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Development of biologically active N-terminally substituted, multi-tyrosinated somatostatin analogs
Abstracts from the l l th International Symposium on Regulatory Peptides
DEVELOPMENT OF A NOVEL SOMATOSTATIN ANALOGUE WITH HIGH AFFINITY FOR HUMAN sstl AND sat2 MS O'Dorisio', F Chen', A Albers', EA Woltering =, TM O'Dorisio~, W Murphy=, D Coy2 'The Ohio State University & Children's Hospital Research Foundation, =Louisiana State University, Tulane University Recent studies which have examined the expression of somatostatin receptor subtypes in various tumors have indicated the presence of sstl as well as sst2 in the majority of tumors. Additionally, recent clinical studies using '"ln-pentatreotide to image tumors have documented a false negative rate of 10-20%. A somatostatin analogue which binds with high affinity to both sstl and sst2 may greatly increase the sensitivity of somatostatin receptor based scintigraphy. We have designed and synthesized a series of multi-tyrosinated somatostatin analogues which are pharmacologically and functionally active. The affinity of each compound for human somatostatin receptors subtypes 1 and 2 was examined in competitive binding to viable human neuroblastoma cell lines which had been transfected with human sstl or sst2. Affinities were estimated using LIGAND. Peptide SS. CH275 BIM23052 WO,C3b WOC4 Octreotide
Affinity of Somatoatatln Analogues for SST1 and SST2 SST1 Kn (nM) 3.3 + 0.3 10.2 + 0.7 17.5 _+2.6 35 +. 20 104 .t; 34 > 1000
SST2 Kn(nM) 8.1 + 3.6 > 1000 ND 35.0 11.7 7.6 _+7.8
WOC3b, an N-terminally extended, cyclic analogue of somatostatin bound with high affinity to both sstl and sst2. This analogue has the additional advantage of possessing 3 tyrosines which allows for labeling of the compound to high specific activity using either ~=al-or '=1-. Use of this analogue in somatostatin receptor based imaging may result in greater sensitivity for detection of somatostatin receptor positive tumors.
DEVELOPMENT OF BIOLOGICALLY ACTIVE N-TERMINALLY SUBSTITUTED, MULTI-TYROSINATED SOMATOSTATIN ANALOGS. TM O'Dorisio. MS O'Dorisio, F Cben, WA Murphy, DH Coy, EA Wultering. The Ohio State University, Dept of Internal Medicine, Columbus, OH 43210; Children's Hospital, Dept of Pediatrics, Columbus, OH 43205; Tulane University, New Orleans, LA 70112; Louisiana State University, Dept of Surgery, New Orleans, LA 70112. Mono-t,fosh~ed somatostatin analogs have been used to inhibit peptide release and have been used as substrate for radiolabelling in auton~liography,scintigraphy, and radio-peptide guided surgery. The clinical utility of these radiolabelled compotmds has been limited by their low specific activity (SA) (approximately 2,000 Ci/mM). We Imvedeveloped a series of N-terminally substituted mnhi-tyrnsinated somatostatin-like peptides that demonstrate high binding aff'mities for the somatostatin receptor and maintain high biologic activity. Peptides were solid-phase synthesized and their bindh~g affinity (KD, riM) tested on DriP-,-32membranes. Inhibition of growth hormone (IC~0, ~flVl)was determined in four- to five-day-old prim,'uy rat pituitary cultures. These peptides were tested for antagonism of VIP~nerated cAMP production in neuroblastoma cells. Compound (# Tyrosines)
Somatostatin Rec Binding KD 0dvl) 4- SD
OH ICso (nM) + SEM
hu'aibition of VIPstimulated cAMP production*
Theoretic SA (Ci/mM) Range
Somatostatin
0.23 4- 0.15
0.46 + 0.04
.22 + .07
2,000
WOC -2b
1.22 + 0.08
0.59 4 0.11
.25 + .06
4,000
WOC-3b
1.14 + 0.28
0.66 + 0.06
.25 + .06
6,000
WOC~a
0.68+0.19
0.37 4- 0.12
Not done
8,000
2.46 + 0.69 .25 4- .06 16,000 0.69 + 0.42 WOC -8 *Basal = .08 + .04 and VIP stim = .50 + .15 ,mol cAMP]'2x 1(P cells. These compounds mayprovide clitficallyuseful radiolabelled analogs for in vivo scintigraphy and in situ radiodetection and radiotherapy.
145
DEVELOPMENT OF A NOVEL SOMATOSTATIN ANALOGUE WITH HIGH AFFINITY FOR HUMAN sstl AND sat2 MS O'Dorisio', F Chen', A Albers', EA Woltering =, TM O'Dorisio~, W Murphy=, D Coy2 'The Ohio State University & Children's Hospital Research Foundation, =Louisiana State University, Tulane University Recent studies which have examined the expression of somatostatin receptor subtypes in various tumors have indicated the presence of sstl as well as sst2 in the majority of tumors. Additionally, recent clinical studies using '"ln-pentatreotide to image tumors have documented a false negative rate of 10-20%. A somatostatin analogue which binds with high affinity to both sstl and sst2 may greatly increase the sensitivity of somatostatin receptor based scintigraphy. We have designed and synthesized a series of multi-tyrosinated somatostatin analogues which are pharmacologically and functionally active. The affinity of each compound for human somatostatin receptors subtypes 1 and 2 was examined in competitive binding to viable human neuroblastoma cell lines which had been transfected with human sstl or sst2. Affinities were estimated using LIGAND. Peptide SS. CH275 BIM23052 WO,C3b WOC4 Octreotide
Affinity of Somatoatatln Analogues for SST1 and SST2 SST1 Kn (nM) 3.3 + 0.3 10.2 + 0.7 17.5 _+2.6 35 +. 20 104 .t; 34 > 1000
SST2 Kn(nM) 8.1 + 3.6 > 1000 ND 35.0 11.7 7.6 _+7.8
WOC3b, an N-terminally extended, cyclic analogue of somatostatin bound with high affinity to both sstl and sst2. This analogue has the additional advantage of possessing 3 tyrosines which allows for labeling of the compound to high specific activity using either ~=al-or '=1-. Use of this analogue in somatostatin receptor based imaging may result in greater sensitivity for detection of somatostatin receptor positive tumors.
DEVELOPMENT OF BIOLOGICALLY ACTIVE N-TERMINALLY SUBSTITUTED, MULTI-TYROSINATED SOMATOSTATIN ANALOGS. TM O'Dorisio. MS O'Dorisio, F Cben, WA Murphy, DH Coy, EA Wultering. The Ohio State University, Dept of Internal Medicine, Columbus, OH 43210; Children's Hospital, Dept of Pediatrics, Columbus, OH 43205; Tulane University, New Orleans, LA 70112; Louisiana State University, Dept of Surgery, New Orleans, LA 70112. Mono-t,fosh~ed somatostatin analogs have been used to inhibit peptide release and have been used as substrate for radiolabelling in auton~liography,scintigraphy, and radio-peptide guided surgery. The clinical utility of these radiolabelled compotmds has been limited by their low specific activity (SA) (approximately 2,000 Ci/mM). We Imvedeveloped a series of N-terminally substituted mnhi-tyrnsinated somatostatin-like peptides that demonstrate high binding aff'mities for the somatostatin receptor and maintain high biologic activity. Peptides were solid-phase synthesized and their bindh~g affinity (KD, riM) tested on DriP-,-32membranes. Inhibition of growth hormone (IC~0, ~flVl)was determined in four- to five-day-old prim,'uy rat pituitary cultures. These peptides were tested for antagonism of VIP~nerated cAMP production in neuroblastoma cells. Compound (# Tyrosines)
Somatostatin Rec Binding KD 0dvl) 4- SD
OH ICso (nM) + SEM
hu'aibition of VIPstimulated cAMP production*
Theoretic SA (Ci/mM) Range
Somatostatin
0.23 4- 0.15
0.46 + 0.04
.22 + .07
2,000
WOC -2b
1.22 + 0.08
0.59 4 0.11
.25 + .06
4,000
WOC-3b
1.14 + 0.28
0.66 + 0.06
.25 + .06
6,000
WOC~a
0.68+0.19
0.37 4- 0.12
Not done
8,000
2.46 + 0.69 .25 4- .06 16,000 0.69 + 0.42 WOC -8 *Basal = .08 + .04 and VIP stim = .50 + .15 ,mol cAMP]'2x 1(P cells. These compounds mayprovide clitficallyuseful radiolabelled analogs for in vivo scintigraphy and in situ radiodetection and radiotherapy.
145