SMARCA2 (DC50 = 6 nM in MV-4-11 cells), SMARCA4 (DC50 = 11 nM in MV-4-11 cells), PBRM1 (DC50 = 32 nM in MV-4-11 cells). [1]
ACBI1 targets SMARCA2 (BRG1), SMARCA4 (BRM), and PBRM1, which are ATPase subunits of the BAF/PBAF chromatin remodeling complex. It recruits the E3 ubiquitin ligase VHL to these target proteins, leading to their poly-ubiquitination and subsequent proteasomal degradation. The compound exhibits PROTAC activity at VHL/SMARCA2 and VHL/SMARCA4 in human A549 and MV4-11 cells.

ACBI1 exhibits anti-proliferative action at 1–10,000 nM over 3–7 days[1]. SK-MEL-5 cells undergo apoptosis in response to ACBI1 (0.3 µM; 100 h)[1]. The E3 ubiquitin ligase von Hippel-Lindau, a linker, and a bromodomain ligand make up ACBI1[1].

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ACBI1 induced complete and potent degradation of SMARCA2 (DC50 6 nM) and SMARCA4 (DC50 11 nM) in MV-4-11 cells, and similar SMARCA2 degradation in SMARCA4-deficient NCI-H1568 cells, with PBRM1 degradation (DC50 32 nM). [1]
Degradation was rapid: half-maximal degradation occurred within 2 hours at 1 μM ACBI1 in MV-4-11 cells. [1]
Antiproliferative activity: IC50 of 28 nM in MV-4-11 cells, 77 nM in SK-MEL-5 cells, and 68 nM in NCI-H1568 cells; no effect in SMARCA2/4-null NCI-H1703 cells. [1]
Induced apoptosis in SK-MEL-5 cells with increased caspase activity comparable to doxorubicin (1 μM), and cleaved PARP appearance after 40 hours. [1]
Unbiased TMT proteomics (333 nM ACBI1, 8 h in MV-4-11 cells) confirmed selective knockdown of SMARCA2, SMARCA4, and PBRM1 with minimal off-target effects. [1]
Immunoprecipitation-mass spectrometry showed that ACBI1 treatment leads to dissociation of some BAF subunits (e.g., ACTL6A, BCL7A, PHF10) from the complex. [1]
In vitro, ACBI1 shows potent degradation activity with DC₅₀ values of 6 nM for SMARCA2, 11 nM for SMARCA4, and 32 nM for PBRM1 in MV-4-11 cells. It induces anti-proliferative effects and apoptosis in cancer cells. ACBI1 exhibits cooperative degradation of its targets. The compound demonstrates significant degradation of the SMARCA2 and SMARCA4 bromodomains in A549 cells. It is a potent PROTAC degrader for targeted protein degradation experiments.

In vivo activity data for ACBI1 is not extensively reported in the available literature. As a PROTAC degrader targeting chromatin remodeling complex subunits, it is expected to have antitumor activity in vivo by disrupting SWI/SNF complex function. The compound would be evaluated in xenograft or other animal models of cancer for efficacy. Pharmacodynamic studies would assess target degradation in tumor tissues.

Fluorescence Polarization (FP) competition assay: ACBI1 was tested for displacement of a FAM-labeled HIF-1α peptide from VCB in the absence or presence of saturating SMARCA2BD (75 μM), SMARCA4BD (150 μM), or PBRM1BD (100 μM). Assay buffer: 100 mM BIS-TRIS, 100 mM NaCl, 1 mM DTT, pH 7, with 1% DMSO. 5 nM peptide and serial dilutions of ACBI1 (14 points, half-log-fold from 50 μM to 0.016 nM) were incubated, and FP measured at 485/520 nm. IC50 values were fitted, and KD calculated. Cooperativity (α) was ~30 for ACBI1. [1]
TR-FRET competition assay: ACBI1 binding to SMARCA2BD (40 nM) was measured using biotinylated probe 5 (16.6 nM), Lance Eu-W1024 labeled Streptavidin (2.5 nM), and ULight-anti-His6 antibody (50 nM), in 50 mM HEPES, 50 mM NaCl, 2 mM DTT, 0.008% Brij, 0.01% BSA, pH 7.3, with 1% DMSO. Assay performed in the absence or presence of 5 μM VCB (wild-type or R69A mutant). ACBI1 showed binary IC50 of 770 nM and ternary IC50 of 26 nM, cooperativity α~30. The R69A mutation reduced cooperativity to ~3-4. [1]
Caco-2 permeability assay: ACBI1 was dissolved in HTP-4 buffer with 0.25% BSA (0.1-300 μM, 0.5% DMSO). Transport solution applied to apical or basolateral side of Caco-2 cell monolayers (seeded on filter inserts, cultured 10-25 days). Samples collected from donor and receiver sides over 2 h, analyzed by HPLC-MS/MS. Permeability rates: A-B = 2.2×10⁻⁶ cm/s, B-A = 3.8×10⁻⁶ cm/s, efflux ratio = 1.7. [1]
In vitro enzyme/receptor binding protocols for ACBI1 involve assessing its binding to VHL and target proteins (SMARCA2/4 bromodomains). Binding affinity is measured using techniques such as SPR or ITC. The compound is incubated with VHL and target proteins in appropriate buffers. For ternary complex formation assays, ACBI1, VHL, and target protein are incubated together, and complex formation is detected by pull-down or FRET-based methods.

Cell Proliferation Assay[1]
Cell Types: MV-4-11, NCI-H1568 cells
Tested Concentrations: 0-1000 nM
Incubation Duration: 3-7 days
Experimental Results: demonstrated anti-proliferative activity with IC50 of 29, 68 nM for MV-4-11, NCI-H1568 cells, respectively.

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Apoptosis Analysis[1]
Cell Types: SK-MEL-5 cells
Tested Concentrations: 0.3 µM
Incubation Duration: 100 h
Experimental Results: Induced apoptosis in SK-MEL-5 cells.

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Cell viability/antiproliferative assay: Cells (1000/well in 384-well plates) seeded overnight, then ACBI1 added at logarithmic dose series using a digital dispenser, with DMSO normalization. After 1 day and 8 days, cellular ATP content measured via CellTiterGlo. Fold proliferation calculated as day 8/day 1 measurement. For MV-4-11, SK-MEL-5, NCI-H1568, and NCI-H1703 cells. [1]
Protein degradation assay (capillary electrophoresis): Cells (20,000/well in 24-well plates) treated with ACBI1 for indicated times, then lysed (1% Triton, 350 mM KCl, 10 mM Tris pH 7.4, protease/phosphatase inhibitors, 10 mM DTT, benzonase). SMARCA2, SMARCA4, PBRM1 levels determined on a capillary electrophoresis instrument using specific rabbit antibodies and normalized to GAPDH. DC50 values calculated. [1]
Western blot for degradation: Cells (NCI-H1568 in 6-well plates, MV-4-11 in 10 cm plates) treated with ACBI1, lysed (1% Triton X-100, 150 mM NaCl, 1 mM EDTA, 50 mM Tris pH 7.4, protease inhibitors, benzonase). Proteins separated by SDS-PAGE, transferred to nitrocellulose, detected with anti-SMARCA2, anti-SMARCA4, anti-PBRM1, anti-β-actin antibodies, and IRDye secondary antibodies, imaged. [1]
Apoptosis assay (IncuCyte): SK-MEL-5 cells (80,000/well in 12-well plates) seeded with Caspase-3/7 Green Reagent (1:1000), incubated overnight, then treated with ACBI1 (0.3 μM). Phase object confluence and green object confluence measured every 2 h. Apoptosis normalized to proliferation (green area/proliferation area ×1000). PARP cleavage detected by Western blot using anti-PARP antibody. [1]
TMT proteomics: MV-4-11 cells (5×10⁶ per 100 mm plate) treated with 333 nM ACBI1 or cis-ACBI1 for 8 h (triplicate). Cells lysed in 100 mM Tris pH 8.0, 4% SDS with protease inhibitors. 200 μg protein processed by FASP, alkylated, trypsin-digested, desalted, labeled with TMT 10plex, pooled, fractionated by high-pH reverse-phase chromatography (80 fractions concatenated to 20). Analyzed by LC-MS/MS (Q Exactive HF) with data-dependent acquisition. Data searched with MaxQuant (1% FDR). [1]
Immunoprecipitation-MS: MV-4-11 cells treated with 1 μM ACBI1 or cis-ACBI1 for 8 h or 18 h. Anti-SMARCA2 or anti-ARID1A antibodies crosslinked to Dynabeads. Cell lysates (750 μg protein) incubated with beads for 4 h, washed, eluted with 7.5% SDS, reduced, alkylated, cleaned by SP3, trypsin-digested. Peptides analyzed by LC-MS/MS (Orbitrap Velos). [1]
In vitro cell-based assays for ACBI1 involve treating cells (e.g., MV-4-11, A549) with varying concentrations of the compound (typically 0.1 nM to 10 µM) for 4-24 hours. Target protein degradation is assessed by Western blotting for SMARCA2, SMARCA4, and PBRM1. Cell viability is measured using MTT or CellTiter-Glo assays after 72 hours of treatment. Apoptosis is assessed by Annexin V staining or caspase activation assays. DC₅₀ values are calculated from dose-response curves.

In vivo animal study protocols for ACBI1 would involve administration in suitable animal models (e.g., xenograft models). The compound would be formulated in appropriate vehicles (e.g., PEG400, saline) and administered via oral gavage or intraperitoneal injection. Dosing regimens would be determined based on PK data. Tumor growth inhibition, target degradation in tumors, and body weight changes would be monitored. Pharmacodynamic and efficacy endpoints would be assessed at study termination.

Caco-2 permeability: A-B permeability = 2.2×10⁻⁶ cm/s, B-A permeability = 3.8×10⁻⁶ cm/s, efflux ratio = 1.7. [1]
Pharmacokinetic data for ACBI1 is not extensively reported. As a PROTAC molecule (molecular weight typically >700 Da), it may have moderate oral bioavailability and tissue distribution. The compound would be expected to have reasonable plasma stability and cellular permeability. Detailed PK parameters such as half-life, Cmax, and AUC would need to be determined experimentally in relevant species. Formulation optimization may be required for in vivo studies.

Toxicological data for ACBI1 is not extensively reported. As a targeted protein degrader, toxicity would be related to its on-target effects and off-target degradation. Standard toxicology studies in animal models would be required for any therapeutic development. The compound is for research use only. No specific toxicity data is available. Standard safety precautions for handling PROTAC compounds should be observed.

[1]. BAF complex vulnerabilities in cancer demonstrated via structure-based PROTAC design. Nat Chem Biol. 2019 Jul;15(7):672-680.

ACBI1 is a first-in-class chemical tool for acute and profound SMARCA2/4 knockdown, developed via structure-based PROTAC design. It induces cooperative ternary complex formation between VHL and SMARCA2/4, with key protein-protein interactions involving R69 of VHL and the BC loop of SMARCA2BD. Degradation depends on VHL binding, neddylation, and proteasome activity (sensitive to VHL inhibitor and proteasome inhibitor). ACBI1 also degrades PBRM1 (fifth bromodomain) but overexpression of PBRM1 did not rescue antiproliferative effects, while SMARCA2 overexpression attenuated them, indicating that SMARCA2 loss drives the phenotype. The compound demonstrates exquisite selectivity for SMARCA2, SMARCA4, and PBRM1 as shown by unbiased proteomics. [1]
ACBI1 (CAS#: 2375564-55-7) is a VHL-recruiting PROTAC targeting SMARCA2, SMARCA4, and PBRM1. It is used in research on SWI/SNF BAF complex biology and targeted protein degradation. DC₅₀ values are 6 nM, 11 nM, and 32 nM for SMARCA2, SMARCA4, and PBRM1, respectively, in MV-4-11 cells. It induces anti-proliferative effects and apoptosis. It is not approved for therapeutic use and is not in clinical trials.

935.42

C, 62.87; H, 6.25; F, 2.03; N, 13.47; O, 11.96; S, 3.42

White to yellow solid powder

ACBI1; ACBI-1; ACBI 1;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: (1). This product requires protection from light (avoid light exposure) during transportation and storage.  (2). Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture.

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

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Solubility in Formulation 1: ≥ 2.5 mg/mL (2.67 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 25.0 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: ≥ 2.5 mg/mL (2.67 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 25.0 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.)