Tropane-Based Ibogaine Analog Rescues Folding-Deficient Serotonin and Dopamine Transporters

Abstract

Missense mutations that give rise to protein misfolding are rare, but collectively, defective protein folding diseases are consequential. Folding deficiencies are amenable to pharmacological correction (pharmacochaperoning), but the underlying mechanisms remain enigmatic. Ibogaine and its active metabolite noribogaine correct folding defects in the dopamine transporter (DAT), but they rescue only a very limited number of folding-deficient DAT mutant proteins, which give rise to infantile Parkinsonism and dystonia. Herein, a series of analogs was generated by reconfiguring the complex ibogaine ring system and exploring the structural requirements for binding to wild-type transporters, as well as for rescuing two equivalent synthetic folding-deficient mutants, SERT-PG 601,602 AA and DAT-PG 584,585 AA. The most active tropane-based analog (9b) was also an effective pharmacochaperone in vivo in Drosophila harboring the DAT-PG 584,585 AA mutation and rescued 6 out of 13 disease-associated human DAT mutant proteins in vitro. Hence, a novel lead pharmacochaperone has been identified that demonstrates medication development potential for patients harboring DAT mutations.

Functional and Biochemical Consequences of Disease Variants in Neurotransmitter Transporters: A Special Emphasis on Folding and Trafficking Deficits

“…Upon treatment with noribogaine and/or HSP70 inhibitor pifithrin-μ, the mutant transporter reached the axonal compartments and normal sleep time was restored in these flies ( Kasture et al, 2016 ). More importantly, this effect was replicated in flies expressing 2 human DAT variants (V158F and G327R), which give rise to infantile parkinsonism ( Asjad, Kasture, et al, 2017 ) and in the synthetic folding variant DAT-PG 584,585 AA ( Bhat et al, 2020 ). To the best of our knowledge, pharmacochaperoning has not yet been tested in other animal models.…”

“…Since the complex ring system of ibogaine is amenable to structural variations and chemical modifications, a consequent fluorinated analog proved valuable in rescuing the equivalent PG 584,585 AA mutant in DAT, with a higher efficacy than the parent compound ibogaine. This analog also corrected folding defects in 6 DAT mutants associated with DTDS ( Bhat et al, 2020 ).…”

“…Most of these small molecules are thought to facilitate the stabilization of the native state, thus enhancing protein expression ( Marinko et al, 2019 ). Nonetheless, this mechanism does not account for the increased surface expression of monoamine transporters by ibogaine and its congeners ( Bhat et al, 2020 ), which appears to occur via stabilization of a folding trajectory intermediate. In addition, the approved drug tafamidis is a pharmacochaperone, which restores the folded state of transthyretin (mutations in transthyretin cause a form of amyloidosis), by preventing protein aggregation and dissolution of aggregates ( Coelho et al, 2016 ).…”

Thermal Unfolding of the Human Serotonin Transporter: Differential Effect by Stabilizing and Destabilizing Mutations and Cholesterol on Thermodynamic and Kinetic Stability

“…The folding trajectory of SLC transporters is of interest, because folding-deficient mutants are amenable to rescue by pharmacochaperoning (El-Kasaby et al, 2010;Koban et al, 2015;Kasture et al, 2016;Beerepoot et al, 2016;Asjad et al, 2017;Bhat et al, 2017;Bhat et al, 2021c) or by interfering with the endogenous proteinaceous chaperones (Fujiwara et al, 2013;El-Kasaby et al, 2014& 2020. Conceptually, the action of pharmacochaperones can be accounted for by four mechanisms of action (Marinko et al, 2019): binding to and stabilization of the (i) native (=folded) state or (ii) folding intermediate(s); (iii) suppression of aggregate formation; (iii) dissolution of aggregates.…”

“…thermodynamic stabilization of the native state (Marinko et al, 2019). Circumstantial evidence, however, suggests that pharmacochaperones may rescue folding-deficient variants of SERT and of the dopamine transporter (DAT, SLC6A3) by an action on (a) folding intermediate(s) (Bhat et al, 2021c). Progress is hampered by the elusive nature of the folding intermediates.…”