Nalidixic Acid

Katelyn Brubaker

PHS 760

 Nalidixic acid (1-ethyl-1,4-dihydro-7-methyl-4-oxo-1,8-napthyridine-3-carboxylic acid) was first discovered in 1962 as a by-product of the synthesis and purification of the antimalarial agent choloroquine.¹ It is a synthetic, antibacterial napthyridine agent that is generally considered to be the “prototype” fluoroquinolone.²  Nalidixic acid inhibits the synthesis of bacterial DNA gyrase, which is an enzyme responsible for relieving strain on DNA being unwound by helicase via negative supercoiling.³

Nalidixic acid was not introduced for clinical use until 1964. It was FDA-approved for the treatment of urinary tract infections (UTIs) caused by gram-negative organisms, with the exception of Pseudomonas aeruginosa.^4,5 Nalidixic acid had limited indications for clinical use due to its rapid elimination from the body and the lack of accumulation in tissues.^6,7 The primary adverse reactions associated with the use of nalidixic acid include dizziness, drowsiness, GI disturbances, and photosensitive skin rashes.⁸ As more research was conducted on nalidixic acid and its structural analogs, it was discovered that by removing the nitrogen at C-8, the incidence of serious adverse reactions was decreased.⁹ The major pitfalls of nalidixic acid are its limited spectrum of activity and how quickly bacterial resistance developed against it.¹⁰ These issues were addressed in subsequent fluoroquinolone generations.  

The first major alteration to the structure of nalidixic acid was the addition of fluorine at C-6, which increased DNA gyrase inhibition and allowed this new molecule to be more effective against Gram-negative bacteria.⁶ In the second generation, the addition of the fluorine at C-6 was kept and a piperazine group at C-7 was added, which extended the spectrum of coverage to include Gram-positive bacteria. The further addition of a cyclopropyl group at C-1 increased the potency of the drug molecule.¹¹  

An example of the result of the structural alterations to nalidixic acid is ciprofloxacin, a second-generation fluoroquinolone.  In addition to covering Gram-negative organisms, ciprofloxacin also has Pseudomonas aeruginosa coverage and some Gram-positive coverage, both of which the first-generation fluoroquinolones, such as nalidixic acid, do not cover.⁴ These structural changes were important to the development of future fluoroquinolone generations because they ultimately resulted in more potent, more effective drugs.  In fact, nalidixic acid is no longer available in the U.S. due to safer and more effective UTI treatments.²  

As nalidixic acid became more widely used, more organisms began to develop resistance to it.  One of these organisms was Escherischia coli, a Gram-negative bacterium.  Researchers studied its resistance to nalidixic acid and found chromosomal mutations within the A subunit of DNA gyrase.³ This discovery showed that the mechanism of action of nalidixic acid was more specific than previously thought.  It is now known that nalidixic acid functions to inhibit the activity of a specific portion of DNA gyrase, gyrA (previously known as nalA). 

The development of bacterial resistance to nalidixic acid was one of the most important aspects of the drug’s history.  Prior to this, it was not known that nalidixic acid had a more specific mechanism of action towards bacterial DNA gyrase inhibition.  The knowledge that nalidixic acid specifically inhibited the gyrA locus of DNA gyrase allowed researchers to further explore the inhibition of the gyrB locus as a potential mechanism of action for antibacterial drugs.

Nalidixic acid was originally sold under the brand name NegGram, which was manufactured by the French pharmaceutical company Sanofi (now Sanofi-Aventis).¹² The brand name “NegGram” was officially trademarked on February 11, 1964.¹³ The NDA was officially approved by the FDA on March 6, 1964.  The drug was manufactured via the condensation of 2-amino-6-methylpyridine with diethyl ester of malonic acid and then cyclization to ethyl-1,4-dihydro-7-methyl-4-oxo-1,8-naphthyridine-3-carboxylate, followed by ethylation with ethyl bromide & saponification.¹⁴  After the patent expired, two generic versions of nalidixic acid came onto the market in 1988, one from Watson Labs (now Actavis) and another from Sun Pharma Industries.  The FDA approval for NegGram was withdrawn on December 7, 2007, at the request of the manufacturer due to the product no longer being on the market in the United States.¹⁵ The trademark on the brand name “NegGram” expired on April 1, 2016 and was not renewed.

References: 

1.     Lesher Gy, Froelich Ej, Gruett Md, Bailey Jh, Brundage Rp. 1,8 naphthyridine derivatives. A new class of chemotherapeutic agents. J Med Pharm Chem. 1962 Sep;91:1063-5.

2.     Jackson MA, Schutze GE; Committee on Infectious Diseases. The Use of Systemic and Topical Fluoroquinolones. Pediatrics. 2016 Nov;138(5). pii: e20162706. Review.

3.     Gellert M, Mizuuchi K, O'Dea MH, Itoh T, Tomizawa JI. Nalidixic acid resistance: a second genetic character involved in DNA gyrase activity. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4772-6.

4.     Emmerson AM, Jones AM. The quinolones: decades of development and use. J Antimicrob Chemother. 2003 May;51 Suppl 1:13-20. Review.

5.     Barlow AM. Nalidixic acid in infections of urinary tract. Laboratory and clinical investigations. Br Med J. 1963 Nov 23;2(5368):1308-10.

6.     Bisacchi GS. Origins of the Quinolone Class of Antibacterials: An Expanded "Discovery Story". J Med Chem. 2015 Jun 25;58(12):4874-82. doi: 10.1021/jm501881c. Epub 2015 Mar 12. Review.

7.     Mcchesney Ew, Froelich Ej, Lesher Gy, Crain Av, Rosi D. Absorption, excretion,and metabolism of a new antibacterial agent, nalidixic acid. Toxicol Appl Pharmacol. 1964 May;6:292-309.

8.     Autio S, Mäkelä P, Sunila R. Experience with nalidixic acid in the treatment of urinary tract infections of children. Archives of disease in childhood. 1966 Aug;41(218):395

9.     Andriole VT. The quinolones: past, present, and future. Clin Infect Dis. 2005 Jul 15;41 Suppl 2:S113-9. Review.

10.  Kresken M, Wiedemann B. Development of resistance to nalidixic acid and the fluoroquinolones after the introduction of norfloxacin and ofloxacin. Antimicrob Agents Chemother. 1988 Aug;32(8):1285-8. 

11.  Domagala JM. Structure-activity and structure-side-effect relationships for the quinolone antibacterials. J Antimicrob Chemother. 1994 Apr;33(4):685-706. Review. Erratum in: J Antimicrob Chemother 1994 Nov;34(5):851.

12.  Drugs@FDA: FDA Approved Drug Products [Internet]. accessdata.fda.gov. Available:  https://www.accessdata.fda.gov/scripts/cder/daf/index.cfm?event=overview.process &ApplNo=014214

13.  NEGGRAM Trademark - Registration Number 0764565 - Serial Number 72166047:: Justia Trademarks [Internet]. Justia. Available from: https://trademarks.justia.com/721/66/neggram-72166047.html

14.  Kirk-Othmer Encyclopedia of Chemical Technology. 4th ed. Volumes 1: New York, NY. John Wiley and Sons, 1991-Present., p. V20 669

15.  Lederle Laboratories et al.; Withdrawal of Approval of 73 New Drug Applications and 62 Abbreviated New Drug Applications [Internet]. Federal Register. 2007. Available from: https://www.federalregister.gov/documents/2007/11/07/E7-21886/lederle-laboratories-et-al-withdrawal-of-approval-of-73-new-drug-applications-and-62-abbreviated-new