DISCUSSION AND CONCLUSIONS
Our investigation centered on modulating RyR activity, specifically
through a novel class of triazole molecules known as MP compounds that
act as FKBP12 ligands . The primary focus of our study was on MP-010, a
RyR stabilizer with favorable pharmacokinetics and CNS permeability. Our
results in SOD1G93A mice, a widely accepted ALS model,
demonstrated significant benefits following MP-010 administration.
Notably, increased survival, protection against MN loss, muscle
denervation, and enhanced neuromuscular function were observed,
highlighting the potential of MP-010 as a therapeutic intervention for
ALS. The robustness of our findings is emphasized by their replication
in two distinct colonies of SOD1G93A mice with a mixed
hybrid B6SJL genetic background, across two independent laboratories.
This replication is particularly noteworthy considering the potential
genetic drift that hybrid ALS SOD1 colonies may undergo among different
research labs, leading to variability in disease onset or symptom
severity . This potential confounding factor makes the replication of
our findings in two independent laboratories even more significant.
Remarkably, despite differing ages of motor symptom onset between the
two colonies (14 weeks in one laboratory and 17 weeks in the other),
MP-010 demonstrated beneficial effects in both settings at the dose of
61 mg/kg. The compound exhibited remarkable tolerability and appeared
devoid of apparent toxicity even at high doses. Doubling the dose did
not induce a change in body weight, a recognized clinical symptom of
pharmacological toxicity. Nevertheless, the compound’s therapeutic
efficacy was compromised at this higher dose. This unexpected loss of
efficacy could be attributed to off-target effects. These unintended
interactions with non-target proteins could be interfering with the
compound’s ability to reach its intended target and exert its
therapeutic effects. Further investigations are warranted to identify
and characterize specific off-target mechanisms, enabling the
development of strategies to mitigate them and optimize the compound’s
efficacy.
As previously introduced, cytosolic Ca2+ overload
assumes particular importance in the vulnerability of spinal MNs
subjected to ALS toxicity , and deficient Ca2+handling from ER and mitochondrial stores has emerged as a key player in
the pathogenesis of ALS. This sets the stage for a positive feedback
loop that amplifies pathological Ca2+ overload,
intensifying the neurodegenerative processes associated with the disease
. Mutations in genes such as SIGMA1R and VAPB , identified
in ALS patients, underscore the significance of ER and mitochondrial
proteins in Ca2+ homeostasis . In the context of the
present study, the rationale for targeting RyR stemmed from its central
role in Ca2+-induced Ca2+ release
from the ER, aiming to prevent excessive cytosolic
Ca2+ accumulation, especially in the most vulnerable
MNs associated with ALS. The outcomes of the current study,
demonstrating the therapeutic benefits of a drug promoting the
interaction between RyR and its endogenous ligand FKBP12 in the
SOD1G93A mouse model of ALS, are in accordance with
earlier research that has emphasized reductions in FKBP12 in the context
of ALS . Also in the SOD1G93A mouse, various
alternative strategies to reduce the Ca2+ burden on
MNs have been successfully implemented. These strategies include the
application of the AMPA receptor antagonist talampanel and SIGMA1R
agonists , both exerting their effects through the modulation of
extracellular Ca2+ influx to cytosol.
The dysregulation of RyR signaling emerges as a fundamental contributor
to the pathogenesis of various neurodegenerative disorders, notably
Alzheimer’s disease (AD). Specifically, RyRs have been demonstrated to
interact with amyloid-beta (Aβ) peptide, a protein known to play a
pivotal role in AD development. The binding of Aβ to RyRs can modulate
their function, resulting in heightened Ca2+ release
from the ER, subsequently leading to excitotoxicity, neuronal death, and
cognitive deficits . Although our investigation cannot definitively
ascertain the existence of Ca2+ leakage through RyR in
ALS MNs or whether this pathological phenomenon is replicated in the
SOD1G93A mouse model, the administration of the
compound MP-010 emphasizes the potential therapeutic significance of
targeting FKBP12 to safeguard the RyR-dependent proper release of
Ca2+ and alleviate the downstream effects of disrupted
Ca2+ homeostasis. This approach offers a targeted intervention for ALS.
However, while it is important to note that an aberrant increase in
RyR-mediated Ca2+ release can contribute to disrupting
neuronal homeostasis, triggering excitotoxicity, conversely, diminished
RyR function may compromise the crucial Ca2+ signaling
required for proper neuronal operation. This impairment might lead to a
breakdown in synaptic transmission and plasticity, potentially
contributing to the manifestation of motor deficits in individuals with
ALS. Consequently, inhibiting RyR under this premise may not be a
plausible strategy for treating neurons susceptible to ALS. Indeed,
prior research utilizing dantrolene, an RyR1 inhibitor clinically used
to treat malignant hyperthermia, failed to demonstrate significant
improvements when administered to the SOD1G93A mouse
model , confirming the inadequacy of RyR inhibition as a therapeutic
avenue. In contrast, compounds exhibiting RyR-stabilizing activity,
previously recognized under the nomenclature ”rycals” , adopt a distinct
approach by modulating RyR channel opening. These compounds achieve this
modulation by facilitating the interaction between the topoisomerase
FKBP12 and RyR, particularly in pathological scenarios characterized by
persistent Ca2+ leakage . Thus, the strategic
targeting of RyR modulation through rycal compounds offers a more
nuanced and potentially effective therapeutic strategy compared to
direct inhibition.
Comparative analysis with MP-001, another compound characterized by
limited CNS availability, provides additional insights. The lack of
MP-001 effect underscore the significance of targeting
Ca2+ homeostasis at the CNS level to achieve
therapeutic efficacy. Thus, it dismisses the possibility that the
therapeutic effects could be mediated by the modulation of
Ca2+ dynamics in cells of the innate immune system. It
is crucial to consider this aspect, especially given that immune cells
heavily rely on Ca2+-dependent processes and tightly
regulated Ca2+ homeostasis , and that the innate
immune response appears to play an active role in the pathophysiology of
ALS and the SOD1G93A mouse model .
While our study presents promising results, several avenues for future
research emerge. Elucidating the molecular mechanisms underlying the
observed effects of MP-010, particularly its impact on
Ca2+ dynamics and neuronal survival, is critical for a
comprehensive understanding of ALS pathology. Additionally,
considerations of toxicity, dosage optimization, and potential side
effects are paramount before translating these findings into clinical
trials. In conclusion, our study contributes to the evolving landscape
of ALS research by highlighting the therapeutic potential of targeting
intracellular Ca2+ dynamics through the modulation of
RyR activity. MP-010, with its favorable preclinical outcomes, offers a
promising avenue for further exploration and potential clinical
development.