mTORC2 (IC50 = 0.36 μM)
The primary target of JR-AB2-011 is the mTORC2 (mammalian target of rapamycin complex 2) kinase complex. mTORC2 is a multi-subunit complex composed of mTOR, Rictor, mSIN1, and mLST8, and is responsible for phosphorylation and activation of Akt (at Ser473), SGK, and PKC. Unlike mTORC1, mTORC2 is not inhibited by rapamycin and plays a key role in cell survival, metabolism, and cytoskeletal organization. JR-AB2-011 specifically blocks the interaction of the regulatory subunit Rictor with mTOR, thereby inhibiting mTORC2 activity without affecting mTORC1.

Good anti-GBM capabilities are exhibited by JR-AB2-011 (1 μM; 24 hours), which inhibits mTORC2 signaling and Rictor's interaction with mTOR [1]. Compared to CID613034, JR-AB2-011 (0.5–2 μM; 48 hours) exhibits less toxicity to normal neurons at concentrations up to 10 mM with no discernible cytotoxic effects [1].
In vitro, JR-AB2-011 is a selective mTORC2 inhibitor with an IC₅₀ of 0.36 μM. It inhibits mTORC2 activity by blocking the Rictor-mTOR association with a Ki of 0.19 μM. This disruption leads to reduced phosphorylation of Akt and downstream signaling. The compound exhibits cytotoxicity in glioblastoma cells and enhances apoptotic levels in GBM cells. Its selectivity for mTORC2 over mTORC1 makes it a valuable tool for dissecting the specific roles of mTORC2 in cell signaling.

Mouse models receiving JR-AB2-011 (4 mg/kg; i.p. daily for 10 days; 20 mg/kg; i.p. daily for 10 days) in each pilot protocol exhibited substantial effects on tumor development rates. Significant results (74% inhibition at the end of the 4 mg/kg/d treatment period; tumor growth delay of 10.0 days; 80% inhibition at the end of the 20 mg/kg/d drug period; tumor growth delay of 12.0 days) [1].
In vivo, JR-AB2-011 has anti-glioblastoma multiforme (GBM) activity. By specifically targeting mTORC2, the compound provides valuable insights into the role of mTORC2 in cellular processes and offers potential therapeutic applications in cancer and other diseases where mTORC2 is dysregulated. However, specific in vivo efficacy data from animal models is not detailed in the provided sources. The compound's ability to cross the blood-brain barrier would be important for GBM applications.

Examination of mTORC2 activity through in vitro kinase assay in rat primary microglia stimulated with rCCL17 or rCCL17 combined with AZD2098. To elucidate the underlying mechanism, the in vitro kinase assay was performed in primary microglia[2].
For mTORC2 inhibitors, standard cell-free assays involve measuring the kinase activity of immunoprecipitated mTORC2 complexes using recombinant Akt as a substrate. The compound is incubated with the immunoprecipitated mTORC2 complex and ATP, and Akt Ser473 phosphorylation is detected by western blot or ELISA. Alternatively, the compound's ability to disrupt the Rictor-mTOR interaction can be assessed by immunoprecipitation or proximity ligation assays.

Apoptosis analysis [1]
Cell Types: U87 GBM cells; LN229 GBM cells
Tested Concentrations: 1 μM
Incubation Duration: 24 hrs (hours)
Experimental Results: Has good anti-GBM properties and blocks mTORC2 signaling and the association of Rictor with mTOR.

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Cytotoxicity assay [1]
Cell Types: normal mature human neurons
Tested Concentrations: 0.5, 1, 2 μM
Incubation Duration: 48 hrs (hours)
Experimental Results: Minimal toxicity to normal neurons, no obvious cytotoxic effect at concentrations up to 10 mM.
For mTORC2 inhibitors, standard cellular assays involve treatment of cancer cell lines with the test compound for 24-72 hours. mTORC2 activity is assessed by measuring Akt Ser473 phosphorylation by western blot. Downstream signaling (e.g., phosphorylation of FOXO, GSK3β) and cell viability (MTT or CellTiter-Glo) are also measured. Apoptosis is assessed by Annexin V/PI staining or caspase activity assays. The compound's effects on mTORC2-specific substrates (Akt, SGK, PKC) are evaluated.

Animal/Disease Models: Female CB-17-scid (Taconic) mouse LN229 cells [1]
Doses: 4 mg/kg; 20 mg/kg
Route of Administration: daily intraperitoneal (ip) injection; 10-day
Experimental Results: compared with mice receiving vehicle alone Either dosage regimen demonstrated significant inhibition of tumor growth rate compared to mice.

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SAH rat models were assigned to receive recombinant CCL17 (rCCL17) or phosphate buffer saline (PBS). AZD2098 and JR-AB2-011 were applied to investigate the C-C motif chemokine receptor 4 (CCR4)/mammalian target of rapamycin complex 2 (mTORC2) axis in CCL17-mediated neuroprotection. To elucidate the underlying mechanism, the in vitro kinase assay was performed in primary microglia. Microglial-specific Rictor knockdown was administered via intracerebroventricular injection of adenovirus-associated virus. Brain water content, short-term neurobehavioural evaluation, western blot analysis, quantitative RT-PCR and histological staining were performed[2].

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In vivo Xenograft Experiments[3]
Institutional guidelines for animal welfare and experimental conduct were followed for all animal experiments, which were approved by the Institutional Animal Care and Use Committee and the Regional Administrative Authority under protocol G17/151. All animals received food and water ad libitum. Six weeks old male C57BL/6N mice (n = 20) underwent splenic injection of 2.5 × 105 B16 melanoma cells containing firefly luciferase-expressing plasmid pCHMWS_Luciferase under isoflurane anesthesia and 200 mg/kg metamizole pain treatment. Treatment with 20 mg/kg JR-AB2-011 intraperitoneal (i.p.) or solvent control for 13 days was started one day after tumor inoculation (n = 10). Thirteen days after intrasplenic injection, mice were terminated.
For in vivo evaluation of mTORC2 inhibitors, standard animal models include tumor xenograft models (e.g., glioblastoma xenografts). The compound is typically administered orally or intraperitoneally for 2-4 weeks. Tumor growth, Akt phosphorylation, apoptosis markers, and survival are assessed. The compound's ability to penetrate the blood-brain barrier is important for GBM applications.

Detailed pharmacokinetic data for JR-AB2-011 is not extensively provided in the available sources. The compound has a molecular weight of 335.16 and formula C₁₅H₉Cl₂FN₄. It is soluble in DMSO at 90 mg/mL and should be stored at 4°C, protected from light. As a small molecule with moderate lipophilicity, it would be expected to have reasonable cell permeability and oral bioavailability. Comprehensive PK studies would be required for therapeutic development.

Detailed toxicology data for JR-AB2-011 is not provided in the available sources. As a research compound targeting mTORC2, standard preclinical toxicology would be required for therapeutic development. The compound's selectivity for mTORC2 may contribute to a manageable safety profile, though potential effects on mTORC1 or other kinases would need to be assessed. The compound is for research use only and not for therapeutic applications.

[1]. Benavides-Serrato A, et al. Correction: Specific blockade of Rictor-mTOR association inhibits mTORC2 activity and is cytotoxicin glioblastoma. PLoS One. 2019 Feb 6;14(2):e0212160.
[2]. Zhang A, et al. CCL17 exerts neuroprotection through activation of CCR4/mTORC2 axis in microglia after subarachnoid haemorrhage in rats. Stroke Vasc Neurol. 2022 Jul 26;8(1):4–16.
[3]. Guenzle J, et al. Pharmacological Inhibition of mTORC2 Reduces Migration and Metastasis in Melanoma. Int J Mol Sci. 2020 Dec 22;22(1):30.
[4]. Wu M, et al Dioscin ameliorates murine ulcerative colitis by regulating macrophage polarization. Pharmacol Res. 2021 Oct ; 172:105796.

Despite advancements in treatment methods in recent years, the prognosis for liver metastases from melanoma remains poor. While targeting the mTOR signaling pathway has potent antitumor activity, little is known about the effects of mTORC2-specific inhibition on liver metastases. We used the novel mTORC2-specific inhibitor JR-AB2-011 and found that it significantly reduced cell migration and invasion by inhibiting the activation of MMP2 in melanoma cells. Furthermore, blocking mTORC2 induced cell death via non-apoptotic pathways and reduced the proliferation rate of tumor cells in a dose-dependent manner. In addition, in vivo imaging and autopsy confirmed that JR-AB2-011 treatment significantly reduced liver metastases in a homologous mouse liver metastasis model. Therefore, our study highlights for the first time the role of mTORC2 pharmacological blockade as a potent novel anticancer approach in the treatment of liver metastases from melanoma. [3]

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Restoring immune balance by targeting macrophage polarization is a potentially effective strategy for treating ulcerative colitis (UC). Diosgenin is a steroidal saponin with potent anti-inflammatory, immunomodulatory, and lipid-lowering effects. This study investigated the protective effect of diosgenin against colitis (UC) in mice and its potential mechanism. Mice were induced to develop colitis using sodium dextran sulfate (DSS) and simultaneously treated with oral diosgenin. In vitro, RAW264.7 cells were induced to polarize into M1 macrophages using lipopolysaccharide (LPS) and interferon-γ (INF-γ), followed by diosgenin treatment. The results showed that diosgenin improved colitis in mice, reduced M1 macrophage polarization, but significantly promoted M2 polarization in the mouse colon. Diosgenin inhibits the mammalian target of rapamycin complex 1 (mTORC1)/hypoxia-inducible factor-1α (HIF-1α) signaling pathway and suppresses glycolysis in RAW264.7 cells; however, it activates the mammalian target of rapamycin complex 2 (mTORC2)/peroxisome proliferation-activating receptor-γ (PPAR-γ) signaling pathway and promotes fatty acid oxidation (FAO). Regulation of the mTOR signaling pathway may inhibit M1 polarization but promote M2 polarization. Furthermore, the FAO inhibitor etomoxir and the mTORC2 inhibitor JR-AB2-011 can neutralize the effect of diosgenin on M2 polarization. Simultaneously, the mTORC1 agonist L-leucine can alleviate the inhibitory effect of diosgenin on M1 polarization. Both JR-AB2-011 and L-leucine can block the therapeutic effect of diosgenin in mouse ulcerative colitis. Therefore, diosgenin may improve ulcerative colitis in mice by inhibiting M1 polarization of macrophages and promoting M2 polarization. One possible mechanism by which diosgenin inhibits aerobic glycolysis and promotes fatty acid oxidation (FAO) in macrophages is by regulating the mTORC1/HIF-1α and mTORC2/PPAR-γ signaling pathways. In summary, diosgenin protects mice from DSS-induced ulcerative colitis by regulating the mTOR signaling pathway, thereby regulating macrophage metabolism and polarization. [4]
JR-AB2-011 is a selective mTORC2 inhibitor with an IC₅₀ of 0.36 μM that blocks Rictor-mTOR association with a Ki of 0.19 μM. It has anti-glioblastoma multiforme (GBM) activity. The compound was first reported in PLoS One. It is also known as JR-AB2-011 and has the molecular formula C₁₅H₉Cl₂FN₄. No regulatory approval has been identified.

C17H14CL2FN3OS

398.281963825226

397.021

C, 51.27; H, 3.54; Cl, 17.80; F, 4.77; N, 10.55; O, 4.02; S, 8.05

2411853-34-2

2411853-34-2;329182-61-8 (wrong);

138319699

Typically exists as white to off-white solids at room temperature

5.2

1

4

2

25

516

0

CC1CN(C(=NC2=CC=C(C=C2)F)S1)C(=O)NC3=CC(=C(C=C3)Cl)Cl

TWTNZYABDOSOSR-UHFFFAOYSA-N

InChI=1S/C17H14Cl2FN3OS/c1-10-9-21-17(25-10)23(13-5-2-11(20)3-6-13)16(24)22-12-4-7-14(18)15(19)8-12/h2-8,10H,9H2,1H3,(H,22,24)

(Z)-N-(3,4-dichlorophenyl)-2-((4-fluorophenyl)imino)-5-methylthiazolidine-3-carboxamide

JRAB2011 JR AB2 011 JR-AB2-011

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO : ~62.5 mg/mL (~156.92 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (5.22 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (5.22 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)