These symptoms include extreme anxiety which tends to be lifelong, ADHD symptoms which often peak during childhood with the potential for inattention to persist through adulthood, aggression and agitation which tends to peak during adolescence, and sensory hypersensitivity among other concerns (2-5) . Individuals with FXS are also at significantly greater risk than the general population to develop a seizure disorder with the risks generally peaking during childhood and lessening over the course of adulthood. Many individuals with FXS require lifelong around the clock supported care contributing to an overall estimated 4.1- billion-dollar annual direct family healthcare cost associated with FXS in the United States.

Unfortunately, there are no FDA approved treatments for FXS.
Given this, management of behavioral and related concerns in FXS is limited to off label medication strategies that provide little benefit while also presenting risks for adverse effects such as weight gain, abnormal movements, sedation and other drug side effects (4, 6) . All children with FXS are recommended to receive early intervention therapy services, special education services, and when available FXS-informed behavioral therapy support. Unfortunately, in adulthood these various services
often have limited availability for persons with FXS.

Track record of treatment development in Fragile X syndrome
Over twenty years of translational medicine efforts in FXS has resulted in a number of failed late-phase human trials – trials often marked by significant placebo response and limited, if any, study inclusion criteria other than general PCR and Southern Blot defined diagnosis of FXS in participants (6, 8, 9) . We and others have worked tirelessly to develop more rigorous and quantitative measures of potential drug effects in FXS to address the many historic clinical trial challenges in our field. In doing this, we focus on drug target engagement, or what a drug is doing to brain activity and human function with treatment. We do this from the earliest stages of trial development, often pursuing this approach when a new drug mechanism is first trialed in FXS. This approach utilizes a number of modalities including high density EEG that capture neurophysiologic brain activity at rest and when responding to stimuli such as sound and patterns of sounds. In FXS there are well established and highly reproducible parameters for aberrant neurophysiologic brain activity measurement with EEG (10-20) . These parameters often are linked strongly to clinical severity of illness. Throughout this work we have additionally gained appreciation for molecular genetic variation in FXS establishing sex differences in EEG signatures and noting that even within men with FXS, there exists variation in expression of fragile X messenger ribonucleoprotein (FMRP) (21) . This appreciation has led us to approach early phase trials in FXS focusing on who specifically with FXS may be showing positive brain-based target engagement with treatment. This approach has refined our thinking about trials to now focus enrollment specifically on well-defined age, sex, and (at times) molecular FMRP expression-defined groups of people with FXS instead of defaulting to prior approaches of “all comers” enrollment.

Spinogenix Approach to Fragile X syndrome: SPG601
In recent years, dysregulated large conductance, calcium activated (BK) channel activity has been implicated in the pathophysiology of FXS including experiments in mice where rescue of deficient BK channel activity has been associated with normalization of brain and behavioral function (22-32). It has been noted in FXS models that BK channel expression is reduced and that FMRP normally interacts with BK channels to promote their activation. Reduced BK channel activity in FXS is felt to be involved in the anxiety, cognitive, and behavioral deficits associated with the disorder. Spinogenix is developing the BK channel activator SPG601 for use in FXS. SPG601 is a novel small molecule that specifically increases BK channel activity. The novel molecule is orally bioavailable and enters the brain. The safety of SPG601 has already been established in over 240 adult humans from Phase 1 and 2 trials in another disease indication. Additionally, SPG601 has had no evidence of the development of tolerance, which is important in FXS drug development where some hypothesized that the failure of mGluR5 antagonists may have been due to development of tolerance over time. In the FXS knockout (KO) mouse model of FXS, SPG601 normalized mouse aberrant behaviors including hyperactivity, anxiety, and aggression while also improving memory and cognition (31).

With this backdrop thoroughly justifying investigation of BK agonism as a novel drug mechanism in FXS, Cincinnati Children’s Hospital has partnered with Spinogenix to launch the first in human study of SPG601 in FXS. This study, informed by failures of the past and integrating state-of-the art neurophysiological outcome measures of target engagement, is focused specifically on whether SPG601 with a single dose is showing early signs of positively engaging brain function. With this in mind, our crossover double-blind, placebo controlled single-dose trial (NCT06413537) is focused on specific EEG outcomes well validated in FXS in adults including resting power band and connectivity analyses and auditory evoked potentials. We will additionally be evaluating potential treatment-related changes in established measures of eye gaze and pupillary response (33, 34), as well as testing whether performance improves on the NIH toolbox cognitive testing battery (35), a testing construct essentially validated in FXS and related disorder that may gain FDA approval as a potential primary outcome measure in pivotal trials. Our first trial in FXS is enrolling ten men aged 18-45 years with FXS who have a fully methylated CGG repeat expansion in their FMR1 gene. We hope to expand future trials into larger patient samples with a focus on other FXS subgroups such as females and potential in youth in the future. While this appears to be a small trial, we have experience with demonstrating drug target engagement using EEG and performance-based measurement in small samples. With this trial, Spinogenix will be able to gain rapid insights on the potential therapeutic impact of a novel molecule targeting a well-established molecular dysfunction in FXS. SPG601 has a well demonstrated safety profile and is an ideal candidate for the recently started single-dose target engagement study in men with FXS.

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