Mutations in the epidermal growth factor receptor (EGFR) kinase domain are the oncogenic drivers of this adenocarcinoma. Tyrosine kinase inhibitors (TKIs), developed over the last two decades, targeting EGFR mutations, have shown advantages over chemotherapy in treating EGFR-positive NSCLC patients and are now considered the standard method in this field. However, drug resistance has always been a sticking point in cancer therapeutic development. It’s still urgent to develop a new generation of EGFR inhibitors which are supposed to address the problems of the first three generations and satisfy clinical needs.
How are the fourth-generation EGFR inhibitors developed?
1.EGFR inhibitors targeting mutations
Currently, EGFR+/T790M/C797S mutations are generated when patients are treated with Osimertinib and are not treatable with currently available EGFR TKIs. Given the unmet need for next-generation EGFR inhibitors, new BIC inhibitors that do not inhibit WT (wild-type) and are effective against EGFR+/T790M and EGFR+/T790M/C797S mutants need to be developed.
2.Hit revealed by compound screening
Compound 4, a hit screened from Blueprint’s compound library, has potent enzymatic and cellular inhibitory activity against EGFR mutants but not against wild-type EGFR, showing high selectivity (Table 1). The introduction of F substitution in the ortho position of the piperidinol hydroxyl group of compound 4 increases the inhibitory activity against the mutants (2-3 times). This key modification is enlightened by Genentech’s modification idea. 5 to 6 is a common cyclization path, where the pyridine N atom retains the hydrogen bond interaction.
3.Optimization of hit
A SAR study on compound 6 reveals compound 24 (Table 2). Compound 24 has excellent enzymatic and cellular activity against EGFR mutants, but weak inhibition against wild types, showing high selectivity. But its bioavailability is very low with the main problem of poor absorption. PK studies in rats demonstrate that venous clearance is close to the hepatic blood flow rate (about 70 mL/kg/min), resulting in the rapid clearance of compound 24 in rats, and the bioavailability is only 2%. To mitigate the potential risk, researchers add a methyl group to the aziridine ring to produce compound 25. The aim is to increase steric hindrance around the polar substituents to reduce efflux. By replacing a nitrogen atom with a carbon atom to reduce the TPSA of the skeleton, isoquinoline 26 and better MDCK-MDR1 are obtained. Improved passive penetration and reduced efflux result in a greatly improved PK in rats (F=85%, Cl=20 mL/kg/min ), but have little effect on the inhibitory activity of EGFR L858R/T790M cells. Compound 27 further improves the inhibition against mutants and has better selectivity for WT.
4.Problems with compound 27
The pharmacokinetic results of compound 27 show that the pharmacokinetic correlation between in vitro and in vivo is poor (Figure 3). A moderate in vitro clearance rate (Clint=55 μL/mg/min) and a high in vivo clearance rate (Cl=31.8 mL/kg/min) are observed. This suggests that 27 may be prone to biphasic metabolism and that phase II metabolism may be responsible for the high clearance of cyno in vivo. Subsequent experiments reveal that 27 is affected by UGT-mediated glucuronidation in human hepatocytes, but not in rat or dog hepatocytes, and the glucuronidation site is found to be the hydroxyl group of piperidinol.
5.Determination of the PCC
The modification of compound 27 is aimed at reducing phase II metabolism (Figure 4). Substituents are introduced at α and β positions of hydroxyl groups to increase steric hindrance to obtain 29 and 28. The interference of steric hindrance could slow down the binding reaction of phase II metabolic hydroxyl groups. 28 does not work while 29 achieves a significant effect. But the inhibition of 29 against EGFR+/T790M becomes weaker. Hydroxymethylation to block the glucuronidation site of 30 results in compounds with improved structure in vivo while maintaining good activity and selectivity for EGFR WT. 30 (BLU-945) is further characterized to determine its potential as a candidate compound.
6.Validation of efficacy tests
In the preclinical 28-day GLP toxicity study in rats and nonhuman primates (NHP), the appropriate safety window is obtained for BLU-945 to support entry into human trials at an initial dose of 25 mg during the dose-escalation phase. Since compounds at high pH have lower solubility in aqueous solution, the formulation of spray-dried dispersion is chosen by researchers.
Currently, the first human phase Ⅰ dose-escalation study is underway in patients with EGFR-mutated NSCLC who have previously received EGFR-targeted TKI (NCT04862780) at least once. The final results indicate that BLU-945 has a low clearance rate and a long plasma half-life (t1/2).
Visions of the future
Up to 50% of patients treated with first- and second-generation TKIs will develop resistance to T790M mutation. Although the third-generation Osimertinib is effective for T790M mutation, other mutations are still beyond treatment. To improve the prognosis of EGFR-mutated NSCLC patients, it is necessary to develop new inhibitors (the fourth generation, the fifth generation, etc.) for different resistance mechanisms.
Related Tags:
| Tag | Description |
| EGFR | EGFR (The epidermal growth factor receptor; ErbB-1; HER1 in humans) is the cell-surface receptor for members of the epidermal growth factorfamily (EGF-family) of extracellular protein ligands. |
Related Products:
| Tag | Name | CAS | Synonyms | Description |
| EGFR | Osimertinib | 1421373-65-0 | AZD-9291 | Osimertinib is a third-generation EGFR inhibitor, showed promise in preclinical studies and provides hope for patients with advanced lung cancers that have become resistant to existing EGFR inhibitors. |
| EGFR | Gefitinib | 184475-35-2 | ZD-1839 | Gefitinib effectively inhibits all tyrosine phosphorylation sites on EGFR in both the high and low-EGFR-expressing cell lines including NR6, NR6M and NR6W cell lines. |
| EGFR | Afatinib | 439081-18-2 | BIBW-2992 | Afatinib (BIBW2992) irreversibly inhibits EGFR/HER2 including EGFR(wt), EGFR(L858R), EGFR(L858R/T790M) and HER2 with IC50 of 0.5 nM, 0.4 nM, 10 nM and 14 nM, respectively. |
| ALK;EGFR | Brigatinib | 1197953-54-0 | AP26113 | Brigatinib, also known as AP-26113, is an orally active, potent and selective dual ALK/EGFR inhibitor. AP26113 is an orally bioavailable inhibitor of anaplastic lymphoma kinase (ALK; IC50 < 100 nM in Ba/F3 cells). |
| EGFR | EAI045 | 1942114-09-1 | EAI045 is an allosteric, non-ATP competitive inhibitor of mutant EGFR. |
Reference
1.Thomas A. Dineen; et al. Discovery of BLU-945, a Reversible, Potent, and Wild-Type-Sparing Next-Generation EGFR Mutant Inhibitor for Treatment-Resistant Non-Small-Cell Lung Cancer. https://doi.org/10.1021/acs.jmedchem.2c00704.
2.https://clinicaltrials.gov/ct2/home.