical compound C[Si](C)(C)Cl IJOOHPMOJXWVHK-UHFFFAOYSA-N 0.000 description 2
- 230000008878 coupling Effects 0.000 description 2
- 238000010168 coupling process Methods 0.000 description 2
- 238000005859 coupling reaction Methods 0.000 description 2
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 2
- KXKVLQRXCPHEJC-UHFFFAOYSA-N methyl acetate Chemical compound COC(C)=O KXKVLQRXCPHEJC-UHFFFAOYSA-N 0.000 description 2
- 229920005990 polystyrene resin Polymers 0.000 description 2
- 229910002027 silica gel Inorganic materials 0.000 description 2
- 239000000741 silica gel Substances 0.000 description 2
- 229960001866 silicon dioxide Drugs 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- DEQANNDTNATYII-OULOTJBUSA-N (4r,7s,10s,13r,16s,19r)-10-(4-aminobutyl)-19-[[(2r)-2-amino-3-phenylpropanoyl]amino]-16-benzyl-n-[(2r,3r)-1,3-dihydroxybutan-2-yl]-7-[(1r)-1-hydroxyethyl]-13-(1h-indol-3-ylmethyl)-6,9,12,15,18-pentaoxo-1,2-dithia-5,8,11,14,17-pentazacycloicosane-4-carboxa Chemical compound C([C@@H](N)C(=O)N[C@H]1CSSC[C@H](NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](CC=2C3=CC=CC=C3NC=2)NC(=O)[C@H](CC=2C=CC=CC=2)NC1=O)C(=O)N[C@H](CO)[C@H](O)C)C1=CC=CC=C1 DEQANNDTNATYII-OULOTJBUSA-N 0.000 description 1
- ASOKPJOREAFHNY-UHFFFAOYSA-N 1-Hydroxybenzotriazole Chemical group C1=CC=C2N(O)N=NC2=C1 ASOKPJOREAFHNY-UHFFFAOYSA-N 0.000 description 1
- 125000000022 2-aminoethyl group Chemical group [H]C([*])([H])C([H])([H])N([H])[H] 0.000 description 1
- KJJPLEZQSCZCKE-UHFFFAOYSA-N 2-aminopropane-1,3-diol Chemical group OCC(N)CO KJJPLEZQSCZCKE-UHFFFAOYSA-N 0.000 description 1
- JMTMSDXUXJISAY-UHFFFAOYSA-N 2H-benzotriazol-4-ol Chemical compound OC1=CC=CC2=C1N=NN2 JMTMSDXUXJISAY-UHFFFAOYSA-N 0.000 description 1
- SLXKOJJOQWFEFD-UHFFFAOYSA-N 6-aminohexanoic acid Chemical compound NCCCCCC(O)=O SLXKOJJOQWFEFD-UHFFFAOYSA-N 0.000 description 1
- DHMQDGOQFOQNFH-UHFFFAOYSA-M Aminoacetate Chemical compound NCC([O-])=O DHMQDGOQFOQNFH-UHFFFAOYSA-M 0.000 description 1
- 101800001415 Bri23 peptide Proteins 0.000 description 1
- 125000001433 C-terminal amino-acid group Chemical group 0.000 description 1
- 102400000107 C-terminal peptide Human genes 0.000 description 1
- 101800000655 C-terminal peptide Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 description 1
- GUBGYTABKSRVRQ-PICCSMPSSA-N D-Maltose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 1
- 102000038461 Growth Hormone-Releasing Hormone Human genes 0.000 description 1
- 239000000095 Growth Hormone-Releasing Hormone Substances 0.000 description 1
- 101800000736 Growth hormone-releasing factor Proteins 0.000 description 1
- AVXURJPOCDRRFD-UHFFFAOYSA-N Hydroxylamine Chemical compound ON AVXURJPOCDRRFD-UHFFFAOYSA-N 0.000 description 1
- 239000004698 Polyethylene Substances 0.000 description 1
- RHQDFWAXVIIEBN-UHFFFAOYSA-N Trifluoroethanol Chemical compound OCC(F)(F)F RHQDFWAXVIIEBN-UHFFFAOYSA-N 0.000 description 1
- 230000021736 acetylation Effects 0.000 description 1
- 238000006640 acetylation reaction Methods 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 125000003172 aldehyde group Chemical group 0.000 description 1
- 125000002947 alkylene group Chemical group 0.000 description 1
- 125000004202 aminomethyl group Chemical group [H]N([H])C([H])([H])* 0.000 description 1
- 239000012062 aqueous buffer Substances 0.000 description 1
- UORVGPXVDQYIDP-UHFFFAOYSA-N borane Chemical class B UORVGPXVDQYIDP-UHFFFAOYSA-N 0.000 description 1
- 125000004744 butyloxycarbonyl group Chemical group 0.000 description 1
- 125000004432 carbon atom Chemical group C* 0.000 description 1
- 125000005521 carbonamide group Chemical group 0.000 description 1
- PFKFTWBEEFSNDU-UHFFFAOYSA-N carbonyldiimidazole Chemical compound C1=CN=CN1C(=O)N1C=CN=C1 PFKFTWBEEFSNDU-UHFFFAOYSA-N 0.000 description 1
- 150000001732 carboxylic acid derivatives Chemical class 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000007859 condensation product Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 125000000151 cysteine group Chemical group N[C@@H](CS)C(=O)* 0.000 description 1
- LLHRMWHYJGLIEV-UHFFFAOYSA-N desoxy Chemical group COC1=CC(CCN)=CC(OC)=C1C LLHRMWHYJGLIEV-UHFFFAOYSA-N 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000032050 esterification Effects 0.000 description 1
- 238000005886 esterification reaction Methods 0.000 description 1
- 125000000373 fatty alcohol group Chemical group 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 125000004029 hydroxymethyl group Chemical group [H]OC([H])([H])* 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 108091005601 modified peptides Proteins 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 229960002700 octreotide Drugs 0.000 description 1
- 238000010647 peptide synthesis reaction Methods 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000573 polyethylene Polymers 0.000 description 1
- 239000002952 polymeric resin Chemical group 0.000 description 1
- 229920001296 polysiloxane Polymers 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 239000012262 resinous product Substances 0.000 description 1
- 238000004007 reversed phase HPLC Methods 0.000 description 1
- 229910000033 sodium borohydride Inorganic materials 0.000 description 1
- 239000012279 sodium borohydride Substances 0.000 description 1
- 229910000029 sodium carbonate Inorganic materials 0.000 description 1
- NHXLMOGPVYXJNR-ATOGVRKGSA-N somatostatin Chemical class C([C@H]1C(=O)N[C@H](C(N[C@@H](CO)C(=O)N[C@@H](CSSC[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC=2C=CC=CC=2)C(=O)N[C@@H](CC=2C=CC=CC=2)C(=O)N[C@@H](CC=2C3=CC=CC=C3NC=2)C(=O)N[C@@H](CCCCN)C(=O)N[C@H](C(=O)N1)[C@@H](C)O)NC(=O)CNC(=O)[C@H](C)N)C(O)=O)=O)[C@H](O)C)C1=CC=CC=C1 NHXLMOGPVYXJNR-ATOGVRKGSA-N 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- HNKJADCVZUBCPG-UHFFFAOYSA-N thioanisole Chemical compound CSC1=CC=CC=C1 HNKJADCVZUBCPG-UHFFFAOYSA-N 0.000 description 1
- JOXIMZWYDAKGHI-UHFFFAOYSA-N toluene-4-sulfonic acid Chemical compound CC1=CC=C(S(O)(=O)=O)C=C1 JOXIMZWYDAKGHI-UHFFFAOYSA-N 0.000 description 1
- TUQOTMZNTHZOKS-UHFFFAOYSA-N tributylphosphine Chemical compound CCCCP(CCCC)CCCC TUQOTMZNTHZOKS-UHFFFAOYSA-N 0.000 description 1
- 239000005051 trimethylchlorosilane Substances 0.000 description 1
- 125000000026 trimethylsilyl group Chemical group [H]C([H])([H])[Si]([*])(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- FTVLMFQEYACZNP-UHFFFAOYSA-N trimethylsilyl trifluoromethanesulfonate Chemical compound C[Si](C)(C)OS(=O)(=O)C(F)(F)F FTVLMFQEYACZNP-UHFFFAOYSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Landscapes
- Peptides Or Proteins (AREA)
Description
- 1 651 59 SOLID PHASE SYNTHESIS OF PEPTIDE ALCOHOLS The present invention relates to a process for producing in solid phase a peptide alcohol which at the C-terminal end of the peptide chain bears two alcohol groups or one alcohol group and one thiol group.
Patent Application No. 2724/87/ rela^t’es^to α-glucosy1(1-4)I-1 deoxyfructosyl]-DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr-ol. . . _ A ,. . „ 9,,9/Q1 ,(GB-2.,199,831A) u .
Divisional Application No. £44Z/yi/ provides a sugar derivative of a biologically active peptide, which derivative has a prolonged duration of action when compared to the non-sugar modified peptide and contains at least on one of the amino acid units a sugar residue which is attached to an amino group thereof by a coupling other than a direct N-glycosidic bond, and additionally, when it is a condensation product of a carboxyl group containing sugar and a peptide with less than 8 amino acid units, by a coupling other than a direct amide bond, the sugar peptide derivative being other than N"-[a-glucosyl(l-4)-deoxyfructosyl]DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr-ol.
The compounds of these applications may be conveniently prepared by solid phase synthesis.
We have found an especially convenient process for the production of peptide alcohols, which at the C-terminal end of the peptide chain bear two alcohol groups or one alcohol group and one thiol group. The process is especially suitable for the production of peptide alcohols which contain a C-terminal threoninol, serinol or cysteinol radical.
Examples of suitable compounds include some of the somatostatin compounds described herein.
Solid-phase peptide synthesis has proved to be an especially rapid and favourable process for the production of peptides, and has therefore become a generally conventional method. - 2 65159 As is known, first of all an amino acid is bonded by its carhoxyl group, forming an ester or amide group, to a hydroxyl group or amino group of an insoluble synthetic resin; then, the further amino acids are added onto this in the desired sequence, and finally the complete polypeptide is cleaved from the carrier resin.
This synthesis operates without problems for normal polypeptides having C-terminal amino acids. However, polypeptide alcohols, which at the C-end bear an amino-alcohol instead of an amino acid, do not easily form a bond with carrier resins bearing OH- or NHg- groups and/or are not so easily cleaved again when synthesis has ended.
The following have previously been proposed as possible solid-phase processes for the production of peptide alcohols : a) conventional preparation of the corresponding polypeptide containing at the C-end an amino acid (as the ester of a resin bearing OH groups) and subsequent cleavage by reduction, using boron hydrides, the carboxyl group being simultaneously converted into an alcohol function. (US Patents 4,254,023/4). b) Addition of the terminal amino alcohol as ether to a hydroxymethyl resin, using carbonyl diimidazole, and finally, after synthesis of the peptide, cleavage using HC’l/TFA or HBr/TFA (Kun-hwa Hsieh and G.R.Marshall, ACS National Meeting, New Orleans, 21-25. 3. 1977).
However, these methods both require drastic cleavage conditions , We have found that the cleavage of the peptide from the resin, whilst simultaneously forming the C-terminal peptide alcohol, is carried out under mild conditions if the C-terminal amino-alcohol is joined to the resin by an acetal bond.
In accordance with the invention, the peptide alcohol which at the C-terminal end of the peptide chain bears 2 alcohol groups or one alcohol group and one thiol group is produced by acid hydrolysis of an acetal of the peptide alcohol and a polymer resin bearing formylphenyl groups. This is referred to as the synthesis of the invention.
The reaction may be illustrated schematically as follows:- CHO CHR,-XH « 1 * Y-CO-NH-CH-CHjOH wherein (p) is the residue of an insoluble synthetic resin Z is a direct bond or a residue which joins the resin with the (acetalised) foraylphenyl group X is 0 or S R^ is hydrogen or methyl, and Y is the residue of a peptide alcohol which e.g. may bear protecting groups, wherein the optionally acetalised CHOr· group is located in the m- or p-position to the radical Z.For simplicity, in formulae I and IX of the above scheae, only one substitution group was indicated on the resin; it should however be clear that a number of such groups are bonded to a molecule of the resin polymer. Cleavage of the peptide alcohol from the resin by hydrolysis of the acetal group takes place as mentioned above under acidic conditions, e.g. with diluted trifluoroacetic acid. Hydrolysis can be effected at room temperature.
If Z in formula is a direct bond, the phenyl radicals bearing acetal groups are directly bonded to the polymer residue belong to the polymer. Examples o£ such compounds of formula Irare the acetals of a formylated polystyrene resin (in formula I.r, (p) is then a polyethylene chain) .
If Z is a residue, then this residue contains a group which is the result of a reaction of a reactive group, that is directly or indirectly bonded to the polymer, with another reactive group, that is directly or indirectly bonded to the (acetalised) formylphenyl group. The radical Z may be represented for example by the following formula IIIr; -(D)p-Q1-Q2-(E)q- Illf wherein = the residue of a reactive group which is bonded to the polymer Q2 « the residue of a reactive group which is bonded to the (acetalised) formylphenyl group D » a residue which joins the group with the polymer E = a .resιdue which joins the group Q2 with the (acetalised) formylphenyl group p and q, independently of one another, are 0 or 1.
The Q^-Q2 9rouP is preferably an ester or amide group, especially a carbonamide group. is preferably NH and Q2 is preferably CO. and E, independently of one another, are for example alkylene or alkyleneoxy radicals having 1 to 5 C-atoms. Examples of such compounds of formula Ir, wherein Z is a residue of formula IIlr,are compounds wherein ®-D-Q^ is the residue of an aminomethylated polystyrene resin and the residue *1 - X-CH CH ‘CH-NH-CO-Y Q,-E O-CH. is a residue of formula IV r /TVCH CH-NH-CO-Y -C-CH-(0^/07^ / - · o-ch; OR 2 wherein r = hydrogen or methyl and m ® 0 or 1, whereby the acetal group is again located in ra- or p-position.
In this case, Z is thus -CH,-NH-CO-CH-(O) - and • I Hl R ® is polystyrene.
Radical IVris preferably S-y /X-CHR1 -co-ca2-o-/o>-ca ^ch-nh-co-y Instead of the arainoraethylated polystyrene, other polymers can also be used, especially those having free NH2 groups, e.g. polyacrylamides bearing aminoethyl groups.
As mentioned above, the acetalised forayIphenyl group is preferably bonded to the polyraer by an amide bond. This ensures that the bonding of the acetalised foraylphenyl radical to the resin during synthesis of the polypeptide and during cleavage is stable, and that cleavage occurs on the acetal bond as desired, so that on the one hand the peptide alcohol is generated and on the other hand the formylphenyl radical remains on the resin.
If desired, the peptide alcohol can be attached further away from the resin by incorporating so-called spacers between the reactive groups of the polymer (especially amino groups) and the reactive groups of the acetalised formylphenyl derivative (especially carboxyl groups). For certain reactions on the polypeptide alcohol, this may advantageously be before cleavage (e.g. oxidation of cystein radicals). In this case, the radical D or E in formula II/ additionally contains the spacer and or Q2 is the reactive radical of the spacer.
The spacer used can be for example a <*>-aminocar boxy lie acid, such as £-aminocaproic acid.
In a specific case, when using aminomethylated polystyrene, a radical of formula IVY and £-aminocaproic acid as the spacer, Z is -CH2-NH-CO-(CH2)g-NH-CO-CH-(0)mR The compounds of formula can be produced by methods which are usual in solid-phase technology, starting with a compound of formula Vr’l ^zX-CH o-ch2 •CH-NH-A wherein A is a- protecting group of the amino function and the acetal group is in m- or p-position to the radical Z.
For this purpose, first of all the protecting group A is cleaved and then the free amino group is reacted with the next N-protected amino acid etc., until all the amino acids have been added onto the resin in the sequence corresponding to the desired peptide alcohol.
The amino protecting groups to be chosen for the amino acids used or for the amino alcohol must be those which are cleaved under non-acidic conditions, since under acidic conditions 5 hydrolysis of the acetal group takes place. The CF^CO- or the FMOC- group (9-fluorenylmethyloxycarbonyl) can be used e.g. as such amino protecting groups. These protecting groups are cleaved in a basic medium in a manner which is usual for peptide chemistry.
Only protecting groups in the side chains and the amino protecting group of the last administered amino acid may be acid labile and then are simultaneously split off from the resin with the regeneration of the peptide alcohol .
Preferably the Boc groups present as a protecting group.
As bases are preferably used koh or piperidine or NaBfy The building up of the peptide chain may be effected in conventional manner from a peptide moiety having free amino groups and an amino acid with free or activated carboxyl groups.
The reaction may be effected with the addition of e.g. hydroxybenzotriazol and dicyclohexylcarbodiimide.
Compounds of formula Vr may be produced for example by a) reacting a resin carrying an aldehyde group of formula IIr "r wherein the CHO group is in the m or p position to the Z substituent, with an N-protected amino alcohol of formula HX-CHR1-CH(NHA)-CH2OH optionally in activated form, or b) reacting a resin having the formula with a compound of formula VIr CH-HH-A VIr wherein the acetal group is in the m orp position to the "(E) - group and Q-j and are two reactive groups which react together to form a Q-j - Qg bridge.
The acetalization of process a) may be effected in the presence of an acid as catalyst. Suitable acids include p-toluene sulphonic acid and p-trifluoromethylsulphonic acid.
If desired a trimethylsilyl group may be used as a protecting group for a free alcohol.
The esterification process b) may be effected under very mild conditions, e.g. by reaction of a carboxylic acid derivative with an OH or NH2 group carrying polymer.
The compounds of formula VIr may be produced by acetylation of a compound of formula with a compound of formula HX-CHR^CHiNHAJ-CHgOH The acetalisation may be effected as for process a).
During the building up and the splitting off of the 5 peptide alcohol from the resin, further reactions may be effected, e.g. removal of protecting groups, e.g. S-protecting groups, or oxidation of cystein radicals.
Such reactions may be effected after splitting off of the peptide alcohol in the liquid phase.
According to the synthesis of the invention pharmacologically active and other peptides which on the C-end contain 2 alcohol groups or an alcohol and a thiol group may be simply produced.
In the following examples, all temperatures are given in 20 degrees Celsius and the [α]θ values are uncorrected. The following abbreviations are used:AcOH = acetic acid Boc = tert, butyloxycarbonyl Bu^ = tert, butyl DCCI = dicyclohexylcarbodiimide DMF = dimethylformamide Fmoc = 9-fluorenylmethoxycarbonyl MeOH = methanol NEt^ = triethylamine Thr-ol= threoninol radical = CH^-CHOH-CHiCH^OHJ-NHTFA = trif1uoroacetic acid HOBT = N-hydroxybenzotriazole hpGRF = human pancreatic growth hormone releasing factor HOSu N-hydroxy-succinimide All peptides are obtained as polyacetates-polyhydrates with a peptide content of 70 to 90%.
HPLC analysis shows that the peptides contain less than 5% of other peptides.
The factor F mentioned in the following examples shows the peptide content in the products obtained (F = 1 conforms with 100% peptide content). The difference up to 100% C(l-F) x 100] consists of acetic acid and water.
All sugars have the α-configuration unless otherwise stated. Deoxy = Desoxy.
The synthesis of the invention may be effected as follows:EXAMPLE SI Production of octreotide (= SMS) 1) Production of acetal anchor (n-CF3C0-Threonino! acetal of p-formylphenoxy-acetic acid). 105 g (1.0 mmol)L-Threoninol is added to 200 ml methanol which is stirred by a stream of nitrogen. A clear solution « results. A solution of 200 ml tri fl uoro. acetic acid methyl ester in 250 ml methanol is added to the mixture at 0°. The mixture is maintained at a temperature of about 10°C by cooling with an ice bath.
After 1.5 hours no more free Theeoninol is detectable in the mixture. Concentration at 40°C gives a white crystalline resi due.
The residue is dissolved in 200 ml ethyl acetate at 70°C and precipitated by the addition of hexane. The mixture is cooled to 0°C, washed with hexane and dried at room temperature. N-trifluoroacetyl threoninol results. 50.3 g (0.25 mol) of the resultant product is dissolved in 1.25 litres tetrahydrofuran and 75 ml of trimethylchlorosilane is added dropwise. Immediately thereafter a mixture of 70ml triethylamine and 250 ml tetrahydrofuran is added. A white suspension results which is stirred for 4 hours. The mixture is filtered and the filtrate evaporated at 40°C to give an oil.
The oil is dissolved in 1.5 litres of methylene chloride and treated with portions of 90.4 g p-formyl-phenoxyacetic acid at room temperature. Portions of 9 ml trifluoromethane-sulphonic acid trimethylsilyl ester are added. The mixture is stirred for 24 hours at room temperature, then filtered and the residue is washed well with methylene chloride.
The filtrate is concentrated at 40°C to give an orange red resinous product. This product is chromatographed over silicagel. Elution is effected with ethyl acetate. On concentration of the relevant fractions the heading compound is obtained with a purity of 97% (HPLC). 2) Building up of the protected octa-peptide 17.2 g ami nomethylated polystyrene (Brand Dow 0.7% by weight of N correspond!ng to 0.5 mmol amino-methyl groups per g resin) are suspended in 80 ml methylene chloride/ DMF 4:1. Successively there are added 4.17 g of the end product of step 1, 1.6 g HOBT and 4.0 g DCCI. After the mixture is stirred for 2 hours at room temperature, the Kaiser test is negative. The mixture is filtered and washed.
The washed resin is suspended in 100 ml tetrahydrofuran and methanol· 3:1 and treated with portions of 10.4 g sodium borohydride. The mixture is stirred for 6 hours at room temperature, filtered and the resin washed.
The resin is suspended in methylene chloride/DMF 4.1. .5 g Fmoc-Cys (S-t-Bu)OH, 1.74 g HOBT and 3.6 g DCCI are added. The Fmoc protecting group is split off with piperidine (2 x 20 minutes contact time).
In.analogous manner in successive cycles the following N-Fmoc protected amino acids ar° coupled using HOBT/DCCI-. ThrOH; Lys (BOC)-Otts O-TrpOH,Phe-OH,Cys(S-tBu)OH and D-Phe-OH to give the Fmoc protected octapeptide resin.
Final loading 0.26 mmol/g. 3) Oxidation and splitting off The resultant resin is suspendedin 100 ml trifluoroethanol /methy 1 ene chloride 1:1 and treated with 50 ml tri butylphosphine. The mixture is stirred for 70 hours at room temperature. The mixture is filtered, washed and treated with a 100 ml 1:1 mixture of tetrahydrofuran and IN aminoacetate solution. 1.1 ml of 30% aqueous hydrogen peroxide are added. The mixture is stirred for 24 hours at room temperature. The resin is washed. The mixture is treated with 20 ml trifluoroacetic acid, 80 ml methylene chloride, 10 ml water, and 2 ml thioanisole. The mixture is stirred for 2 hours, then filtered and washed with trifluoroacetic acid and methylene chloride. 200 ml diethyl ether are added to the filtrate. The resultant precipitate is filtered off.
The residue is dissolved in aqueous buffer and demineralised, e.g. using Duolite. The solution is freeze-dried as the acetate to give the title compound as the acetate. All the examples of the above mentioned parent and divisional application, e.g. the compounds of examples 1 and 2 may be produced in analogous manner.
EXAMPLE S2: Production of Na-[α-glucosyl(1-4)-deoxy- j- --<-, Xeh£ ί 12 Σ L· e Z ί y Σ ζ Σ 2Σ L Ε Σ Ly 5 Σ Σ L· Γ Σ 2 V s - T h r - o 1 f 393 g of the octapeptide bonded to the resin are produced according to the above example S2. The cysteine protecting groups are removed reductively. The peptide bond to the resin is oxidized to the cyclic octapeptide by hydrogen peroxide in a mixture of tetrahydrofuran/water.
After washing in tetrahydrofuran and then DMF the peptide resin is shaken in 3600 ml of a mixture of DMF/ACOH .(8:1) The suspension is treated with 526 g D( + )maltose mono·.· hydrate. The mixture is warmed to 60° and stirred for 18 hours at this temperature.
The mixture is cooled and the peptide resin filtered off, and successively washed with DMF and methanol .
Then it is washed with methylene chloride. The peptide is then split over 1 hour from the resin with a mixture of 2900 ml methylene chloride and 716 ml trif1uoroacetic acid with a trace of water.
The filtrate is then stirred and treated with portions of 597 g sodium carbonate, stirred for 30 minutes and filtered.
The residue is washed with methylene chloride and methanol.
The filtrate is concentrated to dryness.
It is demineralised using an unfunctionali2ed polystyrene column like Duolite, or reversed phase HPLC material such as silicagel treated with silicone and bearing long chain fatty alcohol groups (e.g. Labomatic, Switzerland, Brand HB-SIL-18-20-100). The pure title compound is obtained.
Claims (12)
1. A process for the production of a peptide alcohol which at the C-terminal end of the peptide chain bears two alcohol groups or one alcohol group and one thiol group, characterised in that the peptide alcohol is produced by acidic hydrolysis of an acetal of the peptide alcohol and a resin containing formylphenyl residues.
2. A process according to claim 1 for the production of a peptide alcohol of formula CHR.-XH Y-CO-NH-CH 1 \ch 2 oh wherei n Y is the residue of a peptide alcohol bearing optionally protecting groups, R1 is hydrogen or methyl, and X i s 0 or S, characterised in that a resin of formula Ir which bears the peptide alcohol, wherein (?) is the residue of an insoluble synthetic resin, and Z is the direct bond or a residue which joins the resin with the acetalised formylphenyl group, wherein the acetalised CHO-group is located in m- or p-position to the radical Z and a number of acetalised formylphenyl groups are coupled to a molecule of the resin polymer, is hydrolysed under acidic conditions. Process according to claim 2 .wherein is a polystyrene radical.
3. 4. Process according to claim 2 or 3, wherein Z is a group of formula -(0) p -0 1 -Q 2 -(E) q wherei n 10 = the residue of a reactive group which is bonded to the resin Q2 = the residue· of a reactive group which is bonded to the acetalised formyl-phenyl group D = a residue which joins the group with the 15 polymer E = a residue which joins the group Q2 with the acetalised formylphenyl group p and q, independently of one another, are 0 or 1 .
4. 5. Process according to any one of claims 2 to 4,wherein
5. 6. 6.
6. 7. Z is the group -CH.-NH-CO-CH-(O) 2 ' 'm 7.
7. 8. 8. R wherein R = H or CH 3 m = 0 or 1 and © 1S a polystyrene residue. Process according to any one of claims 2 to 5 , wherein X is 0 and R^ is CH^. A process for the production of a peptide alcohol substantially as herein before described with reference to any one of the examples and S 2 · A compound whenever produced by a process of any one of claims 1 to 7.
8. 9. 9. Compounds of formula Ir where i n (?) is the residue of an insoluble synthetic resin Z is a direct bond or a residue which joins the resin with the acetalised formylphenyl group is hydrogen or methyl, and Y is the residue of a peptide alcohol which may bear protecting groups wherein the acetalised CHO-group is located in mor p-position to the radical Z and a number of (acetalised) formylphenyl groups are bonded to a molecule of the resin polymer.
9.
10. Compounds of formula Vr Vr wherein (?) is the residue of an insoluble synthetic resin Z is the direct bond or a residue which joins the resin with the acetalised formylphenyl group 5 X is 0 or S R 1 is hydrogen or methyl and A is a protecting group of the amino function, and the acetal group is located in m- or p-position to the residue· 2, and the resin-bears a number of acetalised 10 formylphenyl groups.
11. Compounds of formula VIr X-CH /—>- CH CH-NH-A 2 q \_y O—CH VIr wherein X is 0 or S 5 R 1 is H or CH 3 A is a protecting group of the amino function E is a residue which joins the group Q 2 with the acetalised formylphenyL group °2 is a reactive group which can react with another reactive group that is bonded to a synthetic resin, and is 0 or 1 whereby the acetal group is located in ra- or p-position to the QI-(E) - radical 2 q
12. .. 14. A solid phase synthesis according to any one of claims 1 to 7, for the production of octreotide Octreotide produced by the process according to any one of claims 1 to 7.
Applications Claiming Priority (6)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE3634797 | 1986-10-13 | ||
DE3634826 | 1986-10-13 | ||
DE3634825 | 1986-10-13 | ||
DE3712626 | 1987-04-14 | ||
CH315387 | 1987-08-17 | ||
IE272487A IE872724L (en) | 1987-10-12 | 1987-10-12 | Peptides and intermediates for their preparation |
Publications (2)
Publication Number | Publication Date |
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IE912443L IE912443L (en) | 1988-04-13 |
IE65159B1 true IE65159B1 (en) | 1995-10-04 |
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Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
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IE244391A IE65159B1 (en) | 1986-10-13 | 1987-10-12 | Solid phase synthesis of peptide alcohols |
Country Status (1)
Country | Link |
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IE (1) | IE65159B1 (en) |
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1987
- 1987-10-12 IE IE244391A patent/IE65159B1/en not_active IP Right Cessation
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