A Computational and Conceptual Framework for Rational, Multi-Target Design of Sterol 14α-Demethylase-Directed Therapeutics Against Naegleria fowleri: In Silico Modeling, Polypharmacology, and a Falsifiable Roadmap Toward Experimental Validation

Primary amebic meningoencephalitis (PAM), caused by the thermophilic free-living amoeba Naegleria fowleri, is a fulminant neurotropic infection with a case fatality rate exceeding 97% and fewer than twenty documented survivors worldwide. No therapeutic regimen tested to date has demonstrated reliable efficacy. This manuscript presents a conceptual, hypothesis-generating computational framework for the rational design of a hypothesized small-molecule inhibitor of N. fowleri sterol 14α-demethylase (NfCYP51), a genetically and structurally validated drug target, combined with adjunctive engagement of the mevalonate pathway and glycolytic enolase. Unlike a conventional drug-discovery report, no synthesis, biochemical assay, or animal experiment has been performed; every quantitative claim in this manuscript is explicitly labeled as either (i) a literature-anchored fact, reproduced with its original citation and, where available, its original numerical value, or (ii) a hypothesized model input chosen for illustrative systems-pharmacology simulation and clearly flagged as such. No model of a non-existent compound can support a categorical efficacy claim; instead, this framework offers falsifiable, testable predictions about the parameter regime a real inhibitor would need to occupy to achieve rapid parasite clearance under a defined dosing schedule. A stochastic pharmacokinetic/pharmacodynamic model (Euler–Maruyama scheme, fixed seed, n=1000 realizations) and a from-scratch Saltelli–Jansen Sobol' sensitivity analysis (no external variance-decomposition library) are used to identify which model parameters would most need to be experimentally constrained before any translational claim could be evaluated. All code is included in Appendix B and is fully reproducible. A dedicated risk-assessment section and an explicit falsifiability/roadmap section are provided, consistent with Popperian criteria for scientific testability.