Bioorganic & Medicinal Chemistry Letters 24 (2014) 5455–5459

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Discovery of potent iminoheterocycle BACE1 inhibitors John P. Caldwell ⇑, Robert D. Mazzola, James Durkin, Joseph Chen, Xia Chen, Leonard Favreau, Matthew Kennedy, Reshma Kuvelkar, Julie Lee, Nansie McHugh, Brian McKittrick, Peter Orth, Andrew Stamford, Corey Strickland, Johannes Voigt, Liyang Wang, Lili Zhang, Qi Zhang, Zhaoning Zhu Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USA

a r t i c l e

i n f o

Article history: Received 5 September 2014 Revised 29 September 2014 Accepted 2 October 2014 Available online 23 October 2014 Keywords: Alzheimer’s disease b-Secretase BACE Aspartyl protease inhibitors Structure-based drug design

a b s t r a c t The synthesis of a series of iminoheterocycles and their structure–activity relationships (SAR) as inhibitors of the aspartyl protease BACE1 will be detailed. An effort to access the S3 subsite directly from the S1 subsite initially yielded compounds with sub-micromolar potency. A subset of compounds from this effort unexpectedly occupied a different binding site and displayed excellent BACE1 affinities. Select compounds from this subset acutely lowered Ab40 levels upon subcutaneous and oral administration to rats. Ó 2014 Elsevier Ltd. All rights reserved.

A widely pursued strategy for the potential treatment of Alzheimer’s disease is inhibiting the production of neurotoxic b-amyloid (Ab) peptides, especially Ab40,42. The first step in generation of Ab involves cleavage of amyloid precursor protein (APP) by BACE1.1 Potent inhibitors of BACE1 should halt Ab production thereby reducing the neurotoxic Ab oligomers and plaques. Non-peptidic BACE1 inhibitors have been reported by our labs and others and offer the most promising avenue to the discovery of orally bioavailable, brain penetrant inhibitors.2,3 In earlier accounts, we reported on the rational design of compound 1 from a fragment screening hit.2 The amidine portion of the iminohydantoin core forms a critical hydrogen bond network with the catalytic diad (Asp32 and Asp228) of the BACE1 enzyme. Additionally, the initial design strategy focused on the optimization of four subsites (S20 , S1, S2, and S3) to afford a potent compound, Ki = 27 nM (see Fig. 1).4 Unfortunately, this compound had low plasma exposure in an oral rat pharmacokinetic assay. Consistent with a strategy to maximize the probability of identifying inhibitors with high CNS exposure, we sought to explore variants of 1 that would access S3 directly through S1, eliminating the need for occupation of S2 and inherently lowering the MW and c Log P.2d This work will detail the synthesis of these compounds and highlight the salient structure–activity relationships.

Key to construction of the proposed S1–S3 iminohydantoins was the use of the oxazolidinone intermediate 3 (Scheme 1).2d Elaboration of 3 through alkylation with the appropriate allyl bromide offered many opportunities to access S3 from S1. Scheme 2 shows the approach to S1 piperidine and pyrrolidine systems. For

O

NH N

HN H

N O

N NH

1 BAC 27 nM CE1 K i =27

⇑ Corresponding author. Tel.: +1 908 740 5199. E-mail address: [email protected] (J.P. Caldwell). http://dx.doi.org/10.1016/j.bmcl.2014.10.006 0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.

Figure 1. X-ray of 1 in the active site of BACE1.

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J. P. Caldwell et al. / Bioorg. Med. Chem. Lett. 24 (2014) 5455–5459 Br

Ph HO

O

NH2 a, b, c

O

H3CO2C

H 3CO2C

NHCbz

a, b

NCbz

NHCbz

c, d HO

O 12

2

14 (S)/(R):1/1

13

3

Scheme 1. Reagents and conditions: (a) 5% Rh/C with 50% H2O, 5% Pt/C, MeOH/ H2O/concd HCl aq, 60 psi H2, rt, 97%; (b) CbzOSu, dioxane/H2O, Et3N, rt, 95%; and (c) BF3–OEt2, PhCH(OMe)2, THF, 78 °C then rt, 70%.

NBoc

H 3CO2C e, f

(R)-14

3

TBSO

N

NH 2 g- j

1

NH

O

HO (R)-15

O

O Br

N CH3

f

g,h

4

k, l

(S)-14

BocN

N Boc 5

NBoc

H 3CO2C

NCbz

CO2CH 3 a-e

(R)-16

Ph

HO

3

N

NH 2

1

O

g, h, i

HO

6

(S)-16

(S)- 17

N

O

H 3CO2C i

NH2

H

NH

H

N

O

S

S j

NH

H

(R)-16

NBoc

BocN

(R)- 8

(S)-16

N n, o

NH

O

BocN (S)- 8

7

NBoc

N m

BocN

NH

O

H2N 19

18

O

N

S

NH

H HN

N

O k

S

NH

H

NH

NH

H

RN

Ph NCbz

Br

O

RN (R)-10

(R)-9

N

O l

+

O

20

26, 28-32, 34

steps N Bn 11

H RN 32 and 34

N

O

NH H

NH

p, b, j TBSO

O

21

H 3CO2C

CO2Me

N

HO

q, r, s HO

22

33 and 35

the piperidines, upon deprotonation of 3 with KHMDS and treatment with 5 (generated from 4 in a straightforward manner5,6), 1,10 -oxazolidinone 6 was produced with retention of stereochemistry in a manner consistent with literature precedent.7 Methanolysis and hydrogenation of 6 furnished the aminoester 7 as a diastereomeric mixture of Boc-protected piperidines. After subsequent cyclization to the corresponding thiohydantoins 8, the diastereomerically pure compounds were obtained by silica gel chromatography. The Boc-piperidines were individually deprotected, and the resulting piperidines 9 were elaborated through a variety of standard N-derivatization chemistries (reductive amination, amidation and sulfonylation). These thiohydantoins 10 were subjected to oxidative amination conditions to prepare the final iminohydantoin targets (representative sequence shown for the (R)-isomer in Scheme 2). Derivatives of both diastereomers of the S1-piperidine were explored. The S1-pyrrolidinyl analogs were

NH 2

k, l

RN

Scheme 2. Reagents and conditions: (a) 1-chloroethyl chloroformate, toluene, heat; (b) HCl aq, CH3OH, heat; (c) Boc2O, Et3N, CH2Cl2, rt, a–c: 48% three steps; (d) DIBALH, THF, 78 °C then rt, 73%; (e) PPh3, imidazole, Br2, 76%; (f) KHMDS, 3, THF, 78 °C, 60%; (g) LiOCH3, CH3OH, rt, 92%; (h) Pd(OH)2, H2 1 atm, MeOH, 99%; (i) MeNCS, DIEA, THF, rt, 80%; (j) TFA, CH2Cl2, rt, 97%; (k) N-derivatization; (l) NH4OH, CH3OH, t-BuO2H, rt, 35–60%.

H 3CO2C

NHCbz

NH

NH

NH

O

N O

23 NBoc N NH

t, u

O

NH NH

1

R HN

H 2N 24

Scheme 3. Reagents and conditions: (a) LiHMDS, 3, THF, 78 °C; (b) LiOCH3, CH3OH, rt, a–b: 80% two steps; (c) Pd/C, K2CO3, t-BuO2H, CH2Cl2, 25–60%; (d) NaBH4, CH3OH, 0 °C then rt, 96%; (e) TBSCl, imidazole, rt, 95%; (f) Pd(OH)2/C, H2, rt, 99%; (g) MeNCS, DIEA, THF, rt, 79%; (h) NH4OH, CH3OH, t-BuO2H, rt; (i) Boc2O; CH2Cl2, rt, h– i: 79% two steps; (j) TBAF, THF, rt; (k) [DiPFc-Rh(cod)]BF4, CH2Cl2, 55 psi H2, 77%; (l) 5% Rh/C, 5% Pd/C, 55 psi H2, AcOH, rt, 99%; (m) Dess–Martin reagent, CH2Cl2, rt, 93%; (n) MP-CNBH4, DMBNH2, cat. AcOH, DCE, rt, 96%; (o) 5% Pd(OH)2/C, HCO2NH4, CH3OH, rt, 93%; (p) LiHMDS, THF, 78 °C; (q) BocNHC(S)NHCH3, DIEA, THF, 82%; (r) DIAD, PPh3, 3-pyC(O)N3, THF, rt, 90%;11 (s) 5% Pd/C, H2 CH3OH, rt, 99%; (t) Nderivatization; and (u) TFA, CH2Cl2, rt, 95%.

prepared in an similar fashion starting with the benzyl-protected 2,5-dihydropyrrole ester 11 (Scheme 2, bottom section).8 In a concurrent effort to explore the S1-aminocyclohexane variants (Scheme 3), oxazolidone 3 was alkylated with 12, oxidized to the en-one,9 and then reduced to form a mixture of alcohol diastereomers 14. Since the position of the hydroxyl group as an allyl alcohol offered the opportunity to set the desired C1(R)-stereo-

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chemistry at the cyclohexyl stereocenter, we separated the alcohols to provide pure diastereoisomers 14. Hydrogenation of the TBS-protected derivative of alcohol (R)-14 proceeded cis relative to the bulky silyl protecting group, whereas hydroxyl-directed hydrogenation of (S)-14 provided (S)-17. As desired, both transformations provided the (R)-stereochemistry at the C1 cyclohexyl. Aminoesters (R)-15 and (S)-17 were separately elaborated to the Boc-protected iminohydantoins 16, and the alcohol moieties were then oxidized to give the corresponding cyclohexanone 18. To obtain the (R)-aminocyclohexane of 19, cyclohexanone 18 was treated with 2,4-dimethoxybenzylamine (DMBNH2) under reductive amination conditions. The resulting diastereomers were separated (product d.r. = (S)/(R):1/2), and the DMB group was deprotected under hydrogenation conditions.10 Amine (R)-19 was converted to the corresponding amides (43–45), ureas (46–53), and N-arylated targets (54–55). Once the correct diastereomer was validated through BACE1 Ki determination (vide infra), we developed an alternative streamlined synthesis (Scheme 3, bottom section). In this new route, oxazolidinone 20 was prepared from 2 and then alkylated with chiral allyl bromide 21, which contained the TBS-protected-(S)-hydroxyl such that the C3-cyclcohexyl stereocenter was installed far earlier in the synthesis. After methanolysis of the oxazolidinone and TBAF deprotection, cyclohexenol 22 was subjected to a one-pot sequential hydrogenation to (a) set the C1-cyclohexyl (R)-stereocenter through hydroxyl-directed hydrogenation, (b) deprotect the Nbenzyl carbamate, and (c) reduce the phenethyl to the cyclohexylethyl to produce 23. Aminoester 23 was elaborated to the Boc-protected iminohydantoin, treated with an azide transfer reagent with inversion of stereochemistry at C3, and reduced to the aminocyclohexane 24 now possessing the desired (R)-C3-stereochemistry. Amino derivatives of 24 were produced and Boc-deprotection afforded the final target compounds. Table 1 highlights the SAR in the S1-heterocycloalkyl series. In both the S1 piperidine and S1 pyrrolidine series, there is a preference in terms of BACE1 potency for the (R)-stereochemistry at the tertiary center (cf. 32 vs 33, 34 vs 35, 36 vs 37, and 38 vs 39). Additionally, amides appeared to be the preferred derivative when compared to substituted amines, ureas and sulfonamides (cf. 29,

Table 1 BACE1 affinities of S1-piperidine and pyrrolidine analogs12

(S)/(R)

O

N

30, and 31). When amides were explored further, the n-butyl amide and cyclopentylmethyl amide showed the best potencies with several examples showing sub-micromolar BACE1 Ki values. The S1-pyrolidine was slightly more potent than the S1-piperidine. Table 2 details the results in the S1-cyclohexylamine series. In general, potencies were significantly better in this series in comparison to the heterocycloalkyl series. Reductive amination with a variety of substrates led to compounds (40–42) with sub-micromolar potency. From X-ray crystallographic studies (Fig. 2),13 compounds from both series (e.g., 38 and 41) validated the hypothesis that we can improve potency by occupying S3 directly from S1. Furthermore, significant potency gains were achieved by decreasing the basicity of the exocyclic nitrogen through the amide (43–44) and urea (46–54) derivatives. The benzylamide 45 was five-fold less potent than the phenyl amide analog 43. Once again, X-ray crystallography provided valuable insight into these dramatic potency differences. Surprisingly, these compounds did not

Table 2 BACE1 affinities of S1-aminocyclohexyl analogs12 N O R HN

Compound

R1

BACE-1 Ki (nM)

Cat-D Ki (lM)

40 41 42 43 44 45 46 47 48 49 50 51 52 53 54

–CH2-c-propyl –c-Pentyl –c-Hexyl –C(O)Ph –C(O)-4-pyridyl –C(O)Bn –C(O)NHPh –C(O)NH-2-Cl-Ph –C(O)NH-4-Cl-Ph –C(O)NH-2-MeO-Ph –C(O)NH-4-MeO-Ph –C(O)NH-4-CNPh –C(O)NH-c-hexyl 2-Quinoline– 2-Quinoxaline–

480 430 230 130 39 585 16 301 10 276 10 3 138 22 14

53.3 >100 63.5 3.8 19.3 3.5 7.1 8.5 >100 2.4 2.5 6.9 6.0 25.0 7.5

NH

N

RN

O

n

N

25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 nt = not tested.

R Boc Boc H H n-(CH2)4CH3 –C(O)NH(CH2)2CH3 –SO2(CH2)3CH3 –C(O)(CH2)3CH3 –C(O)(CH2)3CH3 –C(O)CH2-c-pentyl –C(O)CH2-c-pentyl –C(O)(CH2)3CH3 –C(O)(CH2)3CH3 –C(O)CH2-c-pentyl –C(O)CH2-c-pentyl

n 2 2 2 2 2 2 2 2 2 2 2 1 1 1 1

NH

1

NH

Compound

NH

S/R S R S R R R R R S R S R S R S

BACE-1 Ki (lM)

Cat-D Ki(lM)

41 29 66 8.5 7.6 8.5 6.3 3.2 17 0.97 12.4 0.49 17.4 0.64 37

33 >100 >100 >100 >100 2.8 25.5 44.0 28.0 0.87 8.5 23.0 nt 6.0 nt

NH

N

NH

O

NH H NH

HN

O 38

41

Figure 2. X-ray of compound 38 (orange) and 41 (cyan) occupying the S3 subsite directly from S1.

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J. P. Caldwell et al. / Bioorg. Med. Chem. Lett. 24 (2014) 5455–5459 N NH

NH H

N O

N NH H

O

O

NH

O

N

H HN HN

HN H 44

46 NH H N O

NH

N HN H 53

Unfortunately, no cortical Ab40 reduction was observed when 53 was dosed to CRND8 mice.12 Although 53 was potent in the enzymatic assay (Ki = 22 nM) it suffered from a pronounced cell shift when tested in the whole cell assay, WCAb40 EC50 = 258 nM.12 Furthermore, when dosed with a known p-glycoprotein (Pgp) inhibitor, the cellular potency improved to 110 nM, the leftward shift suggesting that 53 was a Pgp substrate. In summary, employment of a modular synthetic strategy allowed us to access structurally distinct iminoheterocycles and enabled rapid SAR exploration of a series of small-molecule BACE1 inhibitors. Our understanding of the SAR was refined through Xray crystallographic studies, which showed that a subset of these inhibitors serendipitously occupied a different subsite of the BACE1 enzyme. Optimization studies exploiting this unique subsite led to the discovery of 53, a compound possessing modest exposure that lowered plasma Ab40 levels upon subcutaneous and oral dosing. Acknowledgments

Figure 3. X-ray of compounds 44 (cyan), 46 (orange), and 53 (yellow). Note that the substituents designed to reside in S3 instead occupy a different hydrophobic region making favorable hydrophobic interactions with the Ile171 and Lys168 residues as well as forming a hydrogen bond with the carbonyl of Phe169.

We would like to thank Dr. William Greenlee, Dr. Eric Parker, and Dr. John Hunter for their support and guidance of this work, Dr. Jesse Wong for preparation of intermediates, Ron Manning for genotyping and maintenance of the CRND8 mice, and Dr. Jared Cumming for suggestions and helpful discussions in the preparation of this Letter. Use of the IMCA-CAT beamline 17-ID at the Advanced Photon Source was supported by the companies of the Industrial Macromolecular Crystallography Association through a contract with the Center for Advanced Radiation Sources at the University of Chicago. References and notes

Table 3 In vivo evaluation of selected compounds Compound

AUC0–6h (nM-h) @ 10 mg/kg

Ab40 lowering plasma (30 mg/kg, sc) (%)

Ab40 lowering plasma (30 mg/kg, po)

44 51 53

0 0 306

63 65 65

Not determined Not determined 55%

occupy the S3 subsite; but, instead occupied a different subsite altogether (Fig. 3).14 This subsite is a narrow hydrophobic region which extends into a solvent-exposed region of the BACE1 enzyme.15 Compounds occupying this subsite make favorable hydrophobic interactions with the side chains of Ile171 and Lys168 as well as take advantage of a hydrogen bond interaction with the carbonyl of Phe169. To maximize the probability of identifying inhibitors with high CNS exposure, urea isosteres 53 and 54 were prepared to reduce the number of hydrogen bond donors present in the urea analogs. In Figure 3, superimposition of the X-ray cocrystal structure of inhibitors 44, 46, and 53 with BACE1 illustrates that the quinolinyl analog 50 binds in a similar manner as both the amide 44 and urea 46 analogs. Selectivities of 44, 46, and 53 for BACE1 versus the closely related aspartyl protease cathepsin D (Cat-D) were 494-, 546-, and 1111-fold, respectively. As a reference, the initial lead compound 1 was not selective versus cathepsin D (cathepsin D Ki = 31 nM). The amide 44 and urea 46 failed to achieve measurable plasma levels in rats when dosed orally, however both compounds exhibited modest levels of plasma Ab40 lowering when administered sub-cutaneously (s.c.) to rats at a dose of 30 mg/kg (Table 3). On the other hand, we were gratified when quinoline 53 displayed modest oral bioavailability in a rat pharmacokinetic study at a dose of 10 mg/kg, p.o.16 Furthermore, compound 53 not only exhibited Ab40 lowering when dosed s.c. but also upon oral administration.

1. Hardy, J.; Selkoe, D. J. Science 2002, 297, 353. 2. (a) Wang, Y.-S.; Strickland, C.; Voigt, J. H.; Kennedy, M. E.; Beyer, B. M.; Senior, M. M.; Smith, E. M.; Nechuta, T. L.; Madison, V. S.; Czarniecki, M.; McKittrick, B. A.; Stamford, A. W.; Parker, E. M.; Hunter, J. C.; Greenlee, W. J.; Wyss, D. F. J. Med. Chem. 2010, 53, 942; (b) Zhu, Z.; Sun, Z.-Y.; Ye, Y.; Voigt, J.; Strickland, C.; Smith, E. M.; Cumming, J.; Wang, L.; Wong, J.; Wang, Y.-S.; Wyss, D. F.; Chen, X.; Kuvelkar, R.; Kennedy, M. E.; Favreau, L.; Parker, E.; McKittrick, B. A.; Stamford, A. W.; Czarniecki, M.; Greenlee, W. J.; Hunter, J. C. J. Med. Chem. 2010, 53, 951; (c) Cumming, J. N.; Smith, E. M.; Wang, L.; Misiaszek, J.; Durkin, J.; Pan, J.; Iserloh, U.; Wu, Y.; Zhu, Z.; Strickland, C.; Voigt, J.; Chen, X.; Kennedy, M. E.; Kuvelkar, R.; Hyde, L. A.; Cox, K.; Favreau, L.; Czarniecki, M. F.; Greenlee, W. J.; McKittrick, B. A.; Parker, E. M.; Stamford, A. W. Bioorg. Med. Chem. Lett. 2012, 22, 2444; (d) Stamford, A. W.; Scott, J. D.; Li, S. W.; Babu, S.; Tadesse, D.; Hunter, R.; Wu, Y.; Misiaszek, J.; Cumming, J. N.; Gilbert, E. J.; Huang, C.; McKittrick, B. A.; Hong, J.; Guo, T.; Zhu, Z.; Strickland, C.; Orth, P.; Voigt, J. H.; Kennedy, M. E.; Chen, X.; Kuvelkar, R.; Hodgson, R.; Hyde, L. A.; Cox, K.; Favreau, L.; Parker, E. M.; Greenlee, W. J. ACS Med. Chem. Lett. 2012, 3, 897. 3. (a) Malamas, M. S.; Erdei; Edwards, P. D.; Albert, J. S.; Sylvester, M.; Aharony, D.; Andisik, D.; Callaghan, O.; Campbell, J. B.; Carr, R. A.; Chessari, G.; Congreve, M.; Frederickson, M.; Folmer, R. H. A.; Geschwindner, S.; Koether, G.; Kolmodin, K.; Krumrine, J.; Mauger, R. C.; Murray, C. W.; Olsson, L. L.; Patel, S.; Spear, N.; Tian, G. J. Med. Chem. 2007, 50, 5912; (b) Barrow, J. C.; Stauffer, S. R.; Rittle, K. E.; Ngo, P. L.; Yang, Z.; Selnick, H. G.; Graham, S. L.; Munshi, S.; McGaughey, G. B.; Holloway, M. K.; Simon, A. J.; Price, E. A.; Sankaranarayanan, S.; Colussi, D.; Tugusheva, K.; Lai, M. T.; Espeseth, A. S.; Xu, M.; Huang, Q.; Wolfe, A.; Pietrak, B.; Zuck, P.; Levorse, D. A.; Hazuda, D.; Vacca, J. P. J. Med. Chem. 2008, 51, 6259; (c) Malamas, M. S.; Erdei, J.; Gunawan, I.; Turner, J.; Hu, Y.; Wagner, E.; Fan, K.; Chopra, R.; Olland, A.; Bard, J.; Jacobsen, S.; Magolda, R. L.; Pangalos, M.; Robichaud, A. J. J. Med. Chem. 2010, 53, 1146; (d) Rueeger, H.; Rondeau, J. M.; McCarthy, C.; Moebitz, H.; Tintelnot-Blomley, M.; Neumann, U.; Desrayaud, S. Bioorg. Med. Chem. Lett. 2011, 21, 1942; (e) Probst, G.; Xu, Y.-Z. Expert Opin. Ther. Patents 2012, 22, 511. 4. Coordinates for the X-ray structure of compound 1 complexed with BACE1 have been deposited in the Protein Data Bank (www.rcsb.org), and can be accessed under PDB 3L5E. 5. Coldham, I.; Crapnell, K. M.; Fernandez, J.-C.; Moseley, J. D.; Rabot, R. J. Org. Chem. 2002, 67, 6185. 6. Winkler, J. D.; Axten, J.; Hammach, A. H.; Kwak, Y.-S.; Lengweiler, U.; Lucero, M. J.; Houk, K. N. Tetrahedron 1998, 54, 7045. 7. (a) Karady, S.; Amato, J. S.; Weinstock, L. M. Tetrahedron Lett. 1984, 25, 4373; (b) Jones, J. B.; Keitz, P.; Cheng, H. J. Org. Chem. 1994, 59, 7671; (c) Eriksson, M.; Napolitano, E.; Xu, J.; Kapadia, S.; Byrne, D.; Nummy, L.; Grinberg, N.; Shen, S.; Lee, H.; Farina, V. Chimia 2006, 60, 566.

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Terao, Y.; Kotaki, H. N.; Achiwa, K. Chem. Pharm. Bull. 1985, 33, 2766. Yu, J.-Q.; Corey, E. J. Org. Lett. 2002, 4, 2727. Gartiser, T.; Selve, C.; Delpuech, J.-J. Tetrahedron Lett. 1983, 24, 1609. Papeo, G.; Posteri, H.; Vianello, P.; Varasi, M. Synthesis 2004, 17, 2886. Inhibitor Ki values at purified human BACE1 were determined using a FRETpeptide substrate hydrolysis assay. Cellular IC50 values for reduction of Ab40 production were determined in stably transfected HEK293-APPswe/lon cells. The protocols for these assays have been previously described in Ref. 2b (Zhu et al.). All values reported are the average of a minimum of two independent determinations. CRND8-APP mice are models for early onset (familial) AD that express human APP containing both Swedish and London mutations that enhance the rate of APP cleavage by BACE1 and favor production of Ab42 over Ab40 in the c-secretase cleavage step as described by Hyde, L. A.; Kazdoba, T.

13.

14.

15.

16.

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M.; Grilli, M.; Lozza, G.; Brussa, R.; Zhang, Q.; Wong, G. T.; McCool, M. F.; Zhang, L.; Parker, E. M.; Higgins, G. A. Behav. Brain Res. 2005, 160, 344. Coordinates for the X-ray structure of compounds 38 and 41 complexed with BACE1 have been deposited in the Protein Data Bank (www.rcsb.org), and can be accessed under PDB 4R8Y and PDB 4R9, respectively. Coordinates for the X-ray structure of compounds 44, 46, and 54 complexed with BACE1 have been deposited in the Protein Data Bank (www.rcsb.org), and can be accessed under PDB 4R92, PDB 4R93 and PDB 4R95, respectively. This subsite was initially reported in a previous account (see Ref. 2b) and termed the ‘A-site’ as this subsite was occupied by ‘Mode A inhibitors’. Serendipitously, we were able to occupy this subsite using ‘Mode B’ inhibitors. Korfmacher, W. A.; Cox, K. A.; Ng, K. J.; Veals, J.; Hsien, Y.; Wainhaus, S.; Broske, L.; Prelusky, D.; Nomeir, A.; White, R. E. Rapid Commun. Mass. Spectrom. 2001, 15, 335.