The history and rationale of using carbonic anhydrase inhibitors in the treatment of peptic ulcers. In memoriam Ioan Puşcaş (1932–2015)

Figures & data

Table 1. Chronology of carbonic anhydrase research1–12.

Year	Author	Location, country, institute	Research	Comment
1909	Christian Bohr (1855–1911)9	University of Copenhagen, Denmark	Effect of CO2 on dissociation of oxygen from haemoglobin (Bohr effect). Formulation of direct combination theory (CO2 is carried in blood in reversible combination with proteins). He described the S-shaped O2 dissociation curve of haemoglobin in 1904.	Father of physicist Niels Bohr (1885–1962).
1922	John Scott Haldane (1860–1936)9	Fellow of New College, Oxford, UK	Promoted the bicarbonate theory: CO2 is carried in blood mainly by bicarbonate. In 1898, the first gas analyser was produced. He described silicosis, gas warfare and proposed the use of O2 therapy in war victims.	Performed breathing experiments on himself and his son.
1924	Carl Faurholt (1890–1972)9	College of Pharmacy, Copenhagen, Denmark	He measured the slow spontaneous dissociation rate of bicarbonate, obtaining values not different from today’s experiments.	A landmark experiment, used extensively by his followers.
1928	Oscar M Henriques (1895–1953)9	Finster Laboratories, Copenhagen	Using a Van Slyke gas analyser, he concluded that CO2 in the blood is transported as haemoglobin carbamate which is formed and dissociated rapidly.	He probably measured CA activity from red blood cells.
1930	Donald Dexter Van Slyke (1883–1971)10	Rockefeller Institute, New York	Devised early equipment for blood gas analysis in 1924. Performed measurements of CO2 transportation in blood but could no differentiate haemoglobin and CA.	He also determined a minimal level of pH = 6.5 below which a coma occurs and showed the relation between renal blood flow and urea clearence.
1932–1933	Norman Urquhart Meldrum (1907–1933)11 and Francis John Worsley Roughton (1899–1972)	Physiological and Biochemical Laboratories, Cambridge, UK	Preparation of CA from erythrocytes of different species and determination of its kinetic properties. They also identified the first CA inhibitors (cyanates, CO and H2S).	Untimely death of Meldrum hindered further experiments. Due to J Physiol policy, authors were listed in alphabetical order and not according to their contribution, although Meldrum was the one who separated CA from haemoglobin.
1939	Horace Willard Davenport (1913–2005)12	WG Kerckhoff Laboratories of Biological Sciences, Pasadena, USA	Discovery of large amounts of CA in the gastric mucosa of cats, rats and rabbits. He identified renal CA and showed that sulphanilamide inhibits it.	Formulated a theory of acid secretion with later was revised by himself and the concept of gastric mucosal barrier.
1939	David Keilin (1887–1963) and Taddeus Mann (1908–1993)4	Molteno Institute, Cambridge	Unsubstituted sulphonamides could inhibit CA and are devoid of antibacterial activity. They showed the presence of zinc atoms in the active CA centre.	Keilin also investigated cytochromes, while Mann contributed to reproductive biology.
1950	Richard Roblin and James W. Clapp5	Cyanamid Lederle Division, Cyanamid, NJ, USA	Discovery of acetazolamide, a compound ∼40 times more potent than sulphanilamide.	Marketed in 1954 and rapidly introduced to therapy of several diseases (Table 2).
1967	Thomas H. Maren (1918–1999)7	University of Florida, Gainesville, USA	Designed a micromethod for determination of CA activity and invented dorzolamide, a topical CA inhibitor for the treatment of glaucoma.	Dedicated 50 years to CA research. His 1967 review article became a citation classic.

Table 2. Timeline of clinical uses of acetazolamide (AAZ).

1955	Glaucoma
1956	Epilepsy
1956	Congestive heart failure
1956	Chronic cor pulmonale
1957	Migraine
1957	Metabolic alkalosis
1960	Peptic ulcer
1968	Mountain sickness

Table 3. Kinetic parameters for CO₂ hydration reaction catalysed by some human α-CA isozymes at 20 °C and pH 7.5, and HpCA isozymes belonging to the α- and β-CA class, and their inhibition data with acetazolamide AAZ (5-acetamido-1,3,4-thiadiazole-2-sulfonamide), a clinically used sulfonamide39–41.

Isozyme	Activity level	kcat (s^−1)	Km (mM)	kcat/Km (M^−1 s^−1)	KI (acetazolamide) (nM)
hCA I	Medium	2.0 × 10^5	4.0	5 × 10^7	250
hCA II	Very high	1.4 × 10^6	9.3	1.5 × 10^8	12
hCA III	Very low	1.0 × 10^4	33.3	3 × 10^5	300 000
hCA VA	Low	2.9 × 10^5	10.0	2.9 × 10^7	63
hCA VB	High	9.5 × 10^5	9.7	9.8 × 10^7	54
hCA XII	Medium	4.2 × 10^5	12.0	3.5 × 10^7	5.7
hCA XIV	Medium	3.1 × 10^5	7.9	3.9 × 10^7	41
HpαCA*	Low	2.5 × 10^5	16.6	1.5 × 10^7	21
HpβCA†	Medium	7.1 × 10^5	14.7	4.8 × 10^7	40

h, human; hp, Helicobacter pylori enzyme.

*At pH 8.9 and 25 °C.

†At pH 8.3 and 20 °C.

Figure 1. Helicobacter pylori CAs and urease and their involvement in the acid acclimation of the pathogen within the gastric mucosa46.

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