 Chemical Structure.png)
Abequolixron (formerly known as SB742881; RG104; SB-742881) is a novel potent and orally bioavailable liver-X nuclear hormone receptor-beta (LXR) agonist with potential immunomodulating and anticancer activities. It modulates innate immunity via transcriptional activation of the ApoE gene. RGX-104 selectively targets and binds to LXRbeta, thereby activating LXRbeta-mediated signaling, leading to the transcription of certain tumor suppressor genes and the downregulation of certain tumor promoter genes. This particularly activates the expression of apolipoprotein E (ApoE), a tumor suppressor protein, in tumor cells and certain immune cells. This activates the innate immune system, resulting in depletion of immunosuppressive myeloid-derived suppressor cells (MDSCs), tumor cells and endothelial cells in the tumor microenvironment. This reverses immune evasion, enhances anti-tumor immune responses and inhibits proliferation of tumor cells.
Physicochemical Properties
| Molecular Formula | C34H33CLF3NO3 | |
| Molecular Weight | 596.078939199448 | |
| Exact Mass | 595.21 | |
| Elemental Analysis | C, 68.51; H, 5.58; Cl, 5.95; F, 9.56; N, 2.35; O, 8.05 | |
| CAS # | 610318-54-2 | |
| Related CAS # | RGX-104 hydrochloride;610318-03-1 | |
| PubChem CID | 10218693 | |
| Appearance | White to off-white solid powder | |
| LogP | 6.3 | |
| Hydrogen Bond Donor Count | 1 | |
| Hydrogen Bond Acceptor Count | 7 | |
| Rotatable Bond Count | 13 | |
| Heavy Atom Count | 42 | |
| Complexity | 783 | |
| Defined Atom Stereocenter Count | 1 | |
| SMILES | C[C@H](CCOC1=CC=CC(=C1)CC(=O)O)N(CC2=C(C(=CC=C2)C(F)(F)F)Cl)CC(C3=CC=CC=C3)C4=CC=CC=C4 |
|
| InChi Key | ZLJZDYOBXVOTSA-XMMPIXPASA-N | |
| InChi Code | InChI=1S/C34H33ClF3NO3/c1-24(18-19-42-29-16-8-10-25(20-29)21-32(40)41)39(22-28-15-9-17-31(33(28)35)34(36,37)38)23-30(26-11-4-2-5-12-26)27-13-6-3-7-14-27/h2-17,20,24,30H,18-19,21-23H2,1H3,(H,40,41)/t24-/m1/s1 | |
| Chemical Name |
|
|
| Synonyms |
|
|
| HS Tariff Code | 2934.99.9001 | |
| Storage |
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. |
|
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets |
LXR[1] Cancer immunotherapy is restricted to immune resistance caused by immunosuppressive tumor microenvironment. Pyroptosis involved in antitumor immunotherapy as a new schedule is prospective to reverse immunosuppression. Herein, acidic tumor microenvironment (TME)-evoked MRC nanoparticles (MRC NPs) co-delivering immune agonist RGX-104 and photosensitizer chlorine e6 (Ce6) are reported for pyroptosis-mediated immunotherapy. RGX-104 remodels TME by transcriptional activation of ApoE to regress myeloid-derived suppressor cells' (MDSCs) activity, which neatly creates foreshadowing for intensifying pyroptosis. Considering Ce6-triggered photodynamic therapy (PDT) can strengthen oxidative stress and organelles destruction to increase immunogenicity, immunomodulatory-photodynamic MRC nanodrugs will implement an aforementioned two-pronged strategy to enhance gasdermin E (GSDME)-dependent pyroptosis. RNA-seq analysis of MRC at the cellular level is introduced to first elucidate the intimate relationship between RGX-104 acting on LXR/ApoE axis and pyroptosis, where RGX-104 provides the prerequisite for pyroptosis participating in antitumor therapy. Briefly, MRC with favorable biocompatibility tackles the obstacle of hydrophobic drugs delivery, and becomes a powerful pyroptosis inducer to reinforce immune efficacy. MRC-elicited pyroptosis in combination with anti-PD-1 blockade therapy boosts immune response in solid tumors, successfully arresting invasive metastasis and extending survival based on remarkable antitumor immunity. MRC may initiate a new window for immuno-photo pyroptosis stimulators augmenting pyroptosis-based immunotherapy.[2] Adv Healthc Mater . 2022 Nov;11(21):e2201233. |
| ln Vitro |
Cancer immunotherapy is restricted to immune resistance caused by immunosuppressive tumor microenvironment. Pyroptosis involved in antitumor immunotherapy as a new schedule is prospective to reverse immunosuppression. Herein, acidic tumor microenvironment (TME)-evoked MRC nanoparticles (MRC NPs) co-delivering immune agonist RGX-104 and photosensitizer chlorine e6 (Ce6) are reported for pyroptosis-mediated immunotherapy. RGX-104 remodels TME by transcriptional activation of ApoE to regress myeloid-derived suppressor cells' (MDSCs) activity, which neatly creates foreshadowing for intensifying pyroptosis. Considering Ce6-triggered photodynamic therapy (PDT) can strengthen oxidative stress and organelles destruction to increase immunogenicity, immunomodulatory-photodynamic MRC nanodrugs will implement an aforementioned two-pronged strategy to enhance gasdermin E (GSDME)-dependent pyroptosis. RNA-seq analysis of MRC at the cellular level is introduced to first elucidate the intimate relationship between RGX-104 acting on LXR/ApoE axis and pyroptosis, where RGX-104 provides the prerequisite for pyroptosis participating in antitumor therapy. Briefly, MRC with favorable biocompatibility tackles the obstacle of hydrophobic drugs delivery, and becomes a powerful pyroptosis inducer to reinforce immune efficacy. MRC-elicited pyroptosis in combination with anti-PD-1 blockade therapy boosts immune response in solid tumors, successfully arresting invasive metastasis and extending survival based on remarkable antitumor immunity. MRC may initiate a new window for immuno-photo pyroptosis stimulators augmenting pyroptosis-based immunotherapy.[2] Adv Healthc Mater . 2022 Nov;11(21):e2201233. In a transwell MDSC differentiation assay where bone marrow cells were co-cultured with B16F10 melanoma cells and GM-CSF for 6 days, addition of RGX-104 (2 µM) on day 3 significantly reduced the number of Gr-1highCD11b+ cells (granulocytic MDSCs) on day 6, to a level comparable to omission of cancer cells. Treatment of MDSCs isolated from tumor-bearing mice with RGX-104 (1 µM or 2 µM) in vitro significantly reduced MDSC survival and increased the percentage of cleaved caspase-3+ MDSCs, indicating induction of apoptosis. |
| ln Vivo |
When given orally to mice with visible tumors, RGX-104 (100 mg/kg daily) effectively suppressed the growth of several cancer types. The combination of RGX-104 and anti-PD-1 was found to be more effective than either drug administered alone. Significantly, mice that received RGX-104 in addition to anti-PD-1 treatment did so with good tolerance and showed no overt damage [1]. Oral administration of RGX-104 (50 mg/kg/day, 100 mg/kg/day via chow, or 80 mg/kg/day via intraperitoneal injection) significantly suppressed tumor growth in multiple syngeneic and xenograft mouse models, including B16F10 melanoma, GL261 glioblastoma, Lewis Lung Carcinoma (LLC), MC38 colon cancer, and human SKOV3 ovarian cancer in immunodeficient mice. RGX-104 treatment reduced the abundance of both granulocytic (G-MDSC) and monocytic (M-MDSC) myeloid-derived suppressor cells in tumors, spleen, and peripheral blood of tumor-bearing mice. MDSC depletion by RGX-104 was associated with increased activation of tumor-infiltrating cytotoxic T lymphocytes (CTLs), as evidenced by elevated frequencies of IFN-γ+Granzyme B+ CD8+ T cells and PD-1+ CD8+ T cells. In an adoptive T cell therapy model using gp100-specific Pmel CD8+ T cells, co-administration of RGX-104 enhanced anti-tumor activity and improved mouse survival. Combination therapy of RGX-104 with anti-PD-1 antibody showed superior anti-tumor efficacy compared to either agent alone in B16F10 melanoma and LLC models, even in the presence of Gvax-induced MDSC accumulation. In a Phase 1 clinical trial involving cancer patients, oral RGX-104 treatment (once daily for 21 days per 28-day cycle) significantly reduced circulating granulocytic MDSC levels (median 86% decrease) and monocytic MDSC levels in 5 of 6 patients. Patient treatment with RGX-104 was associated with increased activation of circulating CD8+ T cells, particularly within the PD-1+ population, as measured by GITR expression. |
| Cell Assay |
Bone marrow cells are cultured with B16F10 melanoma cells and
GM-CSF for 6 days. On day 3, RGX-104 (2 μM) is added to the culture. The
mean number of Gr-1high CD11b+ cells per 50 mL of culture solution is
assessed by flow cytometry on day 6[1].
MDSC in vitro Proliferation Assay [1] Myeloid-derived suppressor cells were isolated as previously described from splenic tissue of tumorbearing mice. One hundred thousand cells were plated in quadruplicates in poly-L-lysine coated plates. After 3 hours of treatment with 1uM Abequolixron (RGX-104) or DMSO as vehicle, cells were fixed with 4% PFA for 15 minutes and wash 3 times with 1X PBS prior staining. Rabbit monoclonal anti-Ki67 antibody (1:400 dilution) was applied at 4C overnight. Cells were incubated with Alexa Fluor 488 secondary antibody (1:200 dilution, Invitrogen) for one hour at room temperature, counterstained with DAPI (1:1000 dilution) and mounted with Prolong Gold. For the analysis of the percentage of Ki67 positive cells, five fields from each replicate were imaged at 20x magnification using Zeiss Axio Imager fluorescence microscope. Image analysis was performed using CellProfiler software. MDSC Adhesion Assay [1] Myeloid-derived suppressor cells were isolated as previously described from splenic tissue of tumor-bearing mice. One hundred thousand cells were plated in triplicates in poly-L-lysine coated plates. Cells were treated with 1uM Abequolixron (RGX-104) or DMSO as vehicle for 2 hours and shaken at 300 rpm for 30 minutes. After this, cells were fixed with 4% PFA for 15 minutes, wash 3 times with 1X PBS, counterstained with DAPI and mounted using Prolong Gold. For the analysis, ten fields from each replicate were imaged at 20x magnification using Zeiss Axio Imager fluorescence microscope. The number of remaining cells was determined using CellProfiler software. In Vitro MDSC Apoptosis Assay [1] Mouse spleens were isolated from either WT, LXRαβ−/−, ApoE−/− or LRP8−/− mice and homogenized to create a single cell suspension. The cells were treated with 1X ACK Lysing Buffer to lyse and remove erythrocytes. MDSCs were isolated from the resulting cell suspension using the Myeloid-Derived Suppressor Cell Isolation Kit. Isolated MDSCs were plated onto slides and treated with either Abequolixron (RGX-104) or murine recombinant ApoE, at the indicated concentrations and times. The samples were then stained with an antibody against Cleaved Caspase-3. Mice[1] B16F10 cancer cells are subcutaneously injected into C57BL/6 mice. Following tumor growth to 5-10 mm3 in volume, mice are fed either control chow, chow supplemented with GW3965 (100 mg/kg), or chow supplemented with RGX-104 (100 mg/kg)[1]. RGX-104 was administered either through formulated drug chow at 100mg/kg/day or 50mg/kg/day or delivered via intraperitoneal injection (80mg/kg/day) in a vehicle suspension consisting of corn oil and ethanol (2.5% by volume) as indicated in each figure. Control cohorts were treated with either normal chow (Purina 5001) or with vehicle consisting of corn oil and ethanol (2.5% by volume), respectively. Tumor measurements were taken on the days indicted throughout the course of the experiment with calipers. For survival analysis, mice were euthanized when total tumor burden approached IACUC guidelines with a tumor burden exceeding 1,500 mm3 in volume. For the relevant experiments, anti-PD-1 mAb (clone RMP1-14) or a control isotype-matched antibody was administered at 10mg/kg intraperitoneally on days 3, 6, and 9 post-tumor injection. Gvax was generated as previously described and administered at high frequency (every 3 days) during the experiments as indicated.[1] For the MDSC apoptosis assay in vitro, MDSCs were isolated from spleens of tumor-bearing mice using a specific isolation kit. Isolated MDSCs were plated and treated with RGX-104 or vehicle at indicated concentrations (e.g., 1 µM, 2 µM) and times (e.g., 3 hours, 6 hours). Cells were then fixed and stained with an antibody against cleaved caspase-3 for quantification of apoptotic cells. For the in vitro MDSC differentiation assay, bone marrow cells were isolated from mouse femurs and cultured in the bottom of a plate. B16F10 melanoma cells were placed in a transwell insert. Cultures were supplemented with GM-CSF. On day 3, RGX-104 (2 µM) or vehicle was added. On day 6, cells in the basal chamber were harvested, stained for CD11b and Gr-1, and analyzed by flow cytometry to quantify MDSC populations. For T cell suppression assays, CD8+ T cells were isolated from naive mice, labeled with a proliferation dye, and co-cultured with MDSCs isolated from control or RGX-104-treated tumor-bearing mice in the presence of CD3/CD28 activator beads and IL-2. After 24 hours, T cell activation (IFN-γ production) and proliferation (dye dilution) were assessed by flow cytometry. |
| Animal Protocol |
Animal/Disease Models: NOD SCID or RAG mice injected with 1×106 SKOV3 ovarian cancer cells[1]. Doses: 100 mg/kg. Route of Administration: Oral administration daily for about 60 days. Experimental Results: Robustly suppressed tumor growth and progression. Mice[1] B16F10 cancer cells are subcutaneously injected into C57BL/6 mice. Following tumor growth to 5-10 mm3 in volume, mice are fed either control chow, chow supplemented with GW3965 (100 mg/kg), or chow supplemented with RGX-104 (100 mg/kg)[1]. RGX-104 was administered either through formulated drug chow at 100mg/kg/day or 50mg/kg/day or delivered via intraperitoneal injection (80mg/kg/day) in a vehicle suspension consisting of corn oil and ethanol (2.5% by volume) as indicated in each figure. Control cohorts were treated with either normal chow (Purina 5001) or with vehicle consisting of corn oil and ethanol (2.5% by volume), respectively. Tumor measurements were taken on the days indicted throughout the course of the experiment with calipers. For survival analysis, mice were euthanized when total tumor burden approached IACUC guidelines with a tumor burden exceeding 1,500 mm3 in volume. For the relevant experiments, anti-PD-1 mAb (clone RMP1-14) or a control isotype-matched antibody was administered at 10mg/kg intraperitoneally on days 3, 6, and 9 post-tumor injection. Gvax was generated as previously described and administered at high frequency (every 3 days) during the experiments as indicated.[1] For most tumor growth studies, cancer cells were suspended in PBS, mixed 1:1 with Matrigel, and injected subcutaneously into the flanks of 6-8 week old mice. Once tumors reached a specified volume (e.g., 5–10 mm³, 30–40 mm³), mice were randomized into treatment groups. RGX-104 was administered orally via drug-formulated chow at doses of 50 mg/kg/day or 100 mg/kg/day. In some experiments, it was administered via intraperitoneal injection (80 mg/kg/day) in a vehicle consisting of corn oil with 2.5% ethanol by volume. Control groups received normal chow or the corn oil/ethanol vehicle via intraperitoneal injection. Tumor dimensions (length and width) were measured periodically with calipers, and volume was calculated. For combination therapy with anti-PD-1, the antibody was administered intraperitoneally at 10 mg/kg on specified days (e.g., days 3, 6, and 9 post-tumor injection). For adoptive T cell therapy experiments, CD8+ T cells were isolated from Pmel-1 TCR transgenic mice and transferred via retro-orbital injection into tumor-bearing recipients, which also received a gp100 peptide vaccine. |
| Toxicity/Toxicokinetics |
In the Phase 1 clinical trial, RGX-104 was reported to be well tolerated in the first six evaluable patients, with no dose-limiting toxicities observed. Treatment was not associated with significant changes in key hematologic parameters (total white blood cell, absolute neutrophil, absolute lymphocyte, and absolute monocyte counts) within the first two weeks, with changes normalizing over the 4-week cycle. |
| References |
[1]. LXR/ApoE Activation Restricts Innate Immune Suppression in Cancer. Cell. 2018 Feb 8;172(4):825-840.e18. |
| Additional Infomation |
Abequolixron is an orally bioavailable agonist of the nuclear receptor liver X receptor beta (LXRbeta; NR1H2; LXR-b), with potential immunomodulating and antineoplastic activities. Upon oral administration, abequolixron selectively targets and binds to LXRbeta, thereby activating LXRbeta-mediated signaling, leading to the transcription of certain tumor suppressor genes and the downregulation of certain tumor promoter genes. This particularly activates the expression of apolipoprotein E (ApoE), a tumor suppressor protein, in tumor cells and certain immune cells. This activates the innate immune system, resulting in depletion of immunosuppressive myeloid-derived suppressor cells (MDSCs), tumor cells and endothelial cells in the tumor microenvironment. This reverses immune evasion, enhances anti-tumor immune responses and inhibits proliferation of tumor cells. LXRbeta, a member of the oxysterol receptor family, which is in the nuclear receptor family of transcription factors, plays a key role in cholesterol transport, glucose metabolism and the modulation of inflammatory responses; activation of LXRbeta suppresses tumor cell invasion, angiogenesis, tumor progression, and metastasis in a variety of tumor cell types. The expression of the ApoE protein becomes silenced in human cancers as they grow, become invasive, and metastasize; ApoE silencing is related to reduced survival in cancer patients. The LXR-ApoE pathway regulates the ability of cancers to evade the immune system and recruit blood vessels. RGX-104 is an investigational LXRβ agonist undergoing a multicenter Phase 1 a/b clinical trial (NCT02922764) in patients with advanced solid cancers or lymphomas. Its anti-tumor and immunomodulatory effects are mediated through activation of the LXR/ApoE axis. LXR activation transcriptionally upregulates ApoE, which in turn promotes MDSC apoptosis via the LRP8 receptor, leading to reduced innate immune suppression and enhanced cytotoxic T cell activity. The study presents RGX-104 as a first-in-class therapeutic strategy to target MDSCs and reverse immune evasion in cancer. |
Solubility Data
| Solubility (In Vitro) |
|
|||
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.17 mg/mL (3.64 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 21.7 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. Solubility in Formulation 2: 2.08 mg/mL (3.49 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. 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 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 3: ≥ 0.83 mg/mL (1.39 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 8.3 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.6776 mL | 8.3881 mL | 16.7763 mL | |
| 5 mM | 0.3355 mL | 1.6776 mL | 3.3553 mL | |
| 10 mM | 0.1678 mL | 0.8388 mL | 1.6776 mL |