Large-scale validation of skin prion seeding activity as a biomarker for diagnosis of prion diseases

Authors: ZHANG, WEIGUANLIU - Case Western Reserve University (CWRU); ORRU, CHRISTINA - Rocky Mountain Laboratories National Institute Of Allergy And Infectious Diseases (NIAID); FOUTZ, AARON - Case Western Reserve University (CWRU); DING, MINGXUAN - Case Western Reserve University (CWRU); YUAN, JUE - Case Western Reserve University (CWRU); SHAH, SYED ZAHID ALI - Case Western Reserve University (CWRU); ZHANG, JING - Case Western Reserve University (CWRU); KOTOBELLI, KEISI - Case Western Reserve University (CWRU); GERASIMENKO, MARIA - Case Western Reserve University (CWRU); GILLILAND, TRICIA - Case Western Reserve University (CWRU); CHEN, WEI - Case Western Reserve University (CWRU); TANG, MICHELLE - Case Western Reserve University (CWRU); COHEN, MARK - Case Western Reserve University (CWRU); SAFAR, JIRI - Case Western Reserve University (CWRU); XU, BIN - North Carolina Central University; HONG, DAO-JUN - Nanchang University; CIU, LI - Jilin University; HUGHSON, ANDREW - Rocky Mountain Laboratories National Institute Of Allergy And Infectious Diseases (NIAID); SCHONBERGER, LAWRENCE - Centers For Disease Control And Prevention (CDC) - United States; TATSUOKA, CURTIS - University Of Pittsburgh School Of Medicine; CHEN, SHU - Case Western Reserve University (CWRU); Greenlee, Justin; WANG, ZERUI - Case Western Reserve University (CWRU); APPLEBY, BRIAN - Case Western Reserve University (CWRU); CAUGHEY, BYRON - Rocky Mountain Laboratories National Institute Of Allergy And Infectious Diseases (NIAID); ZOU, WEN-QUAN - Case Western Reserve University (CWRU)

Submitted to: Acta Neuropathologica
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/22/2023
Publication Date: 1/17/2024
Citation: Zhang, W., Orru, C.D., Foutz, A., Ding, M., Yuan, J., Shah, S., Zhang, J., Kotobelli, K., Gerasimenko, M., Gilliland, T., Chen, W., Tang, M., Cohen, M., Safar, J., Xu, B., Hong, D., Ciu, L., Hughson, A.G., Schonberger, L.B., Tatsuoka, C., Chen, S.G., Greenlee, J.J., Wang, Z., Appleby, B.S., Caughey, B., Zou, W. 2024. Large-scale validation of skin prion seeding activity as a biomarker for diagnosis of prion diseases. Acta Neuropathologica. https://doi.org/10.1007/s00401-023-02661-2.
DOI: https://doi.org/10.1007/s00401-023-02661-2

Interpretive Summary: Prion diseases are invariably fatal neurologic diseases for which there is no known prevention or cure and early diagnosis is exceedingly difficult. Traditionally, diagnosis can only be confirmed using postmortem samples. This manuscript describes the use of in vitro assays to detect the presence of prions in skin samples from humans with various prion diseases. This work suggests that skin may be a useful tissue for early detection of prion disease. This detection method could have a major impact on the prognosis for prion diseases because it allows for earlier detection, which allows earlier attempts at intervention to delay the onset of neurologic signs.

Technical Abstract: Definitive diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD) relies on the examination of brain tissues for the pathological misfolded prion protein (PrPSc). Our previous study revealed that the PrPSc-seeding activity (PrPSc-SA), a peculiar feature of misfolded PrPSc, can be detected in skin tissues of sCJD with high sensitivity and specificity by an ultrasensitive PrPSc seed-amplification assay (PrPSc-SAA) known as the real-time quaking-induced conversion (RT-QuIC). This study validates PrPSc-SA as a useful biomarker for the diagnosis of sCJD, by utilizing a large number of skin samples collected from cadavers with neuropathologically and biochemically confirmed different subtypes of sCJD. A total of 877 skin samples, were retrospectively and prospectively collected at autopsy, from three body areas of 340 individual cases with varying, neuropathologically confirmed subtypes of sCJD and non-sCJD controls were examined. The RT-QuIC assay was used to detect skin PrPSc-SA blindly in two independent laboratories using recombinant hamster PrP90-231 or bank vole PrP23-230 as a substrate. The results of skin PrPSc-SA were analyzed with demographic information, clinical manifestations, cerebrospinal fluid (CSF) PrPSc-SA, and other lab tests including CSF 14-3-3 and tau in patients with and without sCJD. The end-products of skin PrPSc-SAA were compared among different sCJD subtypes. A total of 340 cases are included in this study composed of both retrospective (n=122) and prospective cohorts (n=218). The former is examined blindly in one lab and validated in a separate lab, while the latter cohort was blindly tested in only one lab. Both blind and un-blind PrPSc-SAAs of the retrospective cases exhibit sensitivity of 84.5% and 88.4%, respectively, with both exhibiting 100% specificity. The blind PrPSc-SAA of the prospective cases shows sensitivity and specificity of 83.2%-89.5% and 97.3%-98.7% based on two different common diagnostic criteria, respectively. Sensitivity and specificity of CSF PrPSc-SAA are 87% and 90.9% in 85 out of 122 retrospective cases with available CSF samples, while the sensitivity and specificity of CSF PrPSc-SAA are 90.8% and 100% in 127 available CSF out of 218 prospective cases. The sensitivity of skin PrPSc-SAA is subtype- and dermatome-dependent with the highest in sCJDVV1-2, followed by VV2, MV1-2, MV1, MV1, MV2, MM1, MM1-2, MM2, and VV1, and the highest in the skin area next to the ear, followed by low back and apex, respectively. Although no significant difference in brain PrPSc-SA was found between the cases with false negative and the case with the true positive skin PrPSc-SA, the disease duration is significantly longer in false negative than in true positive cases [12.04 ± 13.27 (months, SD) vs 6.49 ± 6.41, p = 4.75E-41 < 0.0001]. Finally, PK-resistant PrP C-terminal fragments are always detectable in the RT-QuIC end-products with positive ThT reaction regardless of true and false positive PrPSc-SA, confirming that the positive ThT reaction is associated with the generation of PK-resistant PrP. Our study suggests that skin PrPSc-SA can serve as a novel biomarker for the detection of prion diseases, and its sensitivity and specificity is determined by the PrPSc types, PRNP polymorphisms, dermatome sampled, disease duration, and generation of PK-resistant PrP.