TARGET PREDICTIONS AND PHYSICOCHEMICAL PARAMETERS:

HYDROXYCHLOROQUINE

INTRODUCTION

Hydroxychloroquine (HCQ) belongs to a class of drugs known as 4-aminoquinolines, in which

one of the N-ethyl groups is hydroxylated at position 2. Owing to the presence of a basic side
chain, its flat aromatic core structure makes it weak at the base. The accumulation of
hydroxychloroquine in lysosomal compartments, which appears to be essential for their function
and possible interaction with nucleic acids, is facilitated by the basic side chain (Schrezenmeier,
E., & Dörner, T., 2020). Originally developed as an antimalarial, hydroxychloroquine is now used
to treat autoimmune and rheumatic illnesses, including systemic lupus erythematosus, by acting as
an immunomodulatory and anti-inflammatory drug (Bansal et al., 2021).

TARGET PREDICTIONS AND ITS LITERATURE REVIEW

Predictions by software of target proteins to which HCQ can bind with high affinity is as below:

1) Alpha-1d adrenergic receptors (Family A G protein-coupled receptor)

2) Histamine N-methyltransferase (by homology) (Enzyme)
3) HERG (Voltage Gated Iron Channel)

α1-Adrenergic receptors (ARs) are members of the G-Protein Coupled Receptor superfamily and,
along with other related receptors (β and α2), they are involved in regulating the sympathetic
nervous system through binding and activation by norepinephrine and epinephrine. Alpha-1d
adrenergic receptors demonstrated the highest potential drug target for hydroxychloroquine. Thus,
it was discovered by researchers that α1-Adrenergic receptors activate phospholipase C, which
raises IP3 and DAG and, in turn, raises intracellular calcium concentrations, which causes smooth
muscle contraction and glycogenolysis (Taylor et al.,2023).

The target protein Histamine N-Methyltransferase (HNMT) is another that binds with greater
affinity. According to Wang et al. (2002), it is an enzyme that is involved in the metabolism of
histamine, a neurotransmitter and immune system mediator. Metoprine and diphenhydramine are
examples of histamine N-methyltransferase inhibitors that primarily use hydroxychloroquine to
target HNMT. By preventing histamine from being broken down, HCQ can raise the concentration
of histamine in a variety of tissues (Nakazaka et al., 1994). Therefore, both of these can be used as
likely targets for hydroxychloroquine based on target prediction and literature evaluation.

Last but not least, the potassium ion channel encoded by the human Ether-à-go-go-Related Gene
(hERG) is essential for cardiac repolarization. Medication that blocks the hERG potassium channel
may cause the electrical activity of the heart to lengthen the QT interval, which may cause a disease
called drug-induced Long QT Syndrome (LQTS). Nevertheless, no evidence was discovered to
substantiate the relationship between HERG.

PHYSICOCHEMICAL PROPERTIES

LIPOPHILICITY: Because of its mild lipophilicity, Hydroxychloroquine can dissolve in polar

and nonpolar solvents. Its capacity to diffuse throughout the body's tissues and cross-cell
membranes is facilitated by this characteristic. It is crucial to remember that the lipophilicity of
hydroxychloroquine might also affect its metabolism, pharmacokinetics, and possible interactions
with lipids or other molecules (Da Silva et al.,2021).

MOLECULAR WEIGHT: Hydroxychloroquine, with a low molecular weight of 336 g/mol, is

efficiently absorbed and has fewer off-target effects. Its small molecular weight allows more of the
drug to reach its target site, increasing its effectiveness (Browning et al.,2014)
ROTATABLE BONDS: Hydroxychloroquine, with the chemical formula C18H26ClN3O,
contains 09 rotatable bonds. Single bonds that allow the linked groups to freely revolve around
them are known as rotatable bonds. The flexibility and conformational space of the chemical can
be inferred by counting the rotatable bonds (Badraoui, et al., 2021). Molecules with few or no
rotatable bonds, on the other hand, could have a more stiff structure. Additionally, rotatable bonds
can make a molecule more flexible, which is advantageous when interacting with other molecules.

WATER SOLUBILITY: with normal temperature, hydroxychloroquine dissolves less readily in

water, with a rate of around 0.0261 mg/mL. This restricted water solubility may affect how the
medication dissolves and absorbs throughout the body. Usually used orally, hydroxychloroquine's
absorption can be impacted by several variables, including how soluble it is (Mamontov et al.,
2020).

LIPINSKI RULE: Lipinski's rule of five states that a molecule has fewer than five hydrogen bond
donors, ten hydrogen bond acceptors, a molecular weight of less than 500 Daltons, and a Log P
value of less than five, all of which are indicators that the compound is likely to be orally active
(Lipinski et al., 2001). Lipinski's rule of five is satisfied by hydroxychloroquine. Its computed log
P is 3.7 and its molecular weight is 335.87 g/mol. Three hydrogen bond acceptors and two
hydrogen bond donors are present (Badraoui et al., 2021).

VEBER RULE: The Veber rule is another guideline used in drug design, it predicts the likelihood
of a compound exhibiting high toxicity or low bioavailability based on the number of rotatable
bonds and polar surface area. According to the Veber rule, compounds with no more than ten
rotatable bonds, no more than twelve hydrogen bonds, and a polar surface area of less than 140 Å2
are more likely to have favorable pharmacokinetic properties (Veber et al.,2002).
Hydroxychloroquine satisfies the Veber rule since it has nine rotatable bonds, a total of five
hydrogen bonds, and a polar surface area of 48.39 Å2 (Badraoui, et al.,2021).

EGAN’S RULE:The evaluation of membrane permeability for optimum bioavailability was limited
to two descriptors, namely logP and TPSA. It is important to understand that substances that can
enter cells via carrier-mediated transport or active absorption processes may be removed by
bioavailability filters (Kralj et al.,2023). TPSA less than 131.6 Å2 and logP less than 5.88 are two
of the parameters for Egan's rule. Since hydroxychloroquine has a TPSA of 48.39 Å2 and a Log P
value of 3.7, it complies with Egan's criteria (Image 3).
GHOSE’S and MUEGGE’S RULE:The purpose of this rule is to create combinatorial or
medicinal chemistry libraries for drug discovery. The criteria for this filter were established
through an analysis of drug databases and are outlined as follows: 160 < MW < 480, −0.4 < LogP
< 5.6, 20 < number of atoms < 70 and 40 < molar refractivity < 130 (Egan’s rule paper).
hydroxychloroquine meets Ghose's criteria as its molecular weight is 335.87 g/mol, it has a Log P
value of 3.7, consisting of 23 atoms, and possesses a molecular refractivity of 98.57 m3.mol -1.

On the other hand, Muegge’s rule parameters include 200 <MW< 600, -2 < logP< 5, number of
rings (nRing)< 7, number of carbon atoms > 4, number of heteroatoms (nHet) > 1, nRot <15, nHA
< 10, nHD < 5 (Babalola et al.,2022). hydroxychloroquine satisfies Muegge’s rule as its MW is
335.87 g/mol, Log P value of 3.7 along with 2 rings. It also has 18 carbon atoms and 5 heteroatoms.
Additionally, it has 9 rotational bonds along with 3nHA and 2nHD (Image 3).

All five conditions—the Veber, Ghose, Muegge, and Lipinski rules of five—are satisfied by
hydroxychloroquine, indicating that it has good drug-like qualities and is probably going to have
good oral bioavailability.

APPENDIX
 Image 1: Swiss Target Prediction for the drug- Hydroxychloroquine

Image2: List of Swiss Target Prediction for the drug- Hydroxychloroquine

Image 3: Swiss ADME Data Including Radar, Solubility, Physicochemical Properties,
Lipophilicity, Pharmacokinetics, etc.

Image 4: Canonical Smile from PubChem

REFERENCES
1) https://pubchem.ncbi.nlm.nih.gov/compound/3652
2) Lipinski CA, Lombardo F, Dominy BW, Feeney PJ. Experimental and computational
approaches to estimate solubility and permeability in drug discovery and development
settings. Adv Drug Deliv Rev. 2001;46(1–3):3–26. https://doi.org/10.1016/s0169-
409x(00)00129-0.
3) Veber DF, Johnson SR, Cheng HY, Smith BR, Ward KW, Kopple KD. Molecular properties
that influence the oral bioavailability of drug candidates. J Med Chem. 2002;45(12):2615–
23. https://doi.org/10.1021/jm020017n.
4) Schrezenmeier, E., & Dörner, T. (2020). Mechanisms of action of hydroxychloroquine and
chloroquine: implications for rheumatology. Nature Reviews Rheumatology, 16(3), 155-
166.
5) Bansal, P., Goyal, A., Cusick IV, A., Lahan, S., Dhaliwal, H. S., Bhyan, P., ... & Davis III,
J. M. (2021). Hydroxychloroquine: a comprehensive review and its controversial role in
coronavirus disease 2019. Annals of medicine, 53(1), 117-134.
6) Perez, D. M. (2023). α1-Adrenergic Receptors: Insights into Potential Therapeutic
Opportunities for COVID-19, Heart Failure, and Alzheimer’s Disease. International
Journal of Molecular Sciences, 24(4), 4188.
7) Taylor BN, Cassagnol M. Alpha-Adrenergic Receptors. [Updated 2023 Jul 10]. In:
StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan-. Available
from: https://www.ncbi.nlm.nih.gov/books/NBK539830/
8) Furst DE. Pharmacokinetics of hydroxychloroquine and chloroquine during treatment of
rheumatic diseases. Lupus. 1996 Jun;5 Suppl 1:S11-5. PMID: 8803904.
9) Nakazawa H, Sekizawa K, Morikawa M, Yamauchi K, Satoh M, Maeyama K, Watanabe
T, Sasaki H. Viral respiratory infection causes airway hyperresponsiveness and decreases
histamine N-methyltransferase activity in Guinea pigs. Am J Respir Crit Care Med
1994;149:1180–5. https://doi.org/ 10.1164/ajrccm.149.5.8173757.
10) Wang L, Thomae B, Eckloff B, Wieben E, Weinshilboum R. Human histamine N-
methyltransferase pharmacogenetics: gene resequencing, promoter characterization, and
functional studies of a common 5′ -flanking region single nucleotide polymorphism (SNP).
Biochem Pharmacol 2002;64:699–710. https:// doi.org/10.1016/S0006-2952(02)01223-6.
11) Browning, D. J., & Browning, D. J. (2014). Pharmacology of chloroquine and
hydroxychloroquine. Hydroxychloroquine and chloroquine retinopathy, 35-63.
12) Eldanasory OA, Eljaaly K, Memish ZA, Al-Tawfiq JA. Histamine release theory and roles
of antihistamine in the treatment of cytokines storm of COVID-19, Travel Med. Inf Disp
2020;37. https://doi.org/10.1016/j.tmaid.2020.101874
13) Navya, V. B., & Hosur, M. V. (2021). A computational study on hydroxychloroquine
binding to target proteins related to SARS-COV-2 infection. Informatics in Medicine
Unlocked, 26, 100714.
14) da Silva, A. E. A., de Abreu, P. M. B., Geraldes, D. C., & de Oliveira Nascimento, L. (2021).
Hydroxychloroquine: Pharmacological, physicochemical aspects and activity
enhancement through experimental formulations. Journal of Drug Delivery Science and
Technology, 63, 102512.
15) Badraoui, R., Adnan, M., Bardakci, F., & Alreshidi, M. M. (2021). Chloroquine and
hydroxychloroquine interact differently with ACE2 domains reported to bind with the
coronavirus spike protein: mediation by ACE2 polymorphism. Molecules, 26(3), 673.
16) Mamontov, E., Cheng, Y., Daemen, L. L., Keum, J. K., Kolesnikov, A. I., Pajerowski, D.,
... & Stone, M. B. (2020). Effect of hydration on the molecular dynamics of
hydroxychloroquine sulfate. ACS omega, 5(33), 21231-21240.
17) Kralj, S., Jukič, M., & Bren, U. (2023). Molecular Filters in Medicinal
Chemistry. Encyclopedia, 3(2), 501-511.
18) Babalola, S., Igie, N., & Odeyemi, I. (2022). Molecular Docking, Drug-Likeness Analysis,
In Silico Pharmacokinetics, and Toxicity Studies of p-Nitrophenyl Hydrazones as Anti-
inflammatory Compounds against COX-2, 5-LOX, and H+/K+ ATPase. Pharmaceutical
Fronts, 4(04), e250-e266.