Potential drug candidate for AP-4 HSPs identified

AP-4 blood biomarker discovered also

Dr Ebrahimi-Fakhari

A potential drug candidate for AP-4 HSPs has been identified and a blood biomarker of disease severity discovered. Two multinational teams, both led by Darius Ebrahimi-Fakhari of Boston Children’s Hospital and Harvard Medical School, have simultaneously made these important discoveries and significant advances towards establishing treatments for AP-4 HSPs.

Perhaps just as importantly, the approach taken here may well provide a framework for investigating other forms of HSP and other rare and more common disorders of protein trafficking.

Potential drug candidate for AP-4 HSPs identified

Provides proof-of-concept for future studies

A leading drug candidate was identified that restored impaired cellular functions in both skin fibroblasts and iPSC neurons from 2 children (SPG47 and SPG50) with AP-4 deficiency. A unique high-throughput assay was developed, based on in-depth investigations of the potential mechanisms of disease, and used to screen almost 30,000 compounds.


Unbiased phenotypic screens in patient-relevant disease models offer the potential to detect novel therapeutic targets for rare diseases. In this study, we developed a high-throughput screening assay to identify molecules that correct aberrant protein trafficking in adaptor protein complex 4 (AP-4) deficiency, a rare but prototypical form of childhood-onset hereditary spastic paraplegia, characterized by mis-localization of the autophagy protein ATG9A.

Using high-content microscopy and an automated image analysis pipeline, we screened a diverse library of 28,864 small molecules and identified a lead compound, C-01, that restored ATG9A pathology in multiple disease models, including patient-derived fibroblasts and induced pluripotent stem cell-derived neurons. We used multiparametric orthogonal strategies and integrated transcriptomic and proteomic approaches to delineate putative molecular targets of C-01 and potential mechanisms of action.

Our results define molecular regulators of intracellular ATG9A trafficking and characterize a lead compound for the treatment of AP-4 deficiency, providing important proof-of-concept data for future Investigational New Drug (IND)-enabling studies.

SOURCE:  Res Sq. 2023 Jun 12;rs.3.rs-3036166.

doi: 10.21203/rs.3.rs-3036166/v1. Preprint PMID: 37398196

High-Content Small Molecule Screen Identifies a Novel Compound That Restores AP-4-Dependent Protein Trafficking in Neuronal Models of AP-4-Associated Hereditary Spastic Paraplegia

Afshin SaffariBarbara BrechmannCedric BoegerWardiya Afshar SaberHellen JumoDosh WhyeDelaney WoodLara WahlsterJulian AlecuMarvin ZieglerMarlene ScheffoldKellen WindenJed HubbsElizabeth ButtermoreLee BarrettGeorg BornerAlexandra DaviesMustafa SahinDarius Ebrahimi-Fakhari

Blood biomarker of neurodegeneration found for the AP-4 HSPs

Higher levels relate to disease severity

Levels of neurofilament light chain (NfL) in blood plasma samples allow differentiation of AP-4 HSP samples from healthy controls. They further identify more severely affected people within the AP-4 group who experience severe generalized-onset seizures and developmental stagnation

Background: Adaptor protein complex 4-associated hereditary spastic paraplegia (AP-4-HSP) is caused by pathogenic biallelic variants in AP4B1, AP4M1, AP4E1, and AP4S1.

Objective: The aim was to explore blood markers of neuroaxonal damage in AP-4-HSP.

Methods: Plasma neurofilament light chain (pNfL) and glial fibrillary acidic protein (GFAP) levels were measured in samples from patients and age- and sex-matched controls (NfL: n = 46 vs. n = 46; GFAP: n = 14 vs. n = 21) using single-molecule array assays. Patients’ phenotypes were systematically assessed using the AP-4-HSP natural history study questionnaires, the Spastic Paraplegia Rating Scale, and the SPATAX disability score.

Results: pNfL levels increased in AP-4-HSP patients, allowing differentiation from controls (Mann-Whitney U test: P = 3.0e-10; area under the curve = 0.87 with a 95% confidence interval of 0.80-0.94). Phenotypic cluster analyses revealed a subgroup of individuals with severe generalized-onset seizures and developmental stagnation, who showed differentially higher pNfL levels (Mann-Whitney U test between two identified clusters: P = 2.5e-6). Plasma GFAP levels were unchanged in patients with AP-4-HSP.

SOURCE:  Mov Disord. 2023 Jul 22.

doi: 10.1002/mds.29524. Online ahead of print.  PMID: 37482941 © 2023 International Parkinson and Movement Disorder Society.

Plasma Neurofilament Light Chain Is Elevated in Adaptor Protein Complex 4-Related Hereditary Spastic Paraplegia

Julian E Alecu  1   2 Afshin Saffari  1 Marvin Ziegler  1 Catherine Jordan  1 Amy Tam  1 Soyoung Kim  3 Edward Leung  4 Krzysztof Szczaluba  5 Hanna Mierzewska  6 Staci D King  7 Filippo M Santorelli  8 Grace Yoon  9 Bianca Trombetta  10 Pia Kivisäkk  10 Bo Zhang  1   11 Mustafa Sahin  1   12   13 Darius Ebrahimi-Fakhari  1   12   13   14

1. Department of Neurology and F.M. Kirby Neurobiology Center, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts, USA.

2. Friedrich-Alexander-University Erlangen-Nuremberg, Erlangen, Germany.

3. Sozialpaediatrisches Zentrum Frankfurt Mitte, Frankfurt am Main, Germany.

4. Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, Manitoba, Canada.

5. Department of Medical Genetics, Medical University of Warsaw, Warsaw, Poland.

6. Department of Neurology, Institute of Mother and Child, Warsaw, Poland.

7. Department of Neurology, Texas Children’s Hospital, Houston, Texas, USA.

8. Molecular Medicine, Neurogenetics, IRCCS Fondazione Stella Maris, Pisa, Italy.

9. Divisions of Clinical and Metabolic Genetics and Neurology, Department of Pediatrics, The Hospital for Sick Children, University of Toronto, Toronto, Ontario, Canada.

10. Alzheimer’s Clinical and Translational Research Unit, Department of Neurology, Massachusetts General Hospital, Boston, Massachusetts, USA.

11. ICCTR Biostatistics and Research Design Center, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts, USA.

12. Rosamund Stone Zander Translational Neuroscience Center, Boston Children’s Hospital, Boston, Massachusetts, USA.

13. Intellectual and Developmental Disabilities Research Center, Boston Children’s Hospital, Boston, Massachusetts, USA.

14. Movement Disorders Program, Department of Neurology, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts, USA.


    1. Editor’s note: Pre-clinical investigations of Noscapine concluded in September 2020 with computer modelling and mouse studies aimed at determining dose range for clinical trials. With the subsequent lack of success with grant funding applications for clinical trials, the program is on hold. The next logical step would be clinical trials in humans however the main stumbling block remains the lack of valid and accepted markers and measures of disease outcomes that are vital for getting definitive answers in clinical trials – and being successful in attracting the funds to do so.

      Following similar pathways, other candidate drugs for the HSPs have been identified, however they also face the same stumbling block. To the best of our knowledge, none of the other candidate drugs have been identified (they are published with code names) or where they have been identified, they are not compounds that have been approved for therapeutic use.

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