HYL001 is an orally active, selective ALK4/ALK5 inhibitor with K d values of 13 nM and 22.5 nM for ALK4 and ALK5, respectively. HYL001 inhibits GLS expression. HYL001 impairs glutamine metabolism, mitochondrial respiration, the TCA cycle, oxidative phosphorylation, ATP production, and redox balance, while selectively targeting mitochondria, disrupting mitochondrial structure, and inducing Apoptosis and ROS. HYL001 inhibits tumor growth and liver metastasis. HYL001 can be used in research on breast cancer, hepatocellular carcinoma, and microsatellite-stable colorectal cancer metastasis [1] [2] [3] [4] [5] [6] [7].
In Vivo, HYL001 (5 μmol/kg; s.c.; once every three days; five injections) inhibits subcutaneous 4T1 tumor growth and reduces breast cancer stem cell populations in BALB/c mice[2]. HYL001 (5 μmol/kg; s.c.; once every three days; five injections) suppresses lung metastasis and prolongs survival in the metastatic 4T1 tumor model in BALB/c mice[2]. HYL001 (10 μmol/kg; 10 μmol/kg plus PTX 20 μmol/kg; i.v.; once every three days; five times) inhibits orthotopic 4T1 tumor growth, blocks glutamine metabolism, and reduces cancer stem cell populations in BALB/c mice[2]. HYL001 (10 μmol/kg; i.v.; every 2 days; 5 doses) inhibits orthotopic 4T1 tumor growth in female BALB/c mice, with ~28% tumor inhibition by weight, ~57% reduction in intratumoral Glu/Gln ratio, reduced GLS expression, and decreased CD133 CSC staining[6].
In Vitro, HYL001 downregulates GLS expression in 4T1 breast cancer cells[1]. HYL001 exhibits marginal inhibitory activity against purified recombinant GLS[2]. HYL001 displayed IC50 >20 µM in normal NIH-3T3 and HUVEC cells, and IC50 <5 µM in tumor cells[2]. HYL001 (1-10 µM) inhibited 4T1 TRC colony formation, with 100% inhibition at 10 µM, and inhibited preformed 4T1 TRC colonies by 60% at 10 µM[2]. HYL001 (1-10 µM) inhibits HCSC with an IC50 340-fold lower than that of LND, and inhibits H22 TRC colony growth at 1 µM, with an inhibition rate of 58% at 10 µM[2]. HYL001 inhibits breast CSC and HCC stem cells, with IC50 values 380-fold and 340-fold lower than Lonidamine, respectively[4]. HYL001 (1 µM; 12 h) damaged the mitochondria of nearly 100% of BCSCs[2]. HYL001 (5 µM; 12 h) reduced ATP in BCSCs by nearly 100%[2]. HYL001 (1-5 µM; 12 h) induced apoptosis in BCSCs, with apoptosis rates of 30%, 32%, and 55%, respectively[2]. HYL001 (1 µM; 12 h) increases Gln and inhibits the Glu/Gln ratio in BCSCs[2]. HYL001 (1-5 µM; 4 h) enhances mitochondrial ROS in BCSCs, with a 3-fold increase at 5 µM[2]. HYL001 inhibits GLS, reduces GSH, enhances ROS, and impairs the TCA cycle/OXPHOS in 4T1 and H22 cancer cells[4]. HYL001 (0.3-10 μM) is a potent ALK4/ALK5 inhibitor with a Kd of 22.5 nM for ALK5, and shows limited off-target binding at 300 nM, primarily binding to p38α[5]. HYL001 (0.1-10,000 nM; 16 hours) inhibits TGFβ pathway activity in HEK293T reporter cells with an IC50 of 72 nM[5]. HYL001 is a potent inhibitor of ALK4 and ALK5, with Kd values of 13 nM and 22.5 nM, respectively, in cell-free competitive binding assays[7]. HYL001 inhibited ALK5 with an IC50 of 177 nM and p38α with an IC50 of 548 nM in recombinant kinase activity assays[7]. HYL001 (1-320 µg/mL) showed no mutagenic potential in S. typhimurium strains TA98, TA100, TA1535, and TA1537 with or without S9[7].
References:
[1]. Zou W, et al. Targeting glutamine metabolism as a potential target for cancer treatment. Journal of experimental & clinical cancer research: CR. 2025 Jul 01;44(1):180.
[2]. Wang Q, et al. A novel lonidamine derivative targeting mitochondria to eliminate cancer stem cells by blocking glutamine metabolism. Pharmacological research. 2023 Apr;190:106740.
[3]. Fan Z, et al. Boosting oxidative stress with hydroxyethyl starch smart nanomedicines to eliminate cancer stem cells. Accounts of Materials Research. 2024 Oct 24;5(12):1558-70.
[4]. De Los Santos-Jiménez J, et al. GLS and GLS2 Glutaminase Isoenzymes in the Antioxidant System of Cancer Cells. Antioxidants (Basel, Switzerland). 2024 Jun 20;13(6):745.
[5]. Tauriello DVF, et al. New Potent Inhibitor of Transforming Growth Factor-Beta (TGFβ) Signaling that is Efficacious against Microsatellite Stable Colorectal Cancer Metastasis in Combination with Immune Checkpoint Therapy in Mice. ACS pharmacology & translational science. 2025 Jan 10;8(1):97-112.
[6]. Wang Q, et al. Glutaminolysis inhibition boosts photodynamic therapy to eliminate cancer stem cells. Biomaterials. 2024 Apr;306:122497.
[7]. Tauriello DV, et al. HYL001, a new potent TGFβ signaling inhibitor that is efficacious against microsatellite stable CRC metastasis in combination with immune checkpoint therapy in mice. bioRxiv. 2024 May 14:2024-05.
In Vivo, HYL001 (5 μmol/kg; s.c.; once every three days; five injections) inhibits subcutaneous 4T1 tumor growth and reduces breast cancer stem cell populations in BALB/c mice[2]. HYL001 (5 μmol/kg; s.c.; once every three days; five injections) suppresses lung metastasis and prolongs survival in the metastatic 4T1 tumor model in BALB/c mice[2]. HYL001 (10 μmol/kg; 10 μmol/kg plus PTX 20 μmol/kg; i.v.; once every three days; five times) inhibits orthotopic 4T1 tumor growth, blocks glutamine metabolism, and reduces cancer stem cell populations in BALB/c mice[2]. HYL001 (10 μmol/kg; i.v.; every 2 days; 5 doses) inhibits orthotopic 4T1 tumor growth in female BALB/c mice, with ~28% tumor inhibition by weight, ~57% reduction in intratumoral Glu/Gln ratio, reduced GLS expression, and decreased CD133 CSC staining[6].
In Vitro, HYL001 downregulates GLS expression in 4T1 breast cancer cells[1]. HYL001 exhibits marginal inhibitory activity against purified recombinant GLS[2]. HYL001 displayed IC50 >20 µM in normal NIH-3T3 and HUVEC cells, and IC50 <5 µM in tumor cells[2]. HYL001 (1-10 µM) inhibited 4T1 TRC colony formation, with 100% inhibition at 10 µM, and inhibited preformed 4T1 TRC colonies by 60% at 10 µM[2]. HYL001 (1-10 µM) inhibits HCSC with an IC50 340-fold lower than that of LND, and inhibits H22 TRC colony growth at 1 µM, with an inhibition rate of 58% at 10 µM[2]. HYL001 inhibits breast CSC and HCC stem cells, with IC50 values 380-fold and 340-fold lower than Lonidamine, respectively[4]. HYL001 (1 µM; 12 h) damaged the mitochondria of nearly 100% of BCSCs[2]. HYL001 (5 µM; 12 h) reduced ATP in BCSCs by nearly 100%[2]. HYL001 (1-5 µM; 12 h) induced apoptosis in BCSCs, with apoptosis rates of 30%, 32%, and 55%, respectively[2]. HYL001 (1 µM; 12 h) increases Gln and inhibits the Glu/Gln ratio in BCSCs[2]. HYL001 (1-5 µM; 4 h) enhances mitochondrial ROS in BCSCs, with a 3-fold increase at 5 µM[2]. HYL001 inhibits GLS, reduces GSH, enhances ROS, and impairs the TCA cycle/OXPHOS in 4T1 and H22 cancer cells[4]. HYL001 (0.3-10 μM) is a potent ALK4/ALK5 inhibitor with a Kd of 22.5 nM for ALK5, and shows limited off-target binding at 300 nM, primarily binding to p38α[5]. HYL001 (0.1-10,000 nM; 16 hours) inhibits TGFβ pathway activity in HEK293T reporter cells with an IC50 of 72 nM[5]. HYL001 is a potent inhibitor of ALK4 and ALK5, with Kd values of 13 nM and 22.5 nM, respectively, in cell-free competitive binding assays[7]. HYL001 inhibited ALK5 with an IC50 of 177 nM and p38α with an IC50 of 548 nM in recombinant kinase activity assays[7]. HYL001 (1-320 µg/mL) showed no mutagenic potential in S. typhimurium strains TA98, TA100, TA1535, and TA1537 with or without S9[7].
References:
[1]. Zou W, et al. Targeting glutamine metabolism as a potential target for cancer treatment. Journal of experimental & clinical cancer research: CR. 2025 Jul 01;44(1):180.
[2]. Wang Q, et al. A novel lonidamine derivative targeting mitochondria to eliminate cancer stem cells by blocking glutamine metabolism. Pharmacological research. 2023 Apr;190:106740.
[3]. Fan Z, et al. Boosting oxidative stress with hydroxyethyl starch smart nanomedicines to eliminate cancer stem cells. Accounts of Materials Research. 2024 Oct 24;5(12):1558-70.
[4]. De Los Santos-Jiménez J, et al. GLS and GLS2 Glutaminase Isoenzymes in the Antioxidant System of Cancer Cells. Antioxidants (Basel, Switzerland). 2024 Jun 20;13(6):745.
[5]. Tauriello DVF, et al. New Potent Inhibitor of Transforming Growth Factor-Beta (TGFβ) Signaling that is Efficacious against Microsatellite Stable Colorectal Cancer Metastasis in Combination with Immune Checkpoint Therapy in Mice. ACS pharmacology & translational science. 2025 Jan 10;8(1):97-112.
[6]. Wang Q, et al. Glutaminolysis inhibition boosts photodynamic therapy to eliminate cancer stem cells. Biomaterials. 2024 Apr;306:122497.
[7]. Tauriello DV, et al. HYL001, a new potent TGFβ signaling inhibitor that is efficacious against microsatellite stable CRC metastasis in combination with immune checkpoint therapy in mice. bioRxiv. 2024 May 14:2024-05.