Kinases are enzymes that catalyze the transfer of a phosphate group from adenosine triphosphate (ATP), a high-energy phosphate-donating molecule, to the different substrates such as proteins, lipids and carbohydrates, while phosphatases remove a phosphate group from the substrates. Kinases play significant roles in essential cellular functions including cell cycle progression, growth, apoptosis, and metabolism. Consequently, the dysregulation of kinase function is a crucial factor for various diseases, including cancer, immunological, inflammatory, neurodegenerative, metabolic, cardiovascular and infectious diseases. Kinases have emerged as highly promising targets for drug development across diverse disease areas, principally in the field of oncology. The human kinome is composed of around 560 protein kinases, of which around 500 are eukaryotic protein kinases (ePKs). The protein kinases are divided into the sub-subclasses according to the amino acid residue that they phosphorylate, such as serine/threonine protein kinases, tyrosine-specific protein kinases, histidine-specific protein kinases, tryptophan kinases, and aspartyl/glutamyl protein kinase.

The first crystal structure of protein kinase A (PKA) was published in a landmark article [1] in 1991. The structure analysis revealed a conserved structural core present in protein kinases. This core consists of an N-lobe, which comprises a 5-stranded β-sheet (β1–β5) and at least one α-helix. Additionally, there is a C-lobe that is mostly α-helical but with a small yet important β-sheet (β6–β7) (Figure 1A) [2].  An ATP substrate molecule is sandwiched at the interface between the two lobes, and the surfaces of this cleft are formed from the β-sheets on both lobes (Figure 1B) [2]. The phosphates of ATP are positioned underneath the Gly-rich loop, which acts as a connection between β1 and β2. They interact with a conserved Lys residue located on β3 and are further linked to the C-lobe through a divalent cation, typically Mg²⁺. These lobes are joined together by a well-organized and concise linker sequence known as the hinge, which specifically recognizes the adenine base of an ATP molecule through two hydrogen bonds. The molecular interface connecting the two lobes is extensive and enriched in crucial structural and functional features.

Figure 1. Key structural features of a kinase domain

The drug pockets of human kinases exhibit a high degree of similarity, although they are not identical. There are over 80 ligand-binding sites in the kinase catalytic domain. The majority of kinase inhibitors are ATP competitive, deriving potency by occupying the deep hydrophobic pocket at the heart of the kinase domain. The selectivity of these inhibitors is contingent upon exploiting the differences between the amino acids that line the ATP site and exploring the adjacent pockets that are present in the kinase’s inactive states. Recently, there has been a focus on targeting allosteric pockets outside the ATP site to attain enhanced selectivity and combat resistance to existing therapeutics. The interactions of inhibitors with kinases exhibit through the formation of hydrogen bonds, salt bridges, or hydrophobic/hydrophilic interactions. For many years, targeting kinases has been regarded as challenging due to the inherent similarity of the ATP-binding site and the high concentrations of ATP in the cell. One of the primary challenges associated with the development of kinase inhibitors is their limited selectivity towards the binding sites. However, significant progress has been made in overcoming these obstacles, resulting in the development of a good number of potent and relatively selective kinase inhibitors. The majority of approved kinase inhibitors primarily consist of the following heterocyclic rings: Indazole, oxindole, 4-anilino-quinazoline, fused amino-pyrimidine, quinoline, isoquinoline, and 2-anilino-4-aryl-pyrimidine. The chemotype of these kinase inhibitors has been outlined in Table 1.

Table 1: An illustration of the chemotype of approved kinase inhibitors.

Success in drug development via targeting kinase

The first kinase inhibitor received FDA approval in 2001 for the treatment of cancer, chronic myeloid leukaemia (CML), was Imatinib, marketed under the brand names Gleevec and Glivec. Imatinib mesylate functions as a competitive inhibitor of several tyrosine kinases, including BCR-ABL and the platelet-derived growth factor receptors (PDGF-R). Its mechanism of action involves binding to the ATP-binding site of the target kinase, consequently impeding the transfer of phosphate from ATP to tyrosine residues of diverse substrates. Subsequent to Imatinib, four additional small molecule kinase inhibitors (SMKIs) that target ABL have been granted approval, namely Nilotinib, Dasatinib, Bosutinib, and Ponatinib. It is worth noting that the use of SMKIs in the treatment of CML has demonstrated long-lasting effects, with a significant number of patients in the chronic phase of CML showing no relapse after discontinuation of therapy. It is notable that majority (over 80%) of FDA-approved SMKIs are primarily designed for oncology applications.  The compilation of approved kinase inhibitors has been briefly outlined in Figure 2 [3].

Figure 2. Timeline of approved kinase inhibitors

Following the remarkable achievement of Imatinib, there has been a substantial global emphasis on kinase families for drug discovery over the past two decades. More than 70 SMKIs have been approved by FDA and hundreds are in the clinical trials. In the year 2022-2023, a number of kinase inhibitors, namely Abrocitinib, Defactinib, Pacritinib, Deucravacitinib, Futibatinib, and Pirtobrutinib, have received approval for pharmaceutical utilization.

References

[1] D. R. Knighton, J. H. Zheng, et al; Science 1991, 253, 407–414
[2] C. Arter,  L. Trask, et al;  J. Biol. Chem. 2022, 298, 102247
[3] M. M. Attwood, D. Fabbro, et al; Nat. Rev Drug Discov. 2021, 20, 839–861