Molecular docking analysis of sphingosine kinase 1 inhibitors for cancer management

Sphingosine kinase 1 (SK1) catalyses the conversion of sphingosine to the signalling mediator sphingosine 1-phosphate. This is essential for cell survival and proliferation. SK1 is frequently overexpressed in various cancer types, promoting tumor progression. SK1 has been well documented as a promising target for anticancer therapy. In this study, a virtual screening approach was used to screen a total of 1068 natural compounds, with the aim of identifying potential inhibitors of SK1. The top hit compounds, namely CNP0296172, CNP0368143, CNP0380570, and CNP0290815, were selected based on their strong binding affinity and specificity towards the SK1 binding pocket. Notably, these selected hit compounds exhibited a higher affinity towards the SK1 binding pocket when compared to the positive control compound (PF-543). Furthermore, these compounds were found to meet the necessary drug like criteria, thus rendering them suitable candidates for further experimental validation as potential anticancer agents.


Background:
Cancer is a prevalent disease that imposes a significant burden on the healthcare system.It is characterized by unregulated cell proliferation caused by genetic mutations, which results in malignant tumors [1].The incidence and mortality rates of cancer increase exponentially, with an estimated 19.3 million new cases and around 10 million deaths in 2020 [2].Although conventional treatments such as radiation therapy, chemotherapy, and surgical resection are commonly used to manage cancer, their effectiveness is limited due to cellular resistance [3].
Sphingolipids are important components of cellular membranes, contributing to their structure as well as function [4].Sphinganine, ceramide, sphingosine, and sphingosine 1-phosphate (S1P) are metabolites that play important roles in a variety of biological processes, including cell proliferation, differentiation, and apoptosis.Furthermore Through a comprehensive analysis utilizing the Genevisible web tool (http://genevisible.com), a noteworthy expression of SK1 in cancer cases of both humans and mice has been observed (Figure 1).The results indicate that SK1 exhibits abundant expression in various cancer types, including prostate cancer, malignant neoplasms, infiltrating duct carcinoma of the breast, as well as other types of cancers in humans.Similarly, in mice, SK1 demonstrates high expression levels in breast cancer, pancreatic cancer, and infiltrating duct carcinoma.These findings highlight the potential relevance of SK1 as a target for therapeutic interventions in cancer treatment.Drug development is a difficult and time-consuming process that encompasses multiple stages.Drug molecules are initially evaluated for their specificities and activity, followed by an assessment of their pharmacokinetics and toxicities later on.Regardless, many medication candidates fail in the latter stages of research due to unfavorable safety and effectiveness difficulties.Significant advances in computational science have recently provided an alternate method for finding potential bioactive compounds that can target specific diseases of interest.These computational tools present a viable path for speeding up the drug discovery process while reducing the high failure rates associated with traditional drug development methods [12].This study aims to find the novel natural compound SK1 inhibitors to manage the cancers.

Methods:
Protein 3D structure preparation: SK1 was the protein of interest in the study, and its crystal structure (PDB ID: 4V24) complex with PF-543 was obtained from the protein databank [13].The protein was prepared for virtual screening (VS) by removing the coordinates of heteroatoms and PF-543.

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A collection of 1068 natural compounds ranging in molecular weight from 350 to 450 was obtained from the 'coconut database', a highly regarded and systematically annotated natural compounds database.A filtering process was implemented to ensure the selection of compounds that adhere to Lipinski's rule of five.The compounds were downloaded in '.sdf' format and were prepared after energy minimization with the mmff94 force field.The prepared natural compounds were then saved in the 'pdbqt' format, allowing for VS in subsequent steps.

Docking protocol validation:
To validate the molecular docking protocol, a redocking experiment with the bound ligand PF-543 in the active pocket of SK1 was conducted.Following this, the initial x-ray crystallographic conformation of PF-543 was aligned with the active pocket.Remarkably, a precise alignment was attained, demonstrating a perfect match between the x-ray crystallographic conformation of PF-543 and the optimal docked pose.

Virtual screening:
VS are crucial steps in the drug design process, which is used in the study to assess the compounds' affinity for the target SK1 protein.
The active site of SK1, in which the inhibitor PF-543 was cocrystallized, was utilized as positive control for the VS of the prepared coconut database library.The binding affinity calculation was performed using PyRx AutoDock VINA, which is an important factor in determining the strength of binding between the compounds and the target protein [14-16].After that, a comprehensive interaction analysis as well as visual inspections of 2D and 3D interactions was carried out in order to determine the most stable complex based on lower values of binding affinity.

Results and Discussion:
In our pursuit of discovering natural inhibitors of SK1, a structurebased VS approach was employed.This involved screening a meticulously prepared database of 1068 natural compounds, with PF-543 serving as our positive control.It is important to note that PF-543 is widely recognized as a highly effective inhibitor of SK1.Using PF-543 as a benchmark, the goal was to find promising natural compounds that exhibit similar or better inhibitory properties against SK1.
To initiate, the docking protocol was validated in order to ensure its accuracy and reliability.Figure 2 demonstrates the excellent alignment between the redocked complex and the originally downloaded complex from the PDB, confirming the robustness of our molecular docking protocol.Following validation, the prepared compound library was subjected to docking-based VS against SK1.
Natural products are a useful source of bioactive chemicals with medicinal potential due to their high chemical variety.Significant efforts have been made in recent decades to find and isolate novel natural products from a range of sources, including microbes, plants, and other species.These natural chemicals have been intensively studied for their anticancer qualities and underlying mechanisms of action, which has resulted in the invention of various anti-cancer medications [19].Surprisingly, it is projected that between 1981 to 2019 and nearly 25% of newly authorized anticancer medications were derived from natural ingredients [20,21].

Conclusion:
This study shows the molecular interactions between natural compounds and the SK1 protein, with a focus on CNP0296172, CNP0368143, CNP0380570, and CNP0290815, which demonstrated a strong binding affinity for SK1.Furthermore, these compounds were found to meet the essential drug like criteria, making them suitable candidates for additional experimental validation as potential anticancer agents.
, sphingolipids have been linked to the development of a variety of disorders [5], and they have been shown to affect cancer etiology, progression, angiogenesis, proliferation, therapy resistance, and cell death [6].Sphingosine kinases (SKs), specifically SK1 and SK2, catalyze the conversion of sphingosine to S1P and play an important role in controlling the balance of ceramides, sphingosine, and S1P [7].Among these, SK1 has received special attention in cancer research and is considered a bona fide oncogene [8].A considerable body of evidence suggests that SK1 is present in tumor tissues, with overexpression being observed in the majority of investigations [9-11].

Figure 1 :
Figure 1: Expression of SK1 in different cancers.Expression analyses performed by Genevisible showing the SK1 expression in human cancer (A) and mouse cancer (B)

Table 2 :
Molecular descriptor of selected four compounds

Table 1 :
Best 10 screened hits and their binding affinity.