Potent EGFR Inhibitors Research Articles

Design, synthesis and evaluation of the osimertinib analogue (C-005) as potent EGFR inhibitor against NSCLC.

Osimertinib has been approved as a first-line treatment for non-small-cell lung cancer (NSCLC) patients whose tumor carries EGFR activation and / or resistant mutations. To mitigate Osimertinib's toxicity caused by AZ5104, the N-demethylation metabolite of Osimertinib, we designed and synthesized a series of Osimertinib analogs with different headpieces. In vitro and in vivo analysis rendered a potential clinical candidate C-005 which had pyrrolo-pyridine headpiece. Biochemically, C-005 and its main human hepatocyte metabolite showed over 30 fold selectivity of L858R/T790M mutant EGFR over WT EGFR. Such selectivity profile was retained at cellular level. In general, C-005 is 2-14 fold more selective than Osimertinib in a panel of WT EGFR cancer cell lines. Furthermore, C-005 demonstrated robust antitumor efficacy and good tolerability in NCI-H1975, PC-9 and HCC827 xenograft mouse models, making it a potential candidate for human test in clinical.

Abstract 825: Identification of molecular therapeutic targets that enhance the antitumor activity of neratinib in breast cancer cells

Abstract Background: Aberrant expression of members of the ErbB family of receptor tyrosine kinases and its downstream pathways is known to play pivotal roles in breast cancer malignancy. HER2-positive breast cancer has an aggressive nature that is driven by HER2 gene amplification. Inflammatory breast cancer is a rare but the most aggressive clinical subtype of breast cancer. Triple-negative breast cancer is an aggressive receptor subtype of breast cancer. More than 30% of cases of each of these groups express EGFR. Emerging therapeutic resistance continues to be a challenge in recurrent and/or metastatic disease. Neratinib is a potent irreversible EGFR, HER2, and HER4 tyrosine kinase inhibitor that blocks signal transduction generated via these receptors. In this study, we sought to identify the optimal targets in different subtypes of breast cancer and synergistic partners to maximize the antitumor effect of neratinib against breast cancer cells in vitro and in vivo. Methods: Expression of EGFR and HER2 was assessed via Western blot in 31 breast cancer cell lines, including HER2-targeted drug-resistant cell lines (14 triple-negative breast cancer and 10 HER2-positive). The antiproliferative effects of neratinib were measured using cell proliferation and anchorage-independent colony-formation (soft agar) assays. A reverse-phase protein array was used to profile and validate the signaling networks induced by neratinib-based treatment. To identify the synergistic targets, RNA interference screening of a 779-kinome library was performed under multicellular tumor spheroid culture conditions. Results: In vitro proliferation data demonstrated that neratinib had a nanomolar range of half-maximal inhibitory concentrations in most of the tested cell lines, including T-DM1-resistant HER-2 breast cancer cell lines. Neratinib inhibited colony formation in a dose-dependent manner (P = 0.0001). Reverse-phase protein array data revealed that the efficacy of neratinib correlated with phosphorylated EGFR (R2 = 0.6055) or phosphorylated HER2 (R2 = 0.6734) expression level. Statistical analysis identified the 25 most relevant targets in the RNA interference screening, which we will validate for combinational antitumor effects with neratinib in in vitro and in vivo studies. Conclusion: We determined the efficacy of neratinib and identified target molecules to enhance neratinib efficacy in different breast cancer subtype cell lines with ErbB alterations. Our data inform new combined treatment strategies with neratinib for breast cancer that will lead to innovative designs using optimized neratinib combinations in clinical trials. Citation Format: Bora Lim, Jangsoon Lee, Troy Pearson, Liu Huey, Minha Hwang, Kuicheon Choi, Francesca Avogadri-Connors, Richard E. Cutler, Debu Tripathy, Naoto T. Ueno. Identification of molecular therapeutic targets that enhance the antitumor activity of neratinib in breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 825.

Design, synthesis, antiproliferative activity, molecular docking and cell cycle analysis of some novel (morpholinosulfonyl) isatins with potential EGFR inhibitory activity

New series of 5-(morpholinosulfonyl) isatin derivatives were designed and synthesized. The new compounds were characterized on the basis of spectral and elemental analyses. They were examined for their cytotoxic effects using SRB assay on four cancer cell lines HepG2, HCT116, CACO and MCF-7 in addition to the non-cancerous human cell line. They were non cytotoxic towards the normal derived cell line (IC50 value > 130 μM). Compounds 3, 6, 10 and 11 showed IC50 values less than 10 μM on three of the tested cell lines HepG2, HCT116 and CACO. Compounds 2h, 5, and 7b showed IC50 values less than or nearly equal 10 μM on HepG2, CACO and HCT116 respectively. Compounds 3 and 6 revelaed IC50 values less than 12 μM on MCF7. These obtained IC50 values are comparable with that of doxrubicin as it has showed IC50 range from 4.5 to 8.28 μM on the tested cell lines. All these promising derivatives showed inhibitory activity against EGFR with IC50 values less than 2 μM. The most potent EGFR inhibitors 7b (IC50 = 46 nM) and 10 (IC50 = 23 nM) showed to cause cell cycle arrest at G2/M phase and induce apoptosis. Molecular docking studies also were simulated to put insight and make better understanding to their structural features.

Design, synthesis, antiproliferative activity and docking studies of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline as potential EGFR inhibitors

Eight series of quinazoline derivatives bearing 2,3-dihydro-indole or 1,2,3,4-tetrahydroquinoline were designed, synthesized and evaluated for the IC50 values against three cancer cell lines (A549, MCF-7 and PC-3). Most of the forty nine target compounds showed excellent antiproliferative activity against one or several cancer cell lines. The compound 13a showed the best activity against A549, MCF-7 and PC-3 cancer cell lines, with the IC50 values of 1.09 ± 0.04 μM, 1.34 ± 0.13 μM and 1.23 ± 0.09 μM, respectively. Eight selected compounds were further selected to evaluated for the inhibitory activity against EGFR kinase. Three of them showed equal activity against EGFR kinase to positive control afatinib. AnnexinV-FITC, propidium iodide (PI) double staining and acridine orange single staining results indicated that the compound 13a could induce apoptosis of human lung cancer A549 cells.

Synthesis and evaluation of 2,9-disubstituted 8-phenylthio/phenylsulfinyl-9H-purine as new EGFR inhibitors

In present study, we described the synthesis and biological evaluation of a new class of EGFR inhibitors containing 2,9-disubstituted 8-phenylthio/phenylsulfinyl-9H-purine scaffold. Thirty-one compounds were synthesized. Among them, compound C9 displayed the IC50 of 29.4 nM against HCC827 cell line and 1.9 nM against EGFRL858R. Compound C12 showed moderate inhibitory activity against EGFRL858R/T790M/C797S (IC50 = 114 nM). Western bolt assay suggested that compound C9 significantly inhibited EGFR phosphorylation. In vivo test, compound C9 remarkably exhibited inhibitory effect on tumor growth at 5.0 mg/kg by oral administration in established nude mouse HCC827 xenograft model. These results indicate that the 2,9-disubstituted 8-phenylsulfinyl/phenylsulfinyl-9H-purine derivatives can act as potent EGFR(L858R) inhibitors and effective anticancer agents. Additionally, optimization of compound C12 may result in discovering the fourth-generation EGFR-TKIs.

Design, synthesis and anticancer evaluation of novel spirobenzo[h]chromene and spirochromane derivatives with dual EGFR and B-RAF inhibitory activities

A novel series of spirobenzo[h]chromene and spirochromane derivatives was designed, synthesized and evaluated as potential anticancer agents against MCF-7 (human breast carcinoma), HT-29 (human colorectal adenocarcinoma) and A549 (human lung carcinoma) cell lines using MTT assay. Eight compounds 7, 8e, 13a-e and 16 showed a better anticancer activity than that of sorafenib, the multi-kinase inhibitor with IC50 values between 1.78 and 5.47 μM or erlotinib with IC50 values over 20 μM. Representative compounds 8e, 13c and 16 were selected for further mechanistic investigation via EGFR, B-RAF and tubulin polymerization assays. Compound 16 was the most potent EGFR inhibitor (IC50 = 1.2 μM), yet compounds 8e, 13c and 16 displayed moderate tubulin polymerization inhibition effects. Molecular docking studies of those compounds revealed their possible binding modes into the active sites of both EGFR and B-RAF kinases. The newly developed compounds represent a therapeutically promising approach for the treatment of different types of cancer.

Novel quinazoline derivatives bearing various 6-benzamide moieties as highly selective and potent EGFR inhibitors

A series of novel quinazoline derivatives bearing various C-6 benzamide substituents were synthesized and evaluated as EGFR inhibitors, and most showed significant inhibitory potency against EGFR kinase. In particular, compound 6g possessed potent inhibitory activity against EGFR wild-type (IC50 = 5 nM), and strong antiproliferative activity against HCC827 and Ba/F3 (L858R) cell lines. Kinase profiling against a panel of 365 kinases showed that 6g was highly selective for EGFR. Furthermore, 6g showed desirable properties in assays of liver microsome metabolic stability and cytochromes P450 inhibition and preliminary pharmacokinetic study. The overall attractive profile of 6g made it an interesting compound for further development.

Design, synthesis and biological evaluation of novel 4-aminoquinazolines as dual target inhibitors of EGFR-PI3Kα

The overexpression of EGFR correlates with rapidly progressive disease, resistance to chemotherapy and poor prognosis. In certain human cancers, PI3K works synergistically with EGFR to promote proliferation, survival, invasion and metastasis. Development of dual-target drugs against EGFR and PI3K has therapeutic advantage and was an attractive approach against tumors. In this work, based on the molecular docking and previous studies, a series of 4-aminoquinazolines derivatives containing 6-sulfonamide substituted pyridyl group were rationally designed and identified as potent EGFR and PI3K dual inhibitors. The cytotoxicity experiment results showed that this series of compounds could effectively inhibit cell growth. The kinase assay demonstrated that 6c and 6i had high inhibition for EGFR and selectivity for PI3Kα distinguished from other isoforms. Further experiments showed that 6c could induce cell cycle arrest in G1 phase and apoptosis in BT549 cells. The western blot assay indicated that 6c inhibited the proliferation of BT549 cell through EGFR and PI3Kα/Akt signaling pathway. Our study suggested that compound 6c was a potential dual inhibitors of EGFR and PI3Kα.

Design, synthesis, antiproliferative activity and docking studies of quinazoline derivatives bearing oxazole or imidazole as potential EGFR inhibitors

Six series of quinazoline derivatives bearing oxazole or imidazole (8a–f, 9a–f, 10a–d, 11a–f, 12a–d and 13a–i) were designed, synthesized and their IC50 values evaluated against three cancer cell lines (A549, MCF-7 and PC-3).

In silico identification of potent small molecule inhibitors targeting epidermal growth factor receptor 1

The receptor tyrosine kinase of the epidermal growth factor receptor (EGFR, ErbB) family played an important role in multisignaling pathways, which controlled numerous biological activities including proliferation, differentiation, apoptosis, etc. EGFR abnormalities have been associated with a variety of human tumors, which was a well-characterized target for cancer treatment. It was known to all that drug repositioning has been considered as a useful tool to accelerate the process of drug development. Herein, a total of 1408 small molecule drugs approved by the Food and Drug Administration (FDA) were employed to identify potential EGFR inhibitors by a series of bioinformatics approaches, including virtual screening and molecular dynamics (MD) simulations. According to the docking score, five small molecules were chosed for further MD simulations. Following the 5 ns MD simulations, ZINC03830276 (Benzonatate) were finally recognized as "new use" of FDA-approved EGFR-targeting drug. Our findings suggested that the small molecule ZINC03830276 (Benzonatate) could be a promising EGFR inhibitor candidate and may also provide new ideas for designing more potent EGFR inhibitors for the future study.

Discovery of Potent EGFR Inhibitors through the Incorporation of a 3D-Aromatic-Boron-Rich-Cluster into the 4-Anilinoquinazoline Scaffold: Potential Drugs for Glioma Treatment.

New 1,7-closo-carboranylanilinoquinazoline hybrids have been identified as EGFR inhibitors, one of them with higher affinity than the parent compound erlotinib. The comparative docking analysis with compounds bearing bioisoster-substructures, demonstrated the relevance of the 3D aromatic-boron-rich moiety for interacting into the EGFR ATP binding region. The capability to accumulate in glioma cells, the ability to cross the blood-brain barrier and the stability on simulated biological conditions, render these molecules as lead compounds for further structural modifications to obtain dual action drugs to treat glioblastoma.

Novel quinoline-3-carboxamides (Part 2): Design, optimization and synthesis of quinoline based scaffold as EGFR inhibitors with potent anticancer activity

EGFR has a key role in cell growth. Its mutation and overexpression share in epithelial malignancies and tumor growth. Quinazoline and quinoline derivatives are common anticancer intracellular inhibitors of EGFR kinase, and their optimization is an important issue for development of potent targeted anticancer agents. Based on these facts, different strategies were used for optimizing our reported quinoline-3-carboxamide compound III (EGFR IC50 = 5.283 µM and MCF-7 IC50 = 3.46 µM) through different molecular modeling techniques. The optimized compounds were synthesized and subjected to EGFR binding assay and accordingly some more potent inhibitors were obtained. The most potent quinoline-3-carboxamides were the furan derivative 5o; thiophene derivative 6b; and benzyloxy derivative 10 showing EGFR IC50 values 2.61, 0.49 and 1.73 μM, respectively. Furthermore, the anticancer activity of compounds eliciting potent EGFR inhibition (5o, 5p, 6b, 8a, 8b, and 10) was evaluated against MCF-7 cell line where they exhibited IC50 values 3.355, 3.647, 5.069, 3.617, 0.839 and 10.85 μM, respectively. Compound 6b was selected as lead structure for further optimization hoping to produce more potent EGFR inhibitors.

Hit discovery of 4-amino-N-(4-(3-(trifluoromethyl)phenoxy)pyrimidin-5-yl)benzamide: A novel EGFR inhibitor from a designed small library

Searching for hit compounds within the huge chemical space resembles the attempt to find a needle in a haystack. Cheminformatics-guided selection of few representative molecules of a rationally designed virtual combinatorial library is a powerful tool to confront this challenge, speed up hit identification and cut off costs. Herein, this approach has been applied to identify hit compounds with novel scaffolds able to inhibit EGFR kinase. From a generated virtual library, six 4-aryloxy-5-aminopyrimidine scaffold-derived compounds were selected, synthesized and evaluated as hit EGFR inhibitors. 4-Aryloxy-5-benzamidopyrimidines inhibited EGFR with IC50 1.05–5.37 μM. Cell-based assay of the most potent EGFR inhibitor hit (10ac) confirmed its cytotoxicity against different cancerous cells. In spite of no EGFR, HER2 or VEGFR1 inhibition was elicited by 4-aryloxy-5-(thio)ureidopyrimidine derivatives, cell-based evaluation suggested them as antiproliferative hits acting by other mechanism(s). Molecular docking study provided a plausible explanation of incapability of 4-aryloxy-5-(thio)ureidopyrimidines to inhibit EGFR and suggested a reasonable binding mode of 4-aryloxy-5-benzamidopyrimidines which provides a basis to develop more optimized ligands.

Synthesis and evaluation of osimertinib derivatives as potent EGFR inhibitors

Osimertinib has been identified as a promising therapeutic drug targeting for EGFR T790M mutant non-small cell lung cancer (NSCLC). A new series of N-oxidized and fluorinated osimertinib derivatives were designed and synthesized. The cellular anti-proliferative activity, kinase inhibitory activity and the activation of EGFR signaling pathways of 1–6 in vitro were determined against L858R/T790M and wild-type EGFR, the antitumor efficacy in NCI-H1975 xenografts in vivo were further studied. Compound 2, the newly synthesized N-oxide metabolite in N,N,N′-trimethylethylenediamine side chain of osimertinib, showed a comparable kinase selectivity in vitro and a slightly better antitumor efficacy in vivo to osimertinib, making it valuable and suitable for the potential lung cancer therapy.

Design, synthesis and cytotoxic evaluation of a novel series of benzo[d]thiazole-2-carboxamide derivatives as potential EGFR inhibitors

A novel series of benzo[d]thiazole-2-carboxamide derivatives have been de novo designed based on virtual screening methods. The target compounds were synthesized and evaluated for the cytotoxicity against epidermal growth factor receptor high-expressed cancer cell lines (A549, HeLa, and SW480), epidermal growth factor receptor low-expressed cell line (HepG2) and human liver normal cell line (HL7702). Several target compounds have showed moderate to excellent potency against A549, HeLa, and SW480 and weak cytotoxic effects against HepG2, which implies they are probably epidermal growth factor receptor inhibitors. And scarcely did they exhibit any activities against HL7702, which signifies they are likely to overcome the nonspecific toxicity against normal cells. Especially, the compound 6-[2-(diethylamino)-2-oxoethoxy]-N-(furan-2-ylmethyl)benzo[d]thiazole-2-carboxamide (6i) was identified as a promising agent, exhibiting the most potent cytotoxic activities with IC50 values of 4.05, 12.17, 6.76 μM against the A549, HeLa, and SW480 cell lines, respectively.

Trisubstituted Imidazoles with a Rigidized Hinge Binding Motif Act As Single Digit nM Inhibitors of Clinically Relevant EGFR L858R/T790M and L858R/T790M/C797S Mutants: An Example of Target Hopping.

The high genomic instability of non-small cell lung cancer tumors leads to the rapid development of resistance against promising EGFR tyrosine kinase inhibitors (TKIs). A recently detected triple mutation compromises the activity of the gold standard third-generation EGFR inhibitors. We have prepared a set of trisubstituted imidazoles with a rigidized 7-azaindole hinge binding motif as a new structural class of EGFR inhibitors by a target hopping approach from p38α MAPK inhibitor templates. On the basis of an iterative approach of docking, compound preparation, biological testing, and SAR interpretation, robust and flexible synthetic routes were established. As a result, we report two reversible inhibitors 11d and 11e of the clinically challenging triple mutant L858R/T790M/C797S with IC50 values in the low nanomolar range. Furthermore, we developed a kinome selective irreversible inhibitor 45a with an IC50 value of 1 nM against the EGFR L858R/T790M double mutant. Target binding kinetics and metabolic stability data are included. These potent mutant EGFR inhibitors may serve as a basis for the development of structurally novel EGFR probes, tools, or candidates.

Design, synthesis and evaluation of azaacridine derivatives as dual-target EGFR and Src kinase inhibitors for antitumor treatment

Overexpression of EGFR is often associated with advanced stage disease and poor prognosis. In certain cancers, Src works synergistically with EGFR to promote proliferation, survival, invasion and metastasis. Development of dual-target drugs against EGFR and Src is of therapeutic advantage against these cancers. Based on molecular docking and our previous studies, we rationally designed a new series of azaacridine derivatives as potent EGFR and Src dual inhibitors. Most of the synthesized azaacridines displayed good antiproliferative activity against K562 and A549 cells. The representative compound 13b showed nM IC50 values against K562 and A549 cells, and inhibited EGFR at inhibition rate of 33.53% at 10 μM and Src at inhibition rate of 72.12% at 1 μM. Furthermore, compound 13b could inhibit the expression of EGFR, p-EGFR, Src and p-Src. Moreover, 13b efficiently inhibited the invasion of tumor cells and induced cancer cells apoptosis. Our study suggested that azaacridine scaffold can be developed as novel multi-target kinase inhibitors for cancer therapy.

Design, synthesis, and docking studies of quinazoline analogues bearing aryl semicarbazone scaffolds as potent EGFR inhibitors

Two series of quinazoline derivatives bearing aryl semicarbazone scaffolds (9a–o and 10a–o) were designed, synthesized and evaluated for the IC50 values against four cancer cell lines (A549, HepG2, MCF-7 and PC-3). The selected compound 9o was further evaluated for the inhibitory activity against EGFR kinases. Four of the compounds showed excellent cytotoxicity activity and selectivity with the IC50 values in single-digit μM to nanomole range. Two of them are equal to more active than positive control afatinib against one or more cell lines. The most promising compound 9o showed the best activity against A549, HepG2, MCF-7 and PC-3 cancer cell lines and EGFR kinase, with the IC50 values of 1.32±0.38μM, 0.07±0.01μM, 0.91±0.29μM and 4.89±0.69μM, which were equal to more active than afatinib (1.40±0.83μM, 1.33±1.28μM, 2.63±1.06μM and 3.96±0.59μM), respectively. Activity of the most promising compound 9o (IC50 56nM) against EGFR kinase was slightly lower to the positive compound afatinib (IC50 1.6nM) but more active than reference staurosporine (IC50 238nM). The result of flow cytometry, with the dose of compound 9o increasing, which indicated the compound 9o could induce remarkable apoptosis of A549 and cells in a dose dependent manner. Structure–activity relationships (SARs) and docking studies indicated that replacement of the cinnamamide group by aryl semicarbazone scaffolds slightly decreased the anti-tumor activity. The results suggested that hydroxy substitution at C-4 had a significant impact on the activity and replacement of the tetrahydrofuran group by methyl moiety was not beneficial for the activity.

Discovery and characterization of a novel irreversible EGFR mutants selective and potent kinase inhibitor CHMFL-EGFR-26 with a distinct binding mode.

EGFR T790M mutation accounts for about 40-55% drug resistance for the first generation EGFR kinase inhibitors in the NSCLC. Starting from ibrutinib, a highly potent irreversible BTK kinase inhibitor, which was also found to be moderately active to EGFR T790M mutant, we discovered a highly potent irreversible EGFR inhibitor CHMFL-EGFR-26, which is selectively potent against EGFR mutants including L858R, del19, and L858R/T790M. It displayed proper selectivity window between the EGFR mutants and the wide-type. CHMFL-EGFR-26 exhibited good selectivity profile among 468 kinases/mutants tested (S score (1)=0.02). In addition, X-ray crystallography revealed a distinct “DFG-in” and “cHelix-out” inactive binding mode between CHMFL-EGFR-26 and EGFR T790M protein. The compound showed highly potent anti-proliferative efficacy against EGFR mutant but not wide-type NSCLC cell lines through effective inhibition of the EGFR mediated signaling pathway, induction of apoptosis and arresting of cell cycle progression. CHMFL-EGFR-26 bore acceptable pharmacokinetic properties and demonstrated dose-dependent tumor growth suppression in the H1975 (EGFR L858R/T790M) and PC-9 (EGFR del19) inoculated xenograft mouse models. Currently CHMFL-EGFR-26 is undergoing extensive pre-clinical evaluation for the clinical trial purpose.

P2.03b-016 Tesevatinib in NSCLC Patients with EGFR Activating Mutations and Brain Metastases (BM) or Leptomeningeal Metastases (LM): Topic: Brain Meta

Tesevatinib is a potent reversible EGFR inhibitor with strong preclinical evidence of brain penetration: brain:plasma ratios of 1-4 and brain:meninges ratios of 10-15 in rodents (AACR 2015 Abstract 2590). Tesevatinib was previously shown to have significant clinical activity in patients presenting with EGFR mutant NSCLC, but not in patients with T790M mutation. Approximately 25% of patients with EGFR activating mutations progress in the CNS, and metastases there have a low rate (10%) of T790M mutations. Patients with NSCLC driven by activating EGFR mutations who had BM or LM occurring or progressing while receiving erlotinib, gefitinib, or afatinib were treated with 300 mg of tesevatinib daily. Patients with BM had RECIST 1.1 measurable disease in the brain, and RECIST 1.1 evaluated response. Patients with symptomatic LM were diagnosed by either CSF cytology or MRI findings. Response was measured by improvement in symptoms, CSF cytology, and MRI. Patients with both BM and symptomatic LM were enrolled in the LM cohort. Target accrual is 20 patients in each cohort. To date, 7 patients have been enrolled [2M:5F; median age 61 (36-66); 1 Asian], all with CNS symptoms. Four were in the BM cohort and 3 in the LM cohort. All had prior CNS radiotherapy, either WBRT or SRS or both. All had prior systemic therapy (median 3; range 1-6). Three patients had EGFR del 19, 3 had L858R, and 1 had L861Q. Gr ≥ 3 adverse events, regardless of relationship to study drug, have included Gr 3 prolonged QTc, Grade 3 hypokalemia, Gr 3 dehydration, Gr 3 UTI, and Gr 3 ALT elevation. Three patients had dose reductions due to asymptomatic QTc interval prolongation. Six out of the 7 patients had CNS symptom improvement, often occurring within 14 days of tesevatinib initiation. Two patients decreased steroids. One BM patient had marked improvement in right leg strength and a 19% reduction in the target BM on Study Day 23. One patient with BM and LM had resolution of LM symptoms, a 57% reduction in BM target lesion, and clearance of LM enhancement on MRI at Study Day 41. Early data from the first 7 patents in this ongoing clinical trial indicate that tesevatinib has clinical activity in the CNS in EGFR mutant disease manifesting as BM or LM in patients previously treated with erlotinib, gefitinib, or afatinib. An additional cohort of 20 treatment-naïve patients who have initial presentation with brain metastases is being added.