Thapsigargin, Origin, Chemistry, Structure-Activity Relationships and Prodrug Development

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Current Pharmaceutical Design, 2015, 21, 5501-5517 5501

Thapsigargin, Origin, Chemistry, Structure-Activity Relationships and Prodrug
Development
Nhu Thi Quynh Doan and Søren Brøgger Christensen*
Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenha-
gen, Copenhagen, Denmark
Abstract: Thapsigargin was originally isolated from the roots of the Mediterranean umbelliferous plant Thapsia
garganica in order to characterize the skin irritant principle. Characteristic chemical properties and semi-syntheses
are reviewed. The biological activity was related to the subnanomolar affinity for the sarco/endoplasmic reticulum
calcium ATPase. Prolonged inhibition of the pump afforded collapse of the calcium homeostasis and eventually
apoptosis. Structure-activity relationships enabled design of an equipotent analogue containing a linker. Conjuga-
tion of the analogue containing the linker with peptides, which only are substrates for either prostate specific anti-
gen (PSA) or prostate specific membrane antigen (PSMA) enabled design of prodrugs targeting a number of cancer
diseases including prostate cancer (G115) and hepatocellular carcinoma (G202). Prodrug G202 has under the name Søren B. Christensen
of mipsagargin in phase II clinical trials shown promising properties against hepatocellular carcinoma.
Keywords: Thapsigargin, mipsagargin, drug development, clinical trials, structure activity relationships, prostate specific antigen, prostate
specific membrane antigen, prodrug, anti-angiogenesis.
1. INTRODUCTION prodrugs selectively activated in tumors. Mipsagargin a prodrug
Already Theophrastos (372 – 287 B.C.) described the skin irri- designed only to be cleaved in neovascular tumors has in phase II
tant properties of the resin of Mediterranean plant Thapsia gargani- clinical trials showed an encouraging effect against sorafenib re-
ca L. (Apiaceae, previous Umbelliferae). Later scholars like Dios- sistant patients.
corides (approximately 50 A.D.) and Plinius (24 – 79 A.D.) also
mentioned the use of preparations from the plant for treatment of 2. CHEMISTRY
e.g. pulmonary diseases, catarrh and as a counterirritant for relief of
rheumatic pain [1, 2]. Radix thapsiae and resina thapsiae have been 2.1. The Structures and Diversity of Polyoxygenated Guaiano-
included in several pharmacopeias latest in the French pharmacope- lides
ia from 1937. In spite of this extended traditional use of the plant
Thapsigargin (1) was isolated together with the related
the first publication describing the constitution was not published
thapsigargicin (2, Tc, Fig. 1). Besides Thapsigargin (1) several
before 1978 [3]. The relative configuration of the major skin irritat-
hexaoxygenated guaianolides only differing from Tg (1) by the acyl
ing principle of thapsigargin (1, Tg, Fig. 1) was published in 1980
groups at O-2 and O-8 have been found in the genus Thapsia (2-13,
[4] and 1982 [5] and the absolute configuration in 1985 [6]. A
Fig. 1) [14, 15]. Two additional thapsigargins have been found in
number of cellular assays revealed that the compound was a very
Laser trilobum Borkh. (14, 15, Fig. 2) [16]. As well the hexa- as the
potent secretagogue for histamine release from peritoneal mast cells
pentaoxygenated guaianolides are only present in either Thapsia
[7, 8] and activated a number of cells belonging to the humane in- species or in L. trilobum (16-18, Fig. 3). Comparison of the 1H and
flammatory response [9]. In addition to being a skin irritant Tg (1) 13C NMR spectra of Tg (1) and the related Tc (2) revealed that both
also showed to be a cocarcinogen facilitating skin cancer develop-
compounds possessed an acetoxy, a butanoyloxy and an an-
ment in mice [10]. Intensive interest for Tg (1), however, first ap-
geoyloxy group present on the hydrogen poor guaianolide skeleton.
peared when the potent biological activities were related to the
In addition the presence of an octanoyloxy residue was present in
ability of inhibiting the Sarco/Endoplasmic Reticulum Calcium
Tg (1) whereas a hexanoyloxy group was present in Tc (2). Com-
ATPase (SERCA) [11]. Today Tg (1) has become a positive stand-
parison with the spectra of trilobolide (16, Fig. 3) [17], which just
ard in all experiments for calcium homeostasis in cells [12]. Pro-
had been published at this time, revealed the absence of the two
longed inhibition of the SERCA pump affords a persistent high
protons at C-2, whereas an additional acyloxy group was present.
concentration of calcium ions in the cytosol, which after 12 – 24
A combination of these observations led to the suggestion of the
hours induces apoptosis [13]. The ubiquitous presence of SERCA
constitution of Tg (1) [4]. The poor number of protons at the guaia-
in all living cells and the subnanomolar affinity for the SERCA
nolide skeleton prevented establishment of the relative configura-
pump therefore makes Tg (1) a potent universal cell toxin. Howev-
tion. Fortunately, treatment of Tg (1) with thionyl chloride afforded
er, the overexpression of proteolytic enzyme in neovascular tissue
a crystalline epoxide 19 (Scheme 1) the structure of which was
in tumors and the presence of prostate specific antigen only in the
solved through an X-ray crystallographic analysis [18]. The X-ray
prostate gland or in prostate cancer tumors have enabled design of
analysis did neither enable establishment of the absolute configura-

tion nor of the relative configuration at C-7 or C-11.

Resistance towards periodic acid of the debutanoyl thapsigargin
*Address correspondence to this author at the Department of Drug Design 20 (Scheme 2), however, revealed that the three hydroxyl groups
and Pharmacology, Faculty of Health and Medical Sciences University of had to be trans disposed.
Copenhagen, Universitetsparken 2, DK-2100 Copehagen Ø, Denmark;
Tel/Fax: [phone];
E-mail: [email protected]

1381-6128/15 $58.00+.00 © 2015 Bentham Science Publishers
5502 Current Pharmaceutical Design, 2015, Vol. 21, No. 38 Doan and Christensen
R1O H14 O f romT hsep e1c(cid:1)ie-ds isbpeolosendg iHng-1 tios tAyppiiaccaellaye fo(uumndb einll igfeuraoiauns oplildaenst si)s obluatte da
O
O O 321 1098 O R2 f(ceowm epxoasmitepsl)e s[ 1h9a]v. e been found in plants belonging to Asteraceae
4 5 6 7 OH
15 O 11 OH 2.2. Chemistry of Polyoxygenated Guaianolides
12 13 From a chemical point of view the polyoxygentaed guaianolides
O
possess three functional groups: ester groups, tertiary alcohols and a
Thapsigargin (1): R1 = Oct, R2 = But double bond between C-4 and C-5. In spite of this, methods have
Thapsigargicin (2): R1 = Hex, R2 = But been developed for selective modifications of the structures.
Thapsitranstagin (3): R1 = iVal, R2 = 2-MeBut 2.2.1. Epoxide Formation
Thapsivillosin A (4): R1 = Ang, R2 = Sen Treatment of as well pentaoxygenated as hexaoxygenated guai-
Thapsivillosin B (5): R1 = Ang, R2 = 2-MeBut anolides converts the 7,11-dihydroxy diol into an epoxide with
Thapsivillosin C (6): R1 = Oct, R2 = 2-Mebut conversion of the stereochemistry at C-11 (Scheme 1). Epoxide
Thapsivillosin D (7): R1 = 6-MeOct, R2 = Sen formation from a diol is a very seldom reaction and probably locked
Thapsivillosin E (8): R1 = 6-MeOct, R2 = 2-Mebut optimal conformation of the 7- and 11-hydroxy groups enables the
Thapsivillosin G (9): R1 = 6-MeHep, R2 = 2-Mebut reaction [17, 18].
Thapsivillosin H (10): R1 or R2 = Ang or Sen
Thapsivillosin I (11): R1 = Ang, R2 = But
Thapsivillosin J (12): R1 = iVal, R2 = But O O
Thapsivillosin K (13): R1 = Sen, R2 = 2-MeBut O O H O O O SOCl2 O O H O O O
O O O O
O O O OH O
Ang But Hex 1 O OH 19O
O O
O O Scheme 1. Conversion of thapsigargin (1) into epoxide (19).
2-MeBut 6-MeHep
2.2.2. Selective Hydrolysis of the Ester Group at O-8
O O
Treatment of as well penta- as hexaoxygenated guaianolides
6-MeOct Non with triethylamine in a protolytic solvent like methanol affords a
selective hydrolysis of the O-8 ester group (Scheme 2) [20]. The
O O O hydrolysis of the labile O-8 ester group might be facilitated by the
Oct Sen iVal juxtaposed OH-11 since the hydrolysis of the ester group in the
epoxide 19 demands harsher reaction conditions. The use of a

stronger base like sodium carbonate results in opening of the 6,12-
Fig. (1). Structures of naturally occurring hexaoxygenated guaianolides
lactone. As a consequence, a mixture of the 6,12- (20) and 8,12-
found in Thapsia [thapsigargins] (1-13) with traditional numbering.
(21) guaianolides are obtained after workup under acidic conditions
(Scheme 2) [18].
R1O H14 O R2 O
O O O
O 324 15 160798 OOH O O OH O OOO Et3N O O O H O OOH
15 O 11 OH OH MeOH OH
12 13 O OH O OH
O 1 O 20 O
2-Hydroxy-10-desacetoxytrilobolide (14): R1 = R2 = H Na2CO3 (aq)
2-Acetoxytrilobolide (15): R1 = R2 = Ac MeOH
O O
fFoiugn. d(2 i)n. LS.t rturicltoubruesm o [ft htahpes nigaaturgrainllsy] (o1c4c-u1r5ri)n. g hexaoxygenated guaianolides O O O H O OOH H 20 O O O H O OO
H-OO OOHH HOHHOOCH3O
14 O 19 O 21
H
O
O O 32 11098 OR Scheme 2. Formation of 6,12- (20) and 8,12- (21) guaianolides.

4 5 6 7 OH 2.2.3. Selective Substitutions of the Ester Group at O-2, O-10 and
15 O 11 OH O-8
12 13
O Masking of the OH-8 and OH-11 by reaction with 2,2-dime-
thoxypropane yields an isopropylidende derivative (22, Scheme 3)
in which the polycyclic nature prevents relactonization. Treatment
Trilobolide (16): R = 2-MeBut
Nortrilobolide (17): R = But of 22 with strong base affords 23 and traces of 24 (Scheme 3) [20,
Thapsivillosin F (18): R = Sen 21].
Fig. (3). Structures of naturally occurring pentaoxygenated guaianolides
[trilobolides] (16-18) found T. garganica or L.trilobum.
Thapsigargin, Origin, Chemistry, Structure-Activity Relationships Current Pharmaceutical Design, 2015, Vol. 21, No. 38 5503
O O
O O H O O MeO OMeH O O H O O O H O O MeO OMeH H O O
O OOHH Acetone O OOH O OOHH Acetone OOHH
O OH O O O OH O OH
20 O 22 O
29 O 32 O
HO H OH HO H OH
KOH O O O
MeOH O OOH HO OOH O H O H O
O O O O O OOH MeO OOH
23 O 24 O O O O O
30O 31 O
Scheme 3. Masking OH-8 and OH-11. Minor Major

As expected the (cid:1),(cid:2)-unsaturated angeloyl group at C-3 is more Scheme 5. Formation of isopropylidene derivatives 30 and 31.
stable towards basic hydrolysis than the other acyl groups in the
molecule. Secondary alcohols are known to react faster with acyl 2.2.4. Selective Substitution of the Angloyl Group at O-3
groups than tertiary alcohols. Consequently acylation of OH-2 pro-
Whereas the double bond between C-2 and C-3 in the angeloyl
ceeds faster than acylation of OH-10 enabling selective acylation to
moiety protects the ester bond from saponification the same group
give 25, which subsequently can be acylated at O-10 to give 26
also enable selective hydrolysis of this group. Oxidation of the
(Scheme 4). Removal of the isopropylidene group with acid in an
group with reagents like osmium tetraoxide or permanganate fol-
aqueous medium affords 27 in which O-8 selectively can be acylat-
lowed by treatment with periodate enables formation of the pyruvic
ed affording 28 (Scheme 4). In total, the procedure enables selec-
ester 33, which can be solvolyzed under mild conditions like pyri-
tive replacement of all the acyl groups in the starting material ex-
dine in methanol to give the 3-hydroxy compound 34 (Scheme 6)
cept for the angeloyl group [20, 21].
[20].
Surprisingly attempts to convert the debutanoyl nortrilobolide

29 into the corresponding isopropylidene derivative 30 only afford-
O O
ewda sth teh eta r3g-emt ectohmylpeothuenrd 3 i1n (tSracchee mame o5u)n. tTs hweh meroesats ltihkee lmy aejxopr lparnoadtiuocnt O O H O O O MnO4- or OsO4 O O H O O O
O O O O
for the different behavior of the hexa- and the pentaoxygenated OH OH
guaianolides is that the voluminous octanoyloxy group in the 2- O OH HOHO O OH
position prevents an attack from the methanol on the intermediary 1 O O
formed carbocation 32, whereas the absence of the 2-subsituent in IO4-
29 favors the competing reaction.
O O

O O H O O O Pyridine O O H O O O
O OHO2 H 10OH8 O (RDCMOA)2PO, O RO O H OH O HO 3O4OOOOHH MeOH O O 3O3 OOOOHH
OH OH Scheme 6. Selective angelate cleavage at O-3.
O O O O

23 O 25 O Again a significant difference between the penta- and the hex-
aoxygenated guaianolide has been observed in the selectivity of
D(RM'CAOP)2O, ogxenidaatetido ng uoaf itahneo laindge,e lnooyrlt rdioloubboleli dbeo n(1d7. )I,n tthhee Cca-4se-C o-f5 t hdeo upbelnet abooxnyd-

is sensitive to oxidation, e.g. ozonolysis, affording a cleavage of the
O R' O R' five-membered ring to form 35 and 36 (Scheme 7). Again it is as-
O RO O H O OOH H O RO O H O OO stahunemo laeitddtea tsch.k a to fth teh es tCer-i4c-aCl -e5f fdecotu bolfe t hbeo n2d-o icnt atnhoe yhle sxuabosxtiytgueennat tperde vgeunatis-
O OOHH MeOH O OOH
27 O 26 O O H O O O 1) O3 then PPh3 O H O O O
O O O
D(RM''CAOP)2O, O OOHH 2) Pyridine, MeOH OHO O OOHH
17 O 35 O
O
R'
O R OH O O O
O O R" O
OH O H O O
O OH O
28O OHO O OOHH
36 O
Scheme 4. Selective substitutions of acyl groups at O-2, O-10 and O-8. Scheme 7. Ozonolysis of nortrilobolide (17) to form 35 and 36.
5504 Current Pharmaceutical Design, 2015, Vol. 21, No. 38 Doan and Christensen
Selective removal of the angeloyl group in the pentaoxygentaed O O
guaianolides, however, might be obtained taking advantage of the O O H O O O DAMc2AOP O OH O O O
possible nucleophilic removal of the angeloyloxy group by reacting O O O O
nortrilobolide (17) with an acidic aqueous medium (Scheme 8). A OH OH
similar reaction has been observed in the hexaoxygenated guaiano- O OH O O
lide [14]. As expected a mixture of the two 3-alcohols 37 is ob- 38 OH O 42O O O
tained. SRcCCNl4 O O H O O O
O O
OH
OH
O O O O
O H O O H+ (aq) H O O 43 N R
O O HO O
OH MeCN OH
O OH O OH Scheme 11. Formation of oxazoles 43 from hemiacetal 38.
17O 37O
2.2.7. Synthesis of Thapsigargin (1)
Scheme 8. Selective angelate cleavage of nortrilobolide (17) under acidic At the present the annual demand of paclitaxel is approximately
aqueous conditions to yield 37. 1 ton per year. A successful outcome of the current clinical trials of
mipsagargin is expected to create an annual demand of Tg (1) in the

same order of magnitude. At the present Tg (1) is only available
2.2.5. Selective Reduction of the Lactone Group
from the roots or fruits of the wild population of T. garganica. A
Treatment of Tg (1) with sodium borohydride affords a selec- total synthesis of Tg (1) in 42 steps affording an overall yield of
tive reduction of the lactone carbonyl group to form the hemiacetal 0.6% has been developed starting form (S)-carvone. (Scheme 12)
38, although only in modest yield (Scheme 9) [20, 22]. Sodium [24, 25].
bis(2-methoxyethoxy)ethoxyaluminium hydride (Red-Al) was

found to be superior to sodium borohydride [22].
O

O OOO H O OOO NRaeoBdr-H A4l O OOO H O OOO H 42 steps O O O H O OO O
OH OH OH
O12 OH O OH O O OH
1 O 38OH (S)-carvone
O O H 1 O
Red-Al Na Al Scheme 12. Total synthesis of thapsigargin (1) in 42 steps from (S)-carvone.
O O H

Scheme 9. Selective reduction of lactone carbonyl at C-12.
In spite of the impressive academic achievement this synthesis,
however, is not commercial feasible. An alternative procedure
2.2.6. Chemistry of the 12-Semiacetal (38) could be semi-synthesis from other available natural products. A
few other studies towards synthesis of closely related compounds
Attempts to O-12-alkylate 38 with ethyl orthoformate only to
have been published [26, 27]. Trilobolide (16) might be a possibil-
some extend afforded the acetal 39, instead the two orthoesteres 40
ity as an alternative starting material since this compound is easily
and 41 were formed as the major products (Scheme 10) [22].
available and can be isolated from L. trilobum, which can be grown
e.g. in The Czech Republic.
O A synthetic route has been developed in order to obtain Tg (1)
O O H O O O in four steps starting from nortrilobolide (17) (Scheme 13).
O O
OH Treatment of nortrilobolide (17) with chromium trioxide and
39O OH aqueous hydrogen fluoride in an one pot reaction affords the ketone
Minor O 44 in excellent yield. Oxidation of 44 with manganese(III) acetate
O OO H O O O EtOOEOtEt, H O OO H O O O ilnec tthivee p trhees e(cid:1)n-c2e- oocft aenxoceyslos xaymgoenutnate odf doecrtiavnaotiivce a 4ci5d. aRfefodrudcst isotne roefo s4e5-
O O O O with zinc borohydride gives a mixture of the two epimeric alcohols
OH OH
O OH O O 46S and 46R, 46S being the major product. Finally, acylation of
38 40
OH Major O O 46S with the mixed anhydride generated from angelic acid and
O 2,4,6-trichlorobenzoyl chloride yields thapsigargin (1).
O O H O O O 3. THE SARCO/ENDOPLASMIC RETICULUM CALCIUM
O O
OH ATPASE AS A DRUG TARGET
41 O O In the resting state any cells maintain a low Ca2+ concentration
Major O O (100 nM) in the cytosol and a high Ca2+ concentration (0.5 mM) in

the sarco/endoplasmic reticulum. In the case of the endo/sarco-
Scheme 10. Alkylation at O-12. plasmic reticulum the Sarco/Endoplasmic Reticulum Calcium
ATPase (SERCA) maintain this gradient by transporting Ca2+ ions
Another unexpected property of the 12-hemiacetal is the reac- from the cytosol into the organelle. For each ATP consumed by the
tion of the diacetate 42, obtained from 38, with nitriles to form pump two Ca2+ ions are transported across the membrane
oxazoles 43 (Scheme 11) [23].
Thapsigargin, Origin, Chemistry, Structure-Activity Relationships Current Pharmaceutical Design, 2015, Vol. 21, No. 38 5505
3.1. Structure-Activity Relationships
O O
O H O O CrO3, HF (aq) H O O A requirement for use of the prodrug approach is an intimate
O O O O
OH MeCN OH knowledge of structure-activity relationships (SAR). Only such
O OH O OH knowledge enables location of the linker to give an agent, which
17 O 44 O still possesses activity. Furthermore SAR analysis might indicate
O how structural changes of the molecule might make the compound
Mn(OAc3) O H O O O Zn(BH4)2 meffoercet sd oruf gcahbalne.g eBs eolof wth ea rset rupcrteusreen toefd TTga (b1l)e.s T1h-e1 0d aitlal ugsitvraetnin ign tthhee
O O
Octanoic acid OH table are mainly taken from the references: [21, 30-35].
O OH 3.1.1. Changes at the 8-Position
O 45 O O When interpreting the relative activities presented in Table 1
O H O O O O H O O O (vaanlualeo ogfu eTsg 2(10), i4s7 s-u7b2n) anito mis oilmarp [o2r3ta, n3t5 ]t om reeamnienmg bthera t tehvaet nt haen aICna50-
HO O HO O
logue 100 times less active than Tg (1) is a potent compound. The
OH OH
O OH O OH IC50 value is the concentration, in which the SERCA pump is inhib-
ited 50%. Considering this fact it can be concluded that even dra-
46S O 42R O
matic changes of the side chain at O-8 only to a minor extent reduc-
2,4,6-trichlorobenzoyl chloride es activity. Even hydrolysis to give the 8-hydroxy analogue 20 still
Et3N, Angelic acid affords a potent compound. From the point of view of designing a
prodrug this finding is encouraging since it means that a broad
O
spectrum of linkers possessing a terminal amino group may be used
O O H O O O as linkers.
O O The key then may be a peptide bond. Noticeable is the analogue
OH
O OH terminating with a tert-butyloxycarbonyl protecting group (Boc) 62.
The significant lower activity of this compound might indicate that
1 O
the presence of terminal hydrophilic groups could be of importance.
Scheme 13. Semi-synthesis of Tg (1) starting from nortrilobolide
Important for defining the pharmacophore of Tg (1) is the finding
(17).
that inversion of the stereochemistry at C-8 dramatically reduces
the potency as seen for analogue 72 (Table 2).
and 2-3 H+ ions released into the cytosol [28]. Blockage of the 3.1.2. Changes at the 3-Position
pump results in a collapse of the Ca2+-gradient. As a consequence
In contrast to changes at O-8 dramatic changes can be seen by
the cytosolic Ca2+ concentration increases to 500 nM for about 3
changes at O-3 (analogues 34, 73-82, Table 3).
hours where after it again decreases to low nM concentration. A
continued blockage of the pump affords a new increase after about No major reduction of activity is observed by replacing the
19 hours, but now the concentration reaches 1500 nM. The later angeloyl with a benzoyl 76 or a flexible octanoyl group 74 (Table
burst induces a cascade reaction, which eventually results in apop- 3). However, introduction of a 4-methylbenzoyl 77 causes a severe
tosis [13]. From a chemotherapeutic point of view the cytotoxicity drop in activity, whereas a 3-methylbenzoyl 78 is tolerated.
is interesting since cells are killed in a proliferation independent A similar trend is seen by replacement with a 2-phenylbenzoate
way [13]. Most presently used chemotherapeutics like paclitaxel, 79 and a 4-phenylbenzoate 80. Adding even a little flexibility into
vincristine and doxorubicin only kill cells during the proliferation the side chain as is seen in biphenylacetoyl 81 again regains activi-
and consequently slowly developing cancer diseases like prostate ty.
cancer are not affected [13]. The drawback by targeting SERCA,
As was the case for C-8, inversion of the stereochemistry at C-3
however, is that this pump is essential for survival of almost all
provokes a dramatic decrease in activity as seen in analogue 82
kind of cells meaning that Tg (1) is a general cell toxin. This is
(Table 4).
supported by the low lethal dosis for mice (0.8 mg/kg) [29]. The
3.1.3 Changes at the 2-Position
use of Tg (1) per se as a drug thus is excluded. However, a prodrug
targeting the compound towards cancer tissue has been developed. The binding site is very tolerant toward substitution in the 2-
A prodrug is a drug that by itself is inactive, but is cleaved near the position since no dramatic shifts in activity occurs by replacements
pharmacological target to release the active agent. The moiety used in this position (analogues 83-86, Table 5 and 6). This conclusion is
to inactivate the agent is named the promoiety. An optimum bond further confirmed by inspecting the naturally occurring hexao-
for conjugating the active drug with the promoiety is a bond that xygentaed guaianloides (Fig. 1), all of which have very similar
only is cleaved in the target tissue (Fig. 4). activities.
3.1.4. Changes in the 10-Position
In contrast to the 2-position, the 10-position is located in an
area in the binding cavity, which severely reacts towards introduc-
tion of voluminous groups by reducing the affinity of the ligand
(analogues 87-92, Table 7).
3.1.5 Changes at the 7- and 11-Positions
Important to notice is that replacement of the hydroxy groups
with an acyl group and thereby preventing the group form being
hydrogen donors only to a limiting extent effect the IC value (ana-
Fig. (4). The prodrug principle. An agent (gray oval) is coupled to an inacti- 50
logues 19, 22, 93-101, Table 8 and 9).
vating group (the promoiety, circle) via a linker (wavy line) and a bond
sensitive to cleavage only in the target tissue (bold binding). At the pharma- Acylation of O-7 even with small acyl group only affords a
cological target the sensitive bond is cleaved and the active agent released. minor decrease in activity but flexible large substituents decrease
activity. The epoxide 19 also still possesses some affinity for the
5506 Current Pharmaceutical Design, 2015, Vol. 21, No. 38 Doan and Christensen
Table 1. Changes at O-8.
O
O H O
O O
Thapsigargin analogues O 8 O R Relative IC50*
OH
O OH
O

20 R = H 5.4/50 [30] [21]

O
H
N
47 R = 327 [21]
O
NH
2

O O
48 R = 81 [21]
N NH
2
H

O O
49 R = 4.4 [21]
HN NH2

O
50 R = 1.7 [21]
NH
2

O
51 R = O 1.9 [21]
N O
H

O
52 R = 1.5 [21]
NH
2

O
53 R = O 1.8 [21]
N O
H

H
N O
O
54 R = 1.3 [21]
O

NH
O 2
55 R = 1.7 [21]

O
O
56 R = N NH2 35 [31]
H

O
57 R = NH 99 [31]
2
Thapsigargin, Origin, Chemistry, Structure-Activity Relationships Current Pharmaceutical Design, 2015, Vol. 21, No. 38 5507
(Table 1) Contd....

O
58 R = 90 [31]
NH
2

O
59 R = NH 17 [31]
2

O
60 R = 3 [31]
NH
2

O
61 R = NH 2.6 [31]
2

O H
62 R = N O 240 [31]
O

O
63 R = H 287 [31]
N
NH
2
O

O H
64 R = 3.4 [31]
N
NH
2
O

O H
65 R = N 1.2 [31]
NH
2
O

OH
O H
66 R = N 0.8 [31]
NH
2
O

O O
67 R = N O 30 [35]
H
OH

O
H
68 R = N O 8.5 [35]
O
O

O O
69 R = H 11.5 [35]
N
O N O
H
O

O O
70 R = O O 44 [35]
O N O
H

O O
H
O O N O
O N
71 R = H 70 [35]
O

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].
5508 Current Pharmaceutical Design, 2015, Vol. 21, No. 38 Doan and Christensen
Table 2. Inversion at C-8.
Thapsigargin analogue Relative IC50*
O O
O H O O
O
72 O 8 O 3124 [32]
OH
O OH
O

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].

Table 3. Changes at O-3.
O
O H O O O
Thapsigargin analogues RO 3 O Relative IC50*
OH
O OH
O

34 R = H 565 [21]

O [32]
OH O O O
73 R = O 3 O 66.5
OH
O OH
O

O
74 R = 11.2 [32]

O
75 R = 1.53 [32]

O
76 R = 4.0 [21]

O
77 R = 220 [21]

O
78 R = 45 [21]

O
79 R = 15 [21]

O
80 R = 350 [21]

81 R = O 90 [21]

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].
Thapsigargin, Origin, Chemistry, Structure-Activity Relationships Current Pharmaceutical Design, 2015, Vol. 21, No. 38 5509
Table 4. Inversion at C-3.
Thapsigargin analogue Relative IC *
50
O
O O H O O
O
82 O 3 O 438 [32]
OH
OH
O
O

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].

Table 5. Changes at O-2.
Thapsigargin analogue Relative IC *
50
O O H O O
83 O O 8 [21]
O O
OH
OH
O
O

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].

Table 6. Changes at O-2.
Thapsigargin analogue Relative IC *
50
O
H O
O O
O O
84 40 [35]
OH
OH
O
O

O
H O O
O
O O
85 0.1/17.5 [33] [35]
OH
H OH
O
O

O
H O
O
O O
86 0.3 [33]
OH
H OH
O
O

* The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].
5510 Current Pharmaceutical Design, 2015, Vol. 21, No. 38 Doan and Christensen
Table 7. Changes at O-10.
O
R
O H O O
O
10 Relative
Thapsigargin analogues O O
OH IC50*
OH
O
O

87 R = 42/135 [32] [21]
H

O
88 R = 12.5 [21]

O
89 R = 80 [21]

O

78 [21]
90 R =

O
91 R = 350 [21]

92 R = O 135 [21]

*The relative IC50 value is the ratio between IC50 value of the analogue and the IC50 value of thapsigargin (1) [IC50(analogue)/IC50(thapsigargin)].

Table 8. Changes at O-7 and O-11.
O
O H O O
O O
Relative
Thapsigargin analogues O O
7 O R1 IC50*
O 11 O R2
O

O
93 R1 = R2 = H 2.8 [32]

O
94 R1 = R2 = H 55 [23]

O
95 R1 = R2 = H 65 [23]

O
96 R1 = R2 = H 42.5 [23]