Discovery of a novel 3,4-dimethylcinnoline carboxamide M4 positive allosteric modulator (PAM) chemotype via scaffold hopping

Bioorganic & Medicinal Chemistry Letters 29 (2019) 126678

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Discovery of a novel 3,4-dimethylcinnoline carboxamide M4 positive allosteric modulator (PAM) chemotype via scaffold hopping

T

Kayla J. Templea,b, Julie L. Engersa,b, Madeline F. Longa,b, Alison R. Gregroa,b, Katherine J. Watsona,b, Sichen Changa,b, Matthew T. Jenkinsa,b, Vincent B. Luscombea,b, Alice L. Rodrigueza,b, Colleen M. Niswendera,b,d,e, Thomas M. Bridgesa,b, P. Jeffrey Conna,b,d,e, ⁎ ⁎ Darren W. Engersa,b, , Craig W. Lindsleya,b,c,e, a

Vanderbilt Center for Neuroscience Drug Discovery, Vanderbilt University, Nashville, TN 37232, USA Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA Department of Chemistry, Vanderbilt University, Nashville, TN 37232, USA d Vanderbilt Kennedy Center, Vanderbilt University School of Medicine, Nashville, TN 37232, USA e Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37232, USA b c

ARTICLE INFO

ABSTRACT

Keywords: M4 Muscarinic acetylcholine receptor Positive allosteric modulator (PAM) Structure activity relationship (SAR)

This Letter details our efforts to replace the 2,4-dimethylquinoline carboxamide core of our previous M4 PAM series, which suffered from high predicted hepatic clearance and protein binding. A scaffold hopping exercise identified a novel 3,4-dimethylcinnoline carboxamide core that provided good M4 PAM activity and improved clearance and protein binding profiles.

Muscarinic acetylcholine receptor subtype 4 (M4) positive allosteric modulators (PAMS) have recently emerged as important drug targets as novel treatments for various neurological disorders such as Parkinson’s disease,1 Huntington’s disease,2 and schizophrenia (both the positive and negative symptom clusters).3–6 Many of the classical M4 PAMs possess a β-amino carboxamide moiety as a key pharmacophore (Figure 1).7–13 While this moiety is essential for M4 PAM activity, such chemotypes have been plagued with poor solubility, varying degrees of Pgp efflux, and potency differences across species. Previously, we reported the development of a new M4 PAM chemotype, a 2,4-dimethylquinoline carboxamide core (5) that was devoid of the classical β-amino carboxamide moiety and still afforded potent and CNS penetrant M4 PAMs.14 Utilizing a similar scaffold hopping approach, we envisioned hybridizing the 3,4-dimethylthienol[2,3-c]pyridazine core of 3 and 4 with our β-amino carboxamide lacking 2,4-dimethylquinoline carboxamide core (Figure 2). This strategy led to the discovery of a novel M4 PAM chemotype containing a 3,4-dimethylcinnoline carboxamide core, 6. Based on our previous findings, the optimal amide moieties in relation to analogs 5 are N-heteroaryl azetidine amides which gave the most potent analogs.15 To begin our endeavor, we first generated a library of N-heteroaryl 3-aminoazetidines 9 and 11 (Scheme 1). The

⁎

synthesis of this library began with nucleophilic aromatic substitutions of commercially available pyridines, pyrimidines, and pyrazines with Boc-protected 3-aminoazetidine, 7. N-Boc-deprotection with TFA then afforded N-heteroaryl 3-aminoazetidines 9 in moderate to good yields. For the aryl- and heteroaryl halides 10 that could not readily undergo nucleophilic substitution, Buchwald-Hartwig amination was employed. With intermediates 9 and 11 in hand, we could readily generate desired analogs 13 by employing HATU coupling with commercially available carboxylic acid 12 (See Scheme 2). Select analogs 13 were screened against human M4 (hM4) to determine potency with results highlighted in Table 1. This exercise resulted in analogs with M4 PAM potencies that rivaled our previous series 5, with nearly a 5-fold increase in rat potency (13o; rat EC50 = 82 nM); unfortunately, 13o displayed low human functional potency (hEC50 = 648 nM). In general, pyrimidines (13a and c) were less potent than pyrazine (13d) which were less potent that functionalized benzene and pyridines (13e-s). This trend was also observed in our previous 2,4-dimethylquinoline series. Generally, the most potent analogs contained at least one electron withdrawing group on the heteroaryl ring (13 m-s). Moreover, several of the N-pyridyl azetidine amides possessed M4 PAM functional potencies < 500 nM in both human and rat: 13p (hEC50 = 464 nM; rEC50 = 174 nM), 13q

Corresponding authors. E-mail addresses: [email protected] (D.W. Engers), [email protected] (C.W. Lindsley).

https://doi.org/10.1016/j.bmcl.2019.126678 Received 19 August 2019; Received in revised form 29 August 2019; Accepted 8 September 2019 Available online 10 September 2019 0960-894X/ © 2019 Elsevier Ltd. All rights reserved.

Bioorganic & Medicinal Chemistry Letters 29 (2019) 126678

K.J. Temple, et al.

Table 1 Structures and activities for analog 13.

Cpd

Figure 1. Structures of representative M4 PAMs possessing the classical βamino carboxamide pharmacophore (circled in red) in either a 3-amino-4,6dimethylthieno[2,3-b]pyridine core (2) or a 5-amino-3,4-dimethylthieno[2,3c]pyridazine core (3 & 4). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Figure 2. Hybridizing the β-amino carboxamide-containing M4 PAMs 3 & 4 with the potent β-amino carboxamide-lacking 2,4-dimethylquinoline core 5 to generate a novel M4 PAM scaffold, 6.

Scheme 1. Synthesis of N-aryl azetidine amides. Reagents and conditions: (a) DIEA or Cs2CO3, NMP, 100 °C, 4–18 h; (b) TFA, DCM, 1–3 h, 43–88%; (c) Pd2dba3, rac-BINAP, Cs2CO3, benzene, 110 °C, 2–18 h; (d) TFA, DCM, 1–3 h, 24–71%.

Het

hM4 EC50 (nM)a [% ACh Max]

13a

> 10,000 [77]

13b

3363 [64]

13c

3204 [88]

13d

2509 [80]

13e

1269 [73]

13f

1031 [81]

13g

1388 [77]

13h

2631 [76]

13i

1460 [81]

13j

1320 [77]

13k

1216 [83]

13l

952 [84]

13m

668 [84]

13n

737 [82]

13o

648 [82]

13p

464 [80]

13q

373 [74]

13r

304 [72]

13s

116 [92]

a Calcium mobilization assays with hM4/Gqi5-CHO cells performed in the presence of an EC20 fixed concentration of acetylcholine, n = 1 experiment performed in triplicate.16

rat plasma:brain level (PBL) IV cassette paradigm. In regard to physicochemical properties, all five analogs possess molecular weights less than or nearly equal to 400 Da, with two having attractive CNS xLogPs (2.26 and 2.38). When compared to our previous 2,4-dimethylquinoline series, which displayed high predicted hepatic clearance in both human and rat assays based on microsomal CLint, the new 3,4-dimethylcinnoline series was a slight improvement displaying moderate predicted hepatic clearance in both human (CLheps of 9.3 – 13 mL/min/kg) and rat (CLheps of 23 – 37 mL/min/kg). In comparison, the previous 2,4-

Scheme 2. Synthesis of M4 PAM analogs 12. Reagents and conditions: (a) HATU, DIEA, DMF, 20 min, 23–90%.

(hEC50 = 373 nM; rEC50 = 356 nM), 13r (hEC50 = 304 nM; rEC50 = 138 nM), and 13s (hEC50 = 116 nM; rEC50 = 230 nM). Of these compounds, 13n, 13p, 13q, 13r, 13o, and 13s were advanced into a battery of in vitro DMPK assays (Table 2) and our standard 2

Bioorganic & Medicinal Chemistry Letters 29 (2019) 126678

K.J. Temple, et al.

Table 2 In vitro DMPK and rat PBL data for select analogs 13. Property

13n VU6015178

MW 368.4 xLogP 2.26 TPSA 58.1 In vitro PK parameters CLINT (mL/min/kg), rat 50 CLHEP (mL/min/kg), rat 29 CLINT (mL/min/kg), human 26 12 CLHEP (mL/min/kg), human 0.002 Rat fu,plasma Human fu,plasma 0.051 Rat fu,brain 0.022 Brain Distribution (0.25 h) (SD Rat; 0.2 mg/kg IV) Kp, brain:plasma < 0.05 Kp,uu, brain:plasma < 0.56

13o VU6015198

13p VU6015187

13q VU6015180

13r VU6015190

13s VU6015191

385.8 1.87 71

385.8 1.87 71

397.9 1.73 80.2

392.8 1.49 94.8

402.3 2.38 71

54 30 12 7.5 0.026 0.032 0.009

79 37 17 9.4 0.015 0.048 0.036

34 23 17 9.3 0.012 0.038 0.023

63 33 17 9.3 0.008 0.030 0.042

60 32 32 13 0.017 0.060 0.036

0.03 0.09

0.12 0.29

0.09 0.02

0.04 0.21

0.03 0.06

dimethylquinoline series was highly bound to both human and rat plasma proteins (fus ~ 0.005) whereas four of the six analogs analyzed in the new 3,4-dimethylcinnoline series (13o, 13p, 13q, 13s) possessed reduced rat plasma protein binding fus (fu,s = 0.012 – 0.026). Additionally, all six compounds analyzed (13n, 13o, 13p, 13q, 13r, and 13s) displayed reduced human plasma protein binding fus (fu,s = 0.030 – 0.060) as well as rat brain binding(fu,s = 0.022–0.042). Unfortunately, five of six compounds (13n, 13o, 13q, 13r, and 13 s) tested proved to have limited CNS penetration while 13p was only modest (brain:plasma Kp ~ 0.12, Kp,uu ~ 0.29). The lower Kp is likely due to the higher dielectric constant and dipole of the 3,4-dimethylcinnoline core versus the analogous quinolone core. In summary, a scaffold hopping exercise based on M4 PAMs 3 and 5 identified a novel 3,4-dimethylcinnoline carboxamide core, 6, that exhibited a 3-fold improvement in rat M4 PAM activity (5 vs. 13o), lower human and rat hepatic clearance, and improved fus. Although this endeavor did not produce PAMs with the desired DMPK profiles to advance as potential development candidates, it did provide us with insights and leads to further our goals, which will be disclosed in due course.

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Acknowledgments We thank the NIH for funding via the NIH Roadmap Initiative 1x01 MH077607 (C.M.N.), the Molecular Libraries Probe Center Network (U54MH084659 to C.W.L.) and U01MH087965 (Vanderbilt NCDDG). We also thank William K. Warren, Jr. and the William K. Warren Foundation who funded the William K. Warren, Jr. Chain in Medicine (to C.W.L.). References 1. Shen W, Plotkin JL, Francardo V, et al. M4 Muscarinic receptor signaling ameliorates striatal plasticity deficits in models of L-DOPA-induced dyskinesia. Neuron. 2015;88:762–773. 2. Pancani T, Foster DJ, Moehle MS, et al. Allosteric activation of M4 muscarinic receptors improve behavioral and physiological alteration in early symptomatic YAC128 mice. Proc Natl Acad Sci USA. 2015;112:14078–14083. 3. Bridges TM, LeBois EP, Hopkins CR, et al. The antipsychotic potential of muscarinic allosteric modulation. Drug News Perspect. 2010;23:229–240. 4. Foster DJ, Wilson JM, Wess J, et al. Antipsychotic-like effects of M4 positive allosteric modulators are mediated by CB2 receptor-dependent inhibition of dopamine release. Neuron. 2016;91:1244–1252. 5. Jones CK, Byun N, Bubser M. Muscarinic and nicotinic acetylcholine receptor agonists and allosteric modulators for the treatment of Schizophrenia. Neuropsychopharmacology. 2012;37:16–42. 6. Farrell M, Roth BL. Allosteric antipsychotics: M4 Muscarinic potentiators as novel

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