293t cell

H1299 and A549 NSCLC cells were obtained from American Type Culture Collection
(ATCC, Manassas, USA) and cultivated in RPMI-1640 medium (Invitrogen) with 10%
fetal calf serum (FCS, Biosera, Boussens, France) as reported.^{18 (link)} Human bronchial 16HBE cell line (Procell, Wuhan, China) was used as a
control of non-tumor cells and propagated using standard protocols provided by
Procell. 293 T cells (ATCC) were cultivated in maintain medium provided by ATCC
for dual-luciferase reporter assays.
DTX-resistant NSCLC cells (A549/DTX and H1299/DTX) were established in our
laboratory by treating A549 and H1299 cells with gradually increasing
concentrations of DTX (Sigma-Aldrich, Steinheim, Germany; starting from 20 ng/L
and progressively increasing the concentration up to 10 µg/L) more than 9 months
until they acquired the ability to grow in the presence of DTX at the same rate
as parental cells in the absence of the drug. To maintain the resistance
phenotype, additional 10 µg/L of DTX was used in the cell medium.

Recombinant AAV8 vectors were generated as previously described⁴⁵ (link) with several modifications.⁴⁶ (link) Plasmids required for AAV packaging, adenoviral helper plasmid pAdDeltaF6 (PL-F-PVADF6), and AAV8 packaging plasmid pAAV2/8 (PL-T-PV0007) were obtained from the University of Pennsylvania Vector Core. Each AAV transgene construct was co-transfected with the packaging constructs into 293T cells (ATCC, CRL-3216) using polyethylenimine (PEI). Cell pellets were harvested and purified using a single cesium chloride density gradient centrifugation. Fractions containing AAV vector genomes were pooled and then dialyzed against PBS using a 100 kD Spectra-Por Float-A-Lyzer G2 dialysis device (Spectrum Labs, G235059) to remove the cesium chloride. Purified AAV were concentrated using a Sartorius Vivaspin Turbo 4 Ultrafiltration Unit (VS04T42) and stored at −80°C until use. AAV titers were calculated after DNase digestion using qPCR relative to a standard curve of the transgene plasmid. Primers used for titer are included in Table S3.

U87 MG cells and A172 cells were purchased from ATCC. U87 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM):F12 media (1:1) (ThermoFisher Scientific) with 10% FBS and 1% Glutamax. A172 cells were cultured in DMEM high glucose media (ThermoFisher Scientific) with 10% fetal bovine serum (FBS) and 1% Glutamax. GBM-110 and GBM 511 FHTC primary pediatric cancer cell lines were purchased from Fred Hutchinson Cancer Research Center and grown in predefined serum-free neural stem cell medium, supplemented with epidermal growth factor and fibroblast growth factor, on laminin (ThermoFisher Scientific) coated plates. Tissue culture plates were coated with 1 mL laminin working solution prepared in phosphate-buffered saline (PBS) (10 μg/mL) and incubated at 37°C for a minimum of 1 h. Plates were washed once with PBS before seeding cells. Membrane-bound interleukin 15 (IL15) and 41BB ligand-expressing K562 cells were kindly provided by Dr. Dario Campana (St. Jude Children’s Research Hospital, Memphis, TN) and cultured in RPMI 1640 media (ThermoFisher Scientific) with 10% FBS and 1% Glutamax. We purchased 293T cells from ATCC and were grown in DMEM high glucose media with 10% FBS and 1% Glutamax. Samples were obtained under the guidelines of the Institutional Review Board at CNH - Pro0004033 and CR00000266.