In Silico Characterization of Structural Distinctions between Isoforms of Human and Mouse Sphingosine Kinases for Accelerating Drug Discovery

Abstract

Alterations in cellular signaling pathways are associated with multiple disease states including cancers and fibrosis. Current research efforts to attenuate cancers, specifically lymphatic cancer, focus on inhibition of two sphingosine kinase isoforms, sphingosine kinase 1(SphK1) and sphingosine kinase 2 (SphK2). Determining differences in structural and physicochemical binding site properties of SphKs is attractive to refine inhibitor potency and isoform selectivity. This study utilizes a predictive in silico approach to determine key differences in binding sites in SphK isoforms in human and mouse species. Homology modeling, molecular docking of inhibitors, analysis of binding pocket residue positions, development of pharmacophore models, and analysis of binding cavity volume were performed to determine isoform-and species-selective characteristics of the binding site and generate a system to rank potential inhibitors. Interestingly, docking studies showed compounds bound to mouse SphK1 in a manner more similar to human SphK2 than to human SphK1, indicating that SphKs in mice have structural properties distinct from humans that confounds prediction of ligand selectivity in mice. Our studies aid in the development and production of new compound classes by highlighting structural distinctions and identifying the role of key residues that cause observable, functional differences in isoforms and between orthologues.

Discovery of a Small Side Cavity in Sphingosine Kinase 2 that Enhances Inhibitor Potency and Selectivity

“…Molecular modeling studies conducted with similar SphK inhibitors possessing a guanidine head group predict that the guanidine moiety in our scaffold hydrogen-bonds to catalytic amino acid residue Asp211, flanking the ATP active site when docked in the Sph binding pocket of SphK2. 32,33 Structure−activity relationship (SAR) studies conducted by Congdon et al on the scaffold of 7 established that deletion of the aryl portion of the octylbenzene tail resulted in a significantly diminished inhibition toward SphK2, demonstrating that the aryl ring is essential for inhibitor efficacy. 21 Further SAR experiments focused on the tail portion led to the development of inhibitors 8 and 9 (SLM6031422) (Table 1).…”

“…The Sph binding site of SphKs can be divided up into three regions, as defined by Worrell et al They are as follows: (1) the head region adjacent to the ATP binding site, (2) the hydrophobic core region, and (3) the tail region. 33 Molecular docking of 8 and 9 demonstrate different ligand orientations in the Sph binding site of SphK2, giving insight into the role and influence the trifluoromethyl group has in regard to SphK2 selectivity (Figure 4). Compound 8 docks in the Sph binding pocket of SphK2 in a position that indicates more interactions in the tail region of the binding site, as influenced by the hydrophobic core of the pocket, in particular, the space for the trifluoromethyl group to sit between hydrophobic residues Phe548, Leu544, and Leu547 (Figure 4A,B).…”

Utilization of computational techniques and tools to introduce or reinforce knowledge of biochemistry and protein structure–function relationships

“…The open pedagogy design approach to UR in biochemistry and utilization of computational techniques reinforces wet‐lab experiences and have been successful in creating meaningful poster presentations and publications. In using this approach, to date, we have supported UR students to present in over 40 poster sessions at 10 different local, national, and international conferences and be authors on 5 peer‐reviewed research articles in notable journals 5–9 . This short communication highlights an open pedagogy design approach to UR in biochemistry and the utilization of computational techniques and tools to both reinforce wet‐lab experiences and provide routes for experiential learning and workforce development when in‐person laboratory work is not feasible.…”

Synthesis and Biological Evaluation of BODIPY-PF-543

“…Therefore, we introduced BODIPY, which generally shows a fluorescence value of 500 nm, into PF-543. Extensive docking studies of SK1 in several laboratories have demonstrated that a structurally bulky group can be docked at the tail of SK1 inhibitors [21]. Therefore, we hypothesized that the bulky structure of BODIPY does not significantly affect its SK inhibitory effect.…”