Evaluation of TLR7 Agonist SZU-101 Activity

The TLR7 agonist SZU-101 was synthesised as described in Additional file 2: Figure S2. HEK-BLUE hTLR7 cells were purchased from InvivoGen. The cells stably expressed human TLR7 and a SEAP reporter, which can be used to detect TLR7 agonism through the activation of NF-kB signalling. The cells were maintained in selective DMEM growth medium with an additional 10 μg/ml blasticidin and 100 μg/ml Zeocin™. After incubation with different doses of SZU-101, the cells were tested using the HEK-BLUE detection kit according to the manufacturer’s instructions. The TLR7 agonist imiquimod and purified mouse TNF-α were used as positive controls. The induction of TLR7 activation can be visualised and assessed by reading the OD at 620–655 nm.