{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,3]],"date-time":"2025-11-03T15:23:17Z","timestamp":1762183397215,"version":"build-2065373602"},"reference-count":26,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2025,11,3]],"date-time":"2025-11-03T00:00:00Z","timestamp":1762128000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["NeuroSci"],"abstract":"<jats:p>Microcephaly with early-onset, intractable seizures, and developmental delay (MCSZ) is a rare inherited neurological disorder caused by biallelic loss-of-function variants in the polynucleotide kinase\/phosphatase (PNKP) gene, which encodes an enzyme critical for DNA repair. Here, we describe the clinical history of two novel patients presenting with microcephaly, epilepsy, growth deficiency, language impairment, and severe intellectual disability. Brain MRI in both cases revealed complex cerebral malformations, including lissencephaly, ventriculomegaly, dysmorphic hippocampi, and cerebellar atrophy. Next-generation sequencing (NGS) analyses identified compound heterozygous PNKP variants in both patients. In case #1, we detected the missense variant p.Gln50Glu (c.148C&gt;G) in exon 2 (rs756746191) and a novel nonsense variant, p.Gln248Ter (c.742C&gt;T), leading to a premature stop codon in exon 7. In case #2, we identified the frameshift variant p.Thr424GlyfsTer49, caused by a 17-nucleotide duplication (c.1253_1269dupGGGTCGCCATCGACAAC) in exon 14 (rs587784365), along with a 15-nucleotide deletion (c.1386+49_1387-33delCCTCCTCCCCTGACCCC) in intron 15 (rs752902474). Over long-term follow-up (20 and 36 years for case #1 and case #2, respectively), seizures persisted in the first patient, while full control was achieved in the second case with combined therapy of valproate and clobazam. Along with a review of the literature, these two novel cases confirm the broad phenotypic spectrum of PNKP-associated disorders and underscore the importance of including PNKP in the genetic screening of patients presenting with developmental and epileptic encephalopathy (DEE) and microcephaly.<\/jats:p>","DOI":"10.3390\/neurosci6040110","type":"journal-article","created":{"date-parts":[[2025,11,3]],"date-time":"2025-11-03T13:15:14Z","timestamp":1762175714000},"page":"110","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Compound Heterozygous PNKP Variants Causing Developmental and Epileptic Encephalopathy with Severe Microcephaly: Natural History of Two New Cases and Literature Review"],"prefix":"10.3390","volume":"6","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1399-8271","authenticated-orcid":false,"given":"Francesca","family":"Ragona","sequence":"first","affiliation":[{"name":"Department of Pediatric Neuroscience, European Reference Network EPIcare, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5889-2342","authenticated-orcid":false,"given":"Giuliana","family":"Messina","sequence":"additional","affiliation":[{"name":"Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9610-7544","authenticated-orcid":false,"given":"Stefania","family":"Magri","sequence":"additional","affiliation":[{"name":"Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2272-7855","authenticated-orcid":false,"given":"Fabio Martino","family":"Doniselli","sequence":"additional","affiliation":[{"name":"Department of Neuroradiology, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2293-8687","authenticated-orcid":false,"given":"Elena","family":"Freri","sequence":"additional","affiliation":[{"name":"Department of Pediatric Neuroscience, European Reference Network EPIcare, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5385-761X","authenticated-orcid":false,"given":"Laura","family":"Canafoglia","sequence":"additional","affiliation":[{"name":"Department of Epileptology, European Reference Network EPIcare, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6657-3328","authenticated-orcid":false,"given":"Roberta","family":"Solazzi","sequence":"additional","affiliation":[{"name":"Department of Pediatric Neuroscience, European Reference Network EPIcare, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Cinzia","family":"Gellera","sequence":"additional","affiliation":[{"name":"Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tiziana","family":"Granata","sequence":"additional","affiliation":[{"name":"Department of Pediatric Neuroscience, European Reference Network EPIcare, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4102-1188","authenticated-orcid":false,"given":"Jacopo C.","family":"DiFrancesco","sequence":"additional","affiliation":[{"name":"Department of Neurology, Fondazione IRCCS S. Gerardo dei Tintori, 20900 Monza, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9098-3041","authenticated-orcid":false,"given":"Barbara","family":"Castellotti","sequence":"additional","affiliation":[{"name":"Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,11,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"24176","DOI":"10.1074\/jbc.274.34.24176","article-title":"Molecular Cloning of the Human Gene, PNKP, Encoding a Polynucleotide Kinase 3\u2019-Phosphatase and Evidence for Its Role in Repair of DNA Strand Breaks Caused by Oxidative Damage","volume":"274","author":"Jilani","year":"1999","journal-title":"J. Biol. 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