Reprogramming the Transcriptional Response to Hypoxia with a Chromosomally Encoded Cyclic Peptide HIF‑1 Inhibitor

Ishna N. Mistry, Ali Tavassoli

The cellular response to hypoxia is orchestrated by HIF-1, a heterodimeric transcription factor composed of an α and a β subunit that enables cell survival under low oxygen conditions by altering the transcription of over 300 genes. There is significant evidence that inhibition of HIF-1 would be beneficial for cancer therapy. We recently reported a cyclic hexapeptide that inhibits the HIF-1α/HIF-1β protein-protein interaction in vitro and prevents HIF-1-mediated hypoxia-response signaling in cells. This cyclic peptide was identified from a library of 3.2 × 10(6) members generated using SICLOPPS split-intein mediated protein splicing. With a view to demonstrating the potential for encoding the production of a therapeutic agent in response to a disease marker, we have engineered human cells with an additional chromosomal control circuit that conditionally encodes the production of our cyclic peptide HIF-1 inhibitor. We demonstrate the conditional production of our HIF-1 inhibitor in response to hypoxia, and its inhibitory effect on HIF-1 dimerization and downstream hypoxia-response signaling. These engineered cells are used to illustrate the synthetic lethality of inhibiting HIF-1 dimerization and glycolysis in hypoxic cells. Our approach not only eliminates the need for the chemical synthesis and targeted delivery of our HIF-1 inhibitor to cells, it also demonstrates the wider possibility that the production machinery of other bioactive compounds may be incorporated onto the chromosome of human cells. This work demonstrates the potential of sentinel circuits that produce molecular modulators of cellular pathways in response to environmental or cellular disease stimuli.