Gelatin

Feeder mouse embryonic fibroblasts (MEF cells) were seeded on gelatin (MP Biomedicals, Illkirch, France) in Dulbecco’s modified Eagle’s medium (DMEM, Sigma Aldrich, St. Lois, MO, USA) 3 h to 1 d prior to human ESC seeding. Human ESCs and library cells were cultured at 37 °C and 5% CO₂ on previously prepared MEF plates in human ES medium containing knockout DMEM (ThermoFisher Scientific, Paisley, UK) supplemented with 15% knockout serum replacement (KSR, ThermoFisher Scientific), 0.1 mM nonessential amino acids, 2 mM L-glutamine (Biological Industries, BI, Kibbutz Beit Haemek, Israel), 50 µg/mL streptomycin (BI), 50 units/mL penicillin (BI), 0.1 mM β mercaptoethanol, and 8 ng/mL basic fibroblast growth factor (bFGF). Alternatively, during the ATMi screen experiment and for FACS or CellTiter-Glo assays, cells were seeded on Matrigel-coated plates (BD Biosciences, Le Pont-de-Claix, France), in mTeSR1 media (STEMCELL Technologies, Vancouver, BC, Canada) supplemented with 1:1000 ROCK inhibitor Y-27632 (ROCKi, Stemgent, Boston, MA, USA) in the absence of feeder cells. HEK293T cells were cultured at 37 °C and 5% CO2 in DMEM supplemented with 10% fetal bovine serum (BI), 2 mM L-glutamine, 50 units/mL penicillin, and 50 µg/mL streptomycin. Cells were detached using trypsin (BI) or TRYPLE select (ThermoFisher Scientific).

Levonorgestrel was purchased from Cayman Chemicals, Bangalore, India. Soya phosphatidylcholine (SPC) and oleic acid were gifted and purchased from VAV life sciences, Mumbai, India, and Merck India Ltd., Mumbai, India, respectively. Gelatin and polyvinyl alcohol were purchased from MP Biomedical, Mumbai, India. Polydimethylsiloxane (PDMS) and elastomer were purchased from DuPont, Mumbai, India. All chemicals and reagents used for the study were analytical grade. All solvents used for analytical study were HPLC-grade.

GelMA hydrogels were prepared as previously described¹⁶ . Briefly, 10% (w/v) Gelatin (901,771; MPBiomedicals, USA) was dissolved in PBS at 50 °C for 1 h under constant stirring, to which 20% (w/v) of methacrylic anhydride (276,685; SigmaAldrich, USA) was added dropwise. Excess of PBS was added to stop the reaction and dialyzed against dH₂0 at 37 °C, followed by freeze-drying (Alpha1-2/LDplus, Martin Christ, Germany) and storage at -20 °C. Freeze-dried GelMA (10% w/v) was dissolved in PBS (10,010,023; Gibco, USA) along with a photo-initiator 2-hydroxy-4′-(2-hydroxyethoxy)-2-methylpropiophenone (Irgacure2959-410,896, Sigma-Aldrich, USA), at a concentration of 0.5 mg/mL followed by exposure to UV-365 for 10 min, to obtain hydrogels. These hydrogels were equilibrated with fresh media containing 10% FBS for 24 h prior to seeding of cells for experiments. Freshly dissociated cells were counted and seeded at a concentration of 5*10^{4 (link)} cells per well of a 24-well plate, on top of the hydrogels are were observed to be migrating inside the hydrogels, and spheroids were formed.

Gold chloride trihydrate (HAuCl₄, Sigma-Aldrich), sodium
citrate tribasic dihydrate (Sigma-Aldrich),
methyl polyethylene glycol thiol (mPEG-SH, MW: 2000, Biochempeg),
bovine serum albumin (BSA, Sigma-Aldrich), caspase-3 (CASP3, Abcam),
paraformaldehyde (PFA, Sigma-Aldrich), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide (MTT, EMD Millipore), dimethyl sulfoxide (DMSO, Sigma-Aldrich),
gelatin (MP Biomedicals), silica (0.25 μm diameter, Sigma-Aldrich),
potassium cyanide (KCN, Sigma-Aldrich), Dulbecco’s modified
Eagle’s medium (DMEM, Cytiva), fetal bovine serum (FBS, Sigma-Aldrich),
and penicillin/streptomycin (Sigma-Aldrich) were used in accordance
with the manufacturer’s instructions.

Gelatin was purchased from MP Biomedicals LLC (Irvine, CA, USA) and hexafluoroisopropanol (HFIP) from Darui Co., Ltd. (Shanghai, China). Then poly (L‐lactide‐co‐caprolactone) (P(LLA‐CL)) was provided by GUNZE Co., Ltd. (Kyoto, Japan).