Enantiomerically pure thrombin inhibitors for exploring the molecular-recognition features of the oxyanion hole

Article abstract of DOI:10.1002/hlca.200490225

A new route via intermediate pseudoenantiomers was developed to synthesize racemic and enantiomerically pure new non-peptidic inhibitors of thrombin, a key serine protease in the blood-coagulation cascade. These ligands feature a conformationally rigid tricyclic core and are decorated with substituents to fill the major binding pockets (distal (D), proximal (P), selectivity (S1), and oxyanion hole) at the thrombin active site (Fig. 1). The key step in the preparation of the new inhibitors is the 1,3-dipolar cycloaddition between an optically active azomethine ylide, prepared in situ from L-(4R)-hydroxyproline and 4-bromobenzaldehyde, and N-piperonylmaleimide (Scheme 1). According to this protocol, tricyclic imide (compounds (±)-15-(±)-18 and (+)-21) and lactam (compound (+)-2) inhibitors with OH or ether substituents at C(7) in the proline-derived pyrrolidine ring were synthesized to specifically explore the binding features of the oxyanion hole (Schemes 2-4). Biological assays (Table) showed that the polar oxyanion hole in thrombin is not suitable for the accommodation of bulky substituents of low polarity, thereby confirming previous findings. In contrast, tricyclic lactam (+)-2 (K_i = 9 nM, K _i(trypsin)/K_i(thrombin) = 1055) and tricyclic imide (+)-21 (K_i = 36 nM, K_i(trypsin)/ K_i(thrombin) = 50) with OH-substituents at the (R)-configured C(7)-atom are among the most-potent and most-selective thrombin inhibitors in their respective classes, prepared today. While initial modeling predicted H-bonding between the OH group at C(7) in (+)-2 and (+)-21 with the H₂O molecule bound in the oxyanion hole (Fig. 2), the X-ray crystal structure of the complex of (+)-21 (Fig. 7,b) revealed a different interaction for this group. The propionate side chain of Glu192 undergoes a conformational change, thereby re-orienting towards the OH group at C(7) under formation of a very short ionic H-bond (O-H... ^-OOC; d(O...O) = 2.4 A). The energetic contribution of this H-bond, however, is negligible, due to its location on the surface of the protein and the unfavorable conformation of the H-bonded propionate side chain.

Full text of DOI:10.1002/hlca.200490225

Helvetica Chimica Acta Vol. 87 (2004)
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Enantiomerically Pure Thrombin Inhibitors for Exploring the Molecular-
Recognition Features of the Oxyanion Hole
by Kaspar Sch‰rer, Martin Morgenthaler, Paul Seiler, and FranÁois Diederich*
Laboratorium f¸r Organische Chemie der Eidgenˆssischen Technischen Hochschule, ETH-Hˆnggerberg, HCI,
CH-8093 Z¸rich (e-mail: diederich@org.chem.ethz.ch)
and
David W. Banner*, Thomas Tschopp, and Ulrike Obst-Sander
Pharma Division, Pr‰klinische Forschung, F. Hoffmann-La Roche AG, CH-4070 Basel
A new route via intermediate pseudoenantiomers was developed to synthesize racemic and enantiomeri-
cally pure new non-peptidic inhibitors of thrombin, a key serine protease in the blood-coagulation cascade.
These ligands feature a conformationally rigid tricyclic core and are decorated with substituents to fill the major
binding pockets (distal (D), proximal (P), selectivity (S1), and oxyanion hole) at the thrombin active site
(Fig. 1). The key step in the preparation of the new inhibitors is the 1,3-dipolar cycloaddition between an
optically active azomethine ylide, prepared in situ from l-(4R)-hydroxyproline and 4-bromobenzaldehyde, and
N-piperonylmaleimide (Scheme 1). According to this protocol, tricyclic imide (compounds (Æ)-15-(Æ)-18 and
(‡)-21) and lactam (compound (‡)-2) inhibitors with OH or ether substituents at C(7) in the proline-derived
pyrrolidine ring were synthesized to specifically explore the binding features of the oxyanion hole (Schemes 2
4). Biological assays (Table) showed that the polar oxyanion hole in thrombin is not suitable for the
accommodation of bulky substituents of low polarity, thereby confirming previous findings. In contrast, tricyclic
lactam (‡)-2 (Ki ˆ 9 nm, Ki(trypsin)/Ki(thrombin) ˆ 1055) and tricyclic imide (‡)-21 (Ki ˆ 36 nm, Ki(trypsin)/
Ki(thrombin) ˆ 50) with OH-substituents at the (R)-configured C(7)-atom are among the most-potent and
most-selective thrombin inhibitors in their respective classes, prepared today. While initial modeling predicted
H-bonding between the OH group at C(7) in (‡)-2 and (‡)-21 with the H2O molecule bound in the oxyanion
hole (Fig. 2), the X-ray crystal structure of the complex of (‡)-21 (Fig. 7,b) revealed a different interaction for
this group. The propionate side chain of Glu192 undergoes a conformational change, thereby re-orienting
towards the OH group at C(7) under formation of a very short ionic H-bond (OÀH ¥¥¥ ÀOOC; d(O ¥¥¥ O) ˆ
2.4 ä). The energetic contribution of this H-bond, however, is negligible, due to its location on the surface of the
protein and the unfavorable conformation of the H-bonded propionate side chain.
1. Introduction.
The globular serine protease thrombin from the blood-
coagulation cascade continues to be an important pharmaceutical target for the
prevention and treatment of thrombotic disorders [1][2]. Early crystal structures
revealed a well-defined active site [3][4], and, accordingly, structure-based design has
been intensively applied to the generation of new inhibitors of this enzyme [5 7] (for
some recent reports, see [8]). By a de novo design strategy, we developed a series of
high-affinity inhibitors with a tricyclic core such as lactam (‡)-1 (Fig. 1; inhibitory
constant Ki ˆ 7 nm, selectivity over the digestive serine protease trypsin Ki(trypsin)/
Ki(thrombin) ˆ 740) and confirmed the binding modes of several derivatives at the
thrombin active site by crystal-structure analysis [9][10]. This analysis revealed that the
active site of thrombin has limited conformational flexibility, thereby making it an
outstanding receptor for biological molecular-recognition studies [11]. As a result of
¹ 2004 Verlag Helvetica Chimica Acta AG, Z¸rich

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the rigidity of the active site, related ligands with functional-group mutations form
complexes of similar geometry, allowing direct correlation of measured changes in the
complexation free enthalpy with the contributions of individual ligand substituents.
Previous investigations with this class of tricyclic inhibitors addressed in greater detail
the molecular-recognition features of the spacious hydrophobic D-pocket (D: distal)
[9a,b][10][12], the tight hydrophobic P-pocket (P: proximal) [9b][10], and the
selectivity pocket S1 (Fig. 1) [9b]. Most recently, an extensive fluorine scan of these
inhibitors was undertaken by systematically replacing one or more H-atoms with F-
atoms to map the fluorophilicity/fluorophobicity of the thrombin active site [13]. In
contrast, the molecular-recognition properties of the oxyanion hole remain rather
unexplored [9b]. In particular, we were intrigued by the possibility to bind to the H2O
molecule found in the oxyanion hole in the crystal structure of (‡)-1, H-bonded to the
NÀH residues of Gly193 and Ser195. Molecular modeling with MOLOC [14] suggested
that substitution of the tricyclic core in position 7 (Fig. 1) with a H-bond donor center
would be particularly suitable for bonding to the H2O molecule. According to this
analysis, the OH group at (R)-configured C(7) in inhibitor (‡)-2 could form a short
intermolecular H-bond (d(O ¥¥¥ O) ˆ 2.9ä) to the bound H 2O molecule (Fig. 2). Here,
we describe the synthesis and biological activity of a series of new tricyclic thrombin
inhibitors bearing substituents of different size and polarity at C(7) to investigate the
molecular-recognition properties of the oxyanion hole in thrombin.
Fig. 1. Inhibitor ( ‡ )-1 in the active site of thrombin, according to crystal-structure analysis [9b][10]

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Fig. 2. Molecular model of ( ‡ )-2 complexed to thrombin, showing the predicted H-bond to the H2O molecule
bound in the oxyanion hole. This model was obtained starting from the crystal structure of (‡)-1 bound to
thrombin, attachment of the OH group to (R)-configured C(7), and minimization of the inhibitor in the active
site, while keeping the protein conformation with the bound H2O molecule unchanged. Color code: C-skeleton
of (‡)-2: cyan, C-skeleton of the protein: grey, O-atoms: red, N-atoms: blue, S-atoms: yellow.
2. Results and Discussion. 2.1. Synthesis of Racemic Tricyclic Imide Inhibitors. A
series of tricyclic imide inhibitors was prepared starting with the 1,3-dipolar cyclo-
addition between the azomethine ylide, formed in situ from l-(4R)-hydroxyproline (3)
and 4-bromobenzaldehyde (4), and N-piperonylmaleimide (5) in DMF [9][12][15]
(Scheme 1). The desired endo,trans-derivatives (‡)-6 and (À)-7 were obtained as an
inseparable 1:1 mixture in 34% yield (for their separation, see Sect. 2.2 below)
together with exo,trans-isomer (À)-8 as a major side product (35% yield; exo and endo
refer to the orientation of the 4-bromophenyl substituent at C(4) with respect to the
bicyclic perhydropyrrolo[3,4-c]pyrrole scaffold, and cis and trans to the position of this
4-bromophenyl ring with respect to the configuration of C(8a) at the fusion of the two
pentagons in the perhydropyrrolizine bicycle). The exo-diastereoisomer (À)-8 was
separated by column chromatography, and its relative configuration was assigned on
the basis of 1D-NOE difference 1H-NMR spectra (NOE ˆ nuclear Overhauser effect).
As discussed in a preliminary way in a previous paper [12], the endo-diastereoisomers

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Scheme 1. Synthesis of the Ketones ( Æ )-11 and ( Æ )-12
a) DMF, 858, 24 h; 17% of (‡)-6, 17% of (À)-7, 35% of (À)-8. b) CuCN, DMF, D, 18 h; 39 % of (‡)-9, 39 % of
(À)-10. c) 1. Me2SO, (COCl)2, À 608, 30 min; 2. NEt3, 08, 3 h; 60% of (Æ)-11; 66% of (Æ)-12. DMF ˆ Di-
methylformamide.
(‡)-6 and (À)-7 can be viewed as pseudoenantiomers, differing at only one stereogenic
center (for pseudoenantiomerism in the cinchona alkaloid family, see [16]). They
cannot be separated by column chromatography and crystallize together, forming a
1:1 mixed crystal whose crystal structure was solved [12], In fact, the two compounds
(for the synthesis of the pure diastereoisomers, see Sect. 2.2 below) even show optical
rotational angles of opposite sign yet of identical magnitude: [a]2D5 ˆ ‡169.4 (c ˆ 1.00,
CHCl3) for (‡)-6 and À 169.4 (c ˆ 1.00, CHCl3) for (À)-7.

Helvetica Chimica Acta Vol. 87 (2004)
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Br/CN Exchange starting from the 1:1 mixture of (‡)-6 and (À)-7 provided the
corresponding mixture of nitriles (‡)-9 and (À)-10. Again, separation by chromatog-
raphy was not possible, and crystal-structure analysis revealed the formation of
1:1 mixed crystals (space group P1). In the crystal lattice, the two diastereoisomers are
aligned pairwise and are related by a pseudo-inversion center (Fig. 3,a). Short
intermolecular contacts include H-bonds between the OH group at C(7) (numbering
adopted from Scheme 1) and the CN group (O ¥¥¥ N distances: 3.01 and 3.20 ä, resp.;
Fig. 3,b) as well as p-p stacking between parallel shifted piperonyl rings (shortest C ¥¥¥ C
distance: 3.67 ä; Fig. 3,c).88
In view of the inseparabilitity of the pseudoenantiomeric bromides (‡)-6/(À)-7 and
nitriles (‡)-9/(À)-10, the mixtures were transformed by Swern oxidation [17] into the
racemic ketones (Æ)-11 and (Æ)-12, respectively. The structures of (Æ)-11 (Fig. 4,a)
and (Æ)-12 (Fig. 4,b), which crystallize both in the space group P21/n, were confirmed
by crystal-structure analyses.
The reduction of ketone (Æ)-11 in CH2Cl2 proceeded diastereoselectively to give
alcohol (Æ)-7 as the major product besides alcohol (Æ)-6 as the minor one (Scheme 2).
Correspondingly, reduction of (Æ)-12 under the same conditions provided alcohol (Æ)-
10 as the major and (Æ)-9 as the minor product. Model analysis suggested an
explanation for the observed diastereoselectivity, showing that the Re-face of the
trigonal centers (C(7)) in (3aR,4S,8aR,8bS)-11/12 (and correspondingly the Si-face in
(3aS,4R,8aS,8bR)-11/12) is sterically less hindered than the opposite face. Accordingly,
by increasing the steric bulk of the reducting agents [18], the diastereoselectivity was
enhanced (superhydride (Li[Et3BH]) at À 788: diastereoisomeric ratio (dr) (Æ)7/(Æ)-6
77:23; L-Selectride (Li[(sec-Bu)3BH] at À 788: dr 89 :11; LS-Selectride (lithium
trisiamylborohydride) at À 958: dr 95 :5). The diastereoselectivity in the reduction of
carbonitrile (Æ)-12 was surprisingly lower, with the highest dr ((Æ)-10/(Æ)-9) 81:19
being obtained with LS-Selectride at À 958. O-Methylation and O-benzylation [19]
provided methyl ether (Æ)-13 and benzyl ether (Æ)-14, respectively, and the relative
configurations assigned were confirmed by the crystal-structure analysis of (Æ)-14,
which, surprisingly, crystallized with both enantiomers in the non-centrosymmetric
space group P212121 (Fig. 5).
The conversion of the carbonitriles (Æ)-10, (Æ)-13, and (Æ)-14 to the phenyl-
amidinium salts (Æ)-15, (Æ)-16, and (Æ)-17, respectively, was readily achieved with the
Pinner reaction [20]. When this transformation was applied to carbonitrile (Æ)-12, the
CˆO group at C(7) was acetalized, thereby providing the phenylamidinium salt as the
remarkably stable O,O-acetal (Æ)-18.
2.2. Synthesis of Enantiomerically Pure Tricyclic Imide Inhibitors. Whereas the
1:1 mixture of hydroxy derivatives (‡)-6 and (À)-7 could not be separated, this was
possible after (t-Bu)Me2Si protection. The two silylated diastereoisomers (‡)-19 and
(À)-20 were isolated in pure form after column chromatography and subsequently
transformed by deprotection into the pure pseudoenantiomers (‡)-6 and (À)-7
(Scheme 3). Both compounds were converted to the enantiomerically pure carbo-
nitriles (‡)-9 and (À)-10. Pinner reaction subsequently provided the phenylamidinium
inhibitor (‡)-21, which we hoped would bind to the H2O molecule in the oxyanion hole
of thrombin (Fig. 2) with its suitably positioned C(7)ÀOH group ((À)-10 was not
transformed into the corresponding amidinium salt).

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Fig. 3. X-Ray crystal structure of a 1:1 mixed crystal of ( ‡ )-9 and ( À )-10. a) View on the pairwise alignment of
the two diastereoisomers in the crystal lattice. Arbitrary numbering. Atomic displacement parameters obtained at
173 K are drawn at the 50% probability level. b) H-Bonding between the two diastereoisomers in the crystal.
Distances in ä. c) p-p-Stacking observed in stacks of each diastereoisomer (see next page). Color code: C-
skeleton: grey, O-atoms: red, N-atoms: blue.
2.3. Synthesis of an Optically Active, High-Affinity Tricyclic Lactam Inhibitor. The
preparation of the inhibitor (‡)-2, with an i-Pr group to favorably fill the P-pocket [9
13], started from silyl-protected (‡)-19, which was converted in a regio- and
stereoselective reduction with superhydride (Li[Et3BH]) to hydroxy lactam (‡)-22,
presumably bearing the OH group in exo-position as observed previously by X-ray
analysis [12] (Scheme 4).
Silyl deprotection, followed by treatment of the resulting diol with 4-toluenesulfinic
acid in the presence of CaCl2, afforded sulfone (‡)-23, probably by attack at an

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Fig. 3 (cont.)
Fig. 4. X-Ray crystal structures of bromide ( Æ )-11 (left) and nitrile ( Æ )-12 (right). Arbitrary numbering.
Atomic displacement parameters obtained at 223 K are drawn at the 50% probability level.
intermediately formed acyliminium ion from the less-hindered exo-face [9b]. Silyl
protection provided (‡)-24, and subsequent nucleophilic displacement of the
arylsulfonyl group according to the protocol (ZnCl2/i-PrMgCl) introduced by Ley
and co-workers [21] gave under retention of configuration resulting from attack of the
intermediate acyliminium ion from the exo-face the i-Pr derivative (‡)-25. Silyl
deprotection led to (‡)-26, and X-ray crystal-structure analysis (space group P212121)
confirmed the predicted configuration (Fig. 6). The crystal packing shows a short
intermolecular H-bond between the C(7)-OH (for numbering, see Scheme 1) and the
lactam CˆO groups of adjacent molecules (d(O ¥¥¥ O) ˆ 2.80 ä). Br/CN Exchange

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Scheme 2. Synthesis of the Inhibitors ( Æ )-15 to ( Æ )-18
a) Li[Et3BH], CH2Cl2, À 788, 2.5 h; 70% of (Æ)-7, dr (Æ)-7/(Æ)-6 77:23. b) L-Selectride, CH2Cl2, À 788, 2.5 h;
63% of (Æ)-7; dr 89 :11. c) LS-Selectride, CH2Cl2, À 788, 2.5 h; 75% of (Æ)-7; dr 9 5 :5 orLS-Selectride, CH2Cl2,
À 958, 2.5 h; 79% of (Æ)-7; dr 9 5 :5; 69 % ofÆ( )-10; dr (Æ)-10/(Æ)-9 81:19. d) 1. HCl (g), MeOH, CHCl3, 48,
24 h; 2. NH3, MeOH, 658, 3.5 h; 80% of (Æ)-15, 38% of (Æ)-16, 50% of (Æ)-17, 13% of (Æ)-18. e) NaH,
[15]crown-5, MeI, THF, r.t., 3.5 h; 38% of (Æ)-13). f) NaH, [15]crown-5, BnBr, THF, r.t., 2 h; 70% of (Æ)-14.
Bn ˆ PhCH2.
((‡)-26 ! (‡)-27) and Pinner reaction finally afforded target compound (‡)-2. All
attempted protocols avoiding the rather tedious silyl protection/deprotection steps in
the synthesis of (‡)-2 were unsuccessful, providing much lower yields in the individual
transformations.
2.4. Biological Activity and Crystal-Structure Analysis of the Complex between
Thrombin and Inhibitor (Æ)-21. All newly synthesized inhibitors were tested for their
activity against thrombin, trypsin, and related enzymes (coagulation factors VIIa and
Xa) as previously described [22]. Their potencies were compared with those of the
previously described, structurally related inhibitors (‡)-1 and (Æ)-28, lacking the
substituent at C(7) [9].

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Fig. 5. X-Ray crystal structure of the carbonitrile ( Æ )-14. Arbitrary numbering. Atomic displacement
parameters obtained at 222 K are drawn at the 50% probability level.
Several conclusions can be drawn from the data summarized in the Table:
i) The affinity for thrombin is not much altered by the introduction of an OH group
at C(7): ligands (‡)-1 (without OH; Ki ˆ 7 nm) and (‡)-2 (with OH; Ki ˆ 9 nm) display
similar activity, and the introduction of the OH group does also not increase the activity
at the stage of the imide derivatives ((‡)-21 vs. (Æ)-28). The comparison of (Æ)-15
(Ki ˆ 100 nm) with (‡)-21 (Ki ˆ 36 nm) shows that the configuration at C(7) does not
play a significant role (similar to (‡)-21, only the (3aS,4R,8aS,8bR)-configured
enantiomer of (Æ)-15 is active [9].
ii) Introduction of less-polar substituents at C(7) (MeO and BnO in (Æ)-16 (Ki
710 nm) and (Æ)-17 (Ki ˆ 1100 nm), resp.) reduces the activity by a factor of 7 11 as
compared with (Æ)-15. The similar activity displayed by (Æ)-18 (Ki ˆ 720 nm) could
suggest that the O,O-acetal might actually be stable under the conditions of the assay: a
higher activity would be expected for the free ketone.
iii) The introduction of the OH substituents at C(7) increases the selectivity for
thrombin over trypsin in both the tricyclic imide and lactam classes of inhibitors. Thus,
(‡)-2 is not only one of the most potent thrombin inhibitors in the tricyclic lactam class
[9 13], but at Ki(trypsin)/Ki(thrombin) ˆ 1055, it is also one of the most-selective ones.
On the other hand, introduction of a larger, less-polar BnO substituent at C(7) lowers
not only the binding affinity but also the selectivity ((Æ)-17; Ki ˆ 1100 nm, Ki(trypsin)/
Ki(thrombin) ˆ 3.6). None of the inhibitors showed any affinity for other serine
proteases in the blood-coagulation cascade below the threshold (69 mm) of the assay.
We had started this project with the modeling-supported hypothesis that the OH
group at C(7) of the inhibitors could bind to a H2O molecule bound in the oxyanion
hole of thrombin (such a H2O molecule is not only seen in the complex of (‡)-1 (Fig. 2)
but also in the complex of (Æ)-28 [9a]). To verify this proposal, the X-ray crystal

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Scheme 3. Synthesis of the Inhibitor ( ‡ )-21
a) t-Bu)Me2SiCl, DMAP, CH2Cl2, 258, 16 h; 43% of (‡)-19); 43% of (À)-20. b) Bu4NF, THF, r.t., 1.5 h; 69% of
(‡)-6, 72% of (À)-7. c) CuCN, DMF, D, 18 h; 55% of (‡)-9; 45% of (À)-10. d) 1. HCl (g), MeOH, CHCl3, 48,
24 h; 2. NH3, MeOH, 658, 3.5 h; 55%. DMAP ˆ 4-(Dimethylamino)pyridine.
structure of (‡)-21 bound to thrombin was solved at a 1.54-ä resolution. A
superimposition of the crystal structures of the complexes of thrombin with (‡)-21
and (‡)-1 (or (Æ)-28, not shown [9a]) proved a nearly identical binding mode for the
two inhibitors (Fig. 7,a). The conformation of the surrounding protein was also similar
with one notable exception. The propionate side chain of Glu192, which, in the complex
of (‡)-1 (and in the predicted binding mode for (‡)-2 and (‡)-21; Fig. 2), was pointing
away from the proline-derived pyrrolidine ring, is now oriented towards the OH group
at C(7) under formation of a very short ionic H-bond (OÀH ¥¥¥ ÀOOC; d(O ¥¥¥ O) ˆ
2.4 ä; Fig. 7,b). An intermolecular interaction between the OH group in (‡)-21 and
the H2O molecule in the oxyanion hole is not observed (measured O ¥¥¥ O distance:
4.6 ä). Clearly, the molecular modeling, which did not take into account possible

Helvetica Chimica Acta Vol. 87 (2004)
Scheme 4. Synthesis of the Inhibitor ( ‡ )-2
2527
a) Li[Et3BH], THF, À 788, 15 min; 94%. b) Bu4NF, THF, r.t., 1 h. c) 4-Me-C6H4SO2H, CaCl2, CH2Cl2, r.t., 2 d;
75% over 2 steps. d) (t-Bu)Me2SiCl, DMAP, CH2Cl2, 258, 16 h; 87%. e) (i-Pr)MgCl, ZnCl2, CH2Cl2, r.t., 16 h;
79%. f) Bu4NF, THF, r.t., 1 h; 75%. g) CuCN, DMF, D, 18 h; 55%. h) 1. HCl (g), MeOH, CHCl3, 48, 24 h; 2.
NH3, MeOH, 658, 3.5 h; 60%.
conformational changes of the protein, such as the move of the Glu192 side chain, was
too crude.
Although the ionic H-bond between the OH group of the ligand and the
carboxylate of Glu192 is very short (d(O ¥¥¥ O) ˆ 2.4 ä), the gain in free enthalpy
resulting from this additional intermolecular interaction is negligible, as shown by the
comparison of the Ki values of (Æ)-28 (without OH group) and (‡)-21 (with OH
group). Two factors account for the absence of a significant energetic contribution of
this short ionic H-bond. First, the conformation of the free propionate side chain
(antiplanar torsional angle q(C(a)ÀCÀCÀC(O2)) ˆ 1688) is energetically more
favorable than the conformation of the H-bonded one (gauche torsional angle
q(C(a)ÀCÀCÀC(O2)) ˆ 618). Second, the observed H-bonding occurs rather at the

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Fig. 6. a) X-Ray crystal structure of ( ‡ )-26. Arbitrary numbering. Atomic displacement parameters obtained
at 163 K are drawn at the 50% probability level. b) Crystal packing of ( ‡ )-26. Distance in ä. Color code: see
caption to Fig. 3; Br-atoms: green.
surface of the protein, meaning that the carboxylate of Glu192 exchanges favorable H-
bonds to surrounding H2O molecules for the intermolecular H-bond to the ligand.
3. Conclusions. A new synthesis to enantiomerically pure thrombin inhibitors via
intermediate pseudoenantiomers was developed. This synthesis starts with the 1,3-
dipolar cycloaddition of the azomethine ylide, formed in situ from l-(4R)-hydroxypro-
line and 4-bromobenzaldehyde, and N-piperonylmaleimide, and provided access to
(‡)-2, one of the most-potent and most-selective tricyclic lactam inhibitors of thrombin
prepared today (Ki ˆ 9 nm, Ki(trypsin)/Ki(thrombin) ˆ 1055). While the initial model-

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Table. Activities of the New Tricyclic Thrombin Inhibitors and Selectivities with Respect to Trypsin
Inhibitor
Ki [nm]a)
Selectivityb)
(Æ)-28c)d)
(‡)-1c)
(‡)-2
90
7
7.8
760
91055
100
710
1100
720
36
(Æ)-15d)
(Æ)-16d)
(Æ)-17d)
(Æ)-18d)
(‡)-21
33
35
3.6
32
50
a) The uncertainty of the measured Ki values is Æ 20%.b) Ki(trypsin)/Ki(thrombin). c) From [9b]. d) Only the
(3aS,4R,8aS,8bR)-configured enantiomer is bound, as determined from the crystal-structure analysis [9].
ing predicted H-bonding between the OH group at C(7) in (‡)-2 and the tricyclic imide
analog (‡)-21 with the H2O molecule that is seen in the oxyanion hole of several
thrombin complexes (e.g., of (‡)-1 and (Æ)-28), the X-ray crystal structure of the
complex of (‡)-21 (Ki ˆ 36 nm, Ki(trypsin)/Ki(thrombin) ˆ 50) revealed a different,
more-favorable internal solvation of this OH group by the enzyme. It was found that
the propionate side chain of Glu192 undergoes a conformational change, thereby re-
orienting towards the OH group at C(7) under formation of a very short ionic H-bond
(OÀH ¥¥¥ ÀOOC; d(O ¥¥¥ O) ˆ 2.4 ä). The incremental free-enthalpy contribution of
this very short, ionic H-bond is negligible as shown by the comparison of the Ki values
for (Æ)-28 (without OH group) and with Ki (‡)-21 (with OH group). This can be
explained by the less-favorable conformation of the H-bonded propionate side chain of
Glu192 and by its location close to the solvated surface of the protein. Investigations
with a series of inhibitors featuring bulkier, less-polar substituents at C(7) confirmed

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Helvetica Chimica Acta Vol. 87 (2004)
Fig. 7. a) Inhibitors ( ‡ )-1 [9b][10] and ( ‡ )-21 in the active site of thrombin as revealed by crystal-structure
analyses. Color code: C-skeleton of (‡)-1: cyan, C-skeleton of (‡)-21: golden, C-skeleton of thrombin in the
complex of (‡)-1: grey, C-skeleton of thrombin in the complex of (‡)-21: green, O-atoms: red, N-atoms: blue, S-
atoms: yellow. The H2O molecules in the oxyanion hole are shown in the color of the corresponding enzyme.
b) Binding mode of inhibitor (‡)-21 in the oxyanion hole as revealed in the crystal structure. Distances in ä.

Helvetica Chimica Acta Vol. 87 (2004)
2531
our previous findings [12] that the polar oxyanion hole of thrombin is not an
appropriate site to accommodate such groups.
We thank F. Hoffmann-La Roche, Chugai Pharmaceuticals, and the ETH Research Council for support of
this work. We thank Olivier Kuster for the measurement of enzyme inhibitor constants and Dr. Joerg Benz and
Martine Stihle for help in the crystal preparation.
Experimental Part
General. Solvents and reagents were reagent-grade, purchased from commercial suppliers, and used
without further purification unless otherwise stated. The following compounds were prepared according to
literature procedures: 4-toluenesulfinic acid [23] and the 1:1 mixture of pseudoenantiomers (‡)-6/(À)-7 [12].
THF was freshly distilled from sodium benzophenone ketyl, CH2Cl2 from CaH2. HCl Gas was dried with conc.
H2SO4. Evaporation in vacuo was conducted at H2O-aspirator pressure. If not mentioned otherwise, all products
were dried under high vacuum (10À2 Torr) before anal. characterization. Column chromatography (CC): SiO2-
60 (230 400 mesh, 0.040 0.063 mm) from Fluka. TLC: SiO2-60 F245, Merck; visualization by UV light at
245 nm. M.p.: B¸chi SMP 20 apparatus; uncorrected. Optical rotations: Perkin-Elmer 241 polarimeter, 1-dm
cell, l ˆ 589nm (Na D-line). IR Spectra [cm À1]: Perkin-Elmer 1600-FTIR spectrometer. NMR Spectra (1H, 13C,
NOE): Varian Gemini-300, Varian Gemini-400, and Bruker AMX-500; spectra were recorded at r.t. with solvent
peak as reference. In some 1H- and 13C-NMR spectra (compounds (À)-7, (‡)-9, (Æ)-18, (‡)-19, (‡)-22, (‡)-24,
and (‡)-25), individual resonances overlap or are buried under the solvent peak. High-resolution (HR)
MALDI mass spectra (HR-MS): IonSpec Ultima, 2,5-dihydroxybenzoic acid (DHB) as matrix); molecular ions
‡
(M ) reported for phenylamidinium salts refer to the corresponding phenylamidine derivatives. Elemental
analyses were performed by the Mikrolabor at the Laboratorium f¸r Organische Chemie, ETH-Z¸rich.
Determination of Inhibition Constants. The affinities of thrombin inhibitors were determined according to
[22][24] (chromogenic substrate S-2238). An exhaustive protocol of the binding assay used in this study is
provided in [24].
General Procedure for the 1,3-Dipolar Cycloaddition (GP 1). A mixture of a-amino acid (1 mmol), 4-
bromobenzaldehyde (1 mmol), and N-piperonylmaleimide (1 mmol) in DMF (3 ml) was heated to 1208 for 16
48 h. The solvent was evaporated in vacuo and the residue purified by CC (AcOEt or AcOEt/hexane 70 :30 or
50 :50).
General Procedure for the Preparation of Amidinium Salts by the Pinner Reaction (GP 2). Dry HCl gas was
bubbled at 08 for 10 min into a soln. of the nitrile (2 mmol) in dry CHCl3 (5 ml) and dry MeOH (1 ml). The
mixture was stored at 48 for 24 h, then Et2O was added. The precipitate formed was isolated by filtration, dried
in high vacuum, and redissolved in MeOH (5 ml). A methanolic soln. of NH3 was slowly added until the NH3
odor persisted; then, the mixture was stirred for 3.5 h at 658. After cooling, NH4Cl was precipitated with acetone
and removed by filtration. The solvent was evaporated in vacuo, the residue was dissolved in EtOH, and the
amidinium salt was slowly precipitated with Et2O and purified by CC (CH2Cl2/MeOH 91:9).
General Procedure for the Conversion of an Aryl Bromide to the Corresponding Arene-Carbonitrile (GP 3).
A suspension of the aryl bromide (1 mmol) and CuCN (4 mmol) in DMF (5 ml, purged with Ar) was heated to
reflux under Ar for 12 48 h. The solvent was partially removed, and CH2Cl2 and conc. aq. NH4OH soln. were
added. The mixture was stirred at r.t. for 1 h, the blue aq. phase was removed, and the org. phase was washed
with conc. aq. NH4OH soln. and H2O. The combined aq. phases were extracted with CH2Cl2, dried (MgSO4),
and evaporated in vacuo to give a residue that was purified by CC (hexane/AcOEt/ 50 :50 or 25 :75).
General Procedure for the Silylation of a OH Group with (t-Bu)Me2SiCl (GP 4). To a soln. of cycloadduct
(10 mmol) and DMAP (17 mmol) in CH2Cl2 (16 ml), (t-Bu)Me2SiCl (15 mmol) was added at r.t. under Ar. The
mixture was stirred for 18 h at 208, then poured into sat. aq. NaHCO3 soln. and extracted with CH2Cl2. The
combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and the resulting residue was
purified by CC (pentane/AcOEt 75 :25).
General Procedure for the Deprotection of a (t-Bu)Me2Si-Protected Hydroxy Group (GP 5). To a soln. of
the silyl-protected compound (3.3 mmol) in dry THF (36 ml), Bu4NF (8 mmol; as soln. in THF) was added
dropwise under cooling with ice. The mixture was stirred for 1 h at 208, then poured into sat. aq. NaCl soln. and
extracted with CH2Cl2. The combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and
the resulting residue was purified by CC (pentane/AcOEt 50 :50).
(3aS,4R,7R,8aS,8bR)-, (3aR,4S,7R,8aR,8bS)-, and (3aS,4S,7R,8aR,8bR)-2-[(Benzo[1,3]dioxol-5-yl)meth-
yl]-4-(4-bromophenyl)-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hydroxy-1H-pyrrolo[3,4-a]pyrrolizin-1,3-dione ((‡)-

2532
Helvetica Chimica Acta Vol. 87 (2004)
6, (À)-7, and (À)-8). GP 1, starting from 3, 4, and 5, gave endo-adducts (‡)-6/(À)-7 as a 1:1 mixture in 34%
yield, which precipitated from AcOEt or MeOH as mixed crystals. In addition, the exo-adduct (‡)-8 was formed
in 35% yield.
Synthesis of (‡)-6 and (À)-7. GP 5, starting from (‡)-19, afforded (‡)-6 in 69% yield and, starting from (‡)-
20, gave (À)-7 in 72% yield.
Synthesis of ( Æ )-7. A 1m soln. of LS-Selectride (3.6 ml, 3.6 mmol) in THF was added to a soln. of (Æ)-11
(1.0 g, 2.07 mmol) in dry CH2Cl2 (10 ml) at À 958 under Ar. The mixture was stirred for 2 h at À 958, then
warmed to 08, neutralized with sat. aq. NaHCO3 soln., and extracted with CH2Cl2. The combined org. phases
were dried (MgSO4), the solvent was removed in vacuo, and the resulting residue was purified by CC (pentane/
EtOAc 50 :50) to give (Æ)-7 (804 mg; 79% as a 95 :5 mixture with (Æ)-6).
Data of ( ‡ )-6: Colorless solid. M.p. 1788. [a]D25 ˆ ‡169.4 (c ˆ 1.00, CHCl3). IR (CHCl3): 3609, 3007, 2931,
1774, 1705, 1607, 1503, 1489, 1446, 1399, 1342, 1090, 1042, 1011. 1H-NMR (400 MHz, CDCl3): 1.88 1.96 (m,
2 H); 2.06 (dd, J ˆ 6.4, 12.7, 1 H); 2.77 (dd, J ˆ 2.9, 13.7, 1 H); 2.94 (dd, J ˆ 6.1, 13.7, 1 H); 3.25 (d, J ˆ 8.3, 1 H);
3.44 (dd, J ˆ 8.3, 8.6, 1 H); 3.97 (d, J ˆ 8.6, 1 H); 4.10 (dd, J ˆ 6.4, 12.7, 1 H); 4.41 (s, 2 H); 4.56 4.59( m, 1 H);
5.93, 5.95 (AB, J ˆ 1.4, 2 H); 6.70 6.78 (m, 3 H); 7.06, 7.37 (AA'BB', J ˆ 8.4, 4 H). 13C-NMR (100 MHz, CDCl3):
39.9; 42.2; 48.4; 50.4; 61.3; 65.8; 69.9; 72.4; 101.1; 108.1; 109.5; 121.7; 122.6; 129.3; 129.7; 131.3; 136.6; 147.2;
‡
‡
147.6; 174.8; 177.5. MALDI-HR-MS: 485.0703 (MH , C23H22BrN2O5 ; calc. 485.0712). Anal. calc. for
C23H21BrN2O5 (485.33): C 56.92, H 4.36, N 5.77; found: C 56.97, H 4.36, N 5.91.
Data of ( Æ )-7: Colorless solid. M.p. 2028.
Data of ( À )-7: Colorless solid. M.p. 1748. [a]D25 ˆ À169.4 (c ˆ 1.00, CHCl3). IR (CHCl3): 3613, 3007, 1774,
1704, 1606, 1503, 1446, 1399, 1342, 1098, 1043. 1H-NMR (400 MHz, CDCl3): 1.69(br. s, 1 H); 1.76 1.83 (m,
1 H); 2.51 2.58 (m, 1 H); 2.66 (d, J ˆ 14.0, 1 H); 2.98 (dd, J ˆ 6.1, 14.0, 1 H); 3.22 (d, J ˆ 8.3, 1 H); 3.55 (dd, J ˆ
8.3, 8.7, 1 H); 3.80 (dd, J ˆ 8.7, 8.9, 1 H); 4.40 (s, 2 H); 4.54 4.58 (m, 1 H); 4.69( d, J ˆ 8.7, 1 H); 5.93, 5.95 (AB,
J ˆ 1.4, 2 H); 6.69 6.80 ( m, 3 H); 7.07, 7.37 (AA'BB', J ˆ 8.3, 4 H). 13C-NMR (100 MHz, CDCl3): 40.7; 42.2;
49.6; 50.3; 60.2; 67.2; 68.8; 74.8; 101.0; 108.1; 109.5; 121.5; 122.6; 129.3; 131.2; 136.9; 147.2; 147.6; 175.1; 177.7.
‡
‡
MALDI-HR-MS: 485.0702 (MH , C23H22BrN2O5 ; calc. 485.0712). Anal. calc. for C23H21BrN2O5 (485.33): C
56.92, H 4.36, N 5.77; found: C 56.92, H 4.44, N 5.82.
Data of ( À )-8: Colorless solid. M.p. 878. [a]D25 ˆ À48.0 (c ˆ 0.50, CHCl3). IR (CHCl3): 3613, 2975, 1703,
1489, 1446, 1394, 1248, 1042. 1H-NMR (400 MHz, CDCl3): 1.78 1.85 (m, 1 H); 1.90 (br. s, 1 H); 1.9 6 (dd, J ˆ 6.7,
14.1, 1 H); 2.63 (dd, J ˆ 5.3, 12.3, 1 H); 2.86 (dd, J ˆ 1.1, 12.3, 1 H); 3.29( dd, J ˆ 5.5, 9.1, 1 H); 3.52 (dd, J ˆ 9.1,
9.1, 1 H); 4.03 (d, J ˆ 5.5, 1 H); 4.09 4.18 ( m, 1 H); 4.26 4.29( m, 1 H); 4.53 (s, 2 H); 5.9 3 (s, 2 H); 6.74 (d, J ˆ
7.7, 1 H); 6.89( d, J ˆ 7.7, 2 H); 7.34, 7.47 (AA'BB', J ˆ 7.6, 4 H). 13C-NMR (100 MHz, CDCl3): 36.7; 42.4; 47.7;
55.7; 61.4; 64.4; 69.5; 72.7; 101.2; 108.3; 109.5; 121.4; 122.8; 128.5; 129.1; 131.8; 140.8; 147.5; 147.8; 176.4; 177.3.
‡
‡
MALDI-HR-MS: 485.0711 (MH , C23H22BrN2O5 ; calc. 485.0712). Anal. calc. for C23H21BrN2O5 (485.33): C
56.92, H 4.36, N 5.77; found: C 57.04, H 4.48, N 5.55.
4-{(3aS,4R,7R,8aS,8bR)- and (3aR,4S,7R,8aR,8bS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-1,3-dioxo-
2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hydroxy-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzonitrile ((‡)-9 and (À)-10).
GP 3, starting from a 1:1 mixture of (‡)-6/(À)-7, afforded (‡)-9 and (À)-10 as a 1:1 mixture, which precipitated
from MeOH as 1:1 mixed crystals in 78% yield. Starting from (‡)-6, (‡)-9 was obtained in 55% yield. Starting
from (À)-7, (À)-10 was isolated in 45% yield.
Data of ( ‡ )-9: Colorless solid. M.p. 2028. [a]D25 ˆ ‡147.4 (c ˆ 0.83, CHCl3). IR (CHCl3): 3612, 3008, 2230,
1774, 1705, 1609, 1490, 1399, 1342, 1171, 1098, 1042. 1H-NMR (400 MHz, CDCl3): 1.90 1.96 (m, 1 H); 2.00 2.13
(m, 2 H); 2.84 (dd, J ˆ 2.9, 13.6, 1 H); 2.87 2.95 (m, 1 H); 3.29( d, J ˆ 8.3, 1 H); 3.52 (dd, J ˆ 8.3, 8.6, 1 H); 4.08
(d, J ˆ 8.6, 1 H); 4.11 4.16 (m, 1 H); 4.39( s, 2 H); 4.60 (m, 1 H); 5.96 (m, 2 H); 6.70 6.77 (m, 3); 7.30 7.33 (m,
2 H); 7.49 7.54 ( m, 2 H). 13C-NMR (100 MHz, CDCl3): 40.1; 42.3; 48.5; 50.5; 61.5; 66.0; 69.0; 72.4; 108.2; 111.6;
‡
118.9; 128.8; 129.3; 131.9; 132.0; 143.8; 147.3; 147.4; 147.7; 174.6; 177.3. MALDI-HR-MS: 432.1550 ( MH ,
‡
C24H22N3O5 ; calc. 432.1559). Anal. calc. for C24H21N3O5 (431.44): C 66.81, H 4.91, N 9.74; found: C 66.83, H 4.94,
N 9.67.
Data of ( À )-10: Colorless solid. M.p. 1898. [a]2D5 ˆ À 178.3 (c ˆ 0.83, CHCl3). IR (CHCl3): 3612, 3008, 2230,
1774, 1705, 1609, 1490, 1399, 1342, 1171, 1098, 1042. 1H-NMR (400 MHz, CDCl3): 1.79 1.87 ( m, 1 H); 1.99 (br. s,
1 H); 2.56 2.63 (m, 1 H); 2.67 (d, J ˆ 13.9, 1 H); 3.02 (dd, J ˆ 5.9, 13.9, 1 H); 3.37 (d, J ˆ 8.3, 1 H); 3.63 (dd, J ˆ
8.3, 8.7, 1 H); 3.85 (dd, J ˆ 8.3, 8.8, 1 H); 4.40 (s, 2 H); 4.58 4.61 (m, 1 H); 4.82 (d, J ˆ 8.7, 1 H); 5.94, 5.96 (AB,
J ˆ 1.4, 2 H); 6.70 6.77 (m, 3 H); 7.32 (m, 2 H); 7.51 (m, 2 H). 13C-NMR (100 MHz, CDCl3): 40.9; 42.3; 49.8;
50.4; 60.3; 67.4; 70.0; 74.8; 101.2; 108.1; 109.5; 111.4; 119.0; 122.6; 128.8; 128.8; 129.3; 131.9; 143.8; 147.7; 174.9;
‡
‡
177.5. MALDI-HR-MS: 432.1550 (MH , C24H22N3O5 ; calc. 432.1559). Anal. calc. for C24H21N3O5 (431.44): C
66.81, H 4.91, N 9.74; found: C 66.71, H 5.00, N 9.60.

Helvetica Chimica Acta Vol. 87 (2004)
2533
(3aSR,4RS,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-4-(4-bromophenyl)-2,3,3a,4,5,6,7,8,8a,8b)-deca-
hydro-1H-pyrrolo[3,4-a]pyrrolizine-1,3,7-trione ((Æ)-11). Me2SO (2.34 ml, 2.58 g, 33 mmol) was added to a
soln. of (COCl)2 (1.41 ml, 2.09g, 16.5 mmol) in dry CH 2Cl2 (50 ml) at À 788 under Ar. After 15 min, a
1:1 mixture of (‡)-6/(À)-7 (4 g, 8.24 mmol) was added and, after 30 min, Et3N (8 ml, 5.84 g, 57.7 mmol), and the
mixture was warmed to 08 and stirred for 2 h. The mixture was diluted with AcOEt (150 ml) and extracted with
sat. aq. NaHCO3 soln. The combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and
the resulting residue purified by CC (AcOEt/pentane 66 :34) to give (Æ)-11 (2.37 g, 60%). Colorless solid. M.p.
1698. IR (CHCl3): 3005, 1750, 1707, 1503, 1490, 1446, 1400, 1343, 1041, 1010, 929. 1H-NMR (400 MHz, CDCl3):
2.33 (dd, J ˆ 11.0, 18.4, 1 H); 2.61 (dd, J ˆ 7.2, 18.4, 1 H); 2.88 (d, J ˆ 18.7, 1 H); 3.17 (d, J ˆ 18.7, 1 H); 3.39( d,
J ˆ 8.3, 1 H); 3.55 (dd, J ˆ 8.3, 8.7, 1 H); 4.08 (d, J ˆ 8.7, 1 H); 4.30 (dd, J ˆ 7.2, 11.0, 1 H); 4.47 (s, 2 H); 5.97 (s,
2 H); 6.73 6.82 (m, 3 H); 7.02, 7.38 (AA'BB', J ˆ 8.4, 4 H). 13C-NMR (100 MHz, CDCl3): 40.9; 44.1; 50.4; 51.3;
60.6; 64.7; 69.6; 100.0; 106.7; 108.1; 120.2; 120.6; 126.7; 127.2; 129.0; 132.3; 144.0; 144.3; 169.5; 172.0; 209.2.
‡
‡
MALDI-HR-MS: 483.0546 (MH , C23H20BrN2O5 ; calc. 483.0556). Anal. calc. for C23H19BrN2O5 (483.31): C
57.16, H 3.96, N 5.80; found: C 57.13, H 4.08, N 5.67.
4{(3aSR,4RS,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-1,3,7-trioxodecahydro-1H-pyrrolo[3,4-a]-
pyrrolizin-4-yl}benzonitrile ((Æ)-12). Me2SO (10.6 ml, 9.38 g, 148 mmol) was added to a soln. of (COCl)2
(6.4 ml, 9.38 g, 74.2 mmol) in dry CH2Cl2 (190 ml) at À 788. After 15 min, a 1:1 mixture of (‡)-9/(À)-10 (16 g,
37.1 mmol) was added and, after 30 min, Et3N (36 ml, 26.28 g, 259mmol), and the mixture was warmed to 0 8 and
stirred for 2 h under Ar. The mixture was diluted with AcOEt (500 ml) and extracted with sat. aq. NaHCO3 soln.
The combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and the resulting residue was
purified by CC (AcOEt/pentane 75 :25) to give (Æ)-12 (10.50 g, 66%). Colorless solid. M.p. 1928. IR (CHCl3):
3473, 3007, 2900, 2231, 1751, 1708, 1609, 1504, 1490, 1399, 1041. 1H-NMR (400 MHz, CDCl3): 2.35 (dd, J ˆ 11.0,
18.4, 1 H); 2.65 (dd, J ˆ 7.2, 18.4, 1 H); 2.86 (d, J ˆ 18.7, 1 H); 3.21 (d, J ˆ 18.7, 1 H); 3.44 (d, J ˆ 8.3, 1 H); 3.63
(dd, J ˆ 8.3, 8.7, 1 H); 4.20 (d, J ˆ 8.7, 1 H); 4.34 (dd, J ˆ 7.2, 11.0, 1 H); 4.38, 4.45 (AB, J ˆ 6.6, 2 H); 5.97, 5.99
(AB, J ˆ 1.4, 2 H); 6.73 6.81 (m, 3 H); 7.27, 7.54 (AA'BB', J ˆ 7.5, 4 H). 13C-NMR (100 MHz, CDCl3): 39.2;
42.6; 49.1; 50.1; 59.8; 64.3; 69.3; 101.2; 108.2; 109.6; 112.3; 118.6; 122.8; 128.8; 129.1; 132.2; 141.6; 147.5; 147.7;
‡
‡
174.0; 176.5; 215.2. MALDI-HR-MS: 430.1402 (MH , C24H20N3O5 ; calc. 430.1403). Anal. calc. for C24H19N3O5
(429.42): C 67.13, H 4.46, N 9.79; found: C 67.13, H 4.65, N 9.71.
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-meth-
oxy-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzonitrile ((Æ)-13). A suspension of NaH (63 mg, 1.75 mmol)
and (Æ)-10 (500 mg, 1.16 mmol) in dry THF (5 ml) was stirred for 10 min. After addition of [15]crown-5
(0.41 ml, 2.09mmol), the soln. was stirred for another 10 min and MeI (0.11 ml, 1.75 mmol) was added. The soln.
was stirred at r.t. for 3 h, and subsequently treated with sat. aq. NH4Cl soln. The aq. phase was removed and
extracted with AcOEt. The combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and
the resulting residue was purified by CC (AcOEt/pentane 67:33) to yield (Æ)-13 (196 mg, 38%). Colorless solid.
M.p. 1698. IR (CHCl3): 3008, 2972, 2230, 1705, 1608, 1503, 1490, 1446, 1369, 1342, 1100, 1042. 1H-NMR
(400 MHz, CDCl3): 1.76 1.83 (m, 1 H); 2.50 2.58 (m, 1 H); 2.76 (d, J ˆ 14.0, 1 H); 2.91 (dd, J ˆ 6.0, 14.0, 1 H);
3.22 (s, 3 H); 3.34 (d, J ˆ 8.3, 1 H); 3.58 (dd, J ˆ 8.3, 8.7, 1 H); 3.84 (dd, J ˆ 8.3, 9.9, 1 H); 4.00 4.05 (m, 1 H);
4.40 (s, 2 H); 4.66 (dd, J ˆ 8.7, 1 H); 5.96 (d, J ˆ 1.4, 2 H); 6.70 6.78 (m, 3 H); 7.31, 7.52 (AA'BB', J ˆ 8.3, 4 H).
13C-NMR (100 MHz, CDCl3): 38.1; 42.3; 49.7; 50.3; 56.7; 57.3; 67.1; 68.6; 84.0; 101.2; 108.1; 109.6; 111.5; 119.0;
‡
‡
122.6; 128.9; 129.3; 131.9; 143.7; 147.4; 147.7; 174.9; 177.4. MALDI-HR-MS: 446.1707 ( MH , C25H24N3O5 ; calc.
446.1716). Anal. calc. for C25H23N3O5 (445.47): C 67.41, H 5.20, N 9.43; found: C 67.40, H 5.29, N 9.35.
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-7-(benzyloxy)-2,3,3a,4,5,6,7,8,8a,8b-
decahydro-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzonitrile ((Æ)-14). A suspension of NaH (63 mg,
1.75 mmol) and (Æ)-10 (500 mg, 1.16 mmol) in dry THF (5 ml) was stirred for 10 min. After addition of
[15]crown-5 (0.41 ml, 2.09mmol), the soln. was stirred for 10 min and BnBr (0.69ml, 5.79mmol) was added.
The soln. was stirred at r.t. for 2 h, and treated with sat. aq. NH4Cl soln. The aq. phase was separated and
extracted with AcOEt. The combined org. phases were dried (MgSO4), the solvent was removed in vacuo, and
the resulting residue was purified by CC (AcOEt/pentane 50 :50) to yield (Æ)-14 (421 mg, 70%). Colorless solid.
1
M.p. 1518. IR (CHCl3): 3007, 2929, 2230, 1774, 1705, 1609, 1503, 1490, 1446, 1399, 1343, 1100, 1042. H-NMR
(400 MHz, CDCl3): 1.85 1.92 (m, 1 H); 2.53 2.60 (m, 1 H); 2.83 (d, J ˆ 13.9, 1 H); 2.92 (dd, J ˆ 5.8, 13.9, 1 H);
3.34 (d, J ˆ 8.3, 1 H); 3.57 (dd, J ˆ 8.3, 8.7, 1 H); 3.84 (dd, J ˆ 8.3, 9.6, 1 H); 4.22 4.26 (m, 1 H); 4.34 4.44 (m,
4 H); 4.75 (d, J ˆ 8.7, 1 H); 5.95 (d, J ˆ 1.4, 2 H); 6.70 6.77 (m, 3 H); 7.26 7.34 (m, 5 H); 7.22, 7.52 (AA'BB', J ˆ
7.3, 4 H). 13C-NMR (100 MHz, CDCl3): 38.4; 42.3; 49.7; 50.3; 56.9; 67.1; 68.6; 71.8; 81.8; 101.2; 108.1; 109.6;
111.5; 118.9; 122.6; 127.6; 127.8; 128.3; 128.5; 128.9; 129.3; 131.9; 132.2; 137.8; 143.7; 147.3; 147.7; 174.9; 177.4.

2534
Helvetica Chimica Acta Vol. 87 (2004)
‡
‡
MALDI-HR-MS: 522.2019( MH , C31H28N3O5 ; calc. 522.2029). Anal. calc. for C31H27N3O5 (521.56): C 71.39, H
5.22, N 8.06; found: C 71.18, H 5.47, N 8.07.
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hy-
droxy-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((Æ)-15). GP 2, starting from
(Æ)-10, afforded (Æ)-15 in 80% yield. Yellowish solid. M.p. 1758. IR (CHCl3): 3502, 2972, 1704, 1503, 1490, 1446,
1
1399, 1242. H-NMR (400 MHz, (CD3)2SO): 1.72 1.79( m, 1 H); 2.35 2.42 (m, 2 H); 2.90 (m, 1 H); 3.34 (s,
1 H); 3.51 (d, J ˆ 8.4, 1 H); 3.61 (dd, J ˆ 7.9, 10.3, 1 H); 3.84 (dd, J ˆ 8.4, 8.6, 1 H); 4.31 4.38 (m, 2 H); 4.81 (d,
J ˆ 8.6, 1 H); 5.06 5.18 (m, 1 H); 6.01 (s, 2 H); 6.63 6.70 (m, 2 H); 6.87 (d, J ˆ 8.0, 1 H); 7.44, 7.74 (AA'BB',
J ˆ 8.4, 4 H); 9.31 (s, 4 H). 13C-NMR (100 MHz, (CD3)2SO): 47.8; 48.5; 48.8; 50.1; 59.6; 66.7; 68.1; 72.8; 100.9;
108.0; 108.1; 121.0; 126.3; 127.4; 128.5; 129.6; 145.5; 146.4; 147.1; 165.4; 175.3; 178.0. MALDI-HR-MS: 449.1825
‡
‡
(MH , C24H25ClN4O5 ; calc. 449.1825).
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-meth-
oxy-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((Æ)-16). GP 2, starting from (Æ)-
13, afforded (Æ)-16 in 38% yield. Yellowish solid. M.p. > 1558 (dec.). IR (CHCl3): 2972, 1774, 1704, 1506, 1486,
1444, 1399, 1246. 1H-NMR (400 MHz, (CD3)2SO): 1.77 1.84 (m, 1 H); 2.40 2.48 (m, 1 H); 2.50 2.55 (m, 1 H);
2.90 (dd, J ˆ 6.4, 11.1, 1 H); 3.13 (s, 3 H); 3.52 (d, J ˆ 8.2, 1 H); 3.66 (dd, J ˆ 8.2, 9.6, 1 H); 3.84 (dd, J ˆ 8.2, 11.1,
1 H); 4.02 (m, 1 H); 4.85, 4.75 (AB, J ˆ 14.7, 2 H); 4.62 (d, J ˆ 9.6, 1 H); 6.02 (s, 2 H); 6.64 6.69( m, 2 H); 6.88
(d, J ˆ 7.9, 1 H); 7.43, 7.75 (AA'BB', J ˆ 8.5, 4 H); 9.34 (s, 4 H). 13C-NMR (100 MHz, (CD3)2SO): 36.9; 48.7; 50.0;
56.0; 62.7; 66.6; 67.7; 83.4; 101.0; 108.0; 108.1; 121.0; 126.4; 127.3; 127.5; 128.5; 129.6; 145.1; 146.5; 147.1; 165.3;
‡
‡
175.2; 177.9. MALDI-HR-MS: 463.1980 (MH , C25H27N4O5 ; calc. 463.1982).
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-7-(benzyloxy)-2,3,3a,4,5,6,7,8,8a,8b-
decahydro-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((Æ)-17). GP 2, starting
from (Æ)-14, afforded (Æ)-17 in 50% yield. Yellowish solid. M.p. 1488. IR (CHCl3): 2972, 1705, 1501, 1494,
1
1439, 1399, 1240. H-NMR (400 MHz, (CD3)2SO): 1.88 1.95 (m, 1 H); 2.46 2.51 (m, 1 H); 2.63 (d, J ˆ 12.8,
1 H); 2.92 (dd, J ˆ 5.8, 12.8, 1 H); 3.38, 3.40 (AB, J ˆ 2.5, 2 H); 3.54 (d, J ˆ 7.8, 1 H); 3.67 (dd, J ˆ 7.8, 9.7, 1 H);
3.87 (dd, J ˆ 8.6, 9.7, 1 H); 4.22 (dd, J ˆ 5.8, 12.8, 1 H); 4.33 (s, 2 H); 4.74 (d, J ˆ 8.6, 1 H); 6.02 (s, 2 H); 6.65
6.69( m, 2 H); 6.88 (d, J ˆ 7.9, 1 H); 7.25 7.35 (m, 5 H); 7.42, 7.76 (AA'BB', J ˆ 8.5, 4 H); 9.40 (s, 4 H). 13C-NMR
(100 MHz, (CD3)2SO): 37.2; 48.7; 50.1; 56.2; 62.7; 66.6; 67.7; 70.7; 81.4; 101.0; 108.0; 108.1; 121.0; 126.4; 127.2;
‡
127.3; 127.5; 128.1; 128.5; 129.6; 138.2; 145.2; 146.5; 147.1; 165.3; 175.3; 177.9. MALDI-HR-MS: 539.2284 (MH ,
‡
C31H31N4O5 ; calc. 539.2295).
4-{(3aSR,4RS,7SR,8aSR,8bRS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7,7-di-
methoxy-1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((Æ)-18). GP 2, starting from
(Æ)-12, afforded (Æ)-18 in 13% yield. Yellowish solid. M.p. > 1568 (dec.). IR (CHCl3): 2972, 1771, 1707, 1502,
1
1489, 1444, 1399, 1238. H-NMR (400 MHz, (CD3)2SO): 2.11 (dd, J ˆ 11.2, 13.3, 1 H); 2.26 (d, J ˆ 13.3, 1 H);
2.49 2.51 ( m, 1 H); 2.57 (d, J ˆ 13.3, 1 H); 2.91 (d, J ˆ 13.3, 1 H); 3.08 (s, 6 H); 3.48 (d, J ˆ 8.7, 1 H); 3.78 3.86
(m, 1 H); 4.33 (d, J ˆ 6.6, 2 H); 4.57 (d, J ˆ 8.7, 1 H); 6.02 (s, 2 H); 6.64 6.69( m, 2 H); 6.87 6.89( m, 1 H); 7.41,
7.72 (AA'BB', J ˆ 8.5, 4 H); 9.26 (s, 4 H). 13C-NMR (100 MHz, (CD3)2SO): 48.4; 48.9; 49.7; 50.0; 58.5; 64.8; 65.5;
67.7; 101.0; 108.0; 108.1; 112.7; 121.1; 126.6; 127.5; 128.5; 129.6; 144.8; 146.5; 147.1; 165.3; 175.1; 177.7. MALDI-
‡
‡
HR-MS: 493.2098 (MH , C26H29N4O6 ; calc. 493.2087).
(3aS,4R,7R,8aS,8bR)- and (3aR,4S,7R,8aR,8bS)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-4-(4-bromophenyl)-
7-[(tert-butyl)dimethylsilyloxy]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-1H-pyrrolo[3,4-a]pyrrolizin-1,3-dione ((‡)-
19) and (À)-20). GP 4, starting from a 1:1 mixture of (‡)-6 and (À)-7, afforded (‡)-19 (43% yield) and (À)-
20 (43% yield).
Data of ( ‡ )-19: Colorless solid. M.p. 818. [a]D25 ˆ ‡112.1 (c ˆ 1.00, CHCl3). IR (CHCl3): 3684, 2929, 2856,
1
1774, 1704, 1602, 1503, 1489, 1446, 1400, 1369, 1342, 1097, 1042. H-NMR (400 MHz, CDCl3): À 0.01 (s, 3 H);
0.02 (s, 3 H); 0.85 (s, 9H); 1.82 1.90 ( m, 1 H); 2.01 (dd, J ˆ 6.4, 12.9, 1 H); 2.74 (dd, J ˆ 3.1, 13.4, 1 H); 2.89( dd,
J ˆ 6.1, 13.4, 1 H); 3.23 (dd, J ˆ 8.5, 8.5, 1 H); 3.41 (dd, J ˆ 8.5, 8.5, 1 H); 3.93 (d, J ˆ 8.5, 1 H); 4.09( dd, J ˆ 8.7,
8.8, 1 H); 4.43 (s, 2 H); 4.50 4.53 (m, 1 H); 5.94, 5.96 (AB, J ˆ 1.4, 2 H); 6.70 6.80 (m, 3 H); 7.07, 7.38 (AA'BB',
J ˆ 8.4, 4 H). 13C-NMR (100 MHz, CDCl3): À 4.9; À 4.8; ‡18.0; 25.8; 40.1; 42.2; 48.3; 50.4; 61.9; 65.9; 72.7;
‡
101.1; 108.1; 109.6; 121.7; 122.6; 129.4; 131.3; 136.8; 147.2; 147.6; 174.9; 177.6. MALDI-HR-MS: 599.1566 (MH ,
‡
C29H36BrN2O5Si ; calc. 599.1577). Anal. calc. for C29H35BrN2O5Si (599.59): C 58.09, H 5.88, N 4.67; found: C
58.19, H 6.02, N 4.59.
Data of ( À )-20: Colorless solid. M.p. 798. [a]D25 ˆ À 116.7 (c ˆ 1.00, CHCl3). IR (CHCl3): 3684, 3007, 2928,
2856, 1773, 1704, 1602, 1503, 1490, 1446, 1399, 1370, 1342, 1098, 1043, 1010. 1H-NMR (400 MHz, CDCl3): À 0.04
(s, 3 H); 0.02 (s, 3 H); 0.85 (s, 9H); 1.73 1.79( m, 1 H); 2.46 2.53 (m, 1 H); 2.62 (d, J ˆ 13.6, 1 H); 2.91 (dd, J ˆ
6.1, 13.4, 1 H); 3.31 (d, J ˆ 8.6, 1 H); 3.55 (dd, J ˆ 8.6, 8.6, 1 H); 3.79( dd, J ˆ 8.6, 8.6, 1 H); 4.41 (s, 2 H);

Helvetica Chimica Acta Vol. 87 (2004)
2535
4.42 4.47 (m, 1 H); 4.71 (d, J ˆ 8.6, 1 H); 5.94, 5.96 (AB, J ˆ 1.4, 2 H); 6.70 6.79( m, 3 H); 7.04, 7.35 (AA'BB',
J ˆ 8.4, 4 H). 13C-NMR (100 MHz, CDCl3): À 5.0; À 4.9; 17.8; 25.7; 41.7; 42.2; 49.9; 50.4; 60.6; 67.1; 68.8; 75.0;
101.1; 108.1; 109.6; 121.5; 122.6; 129.4; 129.8; 131.2; 137.2; 147.2; 147.6; 175.2; 177.8. MALDI-HR-MS: 599.1566
‡
‡
(MH , C29H36BrN2O5Si ; calc. 599.1577). Anal. calc. for C29H35BrN2O5Si (599.59): C 58.09, H 5.88, N 4.67;
found: C 58.22, H 5.75, N 4.62.
4-{(3aS,4R,7R,8aS,8bR)-2-[(Benzo[1,3]dioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hydroxy-
1,3-dioxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((‡)-21). GP 2, starting from (‡)-9,
afforded (‡)-21 in 55% yield. Yellowish solid. M.p. > 2308 (dec.). [a]2D5 ˆ ‡108.9( c ˆ 0.43, CHCl3). IR
1
(CHCl3): 3502, 2972, 1704, 1503, 1490, 1446, 1399, 1242. H-NMR (400 MHz, (CD3)2SO): 1.72 1.79( m, 1 H);
2.35 2.42 (m, 2 H); 2.90 (m, 1 H); 3.34 (s, 1 H); 3.51 (d, J ˆ 8.4, 1 H); 3.61 (dd, J ˆ 7.9, 10.3, 1 H); 3.84 (dd, J ˆ
8.4, 8.6, 1 H); 4.31 4.38 (m, 2 H); 4.81 (d, J ˆ 8.6, 1 H); 5.06 5.18 (m, 1 H); 6.01 (s, 2 H); 6.63 6.70 (m, 2 H);
6.87 (d, J ˆ 8.0, 1 H); 7.44, 7.74 (AA'BB', J ˆ 8.4, 4 H); 9.31 (s, 4 H). 13C-NMR (100 MHz, (CD3)2SO): 47.8; 48.5;
48.8; 50.1; 59.6; 66.7; 68.1; 72.8; 100.9; 108.0; 108.1; 121.0; 126.3; 127.4; 128.5; 129.6; 145.5; 146.4; 147.1; 165.4;
‡
‡
175.3; 178.0. MALDI-HR-MS: 449.1825 (MH , C24H25N4O5 ; Ber. 449.1825).
(3aS,4R,7R,8aS,8bS)-2-[(1,3-Benzodioxol-5-yl)methyl]-4-(4-bromophenyl)-7-[tert-butyldimethylsilyloxy]-
2,3,3a,4,5,6,7,8,8a,8b-decahydro-1-hydroxy-1H-pyrrolo[3,4-a]pyrrolizin-3-one ((‡)-22).
A 1m soln. of
Li[Et3BH] in THF (2.83 ml, 2.83 mmol) was added to (‡)-19 (1.00 g, 1.66 mmol) in dry THF (6 ml) at À 788
under Ar. After 30 min, the mixture was warmed to r.t., neutralized with sat. aq. NaHCO3 soln., and extracted
with CH2Cl2. The org. phase was dried (MgSO4), the solvent removed, and the residue purified by CC (AcOEt/
pentane 50 :50, then 80 :20) to yield (‡)-22 (939 mg, 94%). Colorless solid. M.p. 78 808 (MeOH). [a]2D5 ˆ ‡55
1
(c ˆ 1.00 CHCl3). IR (CHCl3): 3351, 2928, 1673, 1503, 1443, 1243, 1187. H-NMR (400 MHz, CDCl3): 0.02 (d,
J ˆ 9.1, 6 H); 0.86 (s, 9H); 1.80 1.92 ( m, 3 H); 2.81 (dd, J ˆ 3.6, 13.5, 1 H); 3.00 (dd, J ˆ 7.1, 8.5, 1 H); 3.07 (dd,
J ˆ 6.3, 13.5, 1 H); 3.31 (dd, J ˆ 8.5, 10.3, 1 H); 3.76 (d, J ˆ 10.3, 1 H); 3.88 (dd, J ˆ 6.7, 11.1, 1 H); 4.03 (d, J ˆ
14.5, 1 H); 4.44 (d, J ˆ14.5, 1 H); 4.61 6.63 (m, 1 H); 4.9 8 (d, J ˆ 7.1, 1 H); 5.92, 5.93 (AB, J ˆ 1.5, 2 H); 6.71
6.72 (m, 3 H); 7.14, 7.42 (AA'BB', J ˆ 7.3, 4 H). 13C-NMR (100 MHz, CDCl3): 18.0; 25.8; 38.2; 43.8; 49.8; 62.1;
64.0; 68.6; 72.7; 76.7; 82.3; 101.0; 108.0; 109.2; 121.7; 122.1; 129.6; 130.6; 131.4; 136.5; 146.9; 147.7; 171.5.
‡
‡
MALDI-HR-MS: 600.1600 (MH , C29H38BrN2O5Si ; calc. 600.1655).
(1S,3aS,4R,7R,8aS,8bS)-2-[(1,3-Benzodioxol-5-yl)methyl]-4-(4-bromophenyl)-2,3,3a,4,5,6,7,8,8a,8b-deca-
hydro-7-hydroxy-1-[(4-methylphenyl)sulfonyl]-1H-pyrrolo[3,4-a]pyrrolizin-3-one ((‡)-23). GP 5, starting
from (‡)-22, afforded the hydroxy intermediate in 94% yield without further purification. A mixture of 4-
toluenesulfinic acid (69.2 mg, 0.44 mmol) and dry powdered CaCl2 (49.2 mg, 0.44 mmol) in dry CH2Cl2 (0.8 ml)
was stirred for 10 min under Ar, and a soln. of the hydroxy intermediate (80 mg, 0.16 mmol) in CH2Cl2 (0.4 ml)
was added. After stirring for 19h, sat. aq. NaHCO 3 soln. was added, and the mixture was extracted with CH2Cl2.
The org. phase was dried (MgSO4) and evaporated in vacuo. The residue was purified by CC (AcOEt/hexane
50 :50) to yield (‡)-23 (82 mg, 80%). Yellow solid. M.p. 108 1108 (AcOEt). [a]2D5 ˆ ‡209( c ˆ 1.00, CHCl3).
IR (CHCl3): 3421, 2923, 1705, 1595, 1503, 1488, 1445, 1400, 1372, 1292. 1H-NMR (400 MHz, CDCl3): 1.67 1.74
(m, 1 H); 1.9 5 (dd, J ˆ 6.1, 13.5, 1 H); 1.98 (s, 1 H); 2.42 (s, 3 H); 2.52 (t, J ˆ 6.9, 1 H); 2.71 (dd, J ˆ 5.3, 13.1,
1 H); 2.82 (dd, J ˆ 1.9, 13.0, 1 H); 2.96 (dd, J ˆ 3.4, 7.7, 1 H); 3.34 3.39( m, 1 H); 3.81 3.85 (m, 2 H); 4.29( s,
1 H); 4.43 (t, J ˆ 5.3, 1 H); 4.94 (d, J ˆ 14.7, 1 H); 5.90, 5.92 (AB, J ˆ 1.4, 2 H); 6.56 6.72 (m, 3 H); 7.09( d, J ˆ
6.7, 2 H); 7.32 7.37 (m, 4 H); 7.65 (d, J ˆ 8.3, 2 H). 13C-NMR (100 MHz, CDCl3): 21.8; 40.9; 42.1; 45.1; 51.4;
61.5; 69.3; 70.7; 73.2; 80.9; 101.2; 108.4; 108.7; 121.3; 122.0; 128.7; 129.4; 129.6; 130.5; 131.0; 132.5; 136.6; 146.2;
‡
‡
147.4; 148.2; 172.3. MALDI-HR-MS: 627.0986 (MH , C30H30BrN2O6S ; calc. 627.0988).
(1R,3aS,4R,7R,8aS,8bR)-2-[(1,3-Benzodioxol-5-yl)methyl]-4-(4-bromophenyl)-7-[(tert-butyl)dimethylsi-
lyloxy]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-1-[(4-methylphenyl)sulfonyl]-1H-pyrrolo[3,4-a]pyrrolizin-3-one ((‡)-
24). GP 4, starting from (‡)-23, afforded (‡)-24 in 87% yield. Yellow foam. M.p. 109 101 8 (AcOEt).
[a]2D5 ˆ ‡171 (c ˆ 1.00, CHCl3). IR (CHCl3): 2929, 2359, 1711, 1503, 1488, 1398, 1302, 1134. 1H-NMR (400 MHz,
CDCl3): À 0.06 (s, 6 H); 0.79( s, 9H); 1.61 1.69( m, 1 H); 1.87 (dd, J ˆ 6.0, 13.0, 1 H); 2.42 (s, 3 H); 2.45 (t, J ˆ
6.5, 1 H); 2.64 (dd, J ˆ 2.3, 12.8, 1 H); 2.75 (dd, J ˆ 2.3, 12.8, 1 H); 2.96 (dd, J ˆ 3.1, 7.6, 1 H); 3.33 3.37 (m,
1 H); 3.78 (d, J ˆ 6.5, 1 H); 3.83 (d, J ˆ 15.1, 1 H); 4.32 (s, 1 H); 4.34 4.37 (m, 1 H); 4.9 6 (d, J ˆ 15.1, 1 H); 5.91
(s, 2 H); 6.58 6.64 (m, 2 H); 6.71 (d, J ˆ 7.9, 1 H); 7.10 (d, J ˆ 8.3, 2 H); 7.32 7.37 (m, 4 H); 7.65 (d, J ˆ 8.3,
2 H). 13C-NMR (100 MHz, CDCl3): 18.0; 21.8; 25.8; 41.4; 41.9; 45.0; 51.3; 61.8; 69.5; 70.6; 73.3; 81.1; 101.1; 108.4;
108.7; 121.2; 121.8; 128.8; 129.4; 129.7; 130.5; 131.0; 132.5; 136.8; 146.2; 147.4; 148.2; 172.4. MALDI-HR-MS:
‡
‡
739.1863 (MH , C36H44BrN2O6SSi ; calc. 739.1873).
(1S,3aS,4S,7R,8aS,8bS)-2-[(1,3-Benzodioxol-5-yl)methyl]-4-(4-bromophenyl)-7-[(tert-butyl)dimethylsilyloxy]-
2,3,3a,4,5,6,7,8,8a,8b-decahydro-1-(1-methylethyl)-1H-pyrrolo[3,4-a]pyrrolizin-3-one ((‡)-25). (i-Pr)MgCl
(1.35 ml of a 2m soln. in THF/Et2O, 2.7 mmol) was added to a soln. of ZnCl2 (1.48 ml of 1m soln in Et2O,

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Helvetica Chimica Acta Vol. 87 (2004)
1.48 mmol) in dry CH2Cl2 (6 ml) under Ar. After 30 min, a soln. of (‡)-25 (1 g, 1.35 mmol) in dry CH2Cl2
(6.5 ml) was slowly added at 08. The mixture was stirred at r.t. for 23 h, then 1n HCl was added. After 5 min, the
mixture was neutralized with sat. aq. NaHCO3 soln. and extracted with CH2Cl2. The org. phase was dried
(MgSO4), the solvent was removed, and the product was purified by CC (AcOEt/hexane 50 :50) to yield (‡)-26
(668 mg, 79%). Colorless solid. M.p. 63 658 (AcOEt). [a]2D5 ˆ ‡183 (c ˆ 1.00, CHCl3). IR (CHCl3): 2955, 1685,
1487, 1441, 1371, 1094, 1070, 1038, 1009. 1H-NMR (400 MHz, CDCl3): À 0.04 (s, 6 H); 0.64 (d, J ˆ 6.9, 3 H); 0.80
(s, 9H); 0.87 ( d, J ˆ 6.9, 3 H); 1.67 1.74 (m, 1 H); 1.82 (dd, J ˆ 6.3, 13.0, 1 H); 2.00 2.11 (m, 1 H); 2.39 2.43
(m, 1 H); 2.74 2.83 (m, 2 H); 3.17 (t, J ˆ 8.1, 1 H); 3.24 (t, J ˆ 3.9, 1 H); 3.45 3.50 (m, 1 H); 3.64 (d, J ˆ 14.9,
1 H); 3.90 (d, J ˆ 7.6, 1 H); 4.42 4.45 (m, 1 H); 4.73 (d, J ˆ 14.9, 1 H); 5.89, 5.91 (AB, J ˆ 1.5, 2 H); 6.60 (dd, J ˆ
1.6, 7.9, 1 H); 6.65 6.67 (m, 2 H); 7.22, 7.39( AA'BB', J ˆ 8.4, 4 H). 13C-NMR (100 MHz, CDCl3): 14.7; 18.1;
18.5; 25.9; 28.0; 40.3; 41.2; 43.8; 52.7; 62.2; 67.3; 71.0; 71.2; 73.3; 101.0; 108.1; 108.7; 120.9; 121.2; 130.0; 130.4;
‡
‡
131.0; 138.3; 146.9; 148.0; 172.3. MALDI-HR-MS: 627.2256 ( MH , C32H44BrN2O4Si ; calc. 627.2254).
(1R,3aS,4R,7R,8aS,8bR)-2-[(1,3-Benzodioxol-5-yl)methyl]-4-(4-bromophenyl)-2,3,3a,4,5,6,7,8,8a,8b-deca-
hydro-7-hydroxy-1-(1-methylethyl)-1H-pyrrolo[3,4-a]pyrrolizin-3-one ((‡)-26). GP 5, starting from (‡)-25,
afforded (‡)-26 in 75% yield. Colorless crystals. M.p. 153 1558 (MeCN). [a]2D5 ˆ ‡208 (c ˆ 1.00, CHCl3). IR
1
(CHCl3): 2928, 2360, 1682, 1503, 1488, 1442, 1371. H-NMR (400 MHz, CDCl3): 0.70 (d, J ˆ 6.8, 3 H); 0.91 (d,
J ˆ 6.9, 3 H); 1.71 (s, 1 H); 1.78 1.84 (m, 1 H); 2.00 (ddd, J ˆ 1.3, 6.1, 13.4, 1 H); 2.05 2.12 (m, 1 H); 2.47 2.50
(m, 1 H); 2.89 2.90 ( m, 2 H); 3.25 3.31 (m, 2 H); 3.54 3.59( m, 1 H); 3.68 (d, J ˆ 14.9, 1 H); 4.03 (d, J ˆ 7.7,
1 H); 4.55 (s, 1 H); 4.88 (d, J ˆ 14.9, 1 H); 5.95, 5.97 (AB, J ˆ 1.4, 2 H); 6.59( dd, J ˆ 1.7, 7.9, 1 H); 6.67 (d, J ˆ 1.7,
1 H); 6.73 (d, J ˆ 7.9; 1 H); 7.26, 7.44 (AA'BB', J ˆ 8.3, 4 H). 13C-NMR (100 MHz, CDCl3): 14.7; 18.5; 27.9; 40.7;
40.8; 43.8; 53.0; 62.2; 67.2; 71.1; 71.2; 73.4; 101.0; 108.1; 108.7; 121.0; 121.3; 129.9; 130.3; 131.0; 138.2; 146.9;
‡
‡
147.9; 172.1. MALDI-HR-MS: 513.1390 ( MH , C26H30BrN2O4 ; calc. 513.1389). Anal. calc. for C26H29BrN2O4
(513.42): C 60.82, H 5.69, N 5.46; found: C 60.86, H 5.72, N 5.33.
4-{(1S,3aS,4S,7R,8aS,8bS)-2-[(1,3-Benzodioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hydroxy-1-
(1-methylethyl)-3-oxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzonitrile ((‡)-27). GP 3, starting from (‡)-26,
afforded (‡)-27 in 55% yield. Grey solid. M.p. 768. [a]2D5 ˆ ‡285 (c ˆ 0.50, CHCl3). IR (CHCl3): 3472, 2961,
2236, 1681, 1608, 1501, 1486, 1441, 1239, 1039. 1H-NMR (400 MHz, CDCl3): 0.71 (d, J ˆ 6.8, 3 H); 0.92 (d, J ˆ 6.9,
3 H); 1.63 (s, 1 H); 1.76 1.84 (m, 1 H); 2.01 2.12 (m, 2 H); 2.49 2.53 ( m, 1 H); 2.86 (dd, J ˆ 5.3, 12.8, 1 H);
2.95 (dd, J ˆ 2.1, 12.8, 1 H); 3.26 (t, J ˆ 3.0, 1 H); 3.36 (t, J ˆ 8.2, 1 H); 3.57 3.62 (m, 1 H); 3.67 (d, J ˆ 15.1, 1 H);
4.13 (d, J ˆ 7.7, 1 H); 4.58 (s, 1 H); 4.76 (d, J ˆ 15.1, 1 H); 5.96, 5.98 (AB, J ˆ 1.4, 2 H); 6.60 6.62 (m, 2 H); 6.73
(d, J ˆ 8.3, 1 H); 7.50, 7.61 (AA'BB', J ˆ 8.2, 4 H). 13C-NMR (100 MHz, CDCl3): 14.8; 18.5; 28.0; 40.8; 40.8; 43.8;
53.2; 62.3; 67.1; 71.3; 71.5; 73.5; 101.1; 108.1; 108.5; 110.9; 119.3; 121.3; 128.8; 130.2; 131.7; 145.1; 147.0; 148.0;
‡
‡
171.8. MALDI-HR-MS: 460.2229( MH , C27H30N3O4 ; calc. 460.2236).
4-{(1S,3aS,4S,7R,8aS,8bS)-2-[(1,3-Benzodioxol-5-yl)methyl]-2,3,3a,4,5,6,7,8,8a,8b-decahydro-7-hydroxy-1-
(1-methylethyl)-3-oxo-1H-pyrrolo[3,4-a]pyrrolizin-4-yl}benzamidine Hydrochloride ((‡)-2). GP 2, starting
from (‡)-27, afforded (‡)-2 in 60% yield. Yellowish powder. M.p. > 2008 (dec.). [a]2D5 ˆ ‡211 (c ˆ 0.45,
CHCl3). IR (neat): 3276, 2961, 2359, 1667, 1539, 1489, 1441, 1373, 1242. 1H-NMR (300 MHz, CD3OD): 0.74 (d,
J ˆ 6.9, 3 H); 0.97 (d, J ˆ 6.9, 3 H); 1.91 1.96 (m, 2 H); 2.16 2.22 (m, 1 H); 2.66 2.71 (m, 1 H); 2.80 2.86 (m,
2 H); 3.36 (t, J ˆ 2.8, 1 H); 3.45 (t, J ˆ 7.6, 1 H); 3.55 3.63 (m, 1 H); 3.82 (d, J ˆ 14.9, 1 H); 4.27 (d, J ˆ 7.6, 1 H);
4.49 4.52 ( m, 1 H); 4.65 (d, J ˆ 14.9, 1 H); 5.94, 5.96 (AB, J ˆ 1.2, 2 H); 6.72 6.80 (m, 3 H); 7.61, 7.72 (AA'BB',
J ˆ 8.2, 4 H). 13C-NMR (75 MHz, CD3OD): 14.7; 18.4; 29.0; 41.0; 41.2; 44.4; 54.7; 62.5; 69.3; 72.3; 73.0; 73.5;
102.3; 108.9; 109.2; 122.4; 127.6; 128.1; 130.1; 131.1; 148.1; 148.4; 149.2; 168.3; 174.4. MALDI-HR-MS: 477.2490
‡
‡
(MH , C27H33N4O5 ; calc. 477.2502).
X-Ray Crystal Structures. Copies of the data can be obtained free of charge on application to Cambridge
Crystallographic Data Centre (CCDC), 12 Union Road, Cambridge CB21EZ, UK (fax: (‡44)1223-336-033;
e-mail: deposit@ccdc.cam.ac.uk.
Mixed Crystal of ( ‡ )-9 and ( À )-10. Crystal data at 173(2) K for C24H21N3O5 ¥ C24H21N3O5 (Mr 862.88):
triclinic, space group P1, Dc ˆ 1.450 g cm-3, Z ˆ 1, a ˆ 4.7947(1), b ˆ 14.9058(3), c ˆ 15.1606(9) ä, a ˆ
111.174(1), b ˆ 94.024 (1), g ˆ 99.021(1)8, Vˆ 988.36(4) ä3. Bruker-Nonius Kappa-CCD diffractometer,
MoKa radiation, l ˆ 0.7107, linear crystal dimensions ca. 0.15  0.12  0.10 mm. The structure was solved by
direct methods (SIR97) [25] and refined by full-matrix least-squares analysis (SHELXL-97) [26], using an
isotropic extinction correction. All heavy atoms were refined anisotropically, H-atoms isotropically, whereby H-
positions are based on stereochemical considerations. Final R(F) ˆ 0.0402, wR(F2) ˆ 0.0980 for 620 parameters,
3 restraints, and 6971 reflections with I > 2s(I) and 2.91 < q < 27.488 . Deposition No. CCDC-238291.
Compound (Æ)-11. Crystal data at 223(2) K for C23H19BrN2O5 (Mr 483.31): monoclinic, space group P2(1)/
n, Dc ˆ 1.598 g cmÀ3, Z ˆ 4, a ˆ 10.6776(2), b ˆ 17.0057(3), c ˆ 11.0834(2) ä, b ˆ 93.550 (1)8, Vˆ 2008.66(6) ä3.

Helvetica Chimica Acta Vol. 87 (2004)
2537
Bruker-Nonius Kappa-CCD diffractometer, MoKa radiation, l ˆ 0.7107, linear crystal dimensions ca. 0.20 Â
0.20 Â 0.20 mm. The structure was solved by direct methods (SIR97) and refined by full-matrix least-squares
analysis (SHELXL-97), using an isotropic extinction correction. All heavy atoms were refined anisotropically,
H-atoms isotropically, whereby H-positions are based on stereochemical considerations. Final R(F) ˆ 0.0348,
wR(F2) ˆ 0.0839for 300 parameters, 0 restraints, and 3649reflections with I > 2s(I) and 3.51 < q < 27.488.
Deposition No. CCDC-238289.
Compound (Æ)-12. Crystal data at 223(2) K for C24H19N3O5 (Mr 429.42): monoclinic, space group P2(1)/n,
Dc ˆ 1.423 g cm-3, Z ˆ 4, a ˆ 10.7504(4), b ˆ 16.9363(9), c ˆ 11.0123(5) ä, b ˆ 92.045 (2)8, Vˆ 2003.75(16) ä3.
Bruker-Nonius Kappa-CCD diffractometer, MoKa radiation, l ˆ 0.7107, linear crystal dimensions ca. 0.20 Â
0.20 Â 0.18 mm. The structure was solved by direct methods (SIR97) and refined by full-matrix least-squares
analysis (SHELXL-97). All heavy atoms were refined anisotropically, H-atoms isotropically, whereby H-
positions are based on stereochemical considerations. Final R(F) ˆ 0.0590, wR(F2) ˆ 0.1312 for 309parameters,
0 restraints, and 3562 reflections with I > 2s(I) and 1.17 < q < 28.708. Deposition No. CCDC-238290.
Compound (Æ)-14. Crystal data at 222 K for C31H27N3O5 (Mr 521.573): orthorhombic, space group P212121,
Dc ˆ 1.302 g cm-3, Z ˆ 8, a ˆ 13.6217(2), b ˆ 17.3424(2), c ˆ 22.5267(4) ä, Vˆ 5321.55(14) ä3. Bruker-Nonius
Kappa-CCD diffractometer, MoKa radiation, l ˆ 0.7107. The structure was solved by direct methods (SIR97)
and refined by full-matrix least-squares analysis (SHELXL-97), using an isotropic extinction correction. All
heavy atoms were refined anisotropically, H-atoms isotropically, whereby H-positions are based on stereo-
chemical considerations. Final R(F) ˆ 0.0590, wR(F2) ˆ 0.1455 for 707 parameters, and 9532 reflections with I >
2s(I) and 2.70 < q < 27.398. Deposition No. CCDC-238293.
Compound (‡)-26. Crystal data at 163(2) K for C26H29N2O4Br (Mr 513.42): orthorhombic, space group
P212121, Dc ˆ 1.496 g cmÀ3, Z ˆ 4, a ˆ 5.1744(1), b ˆ 18.7267(3), c ˆ 23.5221(5) ä, Vˆ 2279.28(7) ä3. Bruker-
Nonius Kappa-CCD diffractometer, MoKa radiation, l ˆ 0.7107, linear crystal dimensions ca. 0.25  0.23 Â
0.20 mm. The structure was solved by direct methods (SIR97) and refined by full-matrix least-squares analysis
(SHELXL-97), using an isotropic extinction correction. All heavy atoms were refined anisotropically, H-atoms
isotropically, whereby H-positions are based on stereochemical considerations. Final R(F) ˆ 0.0405, wR(F2) ˆ
0.0946 for 328 parameters and 4738 reflections with I > 2s(I) and 1.39 < q < 27.508. Deposition No. CCDC-
238292.
X-Ray Crystal Structure of the Complex of Thrombin with ( ‡ )-21. Crystals of human a-thrombin in
complex with (‡)-21 were prepared and analyzed as described in [13a], except that the crystal-to-detector
distance was 100 mm, and 678 frames were collected. Unit-cell dimensions were a ˆ 70.9ä, b ˆ 71.4 ä, c ˆ
72.4 ä, and b ˆ 100.48. For 306195 observations of 43087 reflections to 1.54-ä resolution, the merging R factor
on intensities was 3.9% and in the outermost data shell (1.54 ä 1.63 ä) was 21.6% with I/sI ˆ 27.0 overall and
8.56 in the outermost shell. Due to ice rings, which had to be eliminated, data completeness was not optimal,
being 82.2% overall and 83.9% in the outermost shell. Structure 1oyt.pdb was taken as starting model for the
non-inhibitor atoms, and the new structure was refined to final overall crystallographic R factors of 18.1
(working) and 20.9% (free), with values in the outer shell of 21.6 and 22.3% resp., for 2296 protein atoms, 35
‡
‡‡
inhibitor atoms, one Na ion, one Ca ion, and 408 H2O molecules. The inhibitor density is very clear, refined
temp. factors for the inhibitor range from 14 ä2 (amidinium moiety) to 22 ä2. The coordinates have been
deposited in the Protein Data Bank as 1vzq.pdb.
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Received June 3, 2004