Compound (SMILES): Cc1ncsc1-c1cnc(C(C)NC(=O)[C@@H]2C[C@@H](O)CN2C(=O)C(c2cc(OCCN3CCC(n4nc(N)c5nnc(-c6ccccc6O)cc54)CC3)no2)C(C)C)cn1
Modality: PROteolysis-TArgeting Chimera (PROTAC)
Target protein (POI): SMARCA4 (UniProt P51532)
POI sequence: MSTPDPPLGGTPRPGPSPGPGPSPGAMLGPSPGPSPGSAHSMMGPSPGPPSAGHPIPTQGPGGYPQDNMHQMHKPMESMHEKGMSDDPRYNQMKGMGMRSGGHAGMGPPPSPMDQHSQGYPSPLGGSEHASSPVPASGPSSGPQMSSGPGGAPLDGADPQALGQQNRGPTPFNQNQLHQLRAQIMAYKMLARGQPLPDHLQMAVQGKRPMPGMQQQMPTLPPPSVSATGPGPGPGPGPGPGPGPAPPNYSRPHGMGGPNMPPPGPSGVPPGMPGQPPGGPPKPWPEGPMANAAAPTSTPQKLIPPQPTGRPSPAPPAVPPAASPVMPPQTQSPGQPAQPAPMVPLHQKQSRITPIQKPRGLDPVEILQEREYRLQARIAHRIQELENLPGSLAGDLRTKATIELKALRLLNFQRQLRQEVVVCMRRDTALETALNAKAYKRSKRQSLREARITEKLEKQQKIEQERKRRQKHQEYLNSILQHAKDFKEYHRSVTGKIQKLTKAVATYHANTEREQKKENERIEKERMRRLMAEDEEGYRKLIDQKKDKRLAYLLQQTDEYVANLTELVRQHKAAQVAKEKKKKKKKKKAENAEGQTPAIGPDGEPLDETSQMSDLPVKVIHVESGKILTGTDAPKAGQLEAWLEMNPGYEVAPRSDSEESGSEEEEEEEEEEQPQAAQPPTLPVEEKKKIPDPDSDDVSEVDARHIIENAKQDVDDEYGVSQALARGLQSYYAVAHAVTERVDKQSALMVNGVLKQYQIKGLEWLVSLYNNNLNGILADEMGLGKTIQTIALITYLMEHKRINGPFLIIVPLSTLSNWAYEFDKWAPSVVKVSYKGSPAARRAFVPQLRSGKFNVLLTTYEYIIKDKHILAKIRWKYMIVDEGHRMKNHHCKLTQVLNTHYVAPRRLLLTGTPLQNKLPELWALLNFLLPTIFKSCSTFEQWFNAPFAMTGEKVDLNEEETILIIRRLHKVLRPFLLRRLKKEVEAQLPEKVEYVIKCDMSALQRVLYRHMQAKGVLLTDGSEKDKKGKGGTKTLMNTIMQLRKICNHPYMFQHIEESFSEHLGFTGGIVQGLDLYRASGKFELLDRILPKLRATNHKVLLFCQMTSLMTIMEDYFAYRGFKYLRLDGTTKAEDRGMLLKTFNEPGSEYFIFLLSTRAGGLGLNLQSADTVIIFDSDWNPHQDLQAQDRAHRIGQQNEVRVLRLCTVNSVEEKILAAAKYKLNVDQKVIQAGMFDQKSSSHERRAFLQAILEHEEQDESRHCSTGSGSASFAHTAPPPAGVNPDLEEPPLKEEDEVPDDETVNQMIARHEEEFDLFMRMDLDRRREEARNPKRKPRLMEEDELPSWIIKDDAEVERLTCEEEEEKMFGRGSRHRKEVDYSDSLTEKQWLKAIEEGTLEEIEEEVRQKKSSRKRKRDSDAGSSTPTTSTRSRDKDDESKKQKKRGRPPAEKLSPNPPNLTKKMKKIVDAVIKYKDSSSGRQLSEVFIQLPSRKELPEYYELIRKPVDFKKIKERIRNHKYRSLNDLEKDVMLLCQNAQTFNLEGSLIYEDSIVLQSVFTSVRQKIEKEDDSEGEESEEEEEGEEEGSESESRSVKVKIKLGRKEKAQDRLKGGRRRPSRGSRAKPVVSDDDSEEEQEEDRSGSGSEED
E3 ligase: VHL (UniProt P40337)
Ligase sequence: MPRRAENWDEAEVGAEEAGVEEYGPEEDGGEESGAEESGPEESGPEELGAEEEMEAGRPRPVLRSVNSREPSQVIFCNRSPRVVLPVWLNFDGEPQPYPTLPPGTGRRIHSYRGHLWLFRDAGTHDGLLVNQTELFVPSLNVDGQPIFANITLPVYTLKERCLQVVRSLVKPENYRRLDIVRSLYEDLEDHPNVQKDLERLTQERIAHQRMGD
Cell line: SW1573 (Homo sapiens)
Cell line description: African=0.36%; Native American=0.18%; East Asian, North=0.45%; East Asian, South=0%; South Asian=0%; European, North=66.92%; European, South=32.09%.
Gene deletion; HGNC; HGNC Gene deletion; HGNC; HGNC Mutation; HGNC; HGNC Mutation; HGNC; HGNC Mutation; HGNC; HGNC Mutation; HGNC; HGNC
A*02
Stable (MSS) (Sanger).
In situ; Lung; 
Caucasian.
Cancer cell line
Assay: In-Cell Western
DC50: > 10000 nM
Assay description: In-cell Western blotting was used to examine the effects of compounds on BRM and BRG1 degradation.
Methods:
Cells were seeded into 384-well plates at varying concentrations (A549: 5000 cells/well, SW1573: 8000 cells/well, both purchased from ATCC) and incubated overnight at 37°C, 5% CO?. Compounds were dissolved in DMSO (initial concentration: 10 mM) and diluted sequentially with DMSO and culture medium (DMEM medium containing 10% FBS and 1% Penicillin-Streptomycin Solution) to a final concentration of 10 μM. Three-fold dilutions were performed for a total of 10 doses. Cells were incubated at 37°C, 5% CO? for 16-18 hours. Wash each well of a 384-well plate with 50 μL of pre-chilled PBS; discard the solution, add 50 μL of 4% paraformaldehyde fixative, and incubate at room temperature for 20 minutes; discard the fixative, add 50 μL of 0.1% Triton X-100 in PBS, and incubate at room temperature for 30 minutes; discard the solution, add 50 μL of Li-Cor blocking buffer, and incubate at room temperature for 1 hour; discard the blocking buffer, add 50 μL of primary antibodies (rabbit anti-BRM/BRG1 (1:500; BRM antibody; BRG1 antibody); mouse anti-α-Tubulin (1:2000)) and incubate overnight at 4°C. The primary antibody was discarded, and the cells were washed four times with 50 μL of PBST. The solution was discarded, and 50 μL of secondary antibody (goatα-rabbit IRDye 800CW; goatα-mouse IRDye 680RD (1:5000)) was added and incubated at room temperature in the dark for 1-1.5 hours. The secondary antibody was discarded, and the cells were washed four times with 50 μL of PBST, and the solution was discarded. Fluorescence scanning was performed using a Sapphire dual-mode multispectral laser imaging system. The fluorescence intensity of each well was converted to a numerical value using Azure Spot analysis software. XLfit was used to calculate the inhibitory effect of each compound concentration on BRM and BRG1 protein degradation based on the compound concentration and fluorescence intensity values.
The degradation activity of the compounds of the present disclosure against BRM and BRG1 was tested according to the above-described assay method. Some activity data are summarized in Table 2.
Note: "-" indicates not tested; "NDE" indicates no degradation was observed at the tested concentration.
Reference: BRM SELECTIVE DEGRADATION AGENT COMPOUND AND USE THEREOF (patent)
Source database: TPDdb