Synthesis of Pyridazine-Based α-Helix Mimetics

Article abstract of DOI:10.1021/acscombsci.6b00111

A versatile synthesis of pyridazine-based small molecule α-helix mimetics (A) is presented. Modular C-C, C-N, and C-O bond-forming reactions allow for the inclusion of a variety of aliphatic, basic, aromatic, and heteroaromatic side chain moieties. This robust synthesis is suitable for the preparation of small pyridazine-based libraries.

Full text of DOI:10.1021/acscombsci.6b00111

Research Article
pubs.acs.org/acscombsci
Synthesis of Pyridazine-Based α‑Helix Mimetics
Allyn T. Londregan, David W. Piotrowski, and Liuqing Wei*
Pfizer Medicine Design, Eastern Point Road, Groton, Connecticut 06340, United States
S
* Supporting Information
ABSTRACT: A versatile synthesis of pyridazine-based small molecule α-helix mimetics (A) is
presented. Modular C−C, C−N, and C−O bond-forming reactions allow for the inclusion of a
variety of aliphatic, basic, aromatic, and heteroaromatic side chain moieties. This robust syn-
thesis is suitable for the preparation of small pyridazine-based libraries.
KEYWORDS: pyridazine, α-helix mimetic, Minisci reaction, cross coupling
ics that have been designed to bind to multiple recognition
surfaces have emerged.4e While multifaced helix mimetics are not
directly addressed in this work, it is clear that the helix mimetic
field could benefit from chemistry that is capable of introducing
both polar and nonpolar substituents to various scaffolds.
Pyridazine-based mimetics offer the same conformational con-
straint and hydrophobic region as the related terpyridyl analogs
(1)4a but also provide a hydrophilic face, rich in hydrogen bond
donors and acceptors that can be directed toward the solvent.
These mimetics offer several vectors to explore structure−
activity relationships, but to date, the range of substituents at
each of the pertinent vectors has been largely limited to side
chains represented in naturally occurring amino acids. Expansion
of substituents on helix mimetics to include non-natural side
chains could be beneficial for discovery of new drug-like mole-
cules. As shown in compound 2,4c these vectors include (1) the
6-position amide (magenta), which can be modified via sim-
ple amidation chemistry from the corresponding acid, (2) the
3-position oxazole (red), which can be substituted with a variety
of C, O, or N linked substituents using SNAr or Suzuki reactions,
and (3) the 4-position (blue) R group, which can be installed
by either alkyl radical-based methods or lengthy de novo syn-
thetic sequences. Since methods to address objectives 1 and 2 are
well-established in the literature, the main focus of this work
(objective 3) was to explore the addition of a wider variety of
pyridazine 4-position substituents to mimic a range of amino acid
side chains at the i + 3/i + 4 position of the helix mimetic.
INTRODUCTION
■
The α-helix is one of the most common secondary protein
structural motifs with roughly 3.6 amino acid residues per turn.
Amino acid side chains that are adjacent in space, at positions
i, i + 3/i + 4, i + 7 of the helix, appear on the same face and define
three distinct recognition elements. The key residues are often
hydrophobic in nature and are frequently involved in crucial
protein protein interactions (PPI). The α-helix conformation is
stabilized by a combination of steric interactions and hydrogen
bonding. Approximately 62% of the PPIs found in the Protein
Data Bank have an α-helix at the interface which makes this struc-
tural motif important in protein−protein recognition.1 Several
relevant biological targets utilize α-helices as key recognition
elements in PPIs.2 Because of the nature of PPIs, it is probable
that chemical matter targeting these interfaces can lie in beyond
rule of five space.3
Several variants of nonpeptidic α-helix mimetics based on
terphenyl, terephthalamide, oligopyridine and pyridazine scaf-
folds are known and are believed to display side chains in a
manner that closely resembles the positions i, i + 3/i + 4, and i + 7
of an α-helix (Figure 1).4 Some of these have been constructed
RESULTS AND DISCUSSION
■
Retrosynthesis. Multiple routes to α-helix mimetics based
on compound A were devised starting from two readily avail-
able intermediates 3 and 4 (Scheme 1). Substitution of
3-chloropyridazines, such as 3, via Suzuki coupling or SNAr are
well-established methods for diversification at this position and
Figure 1. Representative examples of i, i + 3/i + 4, i + 7 α-helix mimetics.
in a modular fashion allowing for the efficient synthesis of
moderate-to-large libraries.4j−m Small molecule mimetics are
expected to have fewer pharmacokinetic issues (e.g., perme-
ability) than the corresponding peptides,5,6 and thus could allow
for discovery of chemical matter well-suited to small molecule
drug discovery projects. More recently, amphiphilic helix mimet-
Received: July 25, 2016
Revised: August 24, 2016
© XXXX American Chemical Society
A
DOI: 10.1021/acscombsci.6b00111
ACS Comb. Sci. XXXX, XXX, XXX−XXX

ACS Combinatorial Science
Research Article
a
Scheme 1. Retrosynthesis
Table 1. Selected Analogues 8, 9, and 10
a
Scheme 2. Expansion of C3 through Suzuki or SNAr Reactions
a
Isolated yield of purified product from two-step reaction sequence
reported in parentheses.
between 3-chloro-6-carboxypyridazine ethyl ester 35 and vari-
ous carbon radical sources via the Minisci reaction.7 Route II
employed 6-chloropyridazin-3-amine 4 as the starting material,
which was subsequently brominated to afford 5 which allowed
for 4-position substitution beyond carbon to include nitrogen
and oxygen linkers.
Synthesis. Our major building block, pyridazine 3, was
alkylated through Minisci coupling with a number of primary
and secondary aliphatic acids. Unlike Negishi coupling with
branched zincates, the Minisci reaction with branched acids,
such as isobutyric acid, did not show any isomerization. In this
well-known and versatile reaction, the purported mechanism
involves a silver-catalyzed radical formation, via oxidative
decarboxylation, and subsequent addition to the protonated
pyridazine to generate a regioisomeric mixture of 4- and 5-alkyl-
pyridazines.8,9 Thus, the homolytic alkylation with 2-phenyl-
acetic acid led to two regioisomers (Scheme 2). The major
regiosiomer 6a, which was readily separated from the minor
isomer 6b by chromatography, was hydrolyzed with aq. NaOH
and coupled with N-Boc-piperazine.10 This first versatile
synthetic intermediate 7 represented an i + 3/i + 4, i + 7 frag-
ment with a Phe side-chain at i + 3/i + 4 that allowed for explo-
ration of the i position. The common intermediate 7 was applied
a
Reagents and conditions: (a) phenylacetic acid, (NH4)2S2O8,
AgNO3, conc. H2SO4, H2O, 70 °C, 37%; (b) (i) NaOH, MeOH,
30 °C, 100%, (ii) N-Boc-piperidine, HATU, DIPEA, DMF, 30 °C,
58%; (c) (i) ArB(OR)2, Pd(dtbpf)Cl2, K3PO4, dioxane, H2O, 140 °C,
(ii) HCl, MeOH, 30 °C; (d) (i) ROH, Cs2CO3, THF, 100 °C, (ii) TFA,
CH2Cl2, 30 °C; (e) (i) amine, DIPEA, tAmOH, 110 °C, (ii) TFA,
CH2Cl2, 30 °C.
were ultimately utilized. First, however, we aimed to diversify the
4-position of the pyridazine core with a variety of side chains
including aliphatic, basic, aromatic, and heteroaromatic residues.
As shown in Scheme 1, two general retrosynthetic approaches
were utilized. Route I initiated with a C−C bond formation
a
Scheme 3. Expansion of C4 through Minisci Reaction Followed by Acylation
a
Reagents and conditions: (a) (NH4)2S2O8, AgNO3, TFA, H2O, 70 °C, 30 min, 35%; (b) (i) 2-benzylpyrrolidine, DIPEA, NMP, 120 °C, 16 h, 94%,
(ii) NaOH, MeOH, 97%, (iii) N-Boc-piperidine, HATU, DIPEA, DMF, 16 h, 74%, (iv) Pd/C, H2, MeOH, 90%; (c) (i) acid, HATU, DIPEA, DMF,
16 h, (ii) TFA, CH2Cl2.
B
DOI: 10.1021/acscombsci.6b00111
ACS Comb. Sci. XXXX, XXX, XXX−XXX

ACS Combinatorial Science
Research Article
a
to three libraries in a 2-step sequence (Table 1): (1) Suzuki
coupling mediated arylation to give 8, after acidic deprotec-
tion, (2) SNAr mediated etherification-deprotection to provide
aryl ethers 9, and (3) amination-deprotection to generate aryl
amines 10.11 Among amine substituents on compounds 10,
2-benzylpyrrolidine and N-methylbenzylamine were favored for
further exploration of the SAR (data not shown).
Table 2. Selected Analogues 13
Using the same strategy, the Cbz-protected aminomethyl
group was introduced to the C4 position by reaction of ethyl
ester 3 with N-carbobenzyloxyglycine to provide 11 in 35%
yield (Scheme 3). After SNAr reaction with 2-benzylpyrroli-
dine, followed by ester hydrolysis, amide formation, and
Cbz-deprotection, 11 was converted into intermediate 12. The
C4 aminomethyl group of 12 was acylated followed by depro-
tection through parallel chemistry to provide 13 (Table 2).
In the second route, 6-chloropyridazin-3-amine 4 was
employed as the starting material (Scheme 4). Substrate 4
allowed for sequential change of all three vectors on the
pyridazine. Selective bromination of 4 gave 5 (R′, R″ = H),12
which was sequentially alkylated with benzyl bromide and
methyl iodide to form intermediate 14. Pyridazine 14, with dif-
ferentiated functional groups, allowed for selective manipulation
of each position. The bromo on pyridazine 14 was used as the
handle to produce three different libraries in a 3-step sequence:
(1) etherification-amino carbonylation-deprotection to give 15,
(2) Suzuki coupling-amino carbonylation-deprotection to
provide 16, and (3) SNAr-amino carbonylation-deprotection to
offer 17 (Table 3).
a
Isolated yield of purified product from two-step reaction sequence
reported in parentheses.
Scheme 4. Expansion of C4 through C−C Bond Cross
Coupling, SNAr Reaction, Followed by C6 Amino
Carbonylation
a
a
Table 3. Selected Analogues 15, 16, and 17
a
a
Reagents and conditions: (a) Br2, NaHCO3, MeOH, 57%; (b) (i)
Isolated yield of purified product from three-step reaction sequence
reported in parentheses.
benzyl bromide, NaH, THF, 70%, (ii) iodomethane, NaH, THF,
65%; (c) (i) ROH, NaH, DMF, 30 °C, (ii) 1-Boc-piperazine, Pd(OAc)2,
dppp, K3PO4, Et3N, DMF, CO (1 atm), 110 °C, (iii) TFA, CH2Cl2; (d)
(i) ArB(OR)2, Pd(dppf)Cl2·CH2Cl2, Cs2CO3, dioxane, H2O, 100 °C,
(ii) 1-Boc-piperazine, Pd(OAc)2, dppp, K3PO4, Et3N, DMF, CO
(1 atm), 110 °C, (iii) TFA, CH2Cl2; (e) amine, CsF, DIPEA, DMSO,
110 °C, (ii) 1-Boc-piperazine, Pd(OAc)2, dppp, K3PO4, Et3N, DMF,
CO (1 atm), 110 °C, (iii) TFA, CH2Cl2.
A comparison of the crystal structure of Konig’s 1,4-dipipera-
̈
zino benzene mimetic4d to 10d showed a similar display of the
substituents in space. Likewise, superposition of a minimized
version of mimetic 10b had a similar geometrical arrangement
(see Supporting Information).
C
DOI: 10.1021/acscombsci.6b00111
ACS Comb. Sci. XXXX, XXX, XXX−XXX

ACS Combinatorial Science
Research Article
(b) Tosovska, P.; Arora, P. S. Oligooxopiperazines as Nonpeptidic
α-Helix Mimetics. Org. Lett. 2010, 12, 1588−1591. (c) Biros, S. M.;
Moisan, L.; Mann, E.; Carella, A.; Zhai, D.; Reed, J. C.; Rebek, J.
Heterocyclic α-Helix Mimetics for Targeting Protein−Protein Inter-
actions. Bioorg. Med. Chem. Lett. 2007, 17, 4641−4645. (d) Maity, P.;
In summary, pyridazine-based α-helix mimetics display side
chains in a manner that closely resembles the i, i + 3/i + 4, and
i + 7 positions of peptide-derived α-helices. Starting from a few
readily accessible versatile synthetic intermediates, we developed
several synthetic sequences that allow for inclusion of a variety of
side chains including aliphatic, basic, aromatic, and heteroaromatic
residues through a number of C−C, C−N, and C−O bond-
forming reactions. The robust synthesis enabled the explora-
tion of the three accessible vectors and was demonstrated by
preparation of several small libraries. The application of this
methodology to relevant medicinal chemistry targets will be
reported in due course.
Konig, B. Synthesis and Structure of 1,4-Dipiperazino Benzenes: Chiral
̈
Terphenyl-type Peptide Helix Mimetics. Org. Lett. 2008, 10, 1473−
1476. (e) Jayatunga, M. K. P.; Thompson, S.; Hamilton, A. D. α-Helix
Mimetics: Outwards and Upwards. Bioorg. Med. Chem. Lett. 2014, 24,
717−724. (f) Moon, H.; Lee, W. S.; Oh, M.; Lee, H.; Lee, J. H.; Im, W.;
Lim, H.-S. Design, Solid-Phase Synthesis, and Evaluation of a Phenyl-
Piperazine-Triazine Scaffold as α-Helix Mimetics. ACS Comb. Sci. 2014,
16, 695−701. (g) Lee, J. H.; Oh, M.; Kim, H. S.; Lee, H.; Im, W.; Lim,
H.-S. Converting One-Face α-Helix Mimetics into Amphiphilic α-Helix
Mimetics as Potent Inhibitors of Protein−Protein Interactions. ACS
Comb. Sci. 2016, 18, 36−42. (h) Drennen, B.; Scheenstra, J. A.; Yap, J. L.;
Chen, L.; Lanning, M. E.; Roth, B. M.; Wilder, P. T.; Fletcher, S.
Structural Re-engineering of the α-Helix Mimetic JY-1−106 into Small
Molecules: Disruption of the Mcl-1−Bak-BH3 Protein−Protein
Interaction with 2,6-Di-Substituted Nicotinates. ChemMedChem 2016,
11, 827−833. (i) Davis, J. M.; Truong, A.; Hamilton, A. D. Synthesis of a
2,3′;6′,3″-Terpyridine Scaffold as an α-Helix Mimetic. Org. Lett. 2005, 7,
5405−5408. (j) Shaginian, A.; Whitby, L. R.; Hong, S.; Hwang, I.;
Farooqi, B.; Searcey, M.; Chen, J.; Vogt, P. K.; Boger, D. L. Design,
Synthesis, and Evaluation of an α-Helix Mimetic Library Targeting
Protein-Protein Interactions. J. Am. Chem. Soc. 2009, 131, 5564−5572.
(k) Peters, M.; Trobe, M.; Tan, H.; Kleineweischede, R.; Breinbauer, R.
A Modular Synthesis of Teraryl-Based α-Helix Mimetics, Part 1:
Synthesis of Core Fragments with Two Electronally Differentiated
Leaving Groups. Chem. - Eur. J. 2013, 19, 2442−2449. (l) Peters, M.;
Trobe, M.; Breinbauer, R. A Modular Synthesis of Teraryl-Based
α-Helix Mimetics, Part 2: Synthesis of 5-Pyridine Boronic Acid Pinacol
Ester Building Blocks with Amino Acid Side Chains in 3-Position. Chem.
- Eur. J. 2013, 19, 2450−2456. (m) Trobe, M.; Peters, M.; Grimm, S. H.;
Breinbauer, R. The Development of a Modular Synthesis of Teraryl-
Based α-Helix Mimetics as Potential Inhibitors of Protein−Protein
Interactions. Synlett 2014, 25, 1202−1214.
ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge on the ACS
Publications website at DOI: 10.1021/acscombsci.6b00111.
■
S
Experimental procedures, characterization data and X-ray
data for the structure determination of 10d (PDF)
Crystallographic information file for compound 10d
(CIF)
AUTHOR INFORMATION
Corresponding Author
*E-mail: liuqing.wei@pfizer.com.
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
The authors thank Steve B. Coffey, Chris Limberakis, and
Benjamin N. Rocke for their chemistry contributions and Justin
Stroh for HRMS. The authors thank Brian Samas for X-ray
structure determination of 10d (CCDC 1485770).
(5) Gellman, S. H. Foldamers: A Manifesto. Acc. Chem. Res. 1998, 31,
173−180.
ABBREVIATIONS
PPI protein−protein interaction
(6) (a) Voisin-Chiret, A. S.; Burzicki, G.; Perato, S.; De Giorgi, M.;
Franchini, C.; Sopkova-de Oliveira Santos, J.; Rault, S. Aromatic
Garlands, as New Foldamers, to Mimic Protein Secondary Structure.
Tetrahedron 2012, 68, 4381−4389. (b) Lee, J. H.; Zhang, Q.; Jo, S.; Chai,
S. C.; Oh, M.; Im, W.; Lu, H.; Lim, H.-S. Novel Pyrrolopyrimidine-
Based α-Helix Mimetics: Cell-Permeable Inhibitors of Protein−Protein
Interactions. J. Am. Chem. Soc. 2011, 133, 676−679.
■
REFERENCES
■
(1) (a) Davis, J. M.; Tsou, L. K.; Hamilton, A. D. Synthetic Non-
Peptide Mimetics of α-Helices. Chem. Soc. Rev. 2007, 36, 326−334.
(b) Yin, H.; Hamilton, A. D. Strategies for Targeting Protein−Protein
Interactions With Synthetic Agents. Angew. Chem., Int. Ed. 2005, 44,
4130−4163. (c) Bullock, B. N.; Jochim, A. L.; Arora, P. S. Assessing
Helical Protein Interfaces for Inhibitor Design. J. Am. Chem. Soc. 2011,
133, 14220−14223. (d) Cummings, C. G.; Hamilton, A. D. Disrupting
Protein−Protein Interactions with Non-Peptidic, Small Molecule
α-Helix Mimetics. Curr. Opin. Chem. Biol. 2010, 14, 341−346.
(2) (a) Azzarito, V.; Long, K.; Murphy, N. S.; Wilson, A. J. Inhibition of
α-Helix-Mediated Protein−Protein Interactions Using Designed
Molecules. Nat. Chem. 2013, 5, 161−173. (b) Wilson, A. J. Helix
Mimetics: Recent Developments. Prog. Biophys. Mol. Biol. 2015, 119,
33−40.
(7) Duncton, M. A. J. Minisci Reactions: Versatile CH-Functionaliza-
tions for Medicinal Chemists. MedChemComm 2011, 2, 1135−1161.
(8) Cowden, C. Use of N-Protected Amino Acids in the Minisci
Radical Alkylation. Org. Lett. 2003, 5 (23), 4497−4499.
(9) Volonterio, A.; Moisan, L.; Rebek, J. Synthesis of Pyridazine-Based
Scaffolds as α-Helix Mimetics. Org. Lett. 2007, 9, 3733−3736.
(10) The regiochemistry of 6a was verified by conversion to 10d
followed by X-ray crystallography. See Supporting Information.
(11) (a) Martinelli, J. R.; Clark, T. P.; Watson, D. A.; Munday, R. H.;
Buchwald, S. L. Palladium-catalyzed aminocarbonylation of aryl
chlorides at atmospheric pressure: the dual role of sodium phenoxide.
Angew. Chem., Int. Ed. 2007, 46, 8460−8463. (b) Kotha, S.; Lahiri, K.;
Kashinath, D. Recent applications of the Suzuki−Miyaura cross-
coupling reaction in organic synthesis. Tetrahedron 2002, 58, 9633−
9695. (c) In Practical Synthetic Organic Chemistry: Reactions, Principles,
and Techniques. Chapter 4: Nucleophilic Aromatic Substitution; Caron, S.
Wiley, John & Sons: NJ, 2011; pp 237−253.
(3) For recent reviews of drug-discovery in beyond rule of five space,
see: (a) Doak, B. C.; Zheng, J.; Dobritzsch, D.; Kihlberg, J. How Beyond
Rule of 5 Drugs and Clinical Candidates Bind to Their Targets. J. Med.
Chem. 2016, 59, 2312−2327. (b) Doak, B. C.; Over, B.; Giordanetto, F.;
Kihlberg, J. Oral Druggable Space beyond the Rule of 5: Insights from
Drugs and Clinical Candidates. Chem. Biol. 2014, 21, 1115−1142.
(c) Kodadek, T.; McEnaney, P. J. Towards vast Libraries of Scaffold-
Diverse, Conformationally Constrained Oligomers. Chem. Commun.
2016, 52, 6038−6059.
(12) For a recent report on this valuable medicinal chemistry template,
see: Wlochal, J.; Bailey, A. Facile Synthesis of 4-Aryl and Alkyl
Substituted, N6-Alkylated Pyridazine-3,6-Diamines. Tetrahedron Lett.
2015, 56, 6791−6794.
(4) (a) Kutzki, O.; Park, H. O.; Ernst, J. T.; Orner, B. P.; Yin, H.;
Hamilton, A. D. Development of a Potent Bcl-xL Antagonist Based on
α-Helix Mimicry. J. Am. Chem. Soc. 2002, 124, 11838−11839.
D
DOI: 10.1021/acscombsci.6b00111
ACS Comb. Sci. XXXX, XXX, XXX−XXX