Target Mapping in Cancer: Ligandable Protein Pockets on 3D OncoPPI Networks

Abstract

Background/Objectives: Studying protein-protein interaction (PPI) networks is crucial in understanding cancer phenotypes and molecular mechanisms. Here, we focus on PPIs involved in 12 different types of cancer (oncoPPIs), highlighting those protein pockets serving as outposts to modulate protein functioning. Methods: To explore these cavities linked to the cancer phenotype changes, we built a comprehensive pocketome of 314 crystallographically solved oncoPPIs. Based on this experimental data, we identified and investigated all ligandable protein pockets by employing 3D geometric and energetic descriptors. These pockets were classified as suitable for designing new oncoPPI modulators or PROTACs. The ligand-bound crystallographic pockets were analyzed to compare their properties across cancer types. Finally, 3D oncoPPI networks were built for each cancer type to identify highly connected proteins acting as hubs. Results: Combining interaction networks with structural pocket data helps identify cancer-relevant proteins and key interacting residues. Using this approach, we present clinical examples (e.g., S100A1, NRP1, CTNNB1, VCP) to show the therapeutic value of targeting ligandable 3D oncoPPIs. We also provide a publicly available reference dataset supporting future research. Conclusions: Notably, this study offers a flexible framework for evaluating and prioritizing novel disease targets.

Keywords: 3D oncoPPI networks; PPIs modulators; PROTACs; ligandable pockets; pocketome analysis; target prioritization in cancer.

Figure 2

(a) Pocket detection on detached partner 1 (cyan), resulting in an allosteric-like pocket (pink surface) that does not include the other partner. (b) Pocket detection on detached partner 2 (orange), resulting in an interface pocket (green surface) that contains the other partner. (c) Pocket detection on the complexed partners, resulting in an equilibrium pocket (violet surface) defined by both partners. In the inset, a zoomed-in detail highlights the following: (d) on the left, the interface pocket (green surface) defined by residues of detached partner 2 and containing an alpha helix from detached partner 1 (cyan sticks), and (e) on the right, the equilibrium pocket (violet surface) defined by residues of both partners (orange and cyan sticks). (f) Histogram showing the number of pockets per cancer for each category.

Figure 1

Successful examples of predicted pockets in validation dataset. (a) Pocket at the interface of HDM2/P53 (PDB ID: 1ycr). (b) Pocket detected on HDM2 co-crystallized with inhibitor HTZ (PDB ID: 6q9l). (c) Pocket at the interface of CRBN/BRD4 with RN6 PROTAC (PDB ID: 6boy). (d) Pocket detected on BRD4 co-crystallized with inhibitor 0S6 (PDB ID: 4f3i). (e) Spider plots reporting the number of activated PPIs in Gulfidan et al. [8] work (2D-act, in grey) and the number of mapped activated interactions in PDB (3D-act, in pink). (f) The crystallographic complex of the most recurrent 3D oncoPPI, namely Erp57 and Tapasin (PDB ID: 3f8u). (g) Histogram of the number of activated 3D oncoPPIs per cancer. Pockets detected are displayed as surface and as mesh in the protein-bound form and in the ligand-bound form, respectively. Protein interactors are colored in cyan and orange cartoon, whereas ligands and PROTAC are colored in yellow and pink, respectively.

Figure 3

Interface (a,b), allosteric-like (c,d), and equilibrium (e–h) pockets. (a) The S100A1 homodimer is shown with two monomers in different shades of orange. The enlarged region highlights a pocket on one monomer that contains residues from the other (light orange sticks) (PDB ID: 5k89). (b) In the HIF1α–CREBBP heterodimer, HIF1α is depicted in blue and CREBBP in cyan. Two enlarged views are provided: (left) one shows a pocket on CREBBP containing HIF1α residues (blue sticks) and (right) the other shows a pocket on HIF1α containing CREBBP residues (cyan sticks) (PDB ID: 1l8c). (c) PKN1 is illustrated with its allosteric-like pocket, shown as a dark green cartoon with a pink surface; the interacting partner is represented in light grey (PDB ID: 1cxz). (d) ARPC2 displays its allosteric-like pockets as a light green cartoon with a pink surface, with the interacting partner in light grey (PDB ID: 6uhc). (e) The RHOA–AKAP13 heterodimer exhibits an equilibrium pocket (violet surface) that includes residues from both proteins. RHOA and AKAP13 are shown in red and yellow, respectively (PDB ID: 6bca). (f) The UBC–ADRM1 heterodimer features an equilibrium pocket (violet surface) encompassing residues from both partners highlighted in black dotted box (PDB ID: 5v1y). (g,h) A zoomed-in view of the UBC–ADRM1 heterodimer highlights the surface portion interacting with the equilibrium pocket: UBC residues interacting with ADRM1 are depicted as a dark cyan surface, while ADRM1 residues interacting with UBC are shown as a dark blue surface.

Figure 4

Examples of ligand-bound pockets showing specific binding regions where ligands interact with the protein. (a) Interface pocket in green detected on the detached partner NRP1 (cyan cartoon) in complex with VEGFA (orange cartoon) (PDB ID: 4deq). The detached partner NRP1 (dark cyan cartoon) is complexed with compound R40 (yellow stick) (PDB ID: 5iyy). (b) Allostericlike pocket in pink detected on the detached partner CTNNB1 (cyan cartoon) in complex with CTNNBIP1 (orange cartoon) (PDB ID: 1t08). The detached partner CTNNB1 (dark cyan cartoon) is complexed with compound R9Q (yellow stick) (PDB ID: 7afw). (c) Equilibrium pocket in violet found in the region between the FKBP1A and FKBP12–rapamycin-associated protein (MTOR) (PDB ID: 3fap) with rapamycin analog ARD (yellow stick) interacting simultaneously with the complexed partners. (d) Venn–Eulero diagram of crystallographic ligands found within interface, allosteric-like, and equilibrium pockets. (e) PCA scores and loadings plots showing the crystallographic ligands found within interface, allosteric-like, and equilibrium pockets. Ligands from interface pockets, such as 6K8, are marked in green; equilibrium pocket ligands, such as Z7T, in purple; and allosteric-like pocket ligands, such as 048, in pink. The lower right corner displays the PCA loadings plot, highlighting descriptors related to shape/size as well as hydrophobicity and hydrophilicity. Explained variance: PC1: 33.66%–PC2: 24.31%. (f) Pie chart reporting the classification of the clinical status of ligands found in the different categories of pockets.

Figure 5

(a) Distribution of ligand-bound pockets across various cancer types. The y-axis shows the number of ligand-bound pockets for each of the three pocket categories: interface (green bars), allosteric-like (pink bars), and equilibrium (violet bars). (b) Data bar representing the percentage of 3D oncoPPIs surfaces where at least one ligand-bound pocket was found, both with interface pocket and with equilibrium pocket.

Figure 6

(a–c) Interface pocket of the PCNA protein dimer interaction (PDB ID: 1u76), with a T2B ligand extracted from a holo structure (PDB ID: 3wgw) and the corresponding MIFs. (b,d) Allosteric-like pocket on BRAF (PDB ID: 6u2h) and the corresponding MIFs. Protein and pocket residues defining the pocket are displayed as cartoon and stick representations, respectively. The ligand bound is displayed as grey stick representations. Hydrophobic, hydrogen bonding donor, and hydrogen bonding acceptor MIFs are displayed as yellow, blue, and red surfaces, respectively. The probes used are as follows: CRY for hydrophobic (energy threshold = −1.0 Kcal/mol), N1 for hydrogen bonding donor (energy threshold = −4.5 Kcal/mol), and O for hydrogen bonding acceptor (energy threshold = −4.5 Kcal/mol).

Figure 7

(a) Distribution of the number of interactors per protein. This histogram illustrates the frequency of proteins based on the number of crystallographic interactors they engage with, highlighting the distinction between single-interactor proteins and multi-interacting hub proteins across the dataset. Network of the most populated 3D oncoPPI crystallographic hub (PCNA). (b) Histogram of hub proteins per cancer type. Each bar represents the number of hub proteins identified in each cancer type. The color gradient within the bars reflects the degree of connectivity, ranging from hubs with fewer interactors (light green) to those with extensive interaction networks (dark green).

Figure 8

Network representations of 12 cancer types. In each network, nodes represent proteins and edges represent interactions. Node color reflects the total number of pockets (interface + allosteric-like): yellow indicates few pockets, and purple indicates many pockets. Node size represents the number of interface pockets, with larger nodes signifying a higher count of interface pockets. Node size is scaled within each individual network and is not comparable across networks. Nodes with red labels are proteins lacking interface pockets. Nodes without any shape indicate proteins with no pockets (neither interface nor allosteric-like pockets).

Figure 9

(a) The VCP interactors are represented in the VCP-centered network, highlighting partners that show at least one pocket at the interface. ASPSCR1 interactor is depicted in yellow (left). VCP hexamer is composed of different domains. One monomer is displayed in cyan cartoon. ATP binding sites and allosteric binding sites are represented in black and dark grey spheres, respectively. The new site identified in our analysis is represented in green spheres (right). (b) Interaction between the two monomers of VCP (PDB ID: 4ko8), shown in purple and cyan cartoon, respectively. A pocket identified at this interaction interface is illustrated as a dark green mesh. (c) Interaction between VCP and ASPSCR1 (PDB ID: 5ifs), depicted in green and yellow cartoon, respectively. The identified pocket on VCP interacting with ASPSCR1 is highlighted as a light green surface. (d) Overlap of the two identified pockets on two different VCP interactions: the pocket involved in homodimerization (represented as a dark green mesh) and the pocket mediating interaction between the monomer and ASPSCR1 (depicted as a light green surface). Residues involved in these interactions are highlighted in stick and line representation.