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. 2023 Jan 3;28(1):446.
doi: 10.3390/molecules28010446.

New Class of Benzodiazepinone Derivatives as Pro-Death Agents Targeting BIR Domains in Cancer Cells

Michele Fiore  1 , Michele Mosconi  2 , Francesco Bonì  2 , Alice Parodi  3 , Annalisa Salis  3 , Bruno Tasso  4 , Eloise Mastrangelo  2 , Enrico Millo  3 , Federica Cossu  2
Affiliations

New Class of Benzodiazepinone Derivatives as Pro-Death Agents Targeting BIR Domains in Cancer Cells

Michele Fiore et al. Molecules. .

Abstract

Inhibitor of Apoptosis Proteins (IAPs) are validated targets for cancer therapy, and the deregulation of their activities within the NF-κB pathway correlates with chemoresistance events, even after treatment with IAPs-antagonists in the clinic (Smac-mimetics). The molecule FC2 was identified as a NF-κB pathway modulator in MDA-MB-231 adenocarcinoma cancer cells after virtual screening of the Chembridge library against the Baculoviral IAP Repeat 1 (BIR1) domain of cIAP2 and XIAP. An improved cytotoxic effect is observed when FC2 is combined with Smac-mimetics or with the cytokine Tumor Necrosis Factor (TNF). Here, we propose a library of 22 derivatives of FC2, whose scaffold was rationally modified starting from the position identified as R1. The cytotoxic effect of FC2 derivatives was evaluated in MDA-MB-231 and binding to the cIAP2- and XIAP-BIR1 domains was assessed in fluorescence-based techniques and virtual docking. Among 22 derivatives, 4m and 4p display improved efficacy/potency in MDA-MB-231 cells and low micromolar binding affinity vs the target proteins. Two additional candidates (4b and 4u) display promising cytotoxic effects in combination with TNF, suggesting the connection between this class of molecules and the NF-κB pathway. These results provide the rationale for further FC2 modifications and the design of novel IAP-targeting candidates supporting known therapies.

Keywords: MDA-MB-231; NF-κB (nuclear factor kappa light chain enhancer of activated B cells); baculoviral IAP repeat (BIR); benzodiazepinone; inhibitor of apoptosis protein (IAP); virtual docking.

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Conflict of interest statement

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Figures

Scheme 1
Scheme 1
Reagents and conditions: (a) EtOH:H2O (1:1) 80 °C, 3 h, (b) R1CHO, EtOH, acetic acid, RT, 24 h. For complete structures see Table 1.
Figure 1
Figure 1
The Baculoviral IAP Repeat (BIR) domain is composed of three to five α-helices and a central three-stranded β-sheet. In type I BIRs, the hotspot targeted by FC2 (grey sticks in cIAP2, deep blue sticks in XIAP) corresponds to the N-terminal α-helices (a). The sequence of the structural hotspot (α1-turn-α2) is conserved among homologous type I BIRs, as shown in the sequence alignment (b) performed with Clustal [22]. Several electrostatic interactions (c, blue lines) and hydrophobic contacts (c, grey dashed lines) between residues (c, points) from α1 (blue shade), the turn (green-yellow shades) and α2 (red shade) stabilize the folding of the structural hotspot in a conservative manner among homologous BIRs (the numbering of the residues is referred to the deposited Protein Data Bank structures 3M0A [15], chain D for cIAP2 and 2POP [16], chain B for XIAP; panel b and c were adapted with permission from Pymol [23] and RING 2.0 [24]).
Figure 2
Figure 2
The FC2 and FC2 derivative portions, the benzoyl, the benzodiazepinone central scaffold and the R1 groups (a) map a set of conserved residues in the target hotspot. Compounds 4m and 4p poses (pink and teal, respectively) in the BIR1 domains of cIAP2 (b) and XIAP (c) maintain the interaction network of FC2 (grey lines) with the residues (highlighted in lines) of the structural hotspot α1-turn-α2 (in rainbow cartoons). In the zoomed view (left side of panel b), R1 substitutions with electron withdrawing groups (4m, 4p) engage novel interactions with key residues of the turn and with E47 (4p) in cIAP2. In XIAP, both 4m and 4p gain novel interactions between R1 and S38 side chain and between the carboxy moiety of the scaffold and S38 backbone (red dashed lines). The structures were drawn with Pymol [23].
See this image and copyright information in PMC

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