Design and Biological Evaluation of Piperazine-Bearing Nitrobenzamide Hypoxia/GDEPT Prodrugs: The Discovery of CP-506

Off-target aerobic activation of PR-104A by human aldo-keto reductase 1C3 (AKR1C3) has confounded the development of this dual hypoxia/gene therapy prodrug. Previous attempts to design prodrugs resistant to AKR1C3 activation have resulted in candidates that require further optimization. Herein we report the evaluation of a lipophilic series of PR-104A analogues in which a piperazine moiety has been introduced to improve drug-like properties. Octanol–water partition coefficients (LogD7.4) spanned >2 orders of magnitude. 2D antiproliferative and 3D multicellular clonogenic assays using isogenic HCT116 and H1299 cells confirmed that all examples were resistant to AKR1C3 metabolism while producing an E. coli NfsA nitroreductase-mediated bystander effect. Prodrugs 16, 17, and 20 demonstrated efficacy in H1299 xenografts where only a minority of tumor cells express NfsA. These prodrugs and their bromo/mesylate counterparts (25–27) were also evaluated for hypoxia-selective cell killing in vitro. These results in conjunction with stability assays recommended prodrug 26 (CP-506) for Phase I/II clinical trial

2‑Aminopyrimidine Derivatives as New Selective Fibroblast Growth Factor Receptor 4 (FGFR4) Inhibitors

A series of 2-aminopyrimidine derivatives were designed and synthesized as highly selective FGFR4 inhibitors. One of the most promising compounds <b>2n</b> tightly bound FGFR4 with a <i>K</i>_d value of 3.3 nM and potently inhibited its enzymatic activity with an IC₅₀ value of 2.6 nM, but completely spared FGFR1/2/3. The compound selectively suppressed proliferation of breast cancer cells harboring dysregulated FGFR4 signaling with an IC₅₀ value of 0.38 μM. Furthermore, <b>2n</b> exhibited extraordinary target specificity in a Kinome-wide screen against 468 kinases, with <i>S</i>(35) and <i>S</i>(10) selectivity scores of 0.01 and 0.007 at 1.0 μM, respectively

Discovery of Isoform-Selective Akt3 Degraders Overcoming Osimertinib-Induced Resistance in Non-Small Cell Lung Cancer Cells

EGFR inhibitor therapies have brought significant benefit to NSCLC patients. However, all patients gradually progress to acquired resistance via diverse mechanisms. Akt3 overexpression but not Akt1/2 is one of the found molecular events that mediate osimertinib (1) resistance in NSCLC patients. Here, we report 12l as the first bona fide isoform-selective Akt3 degrader which potently induced proteasomal degradation of the target both in vitro and in vivo, whereas its effects on Akt1/2 were minimal. Using 12l as a tool, non-canonical function of Akt3 was validated to contribute greatly to survival of 1-resistant H1975OR NSCLC cells. Degrader 12l potently suppressed the growth of H1975OR as well as several NSCLC cell lines with low nanomolar IC50 values and demonstrated promising in vivo antitumor efficacy in nude mice bearing H1975OR or PC9 NSCLC xenograft models. Selective degradation of Akt3 may be considered as a novel strategy for human cancer therapy

Discovery of Isoform-Selective Akt3 Degraders Overcoming Osimertinib-Induced Resistance in Non-Small Cell Lung Cancer Cells

EGFR inhibitor therapies have brought significant benefit to NSCLC patients. However, all patients gradually progress to acquired resistance via diverse mechanisms. Akt3 overexpression but not Akt1/2 is one of the found molecular events that mediate osimertinib (1) resistance in NSCLC patients. Here, we report 12l as the first bona fide isoform-selective Akt3 degrader which potently induced proteasomal degradation of the target both in vitro and in vivo, whereas its effects on Akt1/2 were minimal. Using 12l as a tool, non-canonical function of Akt3 was validated to contribute greatly to survival of 1-resistant H1975OR NSCLC cells. Degrader 12l potently suppressed the growth of H1975OR as well as several NSCLC cell lines with low nanomolar IC50 values and demonstrated promising in vivo antitumor efficacy in nude mice bearing H1975OR or PC9 NSCLC xenograft models. Selective degradation of Akt3 may be considered as a novel strategy for human cancer therapy

Discovery of Isoform-Selective Akt3 Degraders Overcoming Osimertinib-Induced Resistance in Non-Small Cell Lung Cancer Cells

EGFR inhibitor therapies have brought significant benefit to NSCLC patients. However, all patients gradually progress to acquired resistance via diverse mechanisms. Akt3 overexpression but not Akt1/2 is one of the found molecular events that mediate osimertinib (1) resistance in NSCLC patients. Here, we report 12l as the first bona fide isoform-selective Akt3 degrader which potently induced proteasomal degradation of the target both in vitro and in vivo, whereas its effects on Akt1/2 were minimal. Using 12l as a tool, non-canonical function of Akt3 was validated to contribute greatly to survival of 1-resistant H1975OR NSCLC cells. Degrader 12l potently suppressed the growth of H1975OR as well as several NSCLC cell lines with low nanomolar IC50 values and demonstrated promising in vivo antitumor efficacy in nude mice bearing H1975OR or PC9 NSCLC xenograft models. Selective degradation of Akt3 may be considered as a novel strategy for human cancer therapy

Discovery of Isoform-Selective Akt3 Degraders Overcoming Osimertinib-Induced Resistance in Non-Small Cell Lung Cancer Cells

EGFR inhibitor therapies have brought significant benefit to NSCLC patients. However, all patients gradually progress to acquired resistance via diverse mechanisms. Akt3 overexpression but not Akt1/2 is one of the found molecular events that mediate osimertinib (1) resistance in NSCLC patients. Here, we report 12l as the first bona fide isoform-selective Akt3 degrader which potently induced proteasomal degradation of the target both in vitro and in vivo, whereas its effects on Akt1/2 were minimal. Using 12l as a tool, non-canonical function of Akt3 was validated to contribute greatly to survival of 1-resistant H1975OR NSCLC cells. Degrader 12l potently suppressed the growth of H1975OR as well as several NSCLC cell lines with low nanomolar IC50 values and demonstrated promising in vivo antitumor efficacy in nude mice bearing H1975OR or PC9 NSCLC xenograft models. Selective degradation of Akt3 may be considered as a novel strategy for human cancer therapy

Discovery of Isoform-Selective Akt3 Degraders Overcoming Osimertinib-Induced Resistance in Non-Small Cell Lung Cancer Cells

EGFR inhibitor therapies have brought significant benefit to NSCLC patients. However, all patients gradually progress to acquired resistance via diverse mechanisms. Akt3 overexpression but not Akt1/2 is one of the found molecular events that mediate osimertinib (1) resistance in NSCLC patients. Here, we report 12l as the first bona fide isoform-selective Akt3 degrader which potently induced proteasomal degradation of the target both in vitro and in vivo, whereas its effects on Akt1/2 were minimal. Using 12l as a tool, non-canonical function of Akt3 was validated to contribute greatly to survival of 1-resistant H1975OR NSCLC cells. Degrader 12l potently suppressed the growth of H1975OR as well as several NSCLC cell lines with low nanomolar IC50 values and demonstrated promising in vivo antitumor efficacy in nude mice bearing H1975OR or PC9 NSCLC xenograft models. Selective degradation of Akt3 may be considered as a novel strategy for human cancer therapy

Discovery of 6‑Formylpyridyl Urea Derivatives as Potent Reversible-Covalent Fibroblast Growth Factor Receptor 4 Inhibitors with Improved Anti-Hepatocellular Carcinoma Activity

Fibroblast growth factor receptor 4 (FGFR4) has been considered as a potential anticancer target due to FGF19/FGFR4 mediated aberrant signaling in hepatocellular carcinoma (HCC). Several FGFR4 inhibitors have been reported, but none have gained approval. Herein, a series of 5-formyl-pyrrolo[3,2-b]pyridine-3-carboxamides and a series of 6-formylpyridyl ureas were characterized as selective reversible-covalent FGFR4 inhibitors. The representative 6-formylpyridyl urea 8z exhibited excellent potency against FGFR4WT, FGFR4V550L, and FGFR4V550M with IC50 values of 16.3, 12.6, and 57.3 nM, respectively. It also potently suppressed proliferation of Ba/F3 cells driven by FGFR4WT, FGFR4V550L, and FGFR4V550M, and FGFR4-dependent Hep3B and Huh7 HCC cells, with IC50 values of 1.2, 13.5, 64.5, 15.0, and 20.4 nM, respectively. Furthermore, 8z displayed desirable microsomal stability and significant in vivo efficacy in the Huh7 HCC cancer xenograft model in nude mice. The study provides a promising new lead for anticancer drug discovery directed toward overcoming FGFR4 gatekeeper mutation mediated resistance in HCC patients

Discovery of 6‑Formylpyridyl Urea Derivatives as Potent Reversible-Covalent Fibroblast Growth Factor Receptor 4 Inhibitors with Improved Anti-Hepatocellular Carcinoma Activity

Fibroblast growth factor receptor 4 (FGFR4) has been considered as a potential anticancer target due to FGF19/FGFR4 mediated aberrant signaling in hepatocellular carcinoma (HCC). Several FGFR4 inhibitors have been reported, but none have gained approval. Herein, a series of 5-formyl-pyrrolo[3,2-b]pyridine-3-carboxamides and a series of 6-formylpyridyl ureas were characterized as selective reversible-covalent FGFR4 inhibitors. The representative 6-formylpyridyl urea 8z exhibited excellent potency against FGFR4WT, FGFR4V550L, and FGFR4V550M with IC50 values of 16.3, 12.6, and 57.3 nM, respectively. It also potently suppressed proliferation of Ba/F3 cells driven by FGFR4WT, FGFR4V550L, and FGFR4V550M, and FGFR4-dependent Hep3B and Huh7 HCC cells, with IC50 values of 1.2, 13.5, 64.5, 15.0, and 20.4 nM, respectively. Furthermore, 8z displayed desirable microsomal stability and significant in vivo efficacy in the Huh7 HCC cancer xenograft model in nude mice. The study provides a promising new lead for anticancer drug discovery directed toward overcoming FGFR4 gatekeeper mutation mediated resistance in HCC patients

Tyrosine Kinase Inhibitors. 20. Optimization of Substituted Quinazoline and Pyrido[3,4‑<i>d</i>]pyrimidine Derivatives as Orally Active, Irreversible Inhibitors of the Epidermal Growth Factor Receptor Family

Structure–activity relationships for inhibition of erbB1, erbB2, and erbB4 were determined for a series of quinazoline- and pyrido­[3,4-<i>d</i>]­pyrimidine-based analogues of the irreversible pan-erbB inhibitor, canertinib. Cyclic amine bearing crotonamides were determined to provide rapid inhibition of cellular erbB1 autophosphorylation and good metabolic stability in liver microsome and hepatocyte assays. The influence of 4-anilino substitution on pan-erbB inhibitory potency was investigated. Several anilines were identified as providing potent, reversible pan-erbB inhibition. Optimum 4- and 6-substituents with known 7-substituents provided preferred irreversible inhibitors for pharmacodynamic testing in vivo. Quinazoline <b>54</b> and pyrido­[3,4-<i>d</i>]­pyrimidine <b>71</b> were identified as clearly superior to canertinib. Both compounds possess a piperidinyl crotonamide Michael acceptor and a 3-chloro-4-fluoroaniline, indicating these as optimized 6- and 4-substituents, respectively. Pharmacokinetic comparison of compounds <b>54</b> and <b>71</b> across three species selected compound <b>54</b> as the preferred candidate. Compound <b>54</b> (PF-00299804) has been assigned the nomenclature of dacomitinib and is currently under clinical evaluation