Transcription factors are master regulators of cell state. Commonly activated by genetic events or upstream signalling pathways, they mediate the neoplastic phenotype and maintain tissue specification in cancer. As such, they are highly desirable targets for ligand discovery1
. Owing to expansive protein–protein interfaces and a general absence of hydrophobic pockets, transcription factors have proven among the most chemically intractable of all therapeutic targets. With the exception of nuclear hormone receptors that have evolved the ability to bind natural small-molecule ligands, potent and specific inhibitors of human transcription factors have not been realized. Here we report the successful development of a direct-acting antagonist of an oncogenic transcription factor, NOTCH1.
NOTCH proteins participate in conserved pathways that regulate cellular differentiation, proliferation and death2,3
. Mammalian NOTCH receptors (NOTCH1–4 in humans) are single-pass transmembrane proteins that transmit juxtacrine signals initiated by ligands of the Delta, Serrate or Lag-2 family. Ligand binding to the extracellular domain of NOTCH1 initiates sequential proteolytic processing events catalysed respectively by an ADAM family metalloprotease and a γ-secretase complex, resulting in cytoplasmic release of the intracellular domain of NOTCH1 (ICN1)4–6
. ICN1 then translocates to the nucleus and loads onto the DNA-bound transcription factor CSL7
. The engagement of ICN1 with CSL creates a long, shallow groove along the interface of the two proteins that serves as a binding surface for co-activator proteins of the mastermind-like (MAML) family8,9
. The resulting ICN–CSL–MAML ternary complex then recruits the core transcription machinery, effecting activation of NOTCH-dependent target genes.
The duration and strength of NOTCH signalling is normally tightly controlled. Whereas loss-of-function mutations have been observed in a variety of diseases10–12
, gain-of-function mutations in the NOTCH pathway are causally linked with cancer. Indeed, human NOTCH1
was first discovered owing to its involvement in a t(7;9) chromosomal translocation observed in patients with T-ALL13
. Subsequently, various activating mutations in NOTCH1
have been discovered in >50% of patients with T-ALL14
. Recently, further aberrations that potentiate NOTCH signalling have been identified, including loss-of-function of the NOTCH1 E3 ubiquitin ligase FBXW7 and the intracellular NOTCH inhibitor NUMB, in T-ALL and breast adenocarcinoma, respectively15,16
. Activated NOTCH signalling has also been observed in cancers of the lungs, ovaries, pancreas and gastrointestinal tract as well as in melanoma, multiple myeloma and medulloblastoma17–23
Efforts to antagonize the NOTCH pathway have relied on blocking the generation of ICN using small-molecule inhibitors of the γ-secretase complex (GSIs)24,25
. These molecules are not strictly NOTCH-specific, as they indiscriminately block the many signalling pathways downstream of γ-secretase26
. T-ALL patients treated with GSIs suffer dose-limiting gastrointestinal toxicity, the origin of which is uncertain but it may result from chronic blockade of NOTCH1 and NOTCH2 proccessing27
. Some cell lines containing activating NOTCH1
mutations are resistant to GSIs, and those that do respond commonly undergo growth arrest rather than apoptosis. These observations underscore the mechanistic utility and potential therapeutic value of NOTCH antagonists that act by directly targeting the NOTCH transactivation complex.