Advancing targeted protein degradation for cancer therapy

Abstract

The human proteome contains approximately 20,000 proteins, and it is estimated that more than 600 of them are functionally important for various types of cancers, including nearly 400 non-enzyme proteins that are challenging to target by traditional occupancy-driven pharmacology. Recent advances in the development of small-molecule degraders, including molecular glues and heterobifunctional degraders such as proteolysis-targeting chimeras (PROTACs), have made it possible to target many proteins that were previously considered undruggable. In particular, PROTACs form a ternary complex with a hijacked E3 ubiquitin ligase and a target protein, leading to polyubiquitination and degradation of the target protein. The broad applicability of this approach is facilitated by the flexibility of individual E3 ligases to recognize different substrates. The vast majority of the approximately 600 human E3 ligases have not been explored, thus presenting enormous opportunities to develop degraders that target oncoproteins with tissue, tumour and subcellular selectivity. In this Review, we first discuss the molecular basis of targeted protein degradation. We then offer a comprehensive account of the most promising degraders in development as cancer therapies to date. Lastly, we provide an overview of opportunities and challenges in this exciting field.

Fig. 1 |. Key discoveries and developments in the targeted protein degradation field.

The figure shows a timeline beginning from the first publication of the proteolysis-targeting chimera (PROTAC) concept up to the most recent discoveries. AR, androgen receptor; CRBN, cereblon; DCAF15, DDB1- and CUL4-associated factor 15; ER, oestrogen receptor; EZH2, enhancer of Zeste homologue 2; IMiD, immunomodulatory imide drug; POI, protein of interest; STAT3, signal transducer and activator of transcription 3; VHL, von Elippel–Lindau tumour suppressor.

Fig. 2 |. The ubiquitin–proteasome system and classification of E3 ubiquitin ligases.

a | Ubiquitination begins with the ATP-dependent activation of ubiquitin (Ub) by the E1 ubiquitin-activating enzyme (UBA), resulting in Ub~E1 thioester bond formation (the tilde denotes a high-energy thioester bond), followed by the transfer of ubiquitin to an E2 ubiquitin-conjugating enzyme. Finally, an E3 ubiquitin ligase brings the substrate and the E2~Ub conjugate together and promotes ubiquitin ligation to a lysine residue in the substrate protein. Ubiquitin can be ligated either as a single moiety to one lysine residue or multiple lysine residues or as a polyubiquitin chain where successive ubiquitin molecules are connected. The consequences of ubiquitination for the substrate protein depend not only on whether it is monoubiquitinated or polyubiquitinated but also on the topology of the polyubiquitin chain. One of the well-known consequences of polyubiquitination is the proteolytic degradation of substrate proteins by the 26S proteasome. b | The three types of E3 ubiquitin ligases. Traditionally, E3 ubiquitin ligases are separated into homologous to E6AP carboxy terminus (HECT) and RING types. Both types interact with E2~Ub and recognize substrate proteins, but differ in their catalytic mechanisms. While the classical HECT E3 ligases form a thioester intermediate with ubiquitin on an active-site cysteine before transferring it onto its substrates, RING finger proteins do not form an intermediate with ubiquitin. Instead, they promote the direct transfer of ubiquitin from E2 to the substrate, in part by immobilizing the carboxy-terminal glycine of the ubiquitin in an otherwise highly flexible E2~Ub, for attack by the acceptor lysine^–. More recently, a third class of E3 ligases was identified that contains a tripartite RING1-in-between-RING2 (RBR) domain arrangement and catalyses ubiquitination through a RING–HECT hybrid catalytic mechanism. RING1 in the RBR domain forms a typical cross-brace RING structure to interact with E2~Ub. The ubiquitin is then transferred to, and forms a thioester intermediate with, an active-site cysteine in RING2, followed by the transfer of ubiquitin to a substrate. c | Classification of RING-type E3 ligases. RING type E3 ligases can be either a single subunit, which contain an intrinsic RING finger domain, or a multi-subunit complex assembled on the cullin scaffold that does not contain a RING finger domain but instead binds in trans a small RING finger protein, either RING box protein 1 (ROC1) or ROC2. See BOX 1 for details of the cullin–RING E3 ubiquitin ligase assembly. CRBN, cereblon; VHL, von Hippel–Lindau tumour suppressor.

Fig. 3 |. PROTAC degraders.

a | General schematic of proteolysis-targeting chimera (PROTAC) degraders. A protein of interest (POI) can be degraded by the ubiquitin-proteasome system mediated by a PROTAC, which consists of a POI ligand connected to an E3 ligand via a linker. PROTACs are catalytic. The tilde denotes a high-energy thioester bond. b | PROTACs induce new protein–protein interactions between the POI and the E3 ligase. The image shows an overlay of two SMARCA2 bromodomain–PROTAC–von Hippel–Lindau tumour suppressor (VHL)–elongin C–elongin B complex crystal structures (RCSB Protein Data Bank IDs 6HAX and 6HAY) in a ribbon representation. PROTAC 1 and PROTAC 2 share the same VHL ligand and POI ligand but have different linkers, which are specified in the box. The colours within the chemical structure (PROTAC 1 and PROTAC 2) correspond to the colours of the compounds in the crystal (ribbon) structures, with the VHL ligand in blue, the linker in green and the SMARCA2 bromodomain ligand in magenta. c | Chemical structures of several key heterobifunctional degraders highlighted in this Review. ARD-69, SJF-0628 and DT2216 are VHL-based PROTAC degraders, SS47 and SD-36 are cereblon (CRBN)-based PROTAC degraders and MS1943 is a hydrophobic tag-based degrader. AR, androgen receptor; EZH2, enhancer of Zeste homologue 2; STAT3, signal transducer and activator of transcription 3; Ub, ubiquitin.

PDB, Protein Data Bank; ub, ubiquitin.

BLCA, bladder urothelial carcinoma; BRCA, breast invasive carcinoma; COAD, colon adenocarcinoma; FPKM, fragments per kilobase of transcript per million mapped reads; HNSC, head and neck squamous cell carcinoma; iBAQ, intensity Based Absolute Quantification; KICH, kidney chromophobe; KIRC, kidney renal clear cell carcinoma; KIRP, kidney renal papillary cell carcinoma; LIHC, liver hepatocellular carcinoma; LUAD, lung adenocarcinoma; LUSC, lung squamous cell carcinoma; PRAD, prostate adenocarcinoma; READ, rectum adenocarcinoma; STAD, stomach adenocarcinoma; THCA, thyroid carcinoma; UCEC, uterine corpus endometrial carcinoma.