(E)-3-(3-Carboxyphenyl)-2-(4-ethynylstyryl)quinazolin-4(3H)-one

(E)-3-(2-(4-Cyanostyryl)-4-Oxoquinazolin-3(4h)-Yl)benzoic Acid;

1624273-22-8  CAS

NA SALT 1624273-21-7 CAS

INNOVATORS

University Of Notre Dame

Mayland Chang, Shahriar Mobashery, Renee BOULEY INVENTORS

PAPER

Journal of the American Chemical Society (2015), 137(5), 1738-1741.

http://pubs.acs.org/doi/abs/10.1021/jacs.5b00056

Discovery of Antibiotic (E)-3-(3-Carboxyphenyl)-2-(4-cyanostyryl)quinazolin-4(3H)-one

Abstract

In the face of the clinical challenge posed by resistant bacteria, the present needs for novel classes of antibiotics are genuine. In silico docking and screening, followed by chemical synthesis of a library of quinazolinones, led to the discovery of (E)-3-(3-carboxyphenyl)-2-(4-cyanostyryl)quinazolin-4(3H)–one (compound 2) as an antibiotic effective in vivo against methicillin-resistant Staphylococcus aureus (MRSA). This antibiotic impairs cell-wall biosynthesis as documented by functional assays, showing binding of 2 to penicillin-binding protein (PBP) 2a. We document that the antibiotic also inhibits PBP1 of S. aureus, indicating a broad targeting of structurally similar PBPs by this antibiotic. This class of antibiotics holds promise in fighting MRSA infections.

PATENT

WO 2014138302

http://www.google.com/patents/WO2014138302A1?cl=en

Staphylococcus aureus is a common bacterium found in moist areas of the body and skin. S. aureus can also grow as a biofilm, representing the leading cause of infection after implantation of medical devices. Approximately 29% (78.9 million) of the US population is colonized in the nose with S. aureus, of which 1.5% (4.1 million) is methicillin-resistant S. aureus (MRSA). In 2005, 478,000 people in the US were hospitalized with a S. aureus infection, of these 278,000 were MRSA infections, resulting in 19,000 deaths. MRSA infections have been increasing from 2% of S. aureus infections in intensive care units in 1974 to 64% in 2004, although more recent data report stabilization. Approximately 14 million outpatient visits occur every year in the US for suspected S. aureus skin and soft tissue infections. About 76% of these infections are caused by S. aureus, of which 78% are due to MRSA, for an overall rate of 59%. Spread of MRSA is not limited to nosocomial (hospital-acquired) infections, as they are also found in community-acquired infections. Over the years, β-lactams were antibiotics of choice in treatment of S. aureus infections. However, these agents faced obsolescence with the emergence of

MRSA. Presently, vancomycin, daptomycin or linezolid are agents for treatment of MRSA infections, although only linezolid can be dosed orally. Resistance to all three has emerged. Thus, new anti-MRSA therapeutic strategies are needed, especially agents that are orally bioavailable.

Clinical resistance to β-lactam antibiotics by MRSA has its basis predominantly in acquisition of the mecA gene, which encodes penicillin-binding protein 2a (PBP2a). PBP2a, a cell-wall DD- transpeptidase, is refractory to inhibition by essentially all commercially available β-lactams (ceftaroline is an exception), antibiotics that irreversibly acylate the active-site serine of typical PBPs. PBPs catalyze biosynthesis of the bacterial cell wall, which is essential for the survival of the bacterium. Accordingly, new ηοη-β-lactam antibiotics that inhibit PBP2a are needed to combat drug-resistant strains of bacteria. SUMMARY

Staphylococcus aureus is responsible for a number of human diseases, including skin and soft tissue infections. Annually, 292,000 hospitalizations in the US are due to S. aureus infections, of which 126,000 are related to methicillin-resistant Staphylococcus aureus (MRSA), resulting in 19,000 deaths. A novel structural class of antibiotics has been discovered and is described herein. A lead compound in this class shows high in vitro potency against Gram-positive bacteria comparable to those of linezolid and superior to vancomycin (both considered gold standards) and shows excellent in vivo activity in mouse models of MRSA infection.

The invention thus provides a novel class of ηοη-β-lactam antibiotics, the quinazolinones, which inhibit PBP2a by an unprecedented mechanism of targeting both its allosteric and active sites. This inhibition leads to the impairment of the formation of cell wall in living bacteria. The quinazolinones described herein are effective as anti-MRSA agents both in vitro and in vivo. Furthermore, they exhibit activity against other Gram-positive bacteria. The quinazolinones have anti-MRSA activity by themselves. However, these compounds synergize with β-lactam antibiotics. The use of a combination of a quinazolinone with a β-lactam antibiotic can revive the clinical use of β-lactam antibacterial therapy in treatment of MRSA infections. The invention provides a new class of quinazolinone antibiotics, optionally in combination with other antibacterial agents, for the therapeutic treatment of methicillin- resistant Staphylococcus aureus and other bacteria.

The quinazolinone compounds described herein can be prepared using standard synthetic techniques known to those of skill in the art. Examples of such techniques are described by Khajavi et al. (J. Chem. Res. (S), 1997, 286-287) and Mosley et al. (J. Med. Chem. 2010, 53, 5476-5490). A general preparatory scheme for preparing the compounds described herein, for example, compounds of Formula

wherein each of the variables are as defined for one or more of the formulas described herein, such as Formula (A).

EXAMPLES

Example 1. Compound Preparation

Chemistry. Organic reagents and solvents were purchased from Sigma- Aldrich. ^lH and ¹³C NMR spectra were recorded on a Varian INOVA-500. High-resolution mass spectra were obtained using a Bruker micrOTOF/Q2 mass spectrometer.

2-Methyl-4H-benzo[</| [l,3]oxazin-4-one (3). Anthranilic acid (20 g, 146 mmol) was dissolved in triethyl orthoacetate (45 mL, 245 mmol) and refluxed for 2 h. The reaction mixture was cooled on ice for 4 h to crystallize the intermediate. The resulting crystals were filtered and washed with hexanes to give 3 (17 g, 72% yield). ^lH NMR (500 MHz, CDC1₃) δ 2.47 (s, 3H), 7.50 (t, J= 7.38 Hz, 1H), 7.54 (d, J = 7.98 Hz, 1H), 7.80 (t, J= 7.18 Hz, 1H), 8.18 (d, J= 7.78 Hz, 1H). ¹³C NMR (126 MHz, CDCI3) δ 21.59, 1 16.84, 126.59, 128.42, 128.66, 136.77, 146.61, 159.89, 160.45. HRMS (m/z): [M + H]⁺, calcd for C9H8NO2, 162.0550; found , 162.0555.

2-Methyl-3-(3-carboxyphenyl)-quinazolin-4(3//)-one (4). Compound 3 (2 g, 12.4 mmol) and 3- aminophenol (1.7 g, 12.4 mmol) were suspended in glacial acetic acid (8 mL, 140 mmol), and dissolved upon heating. The reaction was refluxed for 5 h, at which point 5 mL water was added to the cooled reaction mixture. The resulting precipitate was filtered and washed with water, followed by cold ethanol and hexane to give 4 (3.19 g, 92% yield). ^lH (500 MHz, DMSO-d₆) δ 2.87 (s, 3H), 7.52 (t, J= 7.38 Hz, 1H), 7.66-7.73 (m, 3H), 7.84 (t, J= 7.38 Hz, 1H), 8.01 (s, 1H), 8.09 (t, J= 7.58 Hz, 2H). ¹³C NMR (126 MHz, DMSO-de) δ 24.13, 120.48, 126.32, 126.47, 126.72, 129.52, 129.83, 130.01, 132.40, 133.07, 134.67, 138.18, 147.37, 154.13, 161.44, 166.58. HRMS (m/z): [M + H]⁺, calcd for C16H13N2O3 ,

281.0921 ; found, 281.0917.

Sodium (£^‘)-3-(3-carboxyphenyl)-2-(4-cyanostyryl)quinazolin-4(3H)-one (2). Compound 4 (1.0 g, 3.6 mmol) and 4-formylbenzonitrile (0.56 g, 4.3 mmol) were suspended in glacial acetic acid (5 mL, 87 mmol), a suspension that dissolved upon heating. The reaction was refluxed for 18 h and 5 mL water was added to the cooled reaction mixture. The resulting precipitate was filtered and washed with water, followed by cold ethanol and hexanes to afford the carboxylic acid (0.77g, 75% yield). HRMS (m/z): [M + H]⁺, calcd for C24H16N3O3, 394.1 186; found 394.1214. The carboxylic acid (0.45 g, 1.1 mmol) was dissolved in hot ethanol, to which sodium 2-ethylhexanoate (0.28 g, 1.7 mmol) was added. The reaction mixture was stirred on ice for 2 h. The precipitate was filtered and washed with cold ethanol. The product was obtained by dissolving the precipitate in about 5 mL of water and subsequent lyophilization of the solution to give 2 as the sodium salt (0.4 g, 85% yield).

¾ NMR (500 MHz, DMSO- de) δ 6.47 (d, J= 15.55 Hz, 1H), 7.59 (m, 3H), 7.74 (d, J= 5.38 Hz, 2H), 7.79 (m, 3H), 7.91 (m, 2H), 8.05 (s, 1H), 8.14 (d, J= 7.78 Hz, 2H).

¹³C NMR (126 MHz, DMSO-de) δ 11 1.56, 1 18.61, 120.76, 123.42, 126.50, 127.01, 127.35, 128.26, 129.99, 130.06, 130.12, 132.33, 132.83, 133.46, 134.89, 136.95, 137.03, 139.25, 147.21, 150.74, 161.25, 166.52.

HRMS (m/z): [M + H]⁺, calcd for C24Hi₅N₃NaO₃, 416.1006; found, 416.0987.

PAPER

http://pubs.acs.org/doi/full/10.1021/acs.jmedchem.6b00372

Structure–Activity Relationship for the 4(3H)-Quinazolinone Antibacterials

We recently reported on the discovery of a novel antibacterial (2) with a 4(3H)-quinazolinone core. This discovery was made by in silico screening of 1.2 million compounds for binding to a penicillin-binding protein and the subsequent demonstration of antibacterial activity againstStaphylococcus aureus. The first structure–activity relationship for this antibacterial scaffold is explored in this report with evaluation of 77 variants of the structural class. Eleven promising compounds were further evaluated for in vitro toxicity, pharmacokinetics, and efficacy in a mouse peritonitis model of infection, which led to the discovery of compound 27. This new quinazolinone has potent activity against methicillin-resistant (MRSA) strains, low clearance, oral bioavailability and shows efficacy in a mouse neutropenic thigh infection model.

NMR

INVENTORS

Renee Bouley

MAYLAND CHANG

http://chemistry.nd.edu/people/mayland-chang/

MAYLAND CHANG

• Research Professor; Director, Chemistry-Biochemistry-Biology Interface (CBBI) Program
• Office: 247 NSH
• Phone: (574) 631-2965

Dr. Chang obtained B.S. degrees in biological sciences and chemistry from the University of Southern California, and a Ph.D. in chemistry from the University of Chicago.  Subsequently, she conducted postdoctoral research at Columbia University as a National Institutes of Health postdoctoral fellow.  She joined the faculty of the University of Notre Dame in 2003.  Previously, Dr. Chang was Chief Operating Officer of University Research Network, Inc., Senior Scientist with Pharmacia Corporation, and Senior Chemist at Dow Chemical Company.  She has characterized the ADME properties of numerous drugs, as well as prepared NDAs, INDs, Investigator’s Brochures, product development plans, and candidate drug evaluations.

Shahriar Mobashery

/////// 1624273-22-8, Antibiotic,  (E)-3-(3-Carboxyphenyl)-2-(4-cyanostyryl)quinazolin-4(3H)-one, methicillin-resistant S. aureus, MRSA, 1624273-21-7, PRECLINICAL

O=C(O)c1cc(ccc1)N3C(=Nc2ccccc2C3=O)/C=C/c4ccc(C#N)cc4