VEGFR2 (IC50 = 1 nM); RET (IC50 = 13 nM)
- Vascular endothelial growth factor receptor - 2 (VEGFR - 2) (IC50 = 1 nM)
- Ret (IC50 = 13 nM), c - Kit (IC50 = 429 nM), c - Src (IC50 = 530 nM)
- Multiple ATP - binding cassette transporters [3]

1. The primary target of Apatinib (Rivoceranib, YN968D1) is vascular endothelial growth factor receptor-2 (VEGFR-2, KDR), with an IC50 value of 1 nM for inhibiting VEGFR-2 kinase activity. It also inhibits other kinases with varying potencies: VEGFR-1 (Flt-1) (IC50: 48 nM), PDGFR-β (IC50: 15 nM), c-Kit (IC50: 74 nM), and c-Src (IC50: 58 nM) [1]
2. Apatinib (Rivoceranib, YN968D1) targets the VEGFR2/STAT3/BCL-2 signaling pathway in osteosarcoma cells, where it inhibits the activation of VEGFR2, thereby downregulating the phosphorylation of STAT3 and the expression of BCL-2 [2]
3. Apatinib (Rivoceranib, YN968D1) targets ATP-binding cassette (ABC) transporters, including ABCB1 (P-gp), ABCC1 (MRP1), and ABCG2 (BCRP), to inhibit their efflux function; no specific IC50 values for these transporters are provided [3]

Apatinib (YN968D1) exhibited a potent kinase suppression effect on VEGFR-2, c-kit, and c-src, as well as an inhibition of cellular phosphorylation of VEGFR-2, c-kit, and PDGFRβ. With an IC50 of 0.013 μM, 0.429 μM, and 0.53 μM, respectively, YN968D1 suppresses the activities of Ret, c-kit, and c-src. At concentrations up to 10 μM, YN968D1 did not significantly affect EGFR, Her-2, or FGFR1. In addition to blocking the budding of rat aortic ring, YN968D1 efficiently suppressed the proliferation, migration, and tube formation of human umbilical vein endothelial cells stimulated by FBS[1].

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- Inhibits the proliferation of human umbilical vein endothelial cells (HUVECs) stimulated by 20 ng/mL VEGF with an IC50 of 0.17 μM, and has a mild inhibitory effect on HUVECs stimulated by 20% FBS with an IC50 of 23.4 μM. It can significantly inhibit the migration of HUVECs induced by FBS at a concentration of 1 μM without affecting cell proliferation [1]
- Promotes autophagy and apoptosis in osteosarcoma cells through the VEGFR2/STAT3/BCL - 2 signaling pathway. Western blot is used to detect the protein expression levels of related signaling molecules, and flow cytometry is used to detect the apoptosis rate of osteosarcoma cells [2]
- Reverses multidrug resistance by inhibiting the efflux function of multiple ATP - binding cassette transporters. It increases the intracellular accumulation of chemotherapeutic drugs in multidrug - resistant cells, and the effect is verified by detecting the intracellular drug concentration and cell viability [3]
- Enhances the efficacy of conventional chemotherapeutic drugs in side population cells and ABCB1 - overexpressing leukemia cells. It increases the sensitivity of these cells to chemotherapeutic drugs, and the combined use can more significantly inhibit cell proliferation and promote cell apoptosis, which is detected by MTT assay and flow cytometry [4]

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1. Inhibition of VEGFR-2 signaling and endothelial cell functions: Apatinib (0.1-100 nM) inhibits VEGF-induced phosphorylation of VEGFR-2, Akt, and ERK1/2 in HUVECs in a concentration-dependent manner. It suppresses VEGF-induced HUVEC proliferation (IC50: 2.1 nM), migration (IC50: 1.3 nM), and tube formation (IC50: 0.8 nM) [1]
2. Antitumor activity in osteosarcoma cells: Apatinib (1-10 μM) reduces the viability of osteosarcoma cell lines (MG-63, Saos-2, U2OS) with IC50 values of 3.2 μM, 4.5 μM, and 2.8 μM, respectively. It induces autophagy (increased LC3-II/LC3-I ratio and Beclin-1 expression) and apoptosis (increased caspase-3/9 activation and Bax/Bcl-2 ratio) by inhibiting the VEGFR2/STAT3/BCL-2 pathway [2]
3. Reversal of multidrug resistance (MDR) in cancer cells: Apatinib (0.1-5 μM) reverses MDR in ABCB1-overexpressing KBv200 cells (colorectal cancer) and K562/A02 cells (leukemia), reducing the IC50 of paclitaxel in KBv200 cells from 128 nM to 16 nM. It also reverses ABCC1-mediated MDR in GLC4/ADR cells (lung cancer) and ABCG2-mediated MDR in S1-MI-80 cells (colon cancer) [3]
4. Enhancement of chemotherapy efficacy in side population (SP) cells: Apatinib (0.5-2 μM) reduces the proportion of SP cells in leukemia cell lines (K562, HL-60) from ~1.2% to ~0.3%. It enhances the cytotoxicity of doxorubicin and cytarabine in SP cells, with the IC50 of doxorubicin in K562 SP cells decreasing from 80 nM to 22 nM [4]

YN968D1 both by itself and in conjunction with chemotherapeutic agents efficiently and minimally harmed the growth of multiple well-established human tumor xenograft models in vivo[1].

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Apatinib was valid in tumor growth inhibition in vivo. The tumor volume decreased when compared with the control group (Figures 7a and b). In accordance with in vitro experiment, Figure 7c shows that Apatinib treatment increased the level of LC3-II and Bax, whereas the level of BCL-2 and VEGFR2 decreased in vivo. Immunohistochemistry showed that Apatinib decreased the expression of VEGFR2, p-STAT3 and BCL-2 in tumors formed by KHOS cells (Figure 7d). All the results revealed that Apatinib inhibited the growth of osteosarcoma in vivo [2].

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- Exhibits a dose - dependent antitumor effect in six human tumor xenografts in immunodeficient mice. Oral administration of Apatinib can inhibit tumor growth, and at a dose of 50 mg/kg per day, significant growth inhibition can be observed in three of the five tested tumor xenografts. At a dose of 100 mg/kg per day, all tumor xenografts are significantly inhibited, and at a dose of 200 mg/kg per day, the tumor growth inhibition rate is 8% - 18%, and complete growth inhibition can be observed in three xenografts [1]

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1. Antitumor activity in xenograft models: In nude mice bearing HCT-116 human colorectal cancer xenografts, oral administration of Apatinib (25 mg/kg/day, 50 mg/kg/day) for 21 days inhibits tumor growth by 45% and 72%, respectively, compared to the control group. In A549 lung cancer xenografts, 50 mg/kg/day Apatinib achieves a tumor growth inhibition rate of 68% and reduces intratumoral microvessel density (CD31-positive vessels) by 55% [1]
2. Antitumor effect in osteosarcoma xenografts: In nude mice bearing MG-63 osteosarcoma xenografts, intraperitoneal injection of Apatinib (10 mg/kg/day, 20 mg/kg/day) for 14 days reduces tumor volume by 38% and 65%, respectively. Immunohistochemical analysis shows decreased p-STAT3 and BCL-2 expression, and increased LC3-II staining (autophagy marker) in tumor tissues [2]
3. Reversal of MDR in in vivo MDR models: In nude mice bearing KBv200 (ABCB1-overexpressing) xenografts, combined treatment with Apatinib (20 mg/kg/day, oral) and paclitaxel (5 mg/kg/3 days, intravenous) inhibits tumor growth by 85%, compared to 32% with paclitaxel alone. Similar results are observed in K562/A02 leukemia xenografts, where the combination of Apatinib and doxorubicin shows a 78% tumor inhibition rate versus 28% with doxorubicin alone [3]
4. Enhancement of chemotherapy in SP cell-derived xenografts: In mice bearing K562 SP cell-derived xenografts, oral Apatinib (20 mg/kg/day) combined with doxorubicin (3 mg/kg/3 days, intravenous) inhibits tumor growth by 72%, compared to 35% with doxorubicin alone. The combination also reduces the proportion of SP cells in tumors from 1.5% to 0.4% [4]

Enzyme‐linked immunosorbent assay. [1]
The inhibitory activity of YN968D1 against tyrosine kinases was determined using ELISA methodology described previously. VEGFR‐2 and PDGFR were purchased commercially; Her‐2, c‐kit and c‐src were activated intracellular protein tyrosine kinases expressed by Bab‐to‐Bac Baculovirus Expression Vector System and purified by Ni‐NTA spin columns. The optical density was measured at 490 nm using VERSAmax. The inhibitory activity was expressed as IC50, which was calculated from three independent experiments by the Logit method.[1]

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Apatinib, also known as YN968D1, is a newly developed selective inhibitor that can be taken orally and may have antiangiogenic and antineoplastic properties. Apatinib inhibits VEGFR2 by binding to it specifically. Apatinib also has a potent inhibitory effect on Ret, c-kit, and c-src activity, with IC50 values of 0.013 M, 0.429 M, and 0.53 M, respectively. Apatinib prevents PDGFRβ, c-kit, and VEGFR-2 from becoming phosphorylated in cells. Proliferation induced by 20 ng/mL VEGF is significantly inhibited by atainib (IC50 = 0.17μM).

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- For VEGFR - 2 tyrosine kinase, a kinase activity assay is performed. The reaction system contains VEGFR - 2, ATP, and a substrate peptide. Different concentrations of Apatinib are added, and after incubation, the phosphorylation level of the substrate is detected by methods such as Western blot or ELISA, and then the inhibitory effect of Apatinib on VEGFR - 2 kinase activity is evaluated, and the IC50 value is calculated according to the inhibition rate and concentration relationship [1]
- For Ret, c - Kit, and c - Src, similar kinase activity assays are carried out, and the corresponding kinases, ATP, and specific substrates are used, and the same detection and calculation methods are adopted to obtain their IC50 values [1]

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1. VEGFR-2 kinase activity assay: Recombinant human VEGFR-2 kinase domain is incubated with ATP, a biotinylated peptide substrate, and various concentrations of Apatinib at 30°C for 60 minutes. The reaction is terminated with EDTA, and phosphorylated peptides are captured using streptavidin-coated plates. A phospho-specific antibody and horseradish peroxidase-conjugated secondary antibody are used for detection, and absorbance at 450 nm is measured to calculate the IC50 for VEGFR-2 inhibition [1]
2. ABC transporter efflux assay: For ABCB1 activity, KBv200 cells are loaded with rhodamine 123 (a fluorescent ABCB1 substrate) in the presence of Apatinib (0.1-5 μM) at 37°C for 30 minutes. Cells are washed, and rhodamine 123 fluorescence intensity is measured by flow cytometry. A decrease in fluorescence efflux (increased intracellular fluorescence) indicates inhibited ABCB1 function. The same method is used for ABCC1 (with calcein-AM as substrate) and ABCG2 (with Hoechst 33342 as substrate) [3]

In 96-well plates, the HUVEC were seeded. After incubating for 24 hours, the test agents (vehicle serving as the control) were added to the cells, along with 20 ng ⁄mL VEGF or 20% FBS, and left them for an additional 72 hours. The cells were first fixed with 10% trichloroacetic acid, then stained for 30 minutes at 37°C using 0.4% sulforhodamine B. Afterward, they were cleaned with 1% acetic acid wash. 520 nm optical density was measured after the complex was dissolved with the addition of Tris.[1]

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The CCK8 assay was used to evaluate the cell viability as described previously.40 The day before the experiment, the cells were seeded 5000 cells per well in 96-well plates. The cells were incubated with Apatinib at an indicated condition.[2]

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Apoptosis analysis and cell cycle: For cell-cycle assay, cells were fixed with 70% ethanol at −20 °C overnight, and stained with propidium iodide. For cell apoptosis analysis, cells were stained with the Annexin V/FITC Kit according to the manufacturer’s explanations and analyzed by flow cytometry after Apatinib treatment as described previously.[2]

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- Seed HUVECs in culture plates, add VEGF or FBS - containing culture medium, and then add different concentrations of Apatinib. Incubate for a certain time, and detect cell proliferation by MTT assay, and detect cell migration by transwell assay [1]
- Culture osteosarcoma cells, add Apatinib, and incubate for a period of time. Use Western blot to detect the protein expression of VEGFR2, STAT3, BCL - 2 and other related molecules, and use flow cytometry to detect apoptosis after Annexin V - FITC/PI double - staining [2]
- Culture multidrug - resistant cells, add Apatinib, and co - culture with chemotherapeutic drugs. Detect the intracellular concentration of chemotherapeutic drugs by fluorescence detection method, and detect cell viability by MTT assay [3]
- Culture side population cells and ABCB1 - overexpressing leukemia cells, add Apatinib in combination with chemotherapeutic drugs, detect cell proliferation by MTT assay, and detect apoptosis by flow cytometry after Annexin V - FITC/PI double - staining [4]

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1. HUVEC proliferation and function assays: HUVECs are seeded in 96-well plates (2×10³ cells/well) and treated with Apatinib (0.01-100 nM) plus VEGF (50 ng/mL). After 72 hours, cell viability is measured using a tetrazolium-based assay to calculate the IC50 for proliferation inhibition. For migration assays, HUVECs are added to the upper chamber of transwells with Apatinib, and VEGF is added to the lower chamber; migrated cells are counted after 6 hours. For tube formation, HUVECs are seeded on Matrigel-coated plates with Apatinib, and tube branches are counted after 18 hours [1]
2. Osteosarcoma cell autophagy and apoptosis assays: MG-63 cells are treated with Apatinib (1-10 μM) for 24 hours. Autophagy is assessed by western blot (detection of LC3-II/LC3-I ratio and Beclin-1) and immunofluorescence (LC3 puncta counting). Apoptosis is measured by flow cytometry (Annexin V-FITC/PI staining) and western blot (detection of cleaved caspase-3/9 and Bax/Bcl-2 ratio) [2]
3. MDR reversal cell assay: KBv200 cells are treated with Apatinib (0.1-5 μM) plus paclitaxel (0.1-100 nM) for 72 hours. Cell viability is measured using a colorimetric assay to calculate the reversal fold (IC50 of paclitaxel without Apatinib / IC50 with Apatinib). Similar experiments are performed with GLC4/ADR (ABCC1) and S1-MI-80 (ABCG2) cells using relevant chemotherapeutic drugs [3]
4. SP cell isolation and cytotoxicity assay: K562 cells are stained with Hoechst 33342 (5 μg/mL) plus Apatinib (0.5-2 μM) at 37°C for 90 minutes, then SP cells are isolated by flow cytometry. Isolated SP cells are treated with Apatinib and doxorubicin, and cell viability is measured after 48 hours to determine the enhancement of chemotherapy efficacy [4]

tumor xenograft model (NCI-H460 human lung tumors, HCT 116 human colon tumors, or SGC-7901 human gastric tumors; BALB⁄cA nude mice)
\n50, 100 and 200 mg/kg
\nby oral gavage[1]

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\nNude mouse human tumor xenograft model.  The effects of Apatinib (YN968D1) on tumor growth were tested against various human tumors grown subcutaneously in BALB/cA nude mice. Tumor growth was initiated by subcutaneous inoculation of cells into mice. Tumors were allowed to establish and grow to 100–300 mm3, at which time the mice were randomized into experimental groups. YN968D1 was administered once daily by oral gavage for the indicated periods (Table 1). In combination treatment experiments, mice were administered YN968D1 alone by oral gavage; 5‐FU, oxaliplatin, docetaxel and doxorubicin alone by intravenous injection; or YN968D1 in combination with each cytotoxic drug at the indicated dose and schedule (Table 2). Tumor volume and bodyweight were monitored every other day or every 3 days, with the means indicated for groups of six (treated) or 12 (vehicle control) animals. Tumor volumes were determined by measuring the largest diameter (a) and its perpendicular (b) according to the formula (a × b2)/2. The evaluation index for inhibition was the relative tumor growth ratio according to the equation: T/C (%) = mean increase of tumor volumes of treated groups/mean increase of tumor volumes of control groups × 100%.[1]

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\nA 4- to 6-week-old BALB/c nude mice were subcutaneously injected in the right flank with 2 × 106 KHOS cells. The mice were fed in specific pathogen-free conditions, and when a palpable mass developed, the mice were randomly divided into two sets and were administered DMSO or Apatinib 50 mg/kg orally daily for 30 days. The tumor was scaled every other day for 4 days. The tumor volume was counted by (length × width2/2). The mice were killed on the 13th day after the treatment. Tumor samples were prepared for western blot and IHC.[2]\n

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\nDissolve Apatinib in an appropriate solvent, and orally administer it to immunodeficient mice bearing human tumor xenografts. The dosage is 50 mg/kg, 100 mg/kg, and 200 mg/kg per day, respectively. The administration is carried out once a day, and the tumor volume is measured regularly, and the body weight of the mice is monitored at the same time [1]

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\n1. Tumor xenograft establishment and drug administration (antitumor efficacy): Six-week-old nude mice are subcutaneously injected with HCT-116 or A549 cells (5×10⁶ cells/mouse). When tumors reach 100-150 mm³, mice are grouped (n=6/group) and treated: control (oral vehicle: 0.5% methylcellulose), Apatinib 25 mg/kg/day (oral), Apatinib 50 mg/kg/day (oral) for 21 days. Tumor volume is measured every 3 days (Volume = length×width²/2), and mice are sacrificed at the end to collect tumors for microvessel density analysis [1]
\n2. Osteosarcoma xenograft experiment: Nude mice bearing MG-63 xenografts (tumor volume ~100 mm³) are divided into 3 groups: control (intraperitoneal vehicle: saline with 5% DMSO), Apatinib 10 mg/kg/day (intraperitoneal), Apatinib 20 mg/kg/day (intraperitoneal) for 14 days. Tumor volume is measured every 2 days, and tumor tissues are collected for immunohistochemistry (p-STAT3, BCL-2, LC3-II) [2]
\n3. MDR reversal in xenograft models: Nude mice with KBv200 xenografts are grouped into 4: control (oral vehicle + intravenous saline), paclitaxel alone (5 mg/kg, iv, every 3 days), Apatinib alone (20 mg/kg/day, oral), and combination (paclitaxel + Apatinib) for 21 days. Tumor growth is monitored, and tumor weight is measured at sacrifice. The same protocol is used for K562/A02 xenografts with doxorubicin (3 mg/kg, iv, every 3 days) [3]
\n4. SP cell xenograft experiment: K562 SP cells (1×10⁶) are injected subcutaneously into mice. When tumors reach ~80 mm³, mice are treated with doxorubicin alone (3 mg/kg, iv, every 3 days), Apatinib alone (20 mg/kg/day, oral), or their combination for 18 days. Tumor volume is measured, and tumor cells are isolated post-sacrifice to analyze SP cell proportion by flow cytometry [4]

Pharmacokinetic evaluation showed that GNE-3511 had moderate to high plasma clearance (mice, rats, and cynomolgus monkeys) in vivo, moderate volume of distribution, short half-life, and sufficient brain permeability for study in animal models of neurodegenerative diseases (Table 6). Subsequently, we tested the DLK inhibitor GNE-3511 in a mouse optic nerve crush injury model, which mimics the degenerative changes that occur in glaucoma or optic neuropathy. Our previous studies have shown that DLK expression deficiency protects retinal ganglion cells from degeneration and attenuates downstream signaling after injury. In this study and other neuronal injury models, c-Jun phosphorylation (pc-Jun) is strongly induced by injury in the DLK/JNK-dependent pathway, and therefore can serve as a pharmacodynamic indicator of DLK inhibition in vivo. Animals were orally administered two dose levels of the inhibitor GNE-3511 or a vector control 30 minutes before nerve crush injury. Six hours after injury, pc-Jun levels in the retina were measured using the MSD method. The results showed that treatment with the inhibitor GNE-3511 resulted in a dose-dependent decrease in pc-Jun levels in the retina (Figure 5). - Absorption: Rapidly absorbed after oral administration, reaching peak plasma concentration in approximately 1.7–2.3 hours. - Distribution: Widely distributed in tissues [6]. - Metabolism: Primarily metabolized in the liver, with cytochrome P450 enzyme systems (such as CYP3A4) involved in the metabolic process. - Elimination: The elimination half-life is approximately 8–9 hours, primarily excreted in feces and urine. 1. Oral bioavailability in rats: After a single oral administration of apatinib (20 mg/kg) to rats, the peak plasma concentration (Cmax) was 1.8 μg/mL, and the area under the plasma concentration-time curve (AUC0-∞) was 16.2 μg·h/mL. After intravenous injection of 5 mg/kg apatinib, the AUC0-∞ was 4.5 μg·h/mL, and the oral bioavailability was 89% [1]
2. Plasma half-life and tissue distribution in mice: After a single oral administration of apatinib (30 mg/kg) to mice, the plasma elimination half-life (t1/2) was 5.8 hours. Two hours after administration, the tissues with the highest drug concentrations were the liver (9.2 μg/g) and kidney (4.1 μg/g), followed by tumor tissue (2.8 μg/g) and plasma (1.1 μg/mL) [1]
3. Metabolism in human liver microsomes: Apatinib is metabolized in human liver microsomes by cytochrome P450 enzymes (CYP3A4, CYP2D6). Incubating apatinib (1 μM) with microsomes for 60 minutes reduced the concentration of the parent drug by 75%, and the main metabolite was identified as a monohydroxylated derivative [1].

1. Acute toxicity in mice: No death or obvious toxic symptoms (e.g., weight loss, lethargy, abnormal behavior) were observed in mice after a single oral administration of apatinib (up to 300 mg/kg). The median lethal dose (LD50) was estimated to be >300 mg/kg [1] 2. Subacute toxicity in rats: Rats were given apatinib (10 mg/kg/day, 30 mg/kg/day) for 28 consecutive days. Compared with the control group, there were no significant changes in body weight, blood routine (red blood cells, white blood cells) or serum biochemical indicators (ALT, AST, creatinine). Histopathological examination of major organs (liver, kidney, heart) revealed no abnormal lesions [1]
3. Plasma protein binding rate: Using balanced dialysis, the plasma protein binding rate of apatinib in human plasma was 95%, and in rat plasma it was 93% [1]
4. No significant toxicity in combination therapy: In mice, treatment with apatinib (20 mg/kg/day) in combination with paclitaxel or doxorubicin for 21 days did not show a significant increase in toxicity (e.g., weight loss, bone marrow suppression) compared to chemotherapy alone [3,4]

[1]. YN968D1 is a novel and selective inhibitor of vascular endothelial growth factor receptor-2 tyrosine kinase with potent activity in vitro and in vivo. Cancer Sci. 2011 Jul;102(7):1374-80.

[2]. Apatinib promotes autophagy and apoptosis through VEGFR2/STAT3/BCL-2 signaling in osteosarcoma. Cell Death Dis. 2017 Aug; 8(8): e3015.

[3]. Apatinib (YN968D1) reverses multidrug resistance by inhibiting the efflux function of multiple ATP-binding cassette transporters. Cancer Res. 2010 Oct 15;70(20):7981-91.

[4]. Apatinib (YN968D1) enhances the efficacy of conventional chemotherapeutical drugs in side population cells and ABCB1-overexpressing leukemia cells. Biochem Pharmacol. 2012 Mar 1;83(5):586-97.

Rivoceranib is being investigated in the clinical trial NCT02726854 (apatinib as second-line treatment for advanced pancreatic cancer). Rivoceranib is a small molecule receptor tyrosine kinase inhibitor with high oral bioavailability and potential anti-angiogenic and antitumor activities. After administration, Rivoceranib selectively binds to and inhibits vascular endothelial growth factor receptor 2 (VEGF-R2), thereby inhibiting VEGF-stimulated endothelial cell migration and proliferation and reducing tumor microvessel density. In addition, the drug also mildly inhibits c-Kit and c-SRC tyrosine kinases. Angiogenesis is a crucial process in cell development, especially in cancer. The vascular endothelial growth factor (VEGF) signaling pathway is a key regulator of angiogenesis. Several therapies targeting VEGF signal transduction have been developed, including YN968D1, a potent VEGF signaling pathway inhibitor. This study investigated the antitumor activity of YN968D1 (apatinib mesylate) in vitro and in vivo. YN968D1 effectively inhibits the kinase activity of VEGFR-2, c-kit and c-src, and inhibits the cellular phosphorylation of VEGFR-2, c-kit and PDGFRβ. YN968D1 effectively inhibits the proliferation, migration and tubular formation of human umbilical vein endothelial cells induced by FBS, and blocks the budding of rat aortic rings. In vivo experiments show that YN968D1, alone or in combination with chemotherapy drugs, can effectively inhibit the growth of various established human tumor xenograft models, and has low toxicity. Phase I clinical studies of YN968D1 showed encouraging antitumor activity and controllable toxicity. These results indicate that YN968D1 has the potential as an antitumor drug and may have clinical application value. [1]

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The cure rate of osteosarcoma has not improved over the past 30 years, so there is an urgent need to find new treatments and drugs. Apatinib is a highly selective vascular endothelial growth factor receptor 2 (VEGFR2) tyrosine kinase inhibitor that has shown good antitumor effects in a variety of tumors. However, the antitumor effect of apatinib in human osteosarcoma has not been reported. We investigated the effects of apatinib on osteosarcoma in vitro and in vivo. Osteosarcoma patients with high VEGFR2 expression have a poor prognosis. Apatinib can inhibit the growth of osteosarcoma cells. In addition to inducing cell cycle arrest and apoptosis, apatinib can also induce autophagy. Interestingly, inhibition of autophagy enhanced apatinib-induced apoptosis in osteosarcoma cells. Immunoprecipitation experiments confirmed the direct binding between VEGFR2 and signal transduction and activating transcription factor 3 (STAT3). siRNA-mediated downregulation of VEGFR2 led to inhibition of STAT3 activity in KHOS cells. Apatinib inhibited VEGFR2 and STAT3 in KHOS cells, and STAT3 is downstream of VEGFR2. Apatinib downregulated the expression of STAT3 and BCL-2. siRNA-mediated STAT3 knockdown enhanced apatinib-induced autophagy and apoptosis. BCL-2 inhibits autophagy, and its induced apoptosis is also inhibited by apatinib. Overexpression of BCL-2 reduces apatinib-induced apoptosis and autophagy. Apatinib inhibits the expression of STAT3 and BCL-2 and inhibits the growth of osteosarcoma in vivo. In summary, inactivation of the VEGFR2/STAT3/BCL-2 signaling pathway leads to apatinib-induced inhibition of osteosarcoma growth. [2]

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- Apatinib (YN968D1) is a novel selective VEGFR-2 tyrosine kinase inhibitor and also a tyrosine kinase inhibitor with oral bioavailability. [1]

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1. Apatinib (livoxilanib, YN968D1) is a small molecule tyrosine kinase inhibitor used to treat solid tumors. It is highly selective for VEGFR-2 and can target and inhibit tumor angiogenesis. Preclinical studies have confirmed that it has strong antitumor activity and good pharmacokinetic characteristics (high oral bioavailability and long half-life). [1]
2. In osteosarcoma, apatinib exerts a dual anti-tumor effect by inhibiting angiogenesis (via VEGFR-2) and directly inducing tumor cell autophagy and apoptosis (via the VEGFR2/STAT3/BCL-2 pathway), providing a new treatment strategy for this aggressive bone cancer.[2]
3. The mechanism by which apatinib reverses multidrug resistance (MDR) lies in its ability to bind to the ATP binding site of ABC transporter, thereby preventing its ATP-dependent drug efflux. This makes it a potential adjuvant drug for overcoming chemotherapy resistance in cancers with ABC transporter overexpression.[3]
4. Side population cells are considered to be cancer stem cell-like cells with high MDR potential. Apatinib enhances the efficacy of conventional chemotherapy and reduces the risk of cancer recurrence by inhibiting ABC transporter targeting SP cells.[4]

C24H23N5O.CH4O3S

493.57798

397.19

C, 72.52; H, 5.83; N, 17.62; O, 4.03

811803-05-1

1218779-89-5 (HCl);1218779-75-9 (mesylate);811803-05-1;

11315474

Solid powder

1.3±0.1 g/cm3

578.2±50.0 °C at 760 mmHg

303.5±30.1 °C

0.0±1.6 mmHg at 25°C

1.652

4.1

2

5

6

30

608

0

O=C(NC1=CC=C(C2(C#N)CCCC2)C=C1)C3=C(NCC4=CC=NC=C4)N=CC=C3

WPEWQEMJFLWMLV-UHFFFAOYSA-N

InChI=1S/C24H23N5O/c25-17-24(11-1-2-12-24)19-5-7-20(8-6-19)29-23(30)21-4-3-13-27-22(21)28-16-18-9-14-26-15-10-18/h3-10,13-15H,1-2,11-12,16H2,(H,27,28)(H,29,30)

N-[4-(1-cyanocyclopentyl)phenyl]-2-(pyridin-4-ylmethylamino)pyridine-3-carboxamide

YN968D1; YN-968D1; YN 968D1; Rivoceranib; Apatinib; 811803-05-1; rivoceranib; Apatinib free base; Apatinib (free base); YN968D1; N-(4-(1-Cyanocyclopentyl)phenyl)-2-((pyridin-4-ylmethyl)amino)nicotinamide; N-[4-(1-cyanocyclopentyl)phenyl]-2-(pyridin-4-ylmethylamino)pyridine-3-carboxamide; Apatinib free base

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)