CA2528375A1 - Macrocyclic antagonists of the motilin receptor - Google Patents

Abstract

The present invention is directed to novel macrocyclic compounds of formula (I) and their pharmaceutically acceptable salts, hydrates or solvates: wherein R1, R2, R3, R4, R5, R6, n1, m, p Z1, Z2, and Z3 are as describe in the specification. The invention also relates to compounds of formula (I) which are antagonists of the motilin receptor and are useful in the treatment of disorders associated with this receptor and with or with motility dysfunction.

Description

MACROCYCLIC ANTAGONISTS OF THE MOTILIN RECEPTOR:--FIELD OF THE INVENTION
The present invention relates to novel conformationally-defined macrocyclic compounds, pharmaceutical compositions comprising same and intermediates used in their manufacture. More particularly, the invention relates to macrocyclic compounds that have been demonstrated to selectively antagonize the activity of the motilin receptor. The invention further relates to macrocyclic compounds useful as therapeutics for a range of 1 o gastrointestinal disorders, in particular those in which malfuncfiion of gastric motility or increased motilin secretion is observed, such as hypermotilinemia, irritable bowel syndrome and dyspepsia.
BACKGROUND OF THE INVENTION
A number of peptide hor~rtones are involved in the control of the different functions in the gastrointestinal (Gi) tract, including absorption, secretion, blood flow and motility (Mulvihill, et al. in Basic and Clinical Endocrinology, 4t" edition, Greenspan, F.S.;
Baxter, J.D., eds., Appleton & Lange: Norwalk, CT, 1994, pp 551-570). Since interactions between the brain 2 o and GI system are critical to the proper modulation of these functions, these peptides can be produced locally in the GI tract or distally in the CNS
One of these peptide hormones, motilin, a linear 22-amino acid peptide, plays a critical regulatory role in the GI physiological system though governing of fasting gastrointestinal motor activity. As such, the peptide is periodically released from the duodenal mucosa during fasting in mammals, including humans. More precisely, motilin exerts a powerful effect on gastric motility through the contraction of gastrointestinal smooth muscle to stimulate gastric emptying, decrease intestinal transit time and initiate phase III of the migrating motor complex in the small bowel (Itoh, Z., Ed., Motilin, Academic Press: San Diego, CA, 1990, ASIN: 0123757304; Nelson, D.K. Dig. Dis. Sci. 1996, 41, 2006-2015;
3o Peeters, T.L.; Vantrappen, G.; Janssens, J. Gastroenterology 1980, 79, 716-719).
Motilin exerts these effects through receptors located predominantly on the human antrum and proximal duodenum, although its receptors are found in other regions of the GI tract as well (Peeters, T.L.; Bormans, V.; Vantrappen, G. Regul. Pept. 1988, 23, 171-182).
Therefore, motilin hormone is involved in motility of both the upper and lower parts of the GI system (Williams et al. Am. J. Physiol. 1992, 262, G50-G55). In addition, motilin and its receptors have been found in the CNS and periphery, suggesting a physiological role in the nervous system that has not yet been definitively elucidated (Depoortere, I.; Peeters, T.L. Am. J. Physiol. 1997, 272, 6994-999 and O'Donohue, T.L et al. Peptides 1981, 2, 467-477). For example, motilin receptors in the brain have been suggested to play a regulatory role in a number of CNS functions, including feeding and drinking behavior, micturition reflex, central and brain stem neuronal modulation and pituitary hormone z o secretion (ttoh, Z. Motilin and Clinical Applications. Peptides 1997, 18, 593-608; Asakawa, A.; Inui, A.; Momose, K.; et al., M. Peptides 1998, 19, 987-990 and Rosenfeld, D.J.;
Garthwaite, T.L. Physiol. Behav. 1987, 39, 753-756). Physiological studies have provided confirmatory evidence that motilin can indeed have an effect on feeding behavior (Rosenfeld, D.J.; Garthwaite, T.L. Phys. Behav. 1987, 39, 735-736).
The recent identification and cloning of the human motilin receptor (WO
99/64436) has simplified and accelerated the search for agents which can modulate its activity for specific therapeutic purposes.
2 o Due to the critical and direct involvement of motilin in control of gastric motility, agents that either diminish (hypomotility) or enhance (hypermotility) the activity at the motilin receptor, are a particularly attractive area for further investigation in the search for new effective pharmaceuticals towards these indications.
Peptidic agonists of the motilin receptor, which have clinical application for the treatment of hypomotility disorders, have been reported (U.S. 5,695,952; 5,721,353;
6,018,037;
6,380,158; 6,420,521, U.S. Appl. 2001/0041791, WO 98/42840; WO 01/00830 and WO
02/059141 ). Derivatives of erythromycin, commonly referred to as motilides, have also been reported as agonists of the motilin receptor (U.S. 4,920,102; 5,008,249;
5,175,150;
5,418,224; 5,470,961; 5,523,401, 5,554,605; 5,658,888; 5,854,407; 5,912,235;
6,100,239;
6,165,985; 6,403,775).

Antagonists of the motilin receptor are potentially extremely useful as therapeutic treatments for diseases associated with hypermotility and hypermotilinemia, including irritable bowel syndrome, dyspepsia, gastroesophogeal reflux disorders, Crohn's disease, ulcerative colitis, pancreatitis, infantile hypertrophic pyloric stenosis, diabetes mellitus, obesity, malabsorption syndrome, carcinoid syndrome, diarrhea, atrophic colitis or gastritis, gastrointestinal dumping syndrome, postgastroenterectomy syndrome, gastric stasis and eating disorders leading to obesity.
1 o A variety of peptidic compounds have been described as antagonists of the motilin receptor (Depoortere, I.; Macielag, M.J.; Galdes, A.; Peeters, T.L. Eur. J.
Pharmacol. 1995, 286, 241-247; US 5,470,830; 6,255,285; 6,586,630; 6,720,433; U.S.
2003/0176643; WO
02/64623). These peptidic antagonists suffer from the known limitations of peptides as drug molecules, in particular poor oral bioavailability and degradative metabolism.
Cyclization of peptidic derivatives is a method employed to improve the properties of a linear peptide both with respect to metabolic stability and conformational freedom. Cyclic molecules tend to be more resistant to metabolic enzymes. Such cyclic tetrapeptide motilin antagonists have been reported (Haramura, M. et al J. ,Med. Chem.
2002, 45, 670-2 0 675, U.S. 2003/0191053; WO 02/16404).
Other motilin antagonists, which are non-peptidic and non-cyclic in nature have also been reported (U.S. 5,972,939; 6,384,031; 6,392,040; 6,423,714; 6,511,980;
6,624,165;
6,667,309; U.S. 2002/0111484; 2001/041701; 2002/0103238; 2001/0056106, 2 5 2002/0013352; 2003/0203906 and 2002/0002192) The macrocyclic motilin antagonists of the present invention comprise elements of both peptidic and non-peptidic structures in a combination which has not been pursued for this application previously.
Indeed, the structural features of antagonists of the present invention are different. In particular, within the known motilin antagonists which are cyclic peptides, it was found that such derivatives containing D-amino acids were devoid of activity. In contrast, for the tripeptidomimetic compounds of the present invention, the D-stereochemistry is required for two of the three building elements.
The motilin antagonists of the present invention are also distinct from the prior art in that they comprise a tether element to fulfill the dual role of controlling conformations and providing additional sites for interaction either fihrough hydrophobic interactions, hydrogen bonding or dipole-dipole interactions.
SUMMARY OF THE INVENTION
In a first aspect, the present invention is directed to compounds of formula (I):

(CH2)n1 O
R "~"

O

"~ryll R
(CH2)m O (CH~)p and pharmaceutically acceptable salts, hydrates or solvates thereof wherein:
Z1, Z2 and Z3 are independently selected from the group consisting of O, N and NRIO , wherein R1o is selected from the group consisting of hydrogen, lower alkyl, and substituted lower alkyl;
R~ is independently selected from the group consisting of lower alkyl substituted with aryl, lower alkyl substituted with substituted aryl, lower alkyl substituted with heteroaryl and lower alkyl substituted with substituted heteroaryl;
5 RZ is hydrogen;
R3 is independently selected from the group consisting of alkyl and cycloalkyl with the proviso that when Z~ is N, R3 can form a four, five, six or seven-membered heterocyclic ring together with Z~;
R4 is hydrogen;
so R5 and R6 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl and substituted heteroaryl, with the proviso that at least one of R5 and R6 is hydrogen;
X is selected from the group consisting of O, NR8, and N(R9)2~;
- wherein R$ is selected from the group consisting of hydrogen, lower alkyl, substituted lower alkyl, formyl, acyl, carboxyalkyl, carboxyaryl, amido, sulfonyl, sulfonamido and amidino; and - R9 is selected from the group consisting of hydrogen, lower alkyl, and substituted lower alkyl;
m, n~ and p are independently selected from 0, 1 or 2; and T is a bivalent radical of formula Il:
-U-(CH2)d-W-Y-z-(CH2)e- (II) wherein d and a are independently selected from 0, 1, 2, 3, 4 or 5;
wherein U is bonded to X of formula (I) and is -CH2- or -C(=O)- ;
wherein Y and Z are each optionally present;
W, Y and Z are independently selected from the group consisting of: -O-, -NR2$-, ~-, -SO-, -SOr, -C(=O)-, -C(=O)-O-, -O-C(=O)-, -C(=O)-NH-, -NH-C(=O)-, -S02-NH-, -NH-S02-, -CR29R3o-, -CH=CH- with a confiiguration Z' or E, and -C=C-, or from a ring structure independently selected from the group R32 ~ R31 K.~''r w G2 G~ ~ G~ Ka G2 G~~ ~ G~~ ~ '~ ~~ I i GZ
Ks Rsa R~
G~W ~~s~
Ra~Ra~ G1 ~~~J R3$ Ras ~/~\ G2 ~~~""~~G2 R3s Ray / R~s Ras R5o (CH2)r wherein any carbon atom contained within said ring structure, can be replaced by a nitrogen atom,, with fihe proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms;
1o G~ and G2 each independently represent a covalent bond or a bivalent radical selected from the group consisting of -O-, -NR4~-, -S-, -SO-, -S02-, -C(=O)-, -C(=O~
O-, -O-C(=O)-, -C(=O)NH-, -NH-C(=O)-, -S02-NH-, -NH-SO2-, -CR42R43-, -CH=CH-with a configuration Z or E, and -C=C-;with the proviso that G~ is bonded closer to U than G2;
K~, K2, K3, K4, K6, K~5 and K~6 are independently selected from the group consisting of O, NR44 and S;
f is selected from 1, ~, 3, 4, 5 or 6;
2 o R3~, R32, R3g, R3g, R4$ and R49 are independently selected from hydrogen, halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido; and R33e R34~ R35, R36~ R37, R47, R5o and R5~ are independently selected from hydrogen, halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido.
In a second aspect, the invention also proposes compounds of formula (1) which are z o antagonists of the motilin receptor .
In a third aspect, the invention proposes a method of treating a disorder associated with the motilin receptor or motility dysfunction in humans and other mammals , comprising administering a therapeutically effective amount of a compound of formula (1).
15 While the invention will be described in conjunction with example embodiments, it will be understood that it is not intended to limit the scope of the invention to such embodiment.
On the contrary, it is intended to cover all alternatives, modifications and equivalents as may be included as defined by the appended claims.
2 o DETAILED DESCRIPTION OF THE INVENTION
Preferably in formula (I), as depicted hereinabove, R~ is selected from the group consisting of -(CH2)qR~~, and -CHR~zR~3 wherein q is 0, 1, 2 or 3; and R~~ and R~2 are independently selected from a ring structure from the following 2 5 group:
F ~ F
F ~ 'F
F

,rwv~ Aa A

/~~ ~~
Ax A3 Ax A3 As ..
i Bl Az~~~i Bz Al\~B3 /Ba Aa \~ ~
Az A3 ~ A3 Az ,r~
Aa wherein any carbon atom in said ring structure can be replaced a nitrogen atom, with the proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms;
A~, A2, A3, A4 and A5 are each optionally present and are independently selected 1o from the group consisting of halogen, alkyl, substituted alkyl, cycloalkyi, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido;
B~, B2, B3, and B4 are independently selected from NR~4, S or O, wherein R~4 is selected from the group consisting of hydrogen, alkyl, substituted alkyl, formyl, acyl, carboxyalkyl, carboxyaryl, amido, sulfonyl and sulfonamido;
2 o R~3 is as defined for as R~~ and R'2 or is selected from the group comprising lower alkyl, substituted lower alkyl, hydroxy, alkoxy, aryloxy, amino, carboxy, carboxyaikyl, carboxyaryl, and amido .

wherein A~, A2, A3, A4 and A5 are most preferably selected from halogen, trifluroromethyl, C~_6 alkyl or C~.6 alkoxy.
Preferably, R~1, R~2 and R~~ are selected from the group consisting of:
~ / \ \ '~.' \ \
\ I ~ / / I / /
\ \ ~ ~~,, / N / ~ / / s s I s I) s \ I ~ l ~~ \ I ~ I ~~ ~ N ~
N ~, wherein Ra and Rb are chosen from the group consisting of CI, F, CF3, OCH3, OH, and C(CH3)3 and CH3.

Also preferably, R3 in formula (I), is selected from the group consisting of:
-(CHZ)SCH3, -CH(CH3)(CH2)tCH3, -CH(OR~5)CHs, -CH2SCH3 -CHzCH2SCH3, -CH~S(=O)CH3, -CH2CH2S(=O)CH3, -CHzS(=O)2CH3, -CH2CH2S(=O)2CH3, -(CH2)uCH(CH3)z, -C(CH3)3, and -(CH~)y-R2~, wherein:
2 0 ~ s and a are independently selected from 0, 1, 2, 3, 4 or 5;
t is independently selected from 1, 2, 3 or 4;
y is selected from 0, 1, 2, 3 or 4;
R~5 is selected from the group consisting of hydrogen, alkyl, substituted alkyl, formyl and acyl;
25 R2~ is selected from a ring structure selected from the following group:

I
-(CH2)Z
%/~
P2 ~ 2 J
wherein any carbon atom in said ring structure can be replaced by a nitrogen atom, with the proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms;
z is selected from 1, 2, 3, 4 or 5;
E~, E2 and E3 are each optionally present and are independently selected from the 1 o group consisting of halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido; and J is optionally present and is selected from the group consisting of alkyl, substituted .
alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, mercapto, sulfinyl, sulfonyl and sulfonamido.
The tether portion (T) of formula (I) is preferably selected from the group consisting of:

/ °~/ (~'~ /
(~ / (~) (~ j ~/(z3) (~3) ~L4 '~(Z3) LS' _ _ (73) L~(~

wherein L~ is O, NH or NMe; L2 is CH or N; L3 is CH or N; L4 is O or CHI; L5 is CH or N
L6 IS CR52R5s or O; R46 IS H or CH3;
R52, R53~ R54, R55, Rss and R5~ are independently selected from hydrogen, lower alkyl, substituted lower alkyl, hydroxy, alkoxy, aryloxy, amino, and oxo; or R52 together with R~3 or R54 together with R55 or R56 together with R57 can independently form a three to seven-membered cyclic ring comprising carbon, oxygen, sulfur and /or nitrogen atoms;
(X) is the site of a covalent bond to X in formula (I); and (Z3) is the site of a covalent bond to Z3 in formula (I).
to In a particularly preferred embodiment of the invention, there are provided compounds of formula (I) wherein m, n and p are 0, X, Z~, Z2 and Z3 are NH and R~, R4 and R5 are 15 hydrogen, represented by formula (III):

R3 ,,, H O
O NH HN Rs H T .,,,.H
Ni wN O
H H
(III) According to another aspect ofi the invention, there are provided compounds of formula (I) wherein when Z~ is a nitrogen atom, R3 forms a flour, five, six or seven-membered heterocyclic ring together with Z~ ,represented by formula (IV):
R7 _~= H O
O

~'''' R
(CH~)m ~~ O (CH2)p R ~X

(IV) wherein said heterocyclic ring may contain a second nitrogen atom, or an oxygen, or sulfur atom;
Z o n~ is selected from 0, 1, 2 or 3 R7 is optionally present and is selected from the group consisting of alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, 15 amidino, mercapto, sulfiinyl, sulfonyl and sulfonamido.
It is to be understood, that in the context of the present invention, the terms amino, guanidine, ureido and amidino encompass substituted derivatives thereof as well.

Preferably, the invention provides a method of treating a disorder associated with hypermotility or hypermotilinemia in humans and other mammals comprising administering a therapeutically effective amount of a compound of formula (1).
DESCRIPTION OF PREFERRED EMBODIMENTS
Although preferred embodiments of the present invention have been described in detail herein and illustrated in the accompanying structures, schemes and tables, it is to be understood that the invention is not limited to these precise embodiments and that various changes and modifications may be effected therein without departing from the scope or z5 spirit of the present invention.
Specifically preferred compounds of the present invention, include, but are not limited to:

--N O
n, N
NH HN i".
~ NH HN
\ / '-O \ ~
'-O
220 \ / 14 ~ /
O~~ ~ ~~ ~O -I O
Y-NH HN~ O~~ /~ p ~O y-NH HN--( N
NH HN ~~'~~ ~O u..
\ / ~O O \ / ~ HN - NH HN O
CI O \ /
O
212 \ / -202 \ / 164 \ /
O ~ ~ O
O~~ ~ NHS O p ~
y-NH HN~ 0 '~~N
n~.~ O NH H ,. O
NH HN "" _ NH HN
CI \ / ~ - NH HN
O \ / ~ ~ O
._ ~ '-O HO
HO
155 \ / 10 \ /
26 \ /
O O~~ ~O
O ~--~ OH Y-NH HN--~NH HN~ ",. 0 ..,. O ~ HN
,I NH HN \ / O
\ /
O
181 2 \ /
\ /
O ~O O ~O O ~O ..J O
~NH~HN~ ~N~H--H~N~ ~NH~-~H(N~ O\\ ~ NH2 .", O n" O n,. O n~.~NH HN
NH HN - NH HN NH HN _ ( ~O
HN
CI \ / ~ / \ / ~ CI \ / ~ CI \ /
O
200 \/ 21 \/ 149 \~N 158 \/N
O ~O O~~ ~O O ~O O ~O
~NH HN--( ' Y-NH HN~ ~NH N
a"~ ~O n"~ ~0 n"~ / ~NH HN
O ~", O
NH HN \ NH HN - NH HN ~ NH HN
\' S ~ O \ /
O
O HO ~ O
190 \ / 191 \ / 126 \ / 1~ \ /

o -/ o O~NH HN-( O N
u,. O n O
NH HN NH HN
O F \ / ~O
HO p _ 222 \ / 193 \ /
O~~ ~O O ~O
y-NH HN-( ~NH HN--( n,.~ ~O n..( ~O i HN \ ' ~ ~ HN
O S O \ /
163 \ / 167 \ / 2' O ~ O
O //
O'\ ~ NH
~NH HN-( "" O
NH HN "
I ~ NH HN CI \ /
O F \ / \ /
O CI _ J'-' 168 \ / 170 \ / F 13:

O
1' Oy-NH HN
_ ' O
NH HN
CI \ / CI \ /
O
CI
192 1q6 \ /

o i~o o i-~o j--NH HN~ y-NH HN-O
NH HN ~ NH HN
O O
F
19 ~ ~ 165 ~O
~O /'~
~NH HN-u.,' ~p u, NH HN
CI ~ ~
CI
131 ~ ~ 2Q
O ~ O
O ~ O\\ ~ OH
~NH HN~ ~NH HN
n,.~ p n~.( p NH HN - NH HN
CI ~ ~ ~O \ / ~O
_ CI
218 ~ ~ 182 O ~O O~~ ~O ~O
NH HN-( y-NH HN~ ~NH HN
u,. ~p n,.~ ~p n,.( ~p NH HN - NH HN - NH HN
~O
CI
16 \ / 141 \ / 128 \

In addition to the preferred tethers (T) illustrated previously, other specific tethers employed for compounds of the invention are shown hereinbelow:
\ NHPG ~ \ NHPG
HO ~ NHPG / OOH ~O~,OH

/
HO~''O ~ \ O~NHPG ~ ~ NHPG
/ ,OOH / S \
O
T10 Tll HO T12 ~G
NH
\ ~G PG'NH \ NHPG I \ ~G
~,OH ~ / OOH / O~
O O
'-OH
T28 T32 T33a [(R)-isomer]
T33b [(S)-isomer) O
NHPG \ NHPG X \ NHPG
O~N%~~OH F~O~'\/OH ~ / OOH
T35 T36 (X = F) ~4 T37 (X = Cl) ~G OH
NHI'G ~ \ NHPG ~ \ NHPG O
OOH / O,~OH / O OH . , HO~ , '~~OH

NHI'G
O
HO

PG and PG' indicate a standard amine protecting group compatible with the synthetic protocol, such as Boc, Ddz, Fmoc, or Alloc In a preferred embodiment, the present invention is directed to a method of treating irritable bowel syndrome, dyspepsia, Crohn's disease, gastroesophogeal reflux disorders, ulcerative colitis, pancreatitis, infantile hypertrophic pyloric, stenosis, carcinoid syndrome, malabsorption syndrome, diarrhea, diabetes mellitus, obesity, postgastroenterectomy syndrome, atrophic colitis or gastritis, gastric stasis, gastrointestinal dumping syndrome, celiac disease and eating disorders leading to obesity in humans and other mammals comprising administering a therapeutically effective amount of a compound of formula (I).
1 o SYNTHETIC METHODS
A. General Information Reagents and solvents were of reagent quality or better and were used as obtained from various commercial suppliers unless otherwise noted. DMF, DCM and THF
used are of DriSolv~ (EM Science, E. Merck) or synfihesis grade quality except for (i) 15 deprotection, (ii) resin capping reactions and (iii) washing. NMP used for the amino acid (AA) coupling reactions is of analytical grade. DMF was adequately degassed by placing under vacuum for a minimum of 30 min prior to use. Tyr(3tBu) was synthesized following the method reported in JP2000 44595. Cpa was made using literature methods (Tetrahedron: Asymmetry 2003, 14, 3575-3580) or obtained 2 o commercially. Boc- and Fmoc-protected amino acids and side chain protected derivatives, including those of N-methyl and unnatural amino acids, were obtained from commercial suppliers or synthesized through standard methodologies known to those in the art. Ddz-amino acids were either synthesized by standard procedures or obtained commercially from Orpegen (Heidelberg, Germany) or Advanced ChemTech 25 (Louisville, KY, USA). Bts-amino acids were synthesized as described in Example 6.
Hydroxy acids were obtained from commercial suppliers or synthesized from the corresponding amino acids by literature methods. Analytical TLC was performed on pre-coated plates of silica gel 60F254 (0.25 mm thickness) containing a fluorescent indicator. The term "concentrated/evaporated under reduced pressure" indicates 3 o evaporation utilizing a rotary evaporator under either water aspirator pressure or the stronger vacuum provided by a mechanical oil vacuum pump as appropriate for the solvent being removed. "Dry pack" indicates chromatography on silica gel that has not been pre-treated with solvent, generally applied on larger scales for purifications where a large difference in Rf exists between the desired product and any impurities. For solid phase chemistry processes, "dried in the standard manner" is that the resin is dried first in air (1 h), and subsequently under vacuum (oil pump usually) until full dryness is attained (~30 min to O/N).
B. Synthetic Methods for Building Blocks of the Invention Example 6: Standard Procedure for the Synthesis of Bts-Amino Acids O BtsNH~C02H
H3N~C02 N Raa, R,q,q ~ ~>--S02CI
S
Zo To a solution of the amino acid or amino acid derivative (0.1 mol, 1.0 eq) in 0.25 N
sodium hydroxide (0.08 mol, 0.8 eq) with an initial pH of approximately 9.5 (pH meter) at rt, solid Bts-CI (0.11 mol, 1.1 eq) was added in one portion. The resulting suspension was stirred vigorously for 2-3 d. The pH of the reaction should be adjusted with 5.0 N sodium hydroxide as required to remain within the range 9.5-10.0 during this 15 time. Typically, the pH has to be adjusted every 20-30 min during the first 5 h. Once the pH stops dropping, it is an indication that the reaction is almost complete. This can . be confirmed by TLC (EtOAc:MeOH, 95:5). Upon completion, the reaction mixture was washed with Et20. Washing is continued until the absence of non-polar impurities in the aqueous layer is confirmed by TLC (typically 3 x 100 mL). The aqueous solution 2 o was then cooled to 0°C, acidified to pH 2.0 with 1 N HCI until no additional cloudiness forms, and extracted with EtOAc (3 x 100 mL). Alternatively, a mixture of DCM
and EfiOAc may be used as the extraction solvent, depending on the solubility of the product obtained from difFerent amino acids or derivatives. Note that DCM
cannot be used solely as solvent because of the emulsion formed during extraction. The 25 combined organic phases were washed with brine (2 x 150 mL), dried over MgS04, filtered and evaporated under reduced pressure. DCM (1x) and hexanes (2x) were evaporated from the residue in order to ensure complete removal of the EtOAc and give the desired compound as a solid in 55-98% yield.
3 o The following are modifications that have proven useful for certain amino acids:
Gly Ala, D-Ala, ~i-Ala and GABA: Use 1.5 eq of amino acid per eq of Bts-CI, in order to prevent dibetsylation.

Met: Carry out the reaction under NZ to prevent oxidation.
Gln and Asn: Due to the solubility of Bts-Gln and Bts-Asn, the work-up required is modified from the standard procedure: Upon completion of the reaction, the reaction mixture was washed with diethyl ether. Washing is continued until the absence of non-5 polar impurities in the aqueous layer is confirmed by TLC (typically 3 x 100 mL). The aqueous phase was then cooled to 0°C and acidified to pH 2.0 with 6 N
HCI. 6 N HCI
was employed to minimize the volume of the solution due to the water solubility of Bts-Gln and Bts-Asn. (They are, in contrasfi, difficult to dissolve in DCM, EtOAc or chloroform.) The solution was maintained at 0°C for 10 min and the product was Zo collected by filtration as a white precipitate. The solid was washed with cold water (1x), cold brine (2x) and water (1x, 25°C). The pH of this wash was taken, if it is not approximately 4, the solid was washed again with water. Finally, the solid was washed with cold EtOAc, then with cold Et20 (2x), and finally dried under vacuum (oil pump) (83-85% yield).
C. General Synthetic Strategy to Conformationally-Defined Macrocycles of the Present Invention Scheme 1 BB3 BBp BB1 L "~ t-BB3 ~ -~ L-BB3~BBz~BB1-X-PlAG
"loading" of 1st sequential building building block* block assembly (X = 0, NHI
(deprotection, coupling) PG-Y TETHER Z deprotection, then cyclizafionl release gg .BB ~BB -X
L BBg~BBa~BB1-X ~ 3 2 1 tether attachment Y TETHER
[Y = 0, NH] PG.Y TETHE~
L
L = polymer with linker (L) for solid phase; pG = protecting group appropriate protecting group for solution phase;
for the latter, the scheme typically starts with the PIAG = protecting andlor protected first building block activating group BBB _- building blocks (amino acids, hydroxy acids) Z = reactive group *BBi, numbered in order from C-terminus in analogy to standard peptide nomenclature, NOT in order of addition 2 o The compounds of Formula I can be synthesized using traditional solution synthesis techniques or solid phase chemistry methods. In either, the construction involves four phases: first, synthesis of the building blocks, including one to four moieties, comprising recognition elements for the biological target receptor, plus one tether moiety, primarily for control and definition of conformation. These building blocks are assembled together, typically in a sequential fashion, in a second phase employing standard chemical transformations. The precursors from the assembly are then cyclized in the third stage to provide the macrocyclic structures. Finally, a post-cyclization processing stage involving removal of protecting groups and optional purification then provides the desired final compounds (Scheme 1). This method has been previously disclosed in WO 01/25257 and U.S. Pat: Appl. Ser. No. 09/679,331.
D. Procedures for the Synthesis of Representative Tethers of the Present Invention The important tether component required for compounds of the invention are synthesized "as described in W001/25257, U.S. Provisional Pat. Appl. Ser. No. 60/491,248 or herein.
l5 Example 16: Standard Procedure for the Synthesis of Tether T8 COOH ~gCl, \ \ COOMe Br/\/OTHP
MeOH ~ ~ / ICZC03, HI, DMF, 70 ° C
OH OH
Collidine; DMF; LiCI; then \ \ COOMe DIBAL, DCM, \ \ OH MsCI 0°C; then NaN3 O~OTHP . 35 . 0 °C ~O/\/OTHP
(1) PPh3;
\ N3 then HzO, 60 ° C \ \ ~z Ddz-OPh \ \ NHDdz / O~OTHP (2) 1N HCh ~ / OOH DMF; 50°C ~ ~ / O/\,~OH
(T8) Ste~~ T8-1: Chlorotrimethylsilane (116 mL, 0.91 mol, 1.5 eq) was added to a suspension of 2-hydroxycinnamic acid (100 g, 0.61 mol, 1.0 eq) in MeOH (500 mL, HPLC
grade) over 30 min afi 0° C. The resulting mixture was stirred at rt O/N. The reaction was monitored by TLC (EtOAc/MeOH: 98/2). Heating the reaction mixture in a hot water can accelerate the process if necessary. After the reaction was completed, the reaction mixture was evaporated under reduced pressure to afford methyl 2-hydroxycinnamate as a white solid (108.5 g) in quantitative yield. The identity of this intermediate compound is confirmed by NMR. This reaction can be carried out on larger (kg) scale with similar results Step T8-2: 3,4-Dihydro-2H pyran (DHP, 140 mL, 1.54 mol, 2.52 eq) was added dropwise s to 2-bromoethanol (108 mL, 1.51 mol, 2.5 eq) in a 2 L three-neck flask with mechanical stirring afi 0° C over 2 h. The resulting mixture was stirred for additional 1 h at rt. Methyl 2-hydroxycinnamate from Step T8-1 (108 g, 0.61 mol, 1.0 eq), potassium carbonate (92.2 g, 0.67 moi, 1.1 eq), potassium iodide (20 g, 0.12 mol, 0.2 eq) and DMF
(300 mL, spectrometric grade) were added to the above flask. The reaction mixture was stirred z o at 70° C (external temperature) for 24 h. The reaction was monitored by TLC
(DCM/Et2O: 95/5). The reaction was allowed to coolto rt and Et20 (450 mL) was added. The inorganic salts were removed by filtration and washed with Et20 (3 x 50 mL). The filtrate was diluted with hexanes (400 mL) and washed with water (3 x mL), dried over MgS04, filtered and the filtrate evaporated under reduced pressure.
The crude ester (desired product and excess Br-C2H4-OTHP) was used for the subsequent reduction without further purification.
Step T8-3: DIBAL (1.525 L, 1.525 moi, 2.5 eq, 1.0 M in DCM) was added slowly to a solution of the above crude ester from Step T8-2 (0.61 mol based on the theoretical 2 o yield) in anhydrous DCM (610 mL) at -35° C with mechanical stirring over 1.5 h. The resulting mixture was stirred for 1.5 h at -35° C, then 1.5 h at 0° C. The reaction was monitored by TLC (hex/EtOAc: 50/50). When complete, Na2S04~10 H2O (100 g, 0.5 eq) was slowly added; hydrogen evolution was observed, when it subsided water was added (100 mL). The mixture was warmed to rt and stirred for 10 min, then warmed 2 5 to 40° C with hot water and stirred under reflux for 20 min. The mixture was cooled to rt, diluted with DCM (600 mL), and the upper solution decanted into a filter.
The solid that remained in the flask was washed with dichloromethane (5 x 500 mL) with mechanical stirring and filtered. The filtrate from each wash was checked by TLC, and additional washes performed if necessary to recover additional product. In an 3 o alternative work-up procedure, after dilution with DCM (600 mL), the mixture was filtered. The resulting solid was then continuously extracted with 0.5% TEA in dichloromethane using a Soxhlet extractor. Higher yield was typically obtained by this alternative procedure, although ifi does require more time. The filtrate was concentrated under reduced pressure and the residue purified by dry pack (EtOAc/hex/Et3N: 20/80/0.5) to give the product alcohol as a yellowish oil (yield: 90%).
The identity and purity were confirmed by NMR.
Step T8-4: To a mixture of the allylic alcohol from Step T8-3 (28 g, 0.100 mol, 1.0 eq) and collidine (0.110 mol, 1.1 eq) in 200 mL of anhydrous DMF under N2 was added anhydrous LiCI (4.26 g, 0.100 mol, 1.0 eq.) dissolved in 100 mL of anhydrous DMF.
The mixture was then cooled to 0°C, and MsCI (12.67 g, 0.110 mol, 1.1 eq., freshly distilled over P~05), was added dropwise. The reaction was allowed to warm to rt and monitored by TLC (3:7 EtOAc/hex). When the reaction was complete, NaN3 (32.7 g, 0.500 mol, 5.0 eq.) was added. The reaction mixture was stirred at rt O/N~with progress followed by NMR. When the reaction was complete, the mixture is poured into an ice-cooled water bath, and extracted with diethyl ether (3x). The combined organic phases were then washed sequentially with citrate buffer (2x), saturated sodium bicarbonate 15 (2x), and finally with brine(1x). The organic layer was dried with MgS04, filtered and the filtrate concentrated under reduced pressure. The allylic azide was obtained in 90% combined yield, and was of sufficient quality to use as such for the following step.
Step T8-5: PPh3 (25.9 g, 0.099 mol, 1.5 eq) was added at 0° C to a solution of the allylic 2 o azide from Step T8-4 (20.0 g, 0.066 mol, 1.0 eq.) in 100 mL of THF. The solution was stirred for 30 min at 0° C and 20 h at rt. Water (12 mL) was then added and the resulting solution was heated at 60° C for 4 h. The solution was cooled to rt, 2N HCI
(15 mL) added and the mixture stirred for 90 min at 50°C. The separated organic phase was extracted with 0.05 N HCI (2 x 100 mL). The combined aqueous phase was 25 washed with Et20 (5 x 150 mL) and toluene (4 x 150 mL) (more extraction could be necessary, follow by TLC), which were combined and back-extracted with 0.05 N
HCI
(1 x 100 mL). This acidic aqueous phase from back-extraction was combined with the main aqueous phase and washed with ether (5 x 150 mL) again. The pH of the aqueous phase was then adjusted to 8-9 by the addition of sodium hydroxide (5 N).
3 o Care must be exercised to not adjust the pH above 9 due to the reaction conditions required by the next step. The aqueous phase was concentrated under reduced pressure (aspirator, then oil pumps or lyophilized to dryness. Toluene (2x) was added to the residue and then also evaporated under reduced pressure to remove traces of water. The crude product (desired amino alcohol along with inorgnic salt) was used for the next reaction without further purification.
Step T8-6: A mixture of the crude amino alcohol from Step T8-5 (0.5 mol based on the theoretical yield), Ddz-OPh (174 g, 0.55 mol, 1.1 eq) and Et3N (70 mL, 0.5 mol, 1.0 eq) in DMF (180 mL) was stirred for 24 h at 50° C. Additional DMF is added if required to solubilize all materials. The reaction was monitored by TLC (hex/EtOAc: 50/50, ninhydrin detection). After the reaction was complete, the reaction mixture was diluted with Et2O (1.5 L) and water (300 mL). The separated aqueous phase was extracted 2 o with Et20 (2 x 150 mL). The combined organic phase was washed with water (3 x 500 mL) and brine (1 x 500 mL), dried over MgS04, filtered and the filtrate concentrated under reduced pressure. The layers were monitored by TLC to ensure no product was lost into the aqueous layer. If so indicated, perform one or more additional extractions with Et20 of the aqueous phase to recover this material. The crude product was 15 purified by dry pack (recommended column conditions: EtOAclhex/Et3N:
35/65/0.5 to 65/35/0.5) to give the tether Ddz-T8 as a pale yellow syrup (yield: ~40%). The identity and purity of the product was confirmed by NMR.
~H NMR (DMSO-d6): 1.6 ppm (s, 6H, 2 x CH3), 3.6-3.8 ppm (wide s, 10 H, 2 x OCH3, 2 x OCH2), 3.95 ppm (triplet, 2H, CH2N), 6-6.2 ppm (m, 2H, 2 x CH), 6.2-2 0 6.5 ppm (m, 3H, 3 x CH, aromatic), 6.6-7.6 ppm (m, 5H, aromatic).
Example 17: Standard Procedure for the Synthesis of Tether T9 O
OH O~p O
~/ , NaH, DMF,O/N ~ OOH
100°C, N
a I I

~NNDdz [Example 18]
Cul, Pd2Cl2(PPh3)2, Ar CH3CN/Et3N (3:1), 4h O 1) Pt(IV)O, Hz, EtOH ~ O
OOH ~ 24-48h, r.t. I OH
/ NHDdz 2) Scavenger resin NHDdz DCM/Toluene 1:1 The yield of Ddz-T9 .from T9-0 on a 65 g scale was 60.9 g (91 %) 'H NMR (CDC13): C7 7.19-7.01, (m, 2H), 6.92-9.83 (m, 2H), 6.53 (bs, 2H), 6.34 (t, 5 1 H), 5.17 (bt, 1 H), 4.08 (m, 2H), 3.98 (m, 2H), 3.79 (s, 6H), 3.01 (bq, 2H), 2.66 (t, 3H), 1.26 (bs, 8H);
~3C NMR (CDC13) ~ 160.9, 156.8, 155.6, 149.6, 130.4, 127.5, 121.2, 111.7, 103.2, 98.4, 80., 69.7, 61.6, 55.5, 40.3, 30.5, 29.3, 27.4 1 o Tether T9 can also be synthesized from T8 by reduction as in step T9-3 or with other appropriate hydrogenation catalysts known to those in the art.
Example 18: Standard Procedure for the Synthesis of Ddz-propargylamine ~NHZ Ddz-N3, DIPEA, TMG ~NHDdz 15 DMF, 50°C, O/N, N2 In a dried three-neck flask, a solution of propargylamine (53.7 g, 0.975 mol, 1.5 eq) in degassed DMF (Drisolv, 388 mL) was treated with Ddz-N3 (170.9 g, 0.65 mol, 1.0 eq), tetramethylguanidine (TMG, 81.4 mL, 0.65 mol, 1.0 eq) and DIPEA (113.1 mL, 0.65 mol, 2 0 1.0 eq) and stirred at 50°C O/N. The reaction was monitored by TLC
(conditions:25/75 EtOAc/hex. Rf: 0.25; detection: UV, ninhydrin). Upon completion, DMF was evaporated under reduced pressure until dryness and the residue dissolved in Et2O (1 L).
The organic solution was washed sequentially with citrate buffer (pH 4.5, 3x), saturated aqueous sodium bicarbonate (2x), and brine (2x), then dried with MgS04, filtered and the filtrate evaporated under reduced pressure. A pale orange solid was obtained. This solid was triturated with 1 % EtOAc in hex, then collected by filtration and dried under vacuum (oil pump) to provide the desired product (153.4 g, 85.2%).
Example 19: Standard Procedure for the Synthesis of Tether T10 1 o Method A
1) PPh3, DIAD, THF, 4 h, rt Bz0 ~ ~ OH HO~NHDdz (Example 9] HO I ~ O~NHpdz 2) KOH, EtOH/N20 (1/1), 0/N, rt (605g ) 666 g (63%) HO'~Br TBAI, K2C03, DMF
85°C, 2 d HO'~O I ~ O~NHDdz 315 g (42% purified) Two alternative routes to this tether have been developed. The first synthetic approach proceeded starting from the commercially available monobenzoate of resorcinol (T10-0).
Mitsunobu reaction under standard conditions with the protected amino alcohol from Example 9, followed by saponification of the benzoate provided T10-1 in good yield after recrystallization. Alkylation of the phenol with 2-bromoethanol using the optimized conditions shown permitted the desired product Ddz-T10 to be obtained after dry pack purification in 42% yield.
2 o TLC (EtOAc/Hexanes 1:1, detection: UV, ninhydrin; Rf = 0.17) ~H NMR (CDCI3) S 7.18, t, 1 H,J = 8.2Hz; 6.51, m, 5H; 6.34, t, 1 H, J = 2.2Hz;
5.19, s, 1 H;
4.05, t, 2H, J = 5.0Hz; 3.94, m, 4H; 3.75, s, 6H; 3.49, d, 2H J = 5.2Hz; 1.73, s, 6H.

~3C NMR (CDC13) S 160.856; 5 160.152; 160.005; 155.410; 149.305; 130.279;
107.438;
107.310; 103.163; 101.877; 98.517; 69.488; 67.382; 61.595; 55.427; 40.420;
29.427.
HPLC,(standard gradient) tR: 7.25 min MS: 420 (M+H) Method 8 The second synthetic route to T10 is presented in the accompanying scheme.
1) PPh3, DIAD
HO'~NHBOC (35-65%) HO ~ OH Hp~O w D~NHBoc 2) PPh3, DIAD ~ , HO'~DTBDMS Igo%~ T10 3) TBAF (95%) Zo From resorcinol, two successive Mitsunobu reactions are conducted with the appropriate two carbon synthons illustrated, themselves derived from 2-aminoethanol and ethylene glycol, respectively, through known protection methodologies. Lastly, deprotection of the silyl ether, also under standard conditions provided Boc-T10.
Although the yields in the two methods are comparable, the first required less mechanical i5 manipulation and is preferred for larger scales.
Example 20: Standard Procedure for the Synthesis of Tether T11 2$
CO Pd(PPh3)4 5 mol%
OH ~ O (0.98 equiv.) Cul 5.2 mol%
OOH Et3N-DMF (1:3) N Br Cs2C03 (20 moi%) N Br (1 eq) DMF, 130 °C, 4 h ° NNBoc T11-1 76 /o (1.25 eq) 50 °C, OIN
OOH Pt02 (4 mol%) Et3N-EtOH (1:51 v/v) ~ ~ O~~OH

T11 ~ H2 (80 psi), rt, overnight ~NHBoc O ~ T11-2 > 98%
97.8%
TLC (15:85 THF/DCM; detection: UV; Rf: 0.33).
~ H NMR (DMSO-d6) c5 8.00, d, 1 H; 7.32, d, 1 H; 7.15, m, 1 H; 6.44, s, 2H;
6.33, s, 1 H; 3.99, t, 2H; 3.71, m, 8H; 2.89, m = 4, 2H; 2.71, t, 2H; 1.71, m = 5, 2H; 1.61, s, 6H.
~3C NMR, solvent DMSO-d6) b 160.879; 153.275; 151.405; 150.447; 140.773;
122.666;
118.934; 103.347; 98.456; 79.778; 70.449; 60.212; 55.717; 55.599; 29.740;
28.592.
HPLC (standard gradient) tR: 5.4 min MS: 419 (M+H) to Example 26: Standard Procedure for the Synthesis of Tether T12 HO NHa HO NHDdz I \ S I \ DdzN3, DIPEA, TMG
/ DMF, 50°C I / I /
(I2-0) Ddz-T12 In a 3-L flame-dried three-neck flask, a solution of (aminomethyl)phenylthiobenzyl alcohol (12-0, 96 g, 0.39 mol) in degassed DMF (1 L, 0.4 M) was prepared. To this was added DdzN3 (0.95 eq), followed by TMG (0.39 mol, 49 mL). The reaction was stirred for 10 min, then DlPEA (68 mL, 0.39 mol) added. The mixture was heated at 50°C
under N2 until TLC

indicated no DdzN3 remained (48 h typically). (TLC eluent: EtOAc:Hex 50:50;
detection:
ninhydrin). Upon completion, to the reaction mixture was added 3 L citrate buffer and the separated aqueous layer extracted with Et20 (3 x 1500 mL). The combined organic phase was washed sequentially with citrate buffer (2 x 200 mL), water (2 x 200 mL) and brine (2 x 200 mL). The organic layer was dried over MgS04, filtered and the filtrate evaporated under reduced pressure. A dark orange oil was obtained, which was purified by dry-pack.
For this procedure, the oil was first dissolved in EtOAc:Hex:DCM:TEA
(20:80:1:0.5, v/v/v/v). At this point, a little extra DCM was sometimes required to ensure complete dissolution. The solution was loaded onto the column, then the column eluted with Zo EtOAc:Hex:DCM:Et3N (20:80:1:0.5) until all the impurities were separated out as indicated by TLC, paying particular attention to that closest to the desired product.
The elution was then continued with EtOAc:Hex:Et3N 30:70:0.5 (v/v/v) and finally with EtOAc:hexanes:Et3N
(50:50:0.5) to elute the desired product. After removal of the solvent from the fractions containing the product under reduced pressure, the residue was dissolved in the minimum amount of DCM, a three-fold larger volume of hexanes added, then the solvents again evaporated under reduced pressure. This treatment was repeated until an oft white foam was obtained. The latter solidified while drying under vacuum (oil pump).
Alternatively, the material yielded a solid after sequential concentration with DCM (1x) and hexanes (2x).
Tether Ddz-T12 was obtained as an off-white solid (85-90% yield).
Examale 29: Standard Procedure for Attachment of Tethers Utilizing the Mitsunobu Reaction Example 29-A: Using PPh3-DIAD Isolated Adduct To a 0.2 M solution of the appropriate tether (1.5 eq) in THF or THF-toluene (1:1) was added the PPh3-DIAD (pre-formed by mixing equivalent amounts of the reagents and isolated by evaporation of solvent, see Example 29-C) adduct (1.0 eq.). The resultant mixture was manually agitated for 10 sec (the solution remained turbid), then added to the resin. Alternatively, the resin was added to the solution. The reaction suspension was agitated O/N (after ~5 min the mixture becomes limpid). The resin was filtered and washed 2x DCM, 1x toluene, 1x EtOH, 1x toluene, 1x (DCM/MeOH), 1x (THF/MeOH), 1x (DCM/MeOH), 1x (THF/MeOH), 2x DCM, then dried in the standard manner.
Example 29-B: Using "PPh3-DIAD In Situ Procedure"

To a 0.2 M solution of the appropriate tether (4 eq) in THF or THF-toluene (1:1) was added triphenylphosphine (4 eq). The resultant mixture was manually shaken until a homogenous solution was obtained, then added to the resin. Alternatively, the resin (or MiniKans containing resin) was added to the solution. To this suspension was then added DIAD (3.9 5 eq) and the reaction agitated O/N. Note: Since the reaction is exothermic, for larger scales, the reaction should be cooled in an ice bath. In addition, an appropriate vent must be supplied to allow any pressure build-up to be released. The resin was filtered and washed DCM (2x), toluene (1x), EtOH (1x), toluene (1x), DCM/MeOH (1x), 1x THF/MeOH
(1x), DCM/MeOH (1x), THF/MeOH (1x), 2x DCM, then dried in the standard manner.
Example 29-C: Procedure for Synthesis of PPh3-DIAD Adduct THF 0 °C
O N=N O + PPh3 O N-N O
30min O O PPh3 ~+
DIAD (1eq.) (1eq.) Adduct DIAD (1 eq) was added dropwise to a well-stirred solution of triphenylphosphine (1 eq) in THF (0.4 M) at 0°C under nitrogen. The mixture was then maintained at 0°C with stirring for 30 min. The white solid obtained was collected by filtration (use medium sized fritted filters), washed with cold anhydrous THF until the washes were colorless, and lastly washed once with anhydrous Et20. The white solid product was then vacuum-dried (oil 2 o pump) and stored under nitrogen. (Note: The PPh3-DIAD adduct can be made in larger than immediately required quantity and stored under nitrogen; it is very important to store this reagent under anhydrous conditions.) 2 s Example 30: Standard Procedure for Attachment of Tethers via Reductive Amination In certain instances, the Mitsunobu process of Example 29 cannot be applied or is not efficient for incorporation of the tether. Hence, reductive amination has been developed as an alternative that can be employed for tether incorporation as illustrated hereinbelow for one of the preferred tethers. Similar chemistry can be used to incorporate other tethers of the present invention.

I
I ~ o~ (3o-z) ' ~ p O p 0 O O ~Ddz / I
O O (1.5 eq) CF3COz H3N
O ~ TMOF-MeOH (1:3) 0 / ~ BH3~Pyr (2 eq) RT, 0/N h!H
Ddz (30-1 ) (30-3) The Tether (30-2) with the amine protected as its Ddz derivative was efficienfily oxidized to the corresponding aldehyde 30-2 using S03~pyr in DMSO-Et3N-DCM. This aldehyde (0.14 mmol, 56 mg, 1.5 eq based upon loading of resin support) was dissolved in a 1:3 mixture of TMOF-MeOH (DriSolv, 4 mL) at rt. To this was added the resin containing the 1o tripeptide (30-1, as its trifluoroacetic acid salt from the deprotection of the terminal amine), the mixture was agitated briefly to wet the resin, and then borane-pyridine complex (as the commercially available 8 M solution, 23 pL, 2 eq) was introduced to~ the suspension. The reaction was agitated O/N, then the resin filtered, washed with DCM (2x), THF
(1x),, DCM/MeOH [3:1] (1x), THF/MeOH [3:1] (1x), DCM (2x) and dried in fihe standard manner.
Care must be taken to ensure that the desired resin bound product 30-3 is not contaminated with the dialkylated material. However, even if the reaction does not proceed to completion or if a small amount of the dialkylation side product is present, the material is of sufficient purity for the macrocyclization reaction.
2 o Example 32: Standard Procedure for the Synthesis of Tether T28 N02 1, NaBH4, THF:MeON 1:1 H CH3N02, AcOH ~ 2. H2, Pd/C ~ NHBoc OH NH40Ac, 110C, o/n I .~ 3. (Boc)20, THF:H20 OH
OH

(79%) (90%) 28-0 28-1 2g_2 TBDMSO(CHa)2Br, KzCO3, KI

DMF, 75C

NHBoc _ TBAF ~ ~ NHBoc (74%) ' /
OH THF OTBDMS

O '~
O O

(95%) Boc-T28 28-3 Henry reaction of 2-hydroxybenzaldehyde 28-0 provided 28-1 in 79% yield. This was followed by reduction first with sodium borohydride, then with catalytic hydrogenation, to give the amine, which was then protected as its Boc derivative, 28-2. Yields of these first two steps were lower on larger scales. Alkylation of 28-2 with the TBDMS ether of 2-bromoethanol, itself synthesized by standard methods, gave 28-3 in 74% yield.
Deprotection of the silyl ether under standard conditions yielded the desired protected tether, Boc-T28. Alternative use of ethylene carbonate for the phenol alkylation to avoid the protection/deprotection steps, gave 73% yield.
Example 36: Standard Procedure for the Synthesis of Tether T32 OH OH Br~OTBDMS
NBS, TfOH / Br (32-A) CH3CN w I K2C03, KI, DMF
(94%) 70°C
CN CN (100%) 1) LiHMDS, THF (Boc)2O, 1 N NaOH
' 2) 1 M citric acid dioxane (67%, 2 steps) NHDdz (32-B) 1 ) Hz, Pt02 Cul, PdCl2(PPh3)2, PPh3 -IDdz 95% EtOH NHDdz HN(iPr)2, 70°C 2) TBAF, THF
(100%) (88%, 2 steps) 32-5 Ddz-T32(Boc) Overall yield 55%
(7 steps) TLC (100% EtOAc; detection: UV, CMA; Rf = 0.24).
1H NMR (CDC13, ppm): 7.74 (1 H, dd), 7.35 (1 H, d), 6.72 (1 H, d), 6.53-6.49 (2H, m), 3.61-3.29 (1 H, m), 5.06 (1 H, t), 4.25-4.01 (2H, m), 3.91-3.89 (2H, m), 3.73 (3H, s), 2.99 (2H, dd), 2.63 (2H, t), 1.71 (8H, broad), 1.53 (9H, s).
13C NMR (CDCI3, ppm): 163.8, 162.2, 161.0, 159.7, 155.9, 149.4, 130.0, 129.1, 128.0, 126.8, 110.8, 98.1, 80.9, 79.3, 69.7, 61.3, 55.5, 39.1, 29.3, 28.5, 26.7.
so Example 37: Standard Procedure for the Synthesis of Tether T33a and T33b HO~OMe O
OH O ~ I ~OH
(33-A) I O~pMe DIBAL 9.0 M ~ O
I DIAD, PPh3 a CH2CI2, -78°C
THF, rt, o/n I (99%) I
33-0 (gg%) 33-1 33-2 ~NHDdz Cul, Et3N/CH3CN (3:1) PdCl2(PPh3)2, Ar, o/n (88%) .,, , ''~OH
O
1) HZ, 95% EtOH , Pt02, o/n I ~ OOH
i NHDdz ~ 2) PS-TMf, CH~CI2 i (93%) ~ NHDdz Ddz-T33a 33-3 Overall yield 77.2%, 4 steps The construction to the (R)-isomer of this tether (T33a) was accomplished from 15 2-iodophenol (33-0) and (S)-methyl lactate (33-A). Mitsunobu reaction of 33-0 and 33-A
proceeded with inversion of configuration in excellent yield to give 33-1.
Reduction of the ester to the corresponding alcohol (33-2) also occurred in high yield and was followed by Sonagashira reaction with Ddz-propargylamine. The alkyne in the resulting coupling product, 33-3, was reduced with catalytic hydrogenation. Workup with scavenger resin provided the desired product, Ddz-T33a.
~H NMR (CDCI3) 8 (ppm) 7.18-7.11 (m, 2H), 6.90 (m, 2H), 6.52 (m, 2H), 6.33(m, 1H), s 5.09 (bt, 1 H), 4.52 (m, 1 H), 3.77 (s, 6H), 3.08 (bq, 2H), 2.64 (bt, 2H), 1.75 (m, 8H); 1.27 (bd, 3H), ~3C NMR (CDC13) 8 160.8, 155.5, 149.5, 131.2, 130.6, 127.4, 121.2, 113.3, 103.2, 98,4, 80.7, 74.8, 66.5, 55,4, 40.2, 30.6, 29.3, 29.2, 27.4, 16.1 HPLC (standard gradient): tR: 7.93 min The synthesis of the (S)-enantiomer (Ddz-T33b) was carried out in an identical manner in comparable yield starting from (R)-methyl lactate (33-B) ~OH
OH - OMe ~ O
+ HO O ~ ~ , NHDdz Ddz-T33b Example 38: Standard Procedure for the Synthesis of Tether T34 OH O O
N ~ I MezS04 Me~~ LiHMDS, oxirane ~ I OH AczO
OAc H O O N Me THF, rt, BF3:OEt2 N pyr, rt O N
HO N Me NaOH 2 O M
(83%) Me (60%) I 3 (98%) I
34.0 34-1 34-2 34-3 la, DCM, AgOCC
(75%) O O / NHDdz O
Ddz-propargylamine (32-B) ~ I
N s NHDdz H~ N I N
OAc ~ ~ OAc PdCl2(PPh3)z Cul, Bu4Nl, ~ I OAc O N 3 PtOz, EtOH O i 3 Et3N K~C03 O N
I (90 /o) I
(80%) 34-6 34-5 34~4 MeONa, MeOH
(85%) ~ ~NHDdz w..
OH pdz-T34 TLC (100% EtOAc; detection: CMA, Rf = 0.5).

MW Calc. for C2q.H35N3~7, 477.55; MS Found (M+H)+ 478.
~H NMR (CDC13) 61.62 (m, 2H), 1.70 (m, 8H), 2.43 (m, 2H), 2.67 (m, 2H), 3.07 (m, 2H), 3.34 (s, 3H), 3.43 (s, 3H), 3.61 (m, 2H), 3.75 (s, 6H), 5.40 (sb, 1 H), 6.31 (s, 1 H), 6.49 (s, 2H) 5 ~3C NMR (CDC13) 623.25 (CH2), 25.97 (CH2), 28.56 (CH3), 39.31 (CH3), 30.09 (CH3), 31.25 (CH2), _32.19 (CH2), 40.16 (CH2), 55.47 (CH3), 61.38 (CH2), 80.65 (Cq), 99.38 (Cq), 103.17 (Cq), _111.01 (Cq), 149, 60 (Cq), 151.33 (Cq), 152.46 (Cq), 160.80 (Cq).
HPLC (standard gradient) tR: 6.68 min.
Example 39: Standard Procedure for the Synthesis of Tether T35 F OH TBDMSO~ Br F O
T35-A _ I ~ ~OTBDMS
Br K2C03, KI, DMF ~ Br T35-0 55°C, O/N, N2 T35-1 100%
~NHDdz Pd(PhCN)2CI2, dioxane Cul, P(Bu)310% hexanes i-Pr2NH, 60°C, O/N
4 steps X5.2%
Overall yield: 56%
F ~ OOH 1) H2, PtO2, EtOH F I ~ O~OTBDMS
/ NHDdz 2) TBAF 1.0 M, THF, 1 h / ~ NHDdz Ddz-T35 74.5% (2 steps) T35-2 TLC (25/75 EtOAc/Hex; detection: UV, ninhydrin; Rf = 0.03) ~H NMR (CDC13): 8 7.06-7.00 (bt, 1 H), 6.61-6.52 (m, 4H), 6.35 (m, 1 H), 5.12 (bt, 1 H), 4.03 (m, 2H), 3.95 (m, 2H), 3.77 (s, 6H), 3.11-3.04 (bq, 2H), 2.60 (bt, 2H), 1.75 (m, 8H) ~3C NMR (CDC13): 8 163.9, 160.9, 160.6, 157.6, 157.5, 155.6, 149.5, 130.8, 130.6, 125.9, 107.26, 106.9, 103.2, 98,4, 80.8, 77.5, 69.9, 61,3, 60.9, 60.6, 55,4, 40.3, 30.4, 2 0 29.3, 26.9, HPLC (standard gradient): tR = 8.37 min Example 40: Standard Procedure for the Synthesis of Tether T36 74.5% (2 steps) ~ Br OH TBDMSO O
T35-A ' I ~ ~OTBDMS
F Br K2C03, KI, DMF F Br T36-0 55°C, O/N, N2 T36-1 96.7% /~
~NHDdz Pd(PhCN)2CI2, dioxane Cul, P(Bu)310% hexanes i-Pr2NH, 60°C, O/N
4 steps 75.2%
Overall yield: 54%
~ OOH 1) H2, Pt02, EtOH I ~ O~OTBDMS
F I / NHDdz 2) TBAF 1.0 M, THF, 1 h F
Ddz-T36 74.5% (2 steps) T36-2 \ NHDdz TLC: (25/75 EtOAc/Hex; detection: UV, ninhydrin; Rf = 0.03) ~ H NMR (CDC13) 8 (ppm): 6.84-6.75 (m, 3H), 6.52 (bs, 2H), 6.34 (m, 1 H), 5.17 (bt, 1 H), 4.01 (m, 2H), 3.93 (m, 2H), 3.77 (s, 6H), 3.10 (bq, 2H), 2.63 (bt, 2H), 1.74 (m, 8H) ~3C NMR (CDC13) 8 160.9, 158.9, 155.8, 155.6, 152.9, 152.9, 149.5, 132.4, 132.3, 2 0 117.1, 116.8, 112.7, 112.6, 103.2, 98.4, 80.8, 70.4, 61.6, 55.5, 40.2, 30.3, 29.3, 27.4.
HPLC (standard gradient): tR = 8.29 min Example 41: Standard Procedure for the Synthesis of Tether T37 OH TBDMSO~ Br O
T35-A I \ ~OTBDMS
CI ~ Br K2C03, Kl, DMF CI ~ Br T37-0 55°C, O/N, N2 T37-1 86.2% /~
~NHDdz Pd(PPh3)2CI2 Cul, PPh3, Argon 4 Steps i-Pr2NH, 55°C, O/N
Overall yield: 43% 61.2%
OOH 1) H2, Pt02, EtOH I \ O~OTBDMS
CI / NHDdz 2) TBAF 1.0 M, THF, 1 h CI ~ ~ NHDdz Ddz-T37 82% (2 steps) T37-2 TLC (25/75 EtOAc/Hex; detection: UV, ninhydrin; Rf = 0.03 ) ~H NMR (CDC13): b 7.12-7.08 (bd, 2H), 6.76-6.73 (d, 1 H), 6.52 (m, 2H), 6.33 (bs, 1 H), 15 (bt, 1 H), 4.02 (m, 2H), 3.95 (m, 2H), 3.79 (s, 6H), 3.09 (bq, 2H), 2.61 (bt, 2H), 1.74 (m, 8H). ~3C NMR (CDC13) 8 160.8, 155.6, 155.4, 149.5, 132.4, 130.1, 127.0, 126.0, 112.8, 103.2, 98.4, 80.8, 70.0, 61.4, 55.5, 40.3, 30.2, 29.3, 24.5, 27.4 HPLC (standard gradient): tR = 9.60 min Example 42: Standard Procedure for the Synthesis of Tether T38 OH ~ ICZC03 \ OOH
Acetone, 75 °C, O/N
I I
T38-0 93% T38-1 ~NHDdz Cul, Pd2Cl2(PPh3)2, Ar CH3CNlEt3N (3:1 ), O/N
100%
O~ Raney Ni, H2 ~ OOH
OH
/ NHDdz rt, O/N / ~ NHDdz 100%
Ddz-T38 T38-2 ~H NMR (CDCI3): & 7.20-7.10, (m, 2H), 6.95-6.80 (m, 2H), 6.55 (bs, 2H), 6.35 (s, 1 H), 5.18 (bt, 1 H), 4.12 (m, 1 H), 3.95 (m, 2H), 3.80 (s, 6H), 3.15 (bq, 2H), 2.65 (t, 2H), 1.98 (bs, 2H), 1.65 (bs, 6H), 1.25 (m, 3H).
~3C NMR (CDC13): s160.8, 156.6, 155.8, 149.6, 130.4, 127.5, 121.3, 111.7, 103.2, 98.4, 80.7, 73.5, 66.6, 55.5, 40.2, 30.5, 29.3, 29.1, 27.3, 19.5.
Chiral T38 can be accessed through the use of asymmetric synthesis methods, resolution or chiral chromatography techniques available in the literature.
HPLC (standard gradient) tR = 8.46 min Chiral material can be accessed by starting with the chiral epoxide. For example, the (S)-isomer of T38 was constructed in 89% overall yield from (S)-propylene oxide.
Example 43: Standard Procedure for the Synthesis of Tether T39 OH ~ OH
\ CO~H TMSCI, MeOH , ~ \ C02Me gr~OTBDMS
99% ~ K2COg, KI, DMF
70°C, 55%

/OTBDMS
O~OTBDMS O O
\ C02Me Cul, MeLi, Et20 / LiAIH4, THF
0°C, 76% I home 0°C, 100%

/OH
1 ) MsCI, Et3N J( 2i°/., i steps 2) NaN3, DMF, 50°C / NHBoc 3) Pd/C, H2, (Boc)~O
EtOAc T39-4 4) TBAF, THF Boc-T39 67%, 4 steps TLC (50% EtOAc, 50% Hex; detection: UV and CMA; Rf = 0.25) ~H NMR (CDCI~, ppm): 7.11-7.08 (2H, m), 6.86 (1 H, t), 6.76 (1 H, d), 5.05 (1 H, broad), 4.26-3.85 (4H, m), 3.22-3.07 (2H, m), 2.71 (1 H, broad), 1.66-1.60 (2H, m), 1.33 (9H, s), 1.17 (3H, d).
~3C NMR (CDC13, ppm): 156.1, 135.0, 127.1, 127.0, 121.4, 111.7, 69.9, 61.5, 39.8, 38.4, 28.7, 20.7.
1o Chiral T39 can be accessed through the use of asymmetric synthesis methods, resolution or chiral chromatography techniques available in the literature.
Example 44: Standard Procedure for the Synthesis of Tether T40 O O NaH
H Br~OTBDMS / H Ph3PCH(CH3)C02Me OH K2COg, ICI, DMF' W I O~OTBS 90-100%
70°C, 100%

C02Me pIBAL, DCM \ I ~ OOH Pd/C, Et3N
' _ o O
~OTBS 87 90 /o ~OTBS 80-90%

1 ) MsCI, Et3N 41 %, 8 steps ~OH 2) NaN3, DMF, 50°C NHBoc O 3) Pd/C, H2, (Boc)20 ~OTBS EtOAc T40-4 4) TBAF, THF Boc-T40 67%, 4 steps TLC (50% EtOAc, 50% Hex; detection: UV and CMA; Rf = 0.25) ~H NMR (CDC13, ppm): 7.11-7.08 (2H, m), 6.86 (1 H, t), 6.76 (1 H, d), 5.05 (1 H, broad), 4.26-3.85 (4H, m), 3.22-3.07 (2H, m), 2.71 (1 H, broad), 1.66-1.60 (2H, m), 1.33 (9H, s), 1.17 (3H, d).
~3C NMR (CDC13, ppm): 156.1, 135.0, 127.1, 127.0, 121.4, 111.7, 69.9, 61.5, 39.8, 38.4, 28.7, 20.7.
1o Chiral T40 can be accessed through the use of asymmetric synthesis methods, resolution or chiral chromatography techniques available in the literature.
Example 45: Standard Procedure for the Synthesis of Tether T41 TLC (100% EtOAc; detection: CMA; Rf= 0.5) ~H NMR (CDCI3) 6.1.23 (s, 3H), 1.49 (s, 3H), 1.69 (s, 3H), 1.74 (s, 3H), 1.90 (m, 2H), 2.35 (m, 1 H), 3.35 (m, 2H), 3.76 (s, 6H), 3.92 (m, 2H), 4.40 (m, 2H), 5.10 (m, 1 H), 6.15 (s, 1 H), 6.25 (s, 2H).
~3C NMR (CDCI3) 525.52 (CH3), 27.53 (CH3), 28.88 (CH3), 29.61 (CH3), 35.92 (CH2), 42.62 (CH2), 55.43 (CH3), 60.60 (CHI), 82.38 (CH), 83.33 (CH), 83.68 (CH), 84.96 (CH), 98.26 (CH), 103.23 (CH), 118.3 (Cq), 149.50 (Cq), 156.20 (Cq), 160, 02 (Cq) 1o HPLC (standard gradient): tR = 6.64 min MS: M+H found: 439 Example 46: Standard Procedure for the Synthesis of Tether T42 OH ~ hots OH OH hots O
Br H2C0 OHC ~ Br CH3PPh3Br Br HO
I i ~ ~, ~ Br MgCl2, TEA I / t-BuOK, THF, DEAD PPh THF /
CH3CN, reflux -78°C-> rt a.
T42-0 T42-1 ~rt, 6h T42-3 ° ~$% T42-2 95 /0 88%
~CI2(Gy3P)2Ru=CHPh DCM(0.2M), rt, 12 h 70%
W _ ~NHDdz 0 off , -' ~ ~ \ OH MeONa ~ w w csoA~ I w w Cul, Pd(PhCN)2C12 Br O MeOH, RT ~ O OAc DMF, 50°C, O/N ~ OTs P(Bu)3, HN(iPr)2, Br T42-7 dioxane, 70°C, O/N 98% 70% Br NHDdz 80%
H~, Pt02 EtOH, rt, O/N
95%
i p~OH
Ddz-T42 NHDdz ~H NMR (300 MHz, CDC13) ~ 6.82-6.98 (m, 2H); 6.80-6.75 (m, 1 H); 6.53 (s, 2H);
6.35 (t, 1 H, 2 Hz); 5.23 (b, 1 H); 4.08 (m, 1 H); 3.90-3.68 (m, 8H); 3.20-2.97 (m, 2H); 2.95-53 (m, 4H); 2.0-1.63 (m, 10H).
~3C NMR (75.5 MHz, CDC13) b160.85; 155.56; 152.55; 149.56; 128.13; 127.77;
120.28;
103.22; 98.43; 80.72; 76.80; 65.76; 55.46; 40.23; 30.45; 29.34; 29.22; 27.10;
24.97; 23.94.
2o E. Examales of Synthetic Strategies for the Macrocyclic Compounds of the Invention Scheme 2: Thioester Strategy for Macrocyclic Compounds of the Present Invention i 1. TrtSCH2CHzCOaH
DIC, DMAP, 18 h 0 PG-AA3-OH
DMF:DCM (1:1), RW ~ (PG = Ddz or Boc) NHZ 2. AczO/DIPEAIDCM I ~ I H SH pygpP
capping, 1 h, RW
3. 10%TFA, Et3SiH --SH ' "C3'-Linker NMp1 8 h 3 x 15min, RW RW
RW = Resin Wash 1. 1 % TFA, 3 x l5min (Ddz) 1. 2% TFA, 15min (Ddz) or 33% TFA, 1 h (Boc) or 33% TFA, 1 h (Boc) O
Et3SiH, DCM, RW Et3SiH, DCM, RW I' S~AA3NHPG
2. Bts-AAi-OH 2. PG-AAZ-OH
HBTU, DIPEA, NMP HBTU, DIPEA, NMP
18h, RW 18h, RW O
°II Ph3P-DIAD preformed betaine, Tether ---S- 'AA3--AAZ--AAA--NBtS
g~AA3__AAZ__p,p,~_NHBts THF or THF-toluene (1:1), 18h, RW
DdzN H-Tether Ag-assisted macrocyclization Method A Mefhod 8 Better yields but may require special purification to remove 1, 2% TFA, Et3SiH, DCM 1. 2% TFA, Et3SiH, DCM
Ag impurities to trace levels 1 h, RW
1h, RW
2. 1 eq Ag(CF3C02), DIPEA Z~ DIPEA, THF, MP-carbonate, THF, 24 h Mp-carbonate, 24 h 0 1. PS-thiophenol resin O
Purification: ~ KOTMS, THF/EtOH (1:1) CombiFlash ~ HN AAs--AA2--~1--NH HN~AA3--AAz--AAA--NBts Ag-scavenger ~ 2. 50% TFA, DCM
ISCO
etc. Tether Et3SiH oriPr3SiH Tether 2h One or more of the amino acids indicated can be replaced by corresponding hydroxy acids and coupled to the next building block utilizing methods known to those in the art.
Example 47: Standard Procedure for Macrocyclization with Thioester Linker The resin containing the cyclization precursor is combined in an appropriate vessel with pre-washed MP-carbonate resin [Argonaut Technologies, Foster City, CA, commercially supplied MP-carbonate resin was treated with 3x THF (1 L per 400 g) and dried O/N at 30°C in a vacuum oven] (1.4 to 1.6 eq relative to the initial loading of the synthesis resin).
A 0.2 M DIPEA solution in THF was then added to the combined resins (1 mL / 60 mg MP-carbonate resin) and the suspension agitated O/N at rt. Subsequently, the resin was filtered and rinsed 2x THF. The combined filtrates are collected together in an appropriate vessel, then the volatile contents evaporated under reduced pressure [in addition to the i5 standard methods, solvent can also be removed in vacuo using centrifugal evaporation (ThermoSavant Discovery, SpeedVac or comparable)] to provide the crude macrocycles.
Example 48: Standard Procedure for Silver-Assisted Macrocyclization with Thioester Linker Except for the cyclization itself and subsequent work-up, this procedure is identical to that of Example 47. The resin containing the cyclization precursor was combined in an appropriate vessel with pre-washed MP-carbonate resin [Argonaut Technologies, commercially supplied MP-carbonate resin was treated with THF (3x, 1 L per 400 g) and dried O/N at 30°C in a vacuum oven] (1.4 to 1.6 eq relative to the initial loading of the synthesis resin). To this was added THF (1 mL per 100 mg resin) and silver trifluoroacetate (1 eq relative to the initial loading of the resin). Finally, an amount of DIPEA sufficient to obtain a 0.2 M solution was added. The reaction mixture was agitated at rt O/N. The solution was then filtered and the resins washed 2x THF. The filtrates are collected together in an appropriate vessel, then evaporated under reduced pressure [(the volatile contents could also be removed in vacuo using centrifugal evaporation (ThermoSavant Discovery, SpeedVac or comparable)] to provide the crude macrocycles.
For this procedure, silver trifluoroacetate should be stored in a dessicator between uses.
In addition, it is recommended fio use a new bottle of THF (or a bottle that has been recently opened under N2 or Ar) to minimize formation of silver oxide.
Additionally, a ring-closing metathesis (RCM) strategy, as developed by Grubbs et al, can also be used to access some of the macrocyclic compounds of the invention (see for example US 5,811,515; Grubbs, R.H. et al. J. Org. Chem. 2001, 66, 5291-5300;
Furstner, A. Angew. Chem. Int. Ed. 2000, 39, 3012-3043).
To access certain derivatives of compounds of the present invention, additional reactions from those in the general scheme were required. For some, it was advantageous to react the functionality to be derivatized prior to the formation of the macrocyclic ring. The cyclic structure can restrict access of reagents to that functionality. For example, in the synthesis of N-methyl and, N-acyl derivatives of macrocycles, where the secondary nitrogen atom of the ring is the site of derivatization, the reaction is preferred to be performed prior to the application of the appropriate cyclization protocol.
In other cases, for example the derivatization of side chain functionality, the reaction was 3 o best performed after formation of the macrocyclic ring. For example, further reaction of amino moieties on side chains examples was typically efficiently done by reaction of the partially protected macrocycle. in this manner, acylation, sulfonylation, alkylation (via reductive amination), guanidine and urea formation were perFormed via standard methods.

Table 1, hereinbelow, shows a representative, but by no means exclusive, summary of the chemical synthesis of several representative compounds of the invention.

Table 1: Synthesis of Representative Compounds of the Present Invention AAA AAZ AA3 Tether ~ Tether Additional Attachment Ste s Bts-D- Boc-D-Val Boc-Nva Ddz-T8 Example 29 none T r tBu Bts-D- Boc-D-Vai Boc-Nva Boc-T8 Example 29 none Phe Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe Bts-D- Boc-D-Val Boc-Nva Ddz-T9 Example 29 none T r tBu Bts-D- Boc-D-Ala Boc-Nva Ddz-T8 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Met Ddz-T8 Example 29 none T r tBu ' Bts-D- Boc-D-Val Boc-Nle Ddz-T8 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Phe Ddz-T8 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Val Ddz-T8 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Leu Ddz-T9 Example29 none T r tBu Bts-D-2-Boc-D-Val Boc-Nva Boc-T8 Example 29 none Nal 12 gts-D- Boc-D-Val Boc-Abu Ddz-T8 Example 29 none T r tBu 13 Bts-D- Boc-D-Val Boc-Leu Boc-T9 Example 29 none Phe Bts-D-2-Boc-D-Val Boc-Leu Boc-T9 Example 29 none Nal 15 Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3C1 Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(4C1 17 gts-D- Boc-D-Val Boc-Nva Ddz-T9 Example 29 none Trp(Boc Bts-D- Boc-D-2- Boc-Nva Ddz-T9 Example 29 none T r Abu _ tgu Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe 2o gts-D- Boc-D-Val Boc-Leu Boc-T8 Example 29 none Phe 21 gts-D-2-Boc-D-Val Boc-Leu Boc-T8 Example 29 none Nal 22 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Tyr(OM

a 23 gts-D-1-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Nal 24 gts-D-2-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Thi 25 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(2C1 Bts-D- Boc-D-Val Boc-Cpa Ddz-T9 Example 29 none T r tBu 27 gts-D-4-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Thz 2$ Bts-D-3-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Pal is gts-D- Boc-D-Val Ddz- Ddz-T9 Example 29 none T r Da Boc tgu 3o gts-D- Hnva(THP) Boc-Nva Ddz-T9 Example 29 none T r tgu sa Bts-D- Ddz-D- Boc-Nva Ddz-T8 Example 29 None T r T r tBu tBu 38 Bts-D- Boc-D-Val Boc-Ala Ddz-T8 Example 29 none T r tBu 39 Bts-D- Boc-D-Val Boc-0-AlaDdz-T8 Example 29 none T r tBu 4o gts-D- Boc-D-Val Boc-Gly Ddz-T8 Example 29 none T r tBu 41 Bts-D- Boc-DPhe Boc-Nva Ddz-T8 Example 29 none T r tgu Bts-D- Boc-D-Val Boc-Phg Ddz-T8 Example 29 none T r tgu 55 Bts-D- Ddz-D-Val Ddz- Ddz-T8 Example 29 none T r L s Boc tBu 56 Bts-D- Ddz-D-Val Ddz- Ddz-T8 Example 29 none T r Orn Boc tgu Bts-D- Ddz-D-Val Ddz- Ddz-T8 Example 29 none T r Ser tgu tBu Bts-D- Ddz-D-Val Ddz- Ddz-T8 Example 29 none T r T r tgu tBu ss -gts-D-Ddz--D-ValDdz- Ddz-T8 Example 29 none T r Tr Boc tBu _ 6 Bts-D- Boc-D-Val Boc- Ddz-T8 Example 29 none T r T r OMe tgu 65 Bts-D- Boc-D-Val Boc-Nva Ddz-T2 Example 29 none T r tBu 71 gts-D- Boc-D-Val Boc-Nva Ddz-T10 Example 29 none T r tBu 72 Bts-D- Boc-D-Val Boc-2-NalDdz-T8 Example 29 none T r tBu 76 Bts-D- Boc-D-2-NalBoc-Nva Ddz-T8 Example 29 none T r tBu 77 Bts-D- Boc-D-Nle Boc-Nva Ddz-T8 Example 29 none T r tBu $ Bts-D- Boc-D-Val Boc-Ile Ddz-T8 Example 29 none T r tBu $5 Bts-D- Boc-D-Val Boc-D-NvaDdz-T8 Example 29 none T r tBu $' Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Bi $$ Bfis-D-Boc-D-Val Boc-Nva Ddz-T9 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Hfe s Bfis-D-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Di Bfis-D-Boc-D-Nva Boc-Nva Ddz-T9 Example 29 none T r tBu s2 gts-D- Boc-D-T(e Boc-Nva Ddz-T9 Example 29 none T r , tBu 96 gts_p_ goc_a_Ala Boc-Nva Ddz-T9 Example 29 none T r tBu Bts-D- Boc-D-Chg Boc-Nva Ddz-T9 Example 29 none T r tBu s8 Bfis-D-Boc-D-Val Boc-Nva Ddz-T18 Example 29 none T r tBu ss Bts-D- Boc-D-Val Boc-Nva Ddz-T15 Example 29 none T r tBu Los gts-D- Boc-D-Val Ddz- Ddz-T9 Example 29 none T r Dab Boc tBu Bts-D- Boc-D-Val Boc-Nva Ddz-T11 Example 29 none T r tBu 111 gts-D- Boc-D-Val Hval(THP)Ddz-T9 Example 29 none T r tBu Bts-D- Boc-D-Val Boc-Nva Ddz-T9 Example 29 none T r tBu ~zo gts-D- Boc-D-Pro Boc-Nva Ddz-T8 Example 29 none ~ ~ ~ -~ - -T r(tBu) ~

Bts-D- Boc-D-Val Boc-Nva Ac-T8-NH2 Example 29 none T r tBu 122 goc-D- Boc-D-Val Boc-Nva Boc-T9 Example 30 none 3-Pal 123 goc-D- Boc-D-Val Boc-Nva Boc-T9 Example 30 none 2-Pal Boc-D- Boc-D-Val Boc-Nva Boc-T9 Example 30 none 4-Pal 125 gts-D- Boc-D-Cpg Boc-Nva Boc-T9 Example 29 none T r tBu Bts-D- Boc-D-Val Boc- Boc-T9 Example 29 none T r NMeLeu tBu 127 goc_p_ Boc-D-Val Boc-Nva Boc-T12 Example 30 none His Mts ~zs gts-D- Boc-D-Val Boc-Leu Boc-T9 Example 29 none Tyr(OM

a gts-D-1-Boc-D-Val Boc-Leu Boc-T9 Example 29 none Nal ~3o gts-D-2-Boc-D-Val Boc-Leu Boc-T9 Example 29 none Thi 131 gts-D- Boc-D-Val Boc-Leu Boc-T9 Example 29 none ~

Phe(3C1 gts-D- Boc-D-Val Boc-Leu Boc-T9 Example 29 none Phe(4CI

133 gts-D- Boc-D-Val Boc-Leu Boc-T9 Example 29 none Phe 134 gts-D- Boc-D-Val Boc-Leu Boc-T2 Example 29 none Phe(3C1 135 gts-D- Boc-D-Val Boc-Leu Boc-T11 Example 29 none Tyr(OM

a gts-D- Boc-D-Val Boc-Leu Boc-T11 Example 29 none 1 Nal 137 gts-D-2-Boc-D-Val Boc-Leu Boc-T11 Example 29 none Thi ~3s gts-D- Boc-D-Val Boc-Leu Boc-T11 Example 29 none Phe(3C1 139 gts-D- Boc-D-Val Boc-Leu Boc-T11 Example 29 none Phe(4C1 ~4o gts-D- Boc-D-Val Boc-Leu Boc-T11 Example 29 none Phe ~4~ Bts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Tyr(O
M

a Bts-D-1-Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Nal 143 gts-D-2-Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Thi ~a4 gts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe(3CI

X45 gts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe(4CI

gas gts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe 147 gts-D- Boc-D-Val Boc-Cpa Boc-T11 Example 29 none Tyr(OM

a Bts-D-1-Boc-D-Val Boc-Cpa Boc-T11 Example 29 none Nal ~4s gts-D- Boc-D-Val Boc-Cpa Boc-T11 Example 29 none Phe(3CI

~5o gts-D- Boc-D-Val Boc-Cpa Boc-T11 Example 29 none Phe(4C1 151 gts-D- Boc-D-Val Boc-Cpa Boc-T11 Example 29 none Phe 152 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Tyr(OM Dap(Boc) a 153 gts-D-1-Ddz-D-Val Ddz- Ddz-T9 Example 29 none Nal Da Boc 154 gts-D-2-Ddz-D-Val Ddz- Ddz-T9 Example 29 none Thi Da Boc 155 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe(3C1 Dap(Boc) 156 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe(4C1 Dap(Boc) 157 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe Da Boc 158 gts-D- Ddz-D-Val Ddz- Ddz-T11 Example 29 none Phe(3CI Dap(Boc) 159 gts-D- Boc-D-Ile Boc-Nva Boc-T9 Example 29 none T r But ~so gts-D- Boc-D- Boc-Nva Boc-T9 Example 29 none Tyr(But)allolle Boc-D- Boc-D-Val Boc-Nva Boc-T9 Example 30 none Phe(4C

moc ~s2 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(2M

a ~s3 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3M

a ~s4 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(4M

a ~s5 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(30 Me gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(20 Me 167 gts-D-3-Boc-D-Val Boc-Nva Boc-T9 Example 29 none benzothi en I

ass gts-D-3-Boc-D-Val Boc-Nva Boc-T9 Example 29 none Thi Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none 0_ HomoP .

he 3C1 ~7o gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3,4 ' diCl 171 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3,4 diF

172 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3,4 diOMe 173 gts-D- Hnva(THP) Boc-Nva Boc-T9 Example 29 none 1 Nal 174 Bts-D- Hnva(THP) Boc-Nva Boc-T9 Example 29 none Tyr(OM

a 175 gts-D- Boc-D-Val Boc-Nva Boc-T33b Example 29 none T r tBu 176 gts-D- Boc-D-Val Boc-Nva Boc-T33a Example 29 none T r tBu 177 gts-D- Boc-D-Val Boc-Nva Boc-T28 Example 29 none Tyr(tBu) ~~s Bts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Tyr(OM Ser(tBu) a 179 gts-D-1-Ddz-D-Val Ddz- Ddz-T9 Example 29 none Nai Ser tBu ~so gts-D-2-Ddz-D-Val Ddz- Ddz-T9 Example 29 none Thi Ser tBu ~s~ Bts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe(3C1 Ser(tBu) ~sz gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe(4C1 Ser(tBu) ~s3 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 none Phe Ser tBu ~sa gts-D~1-Ddz-D-Vai Ddz- Ddz-T11 Example 29 none Nal Da Boc ~s5 gts-D- Ddz-D-Val Ddz- Ddz-T11 Example 29 none Phe(4CI Dap(Boc) gas pdz-D- Ddz-D-Val Ddz- Ddz-T9 Example 30 none T r His Mts tBu Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(3C

ass gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe gas gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Phe(4N

gts-D-3-Boc-D-Val Boc-Cpa Boc-T9 Example 29 none benzothi en I

Bts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe(30 Me 192 gts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe(3,4 diCl gts-D- Boc-D-Val Boc-Cpa Boc-T9 Example 29 none Phe(3,4 diF

~s4 gts-D- Boc-D-Val Boc-Nva Boc-T34 Example 29 none Tyr(OM

a ~s5 gts-D- Boc-D-Val Boc-Nva Boc-T38 Example 29 none T r OM

e) -. -196 gts-D- Boc-D-Val Boc-Cpa Ddz- Example 29 none Phe(3C1 T32(Boc) Bts-D- Boc-D-Val Boc-Cpa Boc-T34 Example 29 none Phe(3C1 Bts-D- Boc-D-Val Boc-Cpa Boc-T38 Example 29 none Phe(3C1 Bts-D- Boc-D-Val Boc-Cpa Boc-T41 Example 29 none Phe(3C1 gts_p_ Boc-D-Val Boc-Cpa Boc-T8 Example 29 none Phe(3C1 Zoo gts_D-1-Boc-D-Val Boc-Nva Boc-T8 Example 29 none Nal zo2 gts-D- Boc-D-Val Boc-Nva Boc-T8 Example 29 none Phe(30 Me zos gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 acetylation Phe(4CI Dap(Boc) zo4 gts-D- Ddz-D-Val Ddz- Ddz-T9 Exam 1e 29 p guanonnylati Phe(4C1 Dap(Boc) zos gts-D- Boc-D-Val Boc- Boc-T9 Example 29 none Phe(3C1 NMeLeu zos gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 mesylation Phe(4C1 Dap(Boc) Zo' gts-D- Ddz-D-Val Ddz- Ddz-T9 . Example 29 TMS-Phe(4C1 Dap(Boc) isocyanate followed by dilute acid zo8 Bts-D- Ddz-D-Val Ddz- Ddz-T9 Exam 1e 29 p guanidinylati T r Da Boc on tBu zos gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 acetylation T r Da Boc tBu zoo gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 reductive Tyr(tBu) Dap(Boc) amination with acetone 211 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 reductive Phe(4CI Dap(Boc) amination with excess formaldeh d a gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 reductive Phe(4C1 Dap(Boc) amination with acetone 213 Bts-D- Boc-D-Va( Boc-Nva Boc-T9 Example 29 none Tyr(3,5d i1 2~a Bts-D- Boc-D-Val Boc- Boc-T9 Example 29 hydrogenoly Tyr(OM Hse(Bzl) sis for e) protecting group removal z~5 gts-D- Ddz-D-Val Ddz- Ddz-T9 Example 29 reductive Tyr(tBu) Dap(Boc) amination with excess formaldehyd a 216 gts-D- Boc-D-Val Boc-Cpa Boc-T40 Example 29 none Phe(3CI

217 gts-D- Boc-D-Val Boc-Cpa Boc-T36 Example 29 none Phe(3C1 218 gts-D- Boc-D-Val Boc-Nva Boc-T39 Example 29 none Phe(3C1 2~s gts-D- Boc-D-Val Boc-Nva Boc-T37 Example 29 none Phe(3CI

ZZO gts-D- Boc-D-Val Boc-Nva Boc-T39 Example 29 none Phe(3CI

221 gts-D- Boc-D-Val Boc-Nva Boc-T35 Example 29 none Phe(3CI

a22 gts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 none Tyr(3tBu 223 Bts-D- Boc-D-Val Boc-Nva Boc-T9 Example 29 acetylation T r But Zz4 gts-D-1-Boc-D-Val Boc-Leu Boc-T9 Example 29 reductive Nal amination with formaldehyd a z2s gts-D-1-Boc-D-Val Boc-Leu Boc-T9 Example 29 acetylation Nal Zzs gts-D-1-Boc-D-Val Boc-Leu Boc-T9 Example 29 reductive Nal amination with aldeh de 227 gts-D-1-Boc-D-Val Boc-Leu Boc-T9 Example 29 reductive Nal amination with benzaldehy de Notes Any amino acid or tether designated as the Boc derivative could be substituted with the corresponding Ddz derivative.

D. Analytical Data for Selected Compounds of the Invention ~H and ~3C NMR spectra were recorded on a Varian Mercury 300 MHz spectrometer and are referenced internally with respect to the residual proton signals of the solvent.
Information about the conformation of the molecules in solution can be determined utilizing appropriate two-dimensional NMR techniques known to those skilled in the art, HPLC
zo purifications were run on a Waters XTerra MS C18 column, using the Waters FractionLynx system. Automated medium pressure chromatographic purifications were performed on an Isco CombiFlash 16x system with disposable silica or C18 cartridges that permitted up to sixteen (16) samples to be run simultaneously. MS spectra were recorded on a Waters Micromass Platform 1l or ZQ system. HRMS spectra were recorded with a VG
Micromass ZAB-ZF spectrometer. Chemical and biological information were stored and analyzed utilizing the ActivityBase database software (IDBS, Guildford, Surrey, UIC).
General Methods for Analytical HPLC Analyses 2 o HPLC analyses are performed on a Waters Alliance system 2695 running at 1 mL/min using an Xterra MS C13 column 4.6 x 50 mm (3.5 p,m). A Waters 996 PDA provided UV
data for purity assessment. An LCPackings splitter (50:40:10) allowed the flow to be separated in three parts. The first part (50%) went to a Micromass Platform II
MS
equipped with an APCI probe for identity confirmation. The second part (40%) went to an evaporative light scattering detector (ELSD, Polymer Laboratories, PL-ELS-1000) for purity assessment and the last portion (10%) to a chemiluminescence nitrogen detector (CLND, Antek Model 8060) for quantitation and purity assessment. Data was captured and processed utilizing the most recent version of the Waters Millenium software package.
5 An example LC method suitable for compounds of the present invention uses MeOH as solvent A, H20 as solvent B and 1 %TFA/ H20 as solvent D. Initial mobile-phase composition is 5% A, 85% B and 10% D. Details of the standard gradient method are shown below:
Time A% B% D% Curve 10 0.00 5 85 10 6 1.00 5 85 10 6 6.00 50 40 10 6 9.00 50 40 10 6 14.00 90 0 10 6 15 17.00 90 0 10 6 17.50 5 85 10 6 20.00 5 85 10 6 Compounds 2-6, 8-10, 56, 65 and 144 are as defined in Table (3), hereinbelow.
2 o Compound 2 Yield: 12 mg pure macrocycle was obtained (CLND quantification).
~ H NMR (300MHz, DMSO-d6) 5 8.83 (m,1 H); 8.53 (m, 1 H); 7.63 (m, 1 H); 7.4-7.08 (m, 7H);
7.00-6.84 (m, 2H); 6.60 (d, 15Hz, 1 H); 6.41 (dt, 15Hz, 5.4 Hz, 1 H); 4.35 (m, 1 H); 4.25-4.05 (m, 3H); 3.94 (dt, 1 H, 6Hz, 15Hz); 3.79 (dd, 1 H, 3.6Hz, 8.4 Hz); 3.60 (m, 1 H); 3.52-3.40 25 (bd, 1H); 3.22-3.06 (m, 4H); 1.88 (m, 2H); 1.54-1.28 (m, 2H); 1.25 (d, 3H, 4.8Hz); 1.22 (d, 3H, 2.7 Hz); 0.92-0.80 (m, 6H).
HRMS calc. for C3pH40N4~4~ 520.3049; found 520.3057 ~ 0.0016 HPLC [standard gradient method (refers to that presented in General Methods for Analytical HPLC Analyses)] tR = 9.55 min.
Compound 4 Yield: 12 mg pure macrocycle was obtained (CLND quantification).

~ H NMR (300 MHz, DMSO-d6) 8 9.35 (b, 1 H); 8.98 (b, 1 H); 5.52 (d, 1 H, 8.4Hz); 8.38 (b, 1 H); 7.25 (b, 1 H); 7.13-7.07 (m, 4H); 6.86 (t, 2H, 7.5Hz); 6.57 (d, 2H, 8.7 Hz); 4.33 (b, 1 H);
4.21-4.02 (m, 3H); 3.78 (dd, 1 H, 3.3Hz; 8.1 Hz); 3.65-3.54 (m, 1 H); 3.31-3.23 (m, 1 H); 3.13-3.02 (m, 4H); 2.78-2.2.28-2.18 (m, 1 H); 2.0-1.80 (m, 2H); 1.50-1.30 (m, 3H);
1.25 (d, 3H, 4.5Hz); 1.22 (d, 3H, 4.5Hz); 1.01 (d, 3H, 6.6Hz); 0.90 (d, 3H, 6.6Hz); (t, 3H, 7.5Hz).
'3C NMR (75.5MHz, DMSO-d6) 5 172.22; 171.37; 157.77; 157.44; 156.04; 131.76;
130.80;
130.70; 127.88; 121.82; 115.83; 111.71; 62.13; 60.62; 54.21; 52.81; 47.13;
42.47; 33.31;
29.69; 29.30; 28.61; 20.36; 19.44; 18.72; 17.60; 13.97.
HRMS calc. for C3pH42N4~5~ 538.3155; found: 538.3145 ~ 0.0016 2o HPLC (standard gradient) tR = 8.12 min.
Compound 5 Yield: 17 mg pure macrocycle was obtained (CLND quantification).
~ H NMR (300 MHz, DMSO-d6) ~ 9.02 (b, 1 H); 8.47 (d, 1 H, 8.4Hz); 7.7 (b, 1 H); 7.58 (d, 1 H, 5.4Hz); 7.28 (dd, 1 H, 7,8Hz, 0.8Hz); 7.20 (t, 1 H, 9.OHz, 0,8Hz); 7.14 (d, 2H, 8.4Hz); 6.98-6.91 (m, 3H); 6.66 (d, 8.7Hz); 6.63 (d, 1 H, 15.OHz); 6.43 (dt, 1 H, 6.OHz, 15.OHz); 4.28-3.86 (m, 6H); 3.60-3.40 (m, 2H); 3.22-3.12 (m, 1 H0; 3.05 (d, 2H, 5.4Hz); 1.92-1.80 (m, 1 H);
1.56-1.40 (m, 1 H); 1.36-1.20 (m, 2H); 1.25 (d, 3H, 6.6Hz); 0.84 (t, 3H, 7.2Hz).
~3C NMR (75.5MHz, DMSO-d6) ~ 172.54; 171.86; 158.97; 158.56; 127.39; 155.84;
131.62;
2 0 129.73; 129.20; 129.02; 128.43; 126.30; 124.51; 122.01; 115.85; 112.88;
61.23; 52.90;
51.23; 47.08; 42.66; 36.13; 33.30; 21.14; 19.57; 17.07; 14.14; 11.49.
HRMS calc. for C28H36N4O5: 508.2685; found: 508.2681 ~ 0.0015 HPLC (standard gradient) tR = 7.67 min.
2 5 Compound 6 Yield: 16 mg pure macrocycle was obtained (CLND quantification).
1H NMR (300MHz, DMSO-d6) ~ 9.37 (b, 1 H); 8.87 (b, 1 H); 8.61 (d, 1 H, 8.7Hz);
7.62 (b, 1 H); 7.27 (d, 1 H, 7.8Hz); 7.21 (t, 1 H, 8.4Hz); 7.14 (d, 2H, 8.4Hz); 6.98-6.87 (m, 3H); 6.64 (d, 2H, 8.1 Hz); 6.70 (d, 1 H, 15.6Hz); 6.39 (dt, 1 H, 6.3Hz, 15.6Hz); 4.44-4..36 (m, 1 H); 4.34-3 0 4.08 (m, 2Hz); 4.45-3.92 (dt, 1 H, 6.9Hz, 15.6Hz); 3.74 (dd, 1 H, 3.6Hz, 8.4Hz); 3.54-3.26 (m, 3H); 3.22-3.02 (m, 3H); 2.60-2.36 (m, 4H); 2.24-2.14 (m, 1 H); 2.02 (s, 3H); 1.96-1.89 (m, 1 H); 1.80-1.66 (m, 1 H); 1.01 (d, 3H, 6.3Hz); 0.90 (d, 3H, 6.6Hz).

~3C NMR (75.5MHz, DMSO-d6) ~ 171.51; 171.26; 158.90; 158.49; 157.38; 155.86;
131.63;
129.82; 129.21; 128.86; 128.63; 126.21; 121.98; 115.83; 112.83; 62.11; 61.06;
51. 97;
47.10; 42.78; 30.91; 30.67; 29.34; 20.37; 19.39; 15.06. °
HRMS calc. for C3pH40N4~5s: 568.2719; found: 568.2711 ~ 0.0017 HPLC Rt (general method) 7.92 min.
Compound 8 Yield: 27 mg pure macrocycle was obtained (CLND quantification).
~H NMR (300MHz, DMSO-d6) b 9.05 (b, 1 H); 8.43 (b, 1 H); 8.34 (d, 1 H, 9.3Hz);
7.40 (b, s o 1 H); 6.97 (d, 1 H, 7.5Hz); 6.92-6.74 (m, 9H); 6.67-6.54 (m, 2H); 6.33-6.25 (m, 3H); 6.10 (dt, 1 H, 5.7Hz, 16.2Hz); 4.22 (dt, 1 H, 0.9Hz, 12Hz); 3.94-6.66 (m, 4H); 3.30 (dd, 1 H, 3.6Hz, 7.8Hz); 3.24 (m, 1 H); 3.18 (m, 1 H); 2.85-2.68 (m, 3H); 2.44-2.23 (m, 2H);
1.32 (o, 1 H, 7.5Hz); 0.97-0.89 (m, 1 H); 0.42 (d, 3H, 6.6Hz); 0.01 (d, 3H, 6.6Hz).
~3C NMR (75.5MHz, DMSO-d6) ~ 171.20; 157.35; 155.88; 139.12; 131.61; 130.87;
129.74;
129.21; 128.77; 128.88; 126.85; 126.19; 121.97; 115.82; 112.84; 62.04; 61.10;
55.07;
50.01; 47.09; 42.85; 37.42; 29.11.
HRMS calc. For C34H42N~.O5 : 586.3155; found: 586.3145 ~ 0.0017 HPLC Rt (general method) 9.34 min.
2 o Compound 9 Yield: 17 mg pure macrocycle was obtained (CLND quantification).
~H NMR (300MHz, DMSO-d6) ~ 9.39 (b, 1 H); 8.83 (b, 1 H); 8.29 (d, 1 H, 9.3Hz);
7.62 (b, 1 H); 7.28 (d, 1 H, 6.6Hz); 7.20 (t, 1 H, 6.9Hz); 7.12 (d, 2H, 7.8Hz); 6.98-6.91 (m, 2H); 6.63 (d, 2H, 8.4Hz); 6.58 (d, 1 H, 16.2Hz); 6.40 (dt, 1 H, 5.7Hz, 16.2Hz); 4.29-4..13 (m, 3H); 4.03-3.92 (m, 2H); 3.52 (m, 1 H); 3.15-3.05 (m, 3H); 2.45-2.37 (m, 1 H);
1.96-1.88 (m, 1 H); 1.25 (dd, 2H, 4.5Hz; 6Hz); 1.01 (d, 3H, 6.3Hz); 0.91 (d, 3H, 6.6Hz); 0.86 (d, 3H, 7.2Hz); 0.81 (d, 3H, 6.6Hz).
~3C NMR (75.5MHz, DMSO-d6) b 171.85; 171.17; 157.37; 155.87; 131.59; 129.88;
129.18;
3 0 128.97; 128.78; 128.51; 126.16; 121.97; 115.83; 112.85; 61.55; 61.18;
58.15; 54.22;
47.08; 42.89; 36.32; 29.35; 29.00; 20.34; 19.56; 18.73; 17.44.
HRMS calc. for C3pHqpN4O5 536.2998; found: 536.2990 ~ 0.0017.

HPLC (standard gradient) tR = 8.15 min.
Compound 10 Yield: 24 mg pure macrocycle was obtained (CLND quantification).
~H NMR (300 MHz, DMSO-d6) 5 9.33 (b, 1 H); 8.82 (b, 1 H); 8.56 (d, 1 H, 8.3Hz); 7.60 (b, 1 H); 7.27 (d, 2H, 7.8Hz); 7.20 (t, 1 H, 7.8Hz); 7.13 (d, 2H, 8.4Hz); 6.95 (t, 2H, 7.8Hz); 6.64 (d, 2H, 8.4Hz); 6.57 (d, 1 H, 15.4Hz); 6.38 (dt, 1 H, 15.4Hz, 5.8Hz); 4.26-4.10 (m, 3H); 3.96 (dt, 1 H, 5.4Hz, 8.4Hz); 3.77 (dd, 1 H, 3.7Hz, 7.8Hz); 3.51-3.24 (m, 3H); 3.18-3.02 (m, 3H);
1.90 (h, 1 H, 6.4Hz); 1.73-1.54 (m, 2H); 1.45 (dt, 1 H, 6.7Hz, 0.9Hz); 0.99 (d, 3H, 6.6Hz);
0.89 (d, 3H, 6.3Hz); 0.87 (d, 3H, 6.OHz); 0.80 (d, 3H, 6.3Hz).
~3C NMR (75.5MHz, DMSO-d6) 5 172.23; 171.17; 157.37; 155.88; 131.62; 129.82;
129.19;
128.95; 128.59; 126.24; 121.99; 115.84; 112.88; 64.23; 61.98; 61.14; 51.43;
61.14; 51.43;
47.07; 42. 81; 29.38; 24.85; 24.11; 21.00; 20.32; 19.30.
HRMS calc. for C3~H42N4~5 550.3155; found: 550.3150 ~ 0.0016.
,15 HPLC (standard gradient) tR = 8.91 min.
Compound 56 Yield: 16 mg pure macrocycle was obtained (CLND quantification).
~H NMR (300 MHz, DMSO-d6) ~ 9.39 (b, 1 H); 8.90 (b, 1 H); 8.67 (d, 1 H, 8.4Hz); 7.74 (b, 2 0 4H); 7.29-7.08 (m, 4H); 6.99-6.87 (m, 2H); 6.64 (d, 2H, 8.1 Hz); 6.61 (d, 1 H, 16.5Hz); 6.40 (dt, 1 H, 5.7Hz, 16.5Hz); 4.40-4..06 (m, 4H); 4.02-3.95 (m, 1 H); 3.79 (dd, 1 H, 3.6Hz, 7.8Hz);
3.55-3.30 (m, 2H); 3.16-3.05 (m, 3H); 2.82-2.69 (m, 2H); 2.02-1.85 (m, 2H);
1.64-1.43 (m, 3H); 1.29-1.23 (m, 1 H); 1.01 (d, 3H, 6.3Hz); 0.91 (d, 3H, 6.3Hz); 0.86-0.84 (m, 2H).
HPLC (standard gradient) tR = 5.71 min.
Compound 65 Yield: 17 mg pure macrocycle was obtained (CLND quantification).
'H NMR (300 MHz, DMSO-d6) ~ 9.60 (b, 1 H); 9.39 (b, 1 H); 8.88 (b, 1 H); 8.70 (d, 1 H, 7.5Hz); 8.57 (d, 1 H, 4.2Hz); 7.27 (t, 6Hz); 6.96 (d, 2H, 8.4Hz); 6.66 (d, 2H, 8.4Hz); 5.78-3 0 5.68 (m, 1 H); 5.42-5.33 (m, 1 H); 3.96-3.89 (m, 1 H); 3.80-3.57 (m, 5H);
3.41-3.34 (m, 1 H);
3.10-2.90 (m, 1 H); 2.78-2.66 (m, 1 H); 2.21-2.10 (m, 1 H); 2.06-1.93 (m, 1 H); 1.70-1.60 (m, 1 H); 1.52-1.41 (m, 1 H); 1.39-1.26 (m, 1 H); 1.25 (d, 3H, 4.8Hz); 1.23 (d, 3H, 4.5Hz); 0.83 (dd, 3H, 3Hz, 4.5Hz).
~3C NMR (75.5MHz, DMSO-d6) S 172.68; 172.63; 159.15; 158.73; 157.38; 157.25;
130.89;
124.99; 116.03; 62.51; 62.12; 54.29; 49.27; 42.47; 32.77; 30.43; 28.85; 20.46;
19.59;
18.72; 17.39; 13.90; 13.09.
HRMS calc. for Ca4H36N404~ 444.2736; found: 444.2726 ~ 0.0013 HPLC (standard gradient) tR = 6.80 min.
Compound 144 ~H NMR (300 MHz, CD30D) 8 7.4 (m, 1 H); 7.27 (dt, 1 H, 1.5 Hz, 6.6 Hz); 7.22-7.14 (m, zo 2H); 7.08-6.98 (m, 2H); 6.78 9t, 2H, 6.6 Hz); 4.45-4.39 (m, 2H); 4.15 (d, 2H, 8.1 Hz);
7.74 (d, 1 H, 9.3 Hz); 3.54 (d, 1 H, 10.8 Hz); 3.35-3.22 (m, 2H); 3.20 (q, 1 H, 1.5 Hz);
2.82-2.71 (m, 1 H); 2.61-2.55 (m, 1 H); 2.21-2.11 (m, 1 h); 2.02-1.94 (m, 1 H); 1.74-1.40 (m, 5H); 1.04 (d, 3H, 6.6 Hz); 0.93 (d, 3H, 6.6 Hz); 0.74-0.64 9m, 1 H)0.45-0.28 (m, 2H); 0.15-0.08 (m, 1 H); 0.06-0.02 (m, 1 H).
Z5 ~3C NMR (75.5 MHz, CD30D) 8 173.29; 172.14; 167.51; 155.47; 134.86; 134.81;
130.38; 130.31; 128.81; 128.25; 127.44; 121.63; 110.39; 107.71; 105.02; 67.10;
66.66;
62.81; 62.06; 60.10; 53.99; 41.44; 36.07; 31.91; 30.01; 29.18; 28.94; 27.79;
23.68;
23.15; 19.08; 18.25; 8.17; 4.98; 3.16.
HRMS: calc. for C3~H4~N404CI 568.2816; found 568.2802 ~ 0.0017 F. Mass Spectral Data for Selected Compounds of the Invention Table 2: Analysis of selected compounds of the invention Molecular FormulaMolecular WeightMonoisotopicM+H
(calculated) Mass Found C30H40N4O5 536.7 536 537 C30H40N4O4 520.7 520 521 C30H42N4O4 522.7 522 523 C30H42N4O5 538.7 538 539 C28H36N4O5 508.6 508 509 C30H40N4O5S 568.7 568 569 C31 H42N405 550.7 550 551 C34H42N405 586.7 586 587 C30H40N405 536.7 536 537 C31 H42N405 550.7 550 551 11 C34H44N404 572.7 572 573 12 C29H38N405 522.6 522 523 13 C31 H44N404 536:7 536 537 C35H46N404 586.8 586 587 15 C30H41 N404C1 557.1 556 557 16 C30H41 N404C1 557.1 556 557 17 C32H43N504 561.7 561 562 C29H40N405 524.7 524 525 C30H41 N404F 540.7 540 541 ao C31 H42N404 534.7 534 535 C35H44N404 584.7 584 585 22 C31 H44N405 552.7 552 553 2a C34H44N404 572.7 572 573 24 C28H40N404S 528.7 528 529 25 C30H41 N404C1 557.1 556 557 2s C31 H42N405 550.7 550 551 C27H39N504S 529.7 529 530 2$ C29H41 N504 523.7 523 524 29 C28H39N505 525.6 525 526 so C30H41 N306 539.7 539 540 34 C34H40N406 600.7 600 601 3$ C28H36N405 508.6 508 509 39 C28H36N405 508.6 508 509 ao C27H34N405 494.6 494 495 C34H40N405 584.7 584 585 52 C33H38N405 570.7 570 571 55 C31 H43N505 565.7 565 566 56 C30H41 N505 551.7 551 552 I

57 C28H36N406 524.6 524 525 5s C34H40N406 600.7 600 601 59 C36H41 N505 623.7 623 624 so C35H42N406 614.7 614 615 65 C24H36N404 444.6 444 445 C29H40N406 540.7 540 541 '2 C38H42N4~5 634.8 634 635 C38H42N405 634.8 634 635 C31 H42N405 550.7 550 551 s C31 H42N405 550.7 550 551 s5 C30H40N405 536.7 536 537 s~ C36H46N404 598.8 598 599 ss C34H50N405 594.8 594 595 s9 C31 H44N404 536.7 536 537 C36H46N404 598.8 598 599 C30H42N405 538.7 538 539 C31 H44N405 552.7 552 553 C28H38N405 510.6 510 511 C33H46N405 578.7 578 579 9s C24H39N504 461.6 461 462 C24H39N504 461.6 461 462 C29H41 N505 539.7 539 540 G2gH41 N505 539.7 539 540 C30H41 N306 539.7 539 540 112 G31 H44N405 552.7 552 553 ~2o C30H38N405 534.6 534 535 C32H45N506 595.7 595 596 ~z2 C31 H43N404CI 571.2 570 571 123 C29H41 N504 523.7 523 524 124 G29H41 N504 523.7 523 524 C30H40N405 536.7 536 537 126 G32H46N405 566.7 566 567 C30H38N603S 562.7 562 563 ~2s C32H46N405 566.7 566 567 C35H46N404 586.8 586 587 ~3o C29H42N404S 542.7 542 543 131 C31 H43N404CI 571.2 570 571 C31 H43N404CI 571.2 570 571 133 C31 H43N404F 554.7 554 555 ~3a C25H;37N403C1 477.0 476 477 135 C31 H45N505 567.7 567 568 136 C34H45N504 587.8 587 588 137 C28H41 N504S 543.7 543 544 ~3s C30H42N504CI 572.1 571 572 139 C30H42N504C1 572.1 571 572 Sao C30H42N504F 555.7 555 556 141 C32H44N405 564.7 564 565 142 C35H44N404 584.7 584 585 ~a3 C29H40N404S 540.7 540 541 144 C31 H41 N404C1 569.1 568 569 145 C31 H41 N404C1 569.1 568 569 146 C31 H41 N404F 552.7 552 553 C31 H43N505 565.7 565 566 gas C34H43N504 585.7 585 586 149 C30H40N504C1 570.1 569 570 ~5o C30H40N504CI 570.1 569 570 151 C30H40N504F 553.7 553 554 152 C29H41 N505 539.7 539 540 153 C32H41 N504 559.7 559 560 154 C26H37N504S 515.7 515 516 155 C28H38N504C1 544.1 543 544 156 C28H38N504C1 544.1 543 544 C28H38N504F 527.6 527 528 ~5s C27H37N604CI 545.1 544 545 159 C31 H44N405 552.7 552 553 ~6o C31 H44N405 552.7 552 553 161 C31 H45N504 551.7 551 552 162 C31 H44N404 536.7 536 537 C31 H44N404 536.7 536 537 164 C31 H44N404 536.7 536 537 165 C31 H44N405 552.7 552 553 166 C31 H44N405 552.7 552 553 C32H42N404S 578.8 578 579 C28H40N404S 528.7 528 529 169 C31 H43N404C1 571.2 570 571 C30H40N404CI2 591.6 590 591 C30H40N404F2 558.7 558 559 C32H46N406 582.7 582 583 C34H43N305 573.7 573 574 C31 H43N306 553.7 553 554 C31 H44N405 552.7 552 553 C31 H44N405, 552.7 552 553 C29H40N405 524.7 524 525 C29H40N406 540.7 540 541 C32H40N405 560.7 560 561 ~so C26H36N405S 516.7 516 517 C28H37N405C1 545.1 544 545 ~s2 C28H37N405C1 545.1 544 545 ~s3 C28H37N405F 528.6 528 529 ~s4 C31 H40N604 560.7 560 561 ~s5 C27H37N604C1 545.1 544 545 ~s6 ~ C31 H40N605 576.7 576 577 ~s~ C31 H41 N404F3 590.7 590 591 ass C30H41 N404F 540.7 540 541 ~s9 C30H41 N506 567.7 567 568 C33H42N404S 590.8 590 591 191 C32H44N405 564.7 564 565 C31 H40N404CI2 603.6 602 603 C31 H40N404F2 570.7 570 571 C32H48N606 612.8 612 613 195 C32H46N405 ~ 566.7 566 567 196 -- C32H43N604C1 611.2 610 611 C32H45N605CI 629.2 628 629 C32H43N404C1 583.2 582 583 C27H39N406CI 551.1 550 551 20o C31 H39N404C1 567.1 566 567 20~ C34H42N404 570.7 570 571 202 C31 H42N405 550.7 550 551 2os C30H40N505C1 586.1 585 586 204 C29H40N704C1 586.1 585 586 205 C32H45N404CI 585.2 584 585 2os C29H40N506SCI 622.2 621 622 207 C29H39N605C1 587.1 586 587 20a C29H41 N705 567.7 567 568 209 C30H41 N506 567.7 567 568 2~o C31 H45N505 567.7 567 568 2~~ C30H42N504C1 572.1 571 572 2~2 C31 H44N504CI 586.2 585 586 213 C30H40N40512 790.5 790 791 214 C30H42N406 554.7 554 555 2~5 C30H43N505 553.7 553 554 2~6 C32H43N404C1 583.2 582 583 ~

2~7 C31H40N404FCI 587.1 586 587 2~8 C31 H43N404C1 571.2 570 571 2~9 C30H40N404CI2 591.6 590 591 22o C31 H43N404F 554.7 554 555 22~ C30H40N404FC1 575.1 574 575 222 C34H50N405 594.8 594 595 22s C32H44N406 580.7 580 581 22a. C36H48N404 600.8 600 601 22s C37H48N405 628.8 628 629 22s C39H49N504S 683.9 683 684 22~ C42H52N404 676.9 676 677 Notes 1. Molecular formulas and molecular weights (MW) are calculated automatically from the structure via ActivityBase software QDBS, Guildford, Surrey, UK) or, for MW only, from the freeware program Molecular Weight Calculator v. 6.32 2. M+H obtained from LC-MS analysis using the General Method as described 3. All analvses conducted on material after preparative HPLC purification BIOLOGICAL METHODS AND RESULTS
5 The compounds of the presenfi invention were evaluated for their ability to interact at the human motilin receptor utilizing a competitive radioligand binding assay as described in Method B1. Further characterization of the interaction can be performed utilizing the functional assays described in Methods B2, B3 and B4. Some of these methods can be conducted, if so desired, in a high throughput manner to permit the simultaneous z o evaluation of many compounds. Other assays have also been described that are suitable for HTS, such as that based upon the stable expression of a synthetic gene for the human motilin receptor.
Results for the examination of representative compounds of the present invention using 15 Method B1 are presented in Table 3. The binding activity is listed as ranges with the following levels: A = 0.001-0.10 pM; B = 0.10-1.0 pM; C = 1.0-10.0 pM. in addition, the assay results of two additional compounds using this Method are shown below.
As can be observed, this demonstrates the activity of a representative bicyclic compound of Formula IV of the invention, which resulted from incorporation of D-proline as the second z o recognition building block. Significantly, the lack of binding activity obtained with compound 121, which is the linear analogue of compound 1 (K; = level B), illustrates the critical importance of the cyclic structure to attaining the desired interaction.
Compound 120 Compound 121 K; = level B K; > 10 ItM

Competitive binding curves for two representative compounds of the invention (Compounds 8 and 11 ) are presented hereinbelow:
Target: hMTL-R
Radioligand: [~25i]motilin = 100 m v ;~ 50 m ~n 25 ~~ -_ 185 nM
_9 _8 _7 _6 _5 Log M of Compound 8 Target: hMTL-R
Radioligand: [~25~motilin ~ 100 c = 75 m v c~
as a m 25 Ki -__ 7g,3 nM
_9 _8 _7 _6 _5 Log M of Compound 11 For determination of functional significance of the binding, the compounds are preferably Z o tested in the Aequorin assay as described in Method B2, although the procedure of Method B3 is also applicable. As can be seen from the data presented in Table 4, the representative compounds examined act as antagonists at the motilin receptor and are devoid of agonist activity at the concentrations studied. The functional activity is listed as ranges with the following levels: A = 0.001-0.10 pM; B = 0.10-1.0 pM. The higher s5 sensitivity of the assay of Method B2, almost 100 times that of Method C, makes it the preferred one for this assessment. This is evident in the ECSO values obtained in each for the positive agonist standard, motilin. Additionally, Method B2 measures the actual signaling event, which makes it more relevant to the effect that is desired, whereas the assay of Method B3 simply measures GTP turnover.
Table 4: Demonstration of Antagonist Activity at the Motilin Receptor Aequorin (Method B2)~

Compound' Binding (K~) 142 A g Motilin 0.6 not applicable (human, porcine)2 'Activity is listed as ranges with the following levels: A = 0.001-0.10 ~M; B
= 0.10-1.0 pM
2Human and porcine motilin are the same peptide.
In addition, a common and scientifically-accepted ex vivo assay for the measurement of agonist or antagonist activity at the motilin receptor is the contraction of rabbit duodenum Zo or other gastrointestinal smooth muscle tissue. AZ-A4 Agonists are defined as compounds that induce >50% contraction relative to the motilin peptide, whereas antagonists are defined as compounds that cause >50% inhibition of the response to motilin.
Compounds of the present invention have shown significant antagonist activity in this assay. For example, compound 144 exhibited a pA2 = 6.95, while compound 165 had a pA2 =
7.17, s5 as calculated from the Schild plots of the response obtained at various concentrations as described in Method B4.
Gastric motility is generally measured in the clinical setting as the time required for gastric emptying and subsequent transit time through the GI tract. Gastric emptying scans are well known to those skilled in the art an, briefly, comprise use of an oral contrast agent, 2 o such as barium, or a radiolabeled meal. Solid and liquids can be measured independently.

A test food or liquid is radiolabeled with an isotope (99mTc) and after ingestion or administration, transit time through the GI tract and gastric emptying are measured by visualization using gamma cameras. These studies are performed before and after the administration of the therapeutic agent to quantify the efficacy of the compound.
Example Method B1: Competitive Radioligand Binding Assay (Motilin Receptor) Materials:
~ Membranes were prepared from CHO cells stably transfected with the human motilin receptor and utilized at a quantity of 1.5 pglassay point. [PerkinEImerTM
SignaIScreen zo Product #6110544]
. [251]-Motilin (PerkinElmer, #NEX-378); final concentration: 0.04-0.06 nM
~ Motilin (BachemTM, #H-4385); final concenfiration: 1 ~,M
~ Multiscreen Harvest plates-GF/B (MilliporeT"", #MAHFB1 H60) ~ Deep-well polypropylene titer plate (Beckman CouIterTM, #267006) ~ TopSeal-A (PerkinElmer, #6005185) ~ Bottom seal (Millipore, #MATAHOP00) ~ MicroScint-0 (PerkinElmer, #6013611) ~ Binding Buffer: 50 mM Tris-HCI (pH 7.4), 10 mM MgCl2, 1 mM EDTA, 0.1 % BSA
2 0 Assay Volumes:
~ 150 ~,L of membranes diluted in binding buffer ~ 10 ~.L of compound diluted in binding buffer ~ 10 p,L of radioligand ([251]-Motilin) diluted in binding buffer 2 5 Final Test Concentrations (N=11 ) for Compounds:
10, 5, 2, 1, 0.5, 0.2, 0.1, 0.05, 0.02, 0.01, 0.005 pM.
Compound Handling:
Compounds were provided frozen on dry ice at a stock concentration of 10 mM
diluted in 3 0 100% DMSO and stored at -20°C until the day of testing. On the test day, compounds were allowed to thaw at room temperature and than diluted in assay buffer according to the desired test concentrations. Under these conditions, the maximum final DMSO
concentration in the assay was 0.5%.
Assa rLProtocol:
In deep-well plates, diluted cell membranes (1.5 ~,g/mL) are combined with 10 pL of either binding buffer (total binding, N=5), 1 p,M motilin (non-specific binding, N=3) or the appropriate concentration of test compound. The reaction is initiated by addition of 10 p,1 of [251]-motilin (final conc. 0.04 - 0.06 nM) to each well. Plates are sealed with TopSeal-A, 1 o vortexed gently and incubated at room temperature for 2 hours. The reaction is arrested by filtering samples through pre-soaked (0.3% polyethyleneimine, 2 h) Multiscreen Harvest plates using a Tomtec Harvester, washed 9 times with 500 ~rL of cold 50 mM
Tris-HCI (pH
7.4), and than plates are air-dried in a fumehood for 30 minutes. A bottom seal is applied to the plates prior to the addition of 25 ~.L of MicroScint-0 to each well.
Plates are than 15 sealed with TopSeal-A and counted for 30 sec per well on a TopCount Microplate Scintillation and Luminescence Counter (PerkinElmer) where results are expressed as counts per minute (cpm).
Data are analyzed by GraphPadT"" Prism (GraphPad Software, San Diego, CA) using a variable slope non-linear regression analysis. K; values were calculated using a Kd value 20 of 0.16 nM for [251]-motilin (previously determined during membrane characterization).
DmaX = 1 - test concentration with maximal displacement - non-specific binding x 100 total binding - non-specific binding 25 where total and non-specific binding represent the cpm obtained in the absence or presence of 1 pM motilin, respectively.
Example Method B2: Aequorin Functional Assay (Motilin Receptor) Materials:
30 ~ Membranes were prepared using AequoScreenT"" (EUROSCREEN, Belgium) cell lines expressing the human motilin receptor (cell tine ES-380-A; receptor accession #AF034632). This cell line is constructed by transfection of the human motilin receptor into CHO-K1 cells co-expressing Ga~6 and the mitochondrially targeted Aequorin (Ref #ES-WT-A5).
~ Motilin (Bachem, #H-4385) ~ Assay buffer: DMEM-F12 (Dulbeccoe's Modified Eagles Medium) with 15 mM HEPES
5 and 0.1 % BSA (pH 7.0) ~ Coelenterazine (Molecular ProbesT"", Leiden, The Netherlands) Final Test Concentrations (N=5) for Compounds:
10, 3.16, 1, 0.316, 0.1 pM.
Compound Handling:
Compounds were provided as dry films at a quantity of approximately 1.2 ~,mol in pre-formatted 96-well plates. Compounds were dissolved in 100% DMSO at a concentration of 10 mM and stored at -20°C until further use. Daughter plates were prepared at a concentration of 500 p.M in 30% DMSO with 0.1 % BSA and stored at -20°C
until testing.
On the test day, compounds were allowed to thaw at room temperature and than diluted in assay buffer according to the desired test concentrations. Under these conditions, the maximum final DMSO concentration in the assay was 0.6%.
2 o Cell Preparation:
Cells are collected from culture plates with Ca2+ and Mg2+-free phosphate buffered saline (PBS) supplemented with 5 mM EDTA, pelleted for 2 minutes at 1000 x g, resuspended in assay buffer (see above) at a density of 5 x 106 cells/mL and incubated overnight in the presence of 5 p,M coelenterazine. After loading, cells were diluted with assay buffer to a 2 5 concentration of 5° x 105 cells/mL.
Assay Protocol:
For agonist testing, 50 p1 of the cell suspension was mixed with 50 p,1 of the appropriate concentration of test compound or motilin (reference agonist) in 96-well plates (duplicate 3 o samples). The emission of light resulting from receptor activation was recorded using the Functional Drug Screening System 6000 'FDSS 6000' (Hamamatsu Photonics K.K., Japan).

For antagonist testing, an approximate EC80 concentration of motilin (i.e. 0.5 nM; 100 ~,L) was injected onto the cell suspension containing the test compounds (duplicate samples) 15-30 minutes after the end of agonist testing and the consequent emission of light resulting from receptor activation was measured as described in the paragraph above.
Results are expressed as Relative Light Units (RLU). Concentration response curves were analyzed using GraphPad Prism (GraphPad Software, San Diego, CA) by non-linear regression analysis (sigmoidal dose-response) based on the equation E=Em~/(1+ECSO/C)n 1o where E is the measured RLU value at a given agonist concentration (C), EmaX is the maximal response, EC5o is the concentration producing 50% stimulation and n is the slope index. For agonist testing, results for each concentration of test compound were expressed as percent activation relative to the signal induced by motilin at a concentration equal to the ECso (i.e. 0.5 nM). For antagonist testing, results for each concentration of 15 test compound were expressed as percent inhibition relative to the signal induced by motilin at a concentration equal to the EC$o (i.e. 0.5 nM).
Example Method B3: FIashPlate Motilin [35S]-GTPyS Functional Assay 2 o Materials:
~ Membranes were prepared from CHO cells stably transfected with the human motilin receptor and utilized at a quantity of 1.5 p,g/assay point.
[PerkinElmer SignaIScreen Product #6110544]
~ GTPyS (Sigma, #G-8634) 25 ~ (35S]-GTPyS (PerkinElmer, #NEX-030H) ~ Motilin (Bachem, #H-4385) ~ 96-well FIashPlate microplates (PerkinElmer, #SMP200) Deep-well polypropylene titer plate (Beckman Coulter, #267006) ~ TopSeal-A (PerkinElmer, #6005185) 3 0 ~ Assay Buffer: 50 mM Tris (pH 7.4), 100 mM NaCI, 10 mM MgCl2, 1 mM EDTA, 1 p.M
GDP, 0.1 % BSA

Assay Volumes:
~ 25 ~L of compound diluted in assay buffer ~ 25 ~,L of assay buffer (agonist assay) or 0.6 ~,M motilin (0.1 ~,M final concentration) diluted in assay buffer (antagonist assay) 100 p,L of [35S]-GTPyS diluted in assay buffer Final Test Concentrations (N=12) for Compounds:
50, 20, 10, 5, 2, 1, 0.5, 0.2, 0.1, 0.05, 0.02, 0.01 pM.
Compound Handling:
Compounds were provided frozen on dry ice at a stock concentration of 10 mM
diluted in 100% DMSO and stored at -20°C until the day of testing. On the test day, compounds were allowed to thaw at room temperature and than diluted in assay buffer according to the desired test concentrations. Under these conditions, the maximum final DMSO
concentration in the assay was 0.5%.
Assay Protocol:
CHO membranes were immobilized into 96-well FIashPlate microplates. Test compound, 2 o GTPyS, motilin and [35S]-GTPyS were combined in each well according to the Assay Volumes described above.
For the assay to measure agonist activity, an additional 25 p,1 of buffer was added to each well in addition to 25 p,L of either buffer (basal value, N=4), 1 ~,M (final cone) motilin (EmaX
value, N=3), 25 p,M (final cone) GTPyS (non-specific value, N=4), or the appropriate concentration of test compound (N=3).
For the assay to measure antagonist activity, an additional 25 p,L of either buffer (unstimulated control) or motilin (0.1 ~.M final cone) is added to each well, in addition to either 25 ~,L of buffer (basal value,.N=3), 1 ~.M (final cone) motilin (Emax value, N=3), 25 ~M (final cone) GTPyS (non-specific value, N=4), or the appropriate concentration of test compound (N=3).

The reaction is initiated by addition of 100 mL of [35S]-GTPyS to each well.
Each plate is sealed (TopSeal-A) and incubated in the dark at room temperature for 150 min.
Then, plates are counted for 30 seconds per well on the TopCount NXT.
Data were analyzed by GraphPad Prism 3.0 (GraphPad Software, San Diego, CA) using non-linear regression analysis (sigmoidal dose-response) for the calculation of ICSO/ECSo values.
EmaX (agonist) or DmaX (antagonist) = Top - Bottom X 100 Bottom Where Top and Bottom correspond to the top and bottom values of the dose-response curve calculated by GraphPad Prism).
Example Method B4: Rabbit Duodenum Contractility Assay Duodenal segments were vertically suspended in organ chambers of 10 mL filled with Krebs buffer and connected to an isotonic force transducer, with a preload of 1 g. After a stabilization period, the muscle strips were challenged with 10-4 M
acetylcholine and washed. This was repeated until a stable maximal contraction was obtained (2-3 times), 2 o with an interval of at least 20 minutes.
After a stable base line was reached, test compounds were added to the bath.
After 15 min incubation, a dose response to motilin was recorded by adding logarithmically increasing concentrations of motilin to the bath (final concentration 10-9 to 10-6 M). A blank experiment (no test compound present) was also performed. At the end of the dose response curve, a supramaximal dose of acetylcholine (10-4 M) was given and this response was used as a reference (100% contraction).
The results of experiments at different concentrations of test compound were combined and analyzed to derive the pA2 value from the Schild plot.
It is appreciated that although specific experimental methods have been described herein for the purposes of illustration, various modifications to these experimental methods as well as alternate methods of experimentation may be used without departing from the scope of this invention.
Table 3: Binding activity of selected compounds R~ R3 Rs T Ki~,a \/ \
1 ~ ~ ~ \ zs °" / ~x B

~ \ za o~x A
3 \ /~ - I Z3 I B
o/\/x zs A
off ~x '-( off \ ~ Z3 CN3 - ~ I / x B
o'~
i ~' ~ ! \ \ z3 B
~x Zs o~X
\ \ Z3 off / \ / ~ o/\/x / °
\ \ za A
,~''' off - / /\/x - - ~ \ ~ Z3 A
11 \ / / - / °~x s _ 4 \ \
12 '~ \ ~ off - /
°~x 13 ~ \ ~ ~ \ zs / o/\/x ~ w ~ Z3 14 _ / ~~x a 15 ~ ~ \ ~ \z3 A
~x °
w 16 ~ ~ ~ '.~ ~ \ z3 A
~x - --~ ~ ~ o/\/x H

18 ~ - \ B
v / ° _ ~ -~~ ~ Z3 'X

19 q 'X
2u ~. \ / ~ \

'X
' ~O
21 ~ ~ \ q - v ~z3 ~x ,, V ,- ~o 22 - \ A
,.a" ~ ~ v ~Z3 /X
' VO

--\, zs A
23 ~ / °~x \ za A
24 ~ ~ °~x ci ~ ~ B
25 ~ / ~x v ~o A
26 ~ - ~ ( ~ / x OH
\, Z3 B
27 ~ ~ ~ ~ / °~x ~s U w za B
28 0 ~ ~ ~ / ~x \ a ~ za B
29 ~'' ° .- ~ / o/\/x w \z B
30 ~'' \ , aH ~ / ~x 57 ~ - - \ \ gzs OH
O / X
O
58 ~,, - _ ~ \ \ zs g OH ~ o/\
59 ~' ~ ~ ~ za g ~\ i o I
60 ~ ~ ~ ~ \ za C
a s5 .,~'' _ g X ~ Z3 71 x/U ~ °~b B
_~
72 ~ / off ~~ ~ ~ ~ / ~ ~ \ z3 g ~x 7s off / ~ ~ \ z3 C
~x 77 _ ~ ~ \ \ zs C
°
\ / °" -~ ~ ~ ~x 80 ~ ~ I \ ~ Z3 a \ / ~ ~ °~x 8s ,~.~ ° -~~ H \
~x °

\ zs \ / \ / ~ °~x as ~ - \ ~ \z3 c \ / ~ -~ ~ / o~x c 89 ~ \ zs \ / , °~x - \ Z3 ~x \ / °
91 ~' \ / - _ ~ ~ \ zs C
~x -~ ~ / X3 ~O~
96 ~,,,, H \ \/ \~3 C
'X
/ ~O
/ ~N
s~ .~'' OFI ~ \ 23 /X
/ ~O
gg ~ C
X v v N v \Z3 99 .~~OH C
X/ v l v \Z3 _ ~NHZ ~ \ Z3 ~X
O
110 ~ N
OH _ - ~ Z3 'X
' VO
111 ~''~ ~ zs U
/ OH
X
v ~ \zs - / ~x i.,,,, \ N \ zs I / ~ ~x o~

N
\~

x 124, B
\ Z3 ~N _~ ~ I / o~x I \ ~ ~ \
~ \ zs ~oH ' a t x / o~

I \ \ zs 'oH ~ -~ / ~ x x Z3 _ - / S \
i .... \ ~ /

/ ~ ~ ~ \ zs / x ',~i \ ~, \ I~
, , -, \ \/ \ zs ._.. _~ . ~ ~ x -.
A

v \ ci ,' \ za / ~x -.
o , ~ \ ~ \ z3 ,,, I \ _~ ~ x o~
, U\~, _.
A

w ~ zs I ~ ~x -; \o a ci 134 ,'~ ~ \ '~ \ CI
, _ -.
135 , \ za _ ~ ~ w x , 136 ~ -' N
\ \ ~ \/ \~
i x ~ , " ~ '~ z3 _; i o G N

_~ . x B

N
~ Z3 ,, I W _~ , / ~x -, i , G ;
s N
~ X3 i i i -n A

~ ~ zs o/ ~ o~ x ~% 'o' A
142 ~
~ z3 o~ x _.

/ /

A
144 \ /
za ~x / / ' 145 - _ - A
~ 1 I\
x ' _.
A
146 ,, ~I ' /~, I \ \

/ . ~ ~ ~x ' o _.

/,,, I \ N
\
' CI X
n ' N
j~ ~ ~ ~ \ ~~Z3 F

N
' s ~
~X
_.
' o N
t \ \ ~ \ Z3 / ~X
o _.

/ _~ I , ~X
F
' ' i i \ - NH2 ~ ~ \ zs / ~X

\ \/ \ X
\ \ /

NHZ
-.-..... ~ \
' - ' ' x ' s NH2 ~ \ \/ \ za ' / -~ ' ~ /x / o~
156i~, ~ \ NHz \ za A
ci / o/~/ x ~ \ - /NHZ \
/ - ' \/ \ zs ' ' ' x / o~

NHZ N
w \ ci -~ I, \ za ' ' x / o~

\ \/ \ Za ' -' x OH

- \ Z3 ' ~ -i v 'OH ~ X
/

\

_ a ~x / ~O

\ \ za / -~ / ~x \ - ~ ~ \ \/\x _ a \ ~ ~ za a ~x ;~ ~ ~ za a x a o~
166 \ -\ za , ~ - I ~ / ~ /x _ . ~ ~ x ._ ;

\ za -~~ / ~ x y;
' ~ ~o ci _ _ ~' I ~ za I / ~X
I
~/ ~/1 ci I
ci ; ~ ' x ' o~

' F
/'\ ~ - ~ \ ~ \ Z3 ~~

/ . / ~X , F i \ ~ ~ \ 23 I
I
/ ~X
~/ v1 B

I ~ \~ \ Z3 I
I
/ ~ /X
I

~ zs -I
I x Is ~ \ \/ \
/ ~x I o OH I

~ ~~za ~ ~ _;~ I / X

' \
, , ' ( . / ~x OH

~ \ zs , x / o~

'' - ~ H ~ za , , / ~x , \ ~ ~ o / /

~ -~ OH ~ zs _____ ~ \/ \ x , /

ci OH
\/ \ za , , / . , ~x 182 , ,,,v \ ~ \ zs / ci -~ % ~OH / ~ x , ~ = ~ H ~ \ zs , / . / ~x F ' l 184 -. V B
-~NHZ ~ ~ za , \ \ . / ~x za ~ ~X
B

z3 X
187 _ A
F
F
- ~ Z3 F ;
i /X

F , i - ~ I ~ 23 / ~X
/ ~O

>~, w - ; \ \/ \ ~
/o- n i ~ ~ ~ O~ X

z3 o~ x i - , _ 191 ' A
\ o~ ~ \ ~ \ zs I
/ . ~ ~ o~ X
192 ' \ Ci A
I ~ ~ Z3 I, X
O~
_, \ F
_ Z3 n X

B
>~, I w , 1 \N

x ,,,, I \
\ \/ \
X
o \ G H
,\
HZN '~~ ~ ~ Z3 ~ ~ ~. x -, G o \N Z3 ~I X
-i O N

,' I \ G
s _ ; ~ o \ G
/,,,~ -X

_n r HOc .~~/~~"OH
A
200 , ci v \X
~ ~/
-~ o °
201 ~- °~ ~ ~ \ z3 ~x \ \ . ~ / o / A
\ ~'' zs ~. ~,~' I
/
B

203 ~ .~ HN \ z3 I ',~
a A
NH

x ~1~ ~. 1 j~N~NHz I ~ . H ~~-\ ~ ~ z3 x / o/
I / ' B
----- o HN-- ~~~ ~ Z3 _.;--l o I /
B
z3 /\~ x s~~NHz B
a ~ 23 N ~ / v X
NHz 209 ~ C
- ~N I \ za / ; / ~x OH ;

. i \ -~ ,~N ~ \
'OH / o~ X

z z3 \c1 / O~ X

i\ \ - j ,\ ~ \ za /
/ a \ 1 ,~ \ \/ \ Za / ~x o / OH
I

zs / o/ ;
~x x ~ \ \N ~ zs off A

~ c1 \
%''', ~ ~ ~ \ za / o~ x - , -I

', ci ~ za - . ~ / ~x \ c' - ~ ( \ z3 ' O' _;~ x o ~' ci ' I ~ ~ \ za s -~ ' x ' o ,.... ~ \ F , \
-' za / - ' ~x \ \/\~
/ ~x 222 ~ '~' \ \ \/ \ z3 / ; / ~x i O

\ za / ' on -~~ ~ ~ x w \ zs \ \ ~ / x ' o~
0 0 -' \ \/ \ z3 \ \ - ~ / ~x / / ' \ Z3 t _ i I , ~ -~ / o~ x . ~ ~~zs \ \ ~ ; ~ ~~/ x a / .
5 Notes Radioligand competitive binding assays performed using Method B1 Values reported as ranges: A = 0.001-0.100 ~,M; B = 0.100-1.0 wM;
~ o C = 1.0-10.0 ~,M
X is NH except for:
Compound 223 and 225, X is:
,,, ...% ';
Compound 224, X is NMe Compound 226, X is:
2 0 .i ,, N
~,i~
("S

Gompound 227, X is -..,i N .~.
\:' Z~, Z2 and Z3 are NH except for compounds 30, 173 and 174 and where Z1 is O
and compound 111 where ZZ is O.
R2, R4 and R5 are hydrogen except for compound 85 where it is ~5 m, n~ and p are zero.

Claims (15)

1. A compound represented by the general formula (I) and pharmaceutically acceptable salts, hydrates or solvates thereof wherein:
Z1, Z2 and Z3 are independently selected from the group consisting of O, N and NR10, wherein R10 is selected from the group consisting of hydrogen, lower alkyl, and substituted lower alkyl;
R1 is independently selected from the group consisting of lower alkyl substituted with aryl, lower alkyl substituted with substituted aryl, lower alkyl substituted with heteroaryl and lower alkyl substituted with substituted heteroaryl;
R2 is hydrogen;
R3 is independently selected from the group consisting of alkyl and cycloalkyl with the proviso that when Z1 is N, R3 can form a four, five, six or seven-membered heterocyclic ring together with Z1;

R4 is hydrogen;

R5 and R6 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl and substituted heteroaryl, with the proviso that at least one of R5 and R6 is hydrogen;
X is selected from the group consisting of O, NR8, and N(R9)2+;
- wherein R8 is selected from the group consisting of hydrogen, lower alkyl, substituted lower alkyl, formyl, acyl, carboxyalkyl, carboxyaryl, amido, sulfonyl, sulfonamido and amidino; and - R9 is selected from the group consisting of hydrogen, lower alkyl, and substituted lower alkyl;
m, n1 and p are independently selected from 0, 1 or 2; and T is a bivalent radical of formula II:

-U-(CH2)d-W-Y-Z-(CH2)e- (II) wherein d and e are independently selected from 0, 1, 2, 3, 4 or 5;
wherein U is bonded to X of formula (I) and is -CH2- or -C(=O)- ;
wherein Y and Z are each optionally present;
W, Y and Z are independently selected from the group consisting of: -O-, -NR28-, -S-, -SO-, -SO2, -C(=O)-, -C(=O)-O-, -O-C(=O)-, -C(=O)NH-, -NH-C(=O)-, -SO2-NH-, -NH-SO2-, -CR29R30-, -CH=CH- with a configuration Z or E, and -C.ident.C-, or from a ring structure independently selected from the group wherein any carbon atom contained within said ring structure, can be replaced by a nitrogen atom, with the proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms:
G1 and G2 each independently represent a covalent bond or a bivalent radical selected from the group consisting of -O-, -NR41-, -S-, -SO-, -SO2-, -C(=O)-, -C(=O)-O-, -O-C(=O)-, -C(=O)NH-, -NH-C(=O)-, -SO2-NH-, -NH-SO2-, -CR42R43-, -CH=CH-with a configuration Z or E, and -C.ident.C-;with the proviso that G1 is bonded closer to U than G2;
K1, K2, K3, K4, K6' K15 and K16 are independently selected from the group consisting of O, NR44 and S;
f is selected from 1, 2, 3, 4, 5 or 6;
R31, R32, R38, R39, R48 and R49 are independently selected from hydrogen, halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido; and R33, R34, R35, R36, R37, R47, R50 and R51 are independently selected from hydrogen, halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido.

2. A compound as claimed in claim 1 wherein R1 is selected from the group consisting of -(CH2)q R11, and -CHR12R13 wherein q is 0, 1, 2 or 3; and R11 and R12 are independently selected from a ring structure from the following group:
wherein any carbon atom in said ring structure can be replaced a nitrogen atom, with the proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms;

A1, A2, A3, A4 and A5 are each optionally present and are independently selected from the group consisting of halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido;

B1, B2, B3, and B4 are independently selected from NR14, S or O, wherein R14 is selected from the group consisting of hydrogen, alkyl, substituted alkyl, formyl, acyl, carboxyalkyl, carboxyaryl, amido, sulfonyl and sulfonamido;

R13 is as defined for as R11 and R12 or is selected from the group comprising lower alkyl, substituted lower alkyl, hydroxy, alkoxy, aryloxy, amino, carboxy, carboxyalkyl, carboxyaryl, and amido.

3. A compound as claimed in claim 1 wherein R3 is selected from the group consisting of:
-(CH2)s CH3, -CH(CH3)(CH2)t CH3, -CH(OR15)CH3, -CH2SCH3 -CH2CH2SCH3, -CH2S(=O)CH3, -CH2CH2S(=O)CH3, -CH2S(=O)2CH3, -CH2CH2S(=O)2CH3, -(CH2)u CH(CH3)2, -C(CH3)3, and -(CH2)y-R21, wherein:
s and a are independently selected from 0, 1, 2, 3, 4 or 5;
t is independently selected from 1, 2, 3 or 4;
y is selected from 0, 1, 2, 3 or 4;
R15 is selected from the group consisting of hydrogen, alkyl, substituted alkyl, formyl and acyl;
R21 is selected from a ring structure selected from the following group:
wherein any carbon atom in said ring structure can be replaced by a nitrogen atom, with the proviso that if said ring structure is a monocyclic ring structure, it does not comprise more than four nitrogen atoms and if said ring structure is a bicyclic ring structure, it does not comprise more than six nitrogen atoms;

z is selected from 1, 2, 3, 4 or 5;
E1, E2 and E3 are each optionally present and are independently selected from the group consisting of halogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido;
amidino, cyano, nitro, mercapto, sulfinyl, sulfonyl and sulfonamido; and J is optionally present and is selected from the group consisting of alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, mercapto, sulfinyl, sulfonyl and sulfonamido.

4. A compound as claimed in claim 1 wherein R5 and R6 are each independently selected from the group consisting of hydrogen, -(CH2)aa CH3, -CH2SCH3 -CH2CH2SCH3, -CH2S(=O)CH3, -CH2CH2S(=O)CH3, -CH2S(=O)2CH3, -CH2CH2S(=O)2CH3, -(CH2)bb CH(CH3)2, -CH(CH3)(CH2)cc CH3, -(CH2)dd-NR22R23, -(CH2)ee R24, -and -CH2CONR26R27, with the proviso that at least one of R5 or R6 is hydrogen;

wherein as and bb are independently selected from 0, 1, 2, 3, 4 or 5;

cc and dd are independently selected from 1, 2, 3, 4 or 5;

ee is selected from 0, 1, 2, 3 or 4;

R22 and R23 are independently selected from the group consisting of hydrogen;
alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl , substituted heteroaryl, formyl, acyl, carboxyalkyl, carboxyaryl, amido, amidino, sulfonyl and sulfonamido; with the proviso that R22 and R23 may form, taken together with a nitrogen atom, a five-or six-membered substituted or unsubstituted heterocyclic ring;

R24 is as defined for R11 and R12; and R25 is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heterocyclic, substituted heterocyclic, heteroaryl and substituted heteroaryl;
R26 and R27 are as defined for R17 and R18.

5. A compound as claimed in claim 2, wherein A1, A2, A3, A4 and A5 are each optionally present and are independently selected from halogen, trifluoromethyl, C1-6 alkyl or C1-6 alkoxy.

6. A compound as claimed in claim 2, wherein R11, R12 and R13 are each independently selected from the group consisting of:
wherein R a and R b are chosen from the group consisting of Cl, F, CF3, OCH3, OH, CH3 and C(CH3)3

7. A compound as claimed in claim 1 wherein T is selected from the group consisting of:

wherein L1 NH or NMe; L2 is CH or N; L3 is CH or N; L4 is O or CH2; L5 is CH
or N
L6 is CR52R53 or O; R46 is H or CH3;
R52, R53, R54, R55, R56 and R57 are independently selected from hydrogen, lower alkyl, substituted lower alkyl, hydroxy, alkoxy, aryloxy, amino, and oxo; or R52 together with R53 or R54 together with R55 or R56 together with R57 can independently form a three to seven-membered cyclic ring comprising carbon, oxygen, sulfur and /or nitrogen atoms;
(X) is the site of a covalent bond to X in formula (I); and (Z3) is the site of a covalent bond to Z3 in formula (I).

8. A compound as claimed in claim 1 wherein m, n and p are 0; X, Z1, Z2 and Z3 are NH;
and R2, R4 and R5 are hydrogen, represented by formula (III):

9. A compound as claimed in claim 1, wherein R3 forms a four, five, six or seven-membered heterocyclic ring together with Z1 ,represented by formula (IV):
wherein said heterocyclic ring may contain a second nitrogen atom, or an oxygen, or sulfur atom;
n2 is selected from 0, 1, 2 or 3 R7 is optionally present and is selected from the group consisting of alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substituted heterocyclic, aryl, substituted aryl, heteroaryl, substituted heteroaryl, hydroxy, alkoxy, aryloxy, oxo, amino, halogen, formyl, acyl, carboxy, carboxyalkyl, carboxyaryl, amido, carbamoyl, guanidino, ureido, amidino, mercapto, sulfinyl, sulfonyl and sulfonamido.

10. A compound as claimed in claim 1 selected from the group consisting of:

<

11.An antagonist of the motilin receptor with a structure as defined in anyone of claims 1 to 10.

12.A method of treating a disorder associated with the motilin receptor or motility disfunction in humans and other mammals comprising administering a therapeutically effective amount of a compound as claimed in anyone of claims 1 to 10.

13. A method of treating a disorder associated with hypermotility or hypermotilinemia in humans and other mammals comprising administering a therapeutically effective amount of a compound as claimed in anyone of claims 1 to 10.

14.A method of treating irritable bowel syndrome and dyspepsia in humans and other mammals, comprising administering a therapeutically effective amount of a compound as claimed in anyone of claims 1 to 10.

15.A method of treating treating Crohn's disease, gastroesophogeal reflux disorders, ulcerative colitis, pancreatitis, infantile hypertrophic pyloric stenosis, carcinoid syndrome, malabsorption syndrome, diarrhea, diabetes mellitus, obesity, atrophic colitis or gastritis, gastric stasis, gastrointestinal dumping syndrome, postgastroenterectomy syndrome, celiac disease and eating disorders leading to obesity in humans and other mammals comprising administering a therapeutically effective amount of a compound as claimed in anyone of claims 1 to 10.