Abstract

{"title":"Abstract","authors":"","doi":"10.1111/ddg.15735_g","DOIUrl":null,"url":null,"abstract":"<p>Rupesh Paudel^1,2*, Lena F Sorger^1*, Anita Hufnagel¹, Mira Pasemann¹, Tamsanqa Hove^1,3, André Marquardt^1,4, Susanne Kneitz⁵, Andreas Schlosser⁶, Caroline Kisker³, Jochen Kuper^3#, Svenja Meierjohann^1,4#</p><p>¹Institute of Pathology, University of Würzburg, 97080 Würzburg, Germany</p><p>²Department of Dermatology, Venereology and Allergology, University Hospital Würzburg, 97080 Würzburg, Germany.</p><p>³Rudolf Virchow Center for Integrative and Translational Bioimaging, Institute for Structural Biology, University of Würzburg, 97080 Würzburg, Germany</p><p>⁴Comprehensive Cancer Center Mainfranken, University of Würzburg, 97080 Würzburg, Germany</p><p>⁵Department of Biochemistry and Cell Biology, University of Würzburg, 97074 Würzburg, Germany</p><p>⁶Rudolf Virchow Center for Integrative and Translational Bioimaging, Mass Spectrometry Division, University of Würzburg, 97080 Würzburg, Germany</p><p>^#corresponding authors:</p><p>J Kuper (<span>[email protected]</span>)</p><p>S Meierjohann (<span>[email protected]</span>)</p><p>*These authors contributed equally to this work</p><p><b>Abstract</b></p><p>Germline mutations in the DNA repair helicase XPD cause the diseases xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome. XP patients bear an increased risk of developing skin cancer including melanoma, even at young age. This is not observed for TTD patients despite DNA repair defects. To examine whether TTD cells harbor features counteracting tumorigenesis, we developed a TTD melanoma cell model containing the XPD variant R722W. Intriguingly, TTD melanoma cells exhibited reduced pro-tumorigenic features and an increased signature of the melanocyte lineage differentiation factor MITF, which went along with a strong basal upregulation of REDD2, an inhibitor of the mTOR/S6K/4EBP1 dependent protein translation machinery. REDD2 levels were partially driven by MITF, increased under UV exposure, and contributed to the reduced melanoma proliferation. To investigate whether similar processes are affected in melanocytes - the progenitor cell type of melanoma - we developed a TTD melanocyte model. Again, the MITF gene signature was increased, this time without affecting REDD2 expression. However, protein translation analyses revealed reduced ribosomal protein synthesis particularly in R722W melanocytes after UV stress, indicating a compromised protein translation machinery. Impaired protein translation was also demonstrated for the TTD XPD variant A725P, but not for the XPD variant D234N that causes XP. Concludingly, although effectors of R722W partially differ between melanoma cells and melanocytes, they result in translation inhibition and therefore reduced fitness, particularly under UV exposure. This may limit the probability of UV-driven tumorigenesis and offers an explanation why TTD patients do not develop melanomas.</p><p>Gaia Veniali¹, Anita Lombardi¹, Massimo Teson², Tiziana Nardo¹, Elena Botta¹, Elena Dell'Ambra², Donata Orioli¹, Manuela Lanzafame¹</p><p>¹CNR-Istituto di Genetica Molecolare, Pavia, Italy</p><p>²Istituto Dermopatico dell'Immacolata (IDI), Rome, Italy</p><p>Nucleotide Excision Repair (NER) is the only DNA repair system in humans dedicated to removing bulky DNA adducts induced by ultraviolet (UV) radiations. NER defects are responsible for the inherited disorder xeroderma pigmentosum (XP), which is characterized by high predisposition of developing melanoma and non-melanoma skin cancers due to the accumulation of unrepaired UV-induced DNA lesions. Interestingly, mutations in the NER-related genes <i>XPB</i> or <i>XPD</i> can also cause trichothiodystrophy (TTD), another rare disorder characterized by NER defects. However, unlike XP, TTD patients do not develop cancer but are characterized by hair abnormalities, pre- or post-natal growth failure, neurodevelopmental and neurological dysfunctions, signs of premature aging. Investigations and comparative gene expression profile studies on skin cells from XP and TTD patients carrying mutations in the same gene, offer a promising tool for identifying molecular pathways that drive or counteract skin carcinogenesis.</p><p>Whole-transcriptome analysis has been performed on our collection of primary skin fibroblasts and keratinocytes from <i>XPD</i>-mutated patients. We found disease-specific expression signatures, with XP cells showing a tumor-like profile characterized by the upregulation of genes involved in inflammation and extracellular matrix remodeling, whereas TTD cells exhibit an anti-tumor signature marked by the upregulation of tumor suppressor genes and the downregulation of oncogenic pathways.</p><p>The relevance of the identified transcription deregulations on skin cancer pathophysiology is currently under investigation by silencing or overexpressing the genes of interests in control skin cells and taking advantage of three-dimensional (3D) culture models that mimic critical aspects of solid cancers, including mechanical regulation of growth, hypoxia, and invasiveness. Our preliminary data on melanoma spheroids indicate that specific gene expression deregulations in the tumor microenvironment influence melanoma invasiveness.</p><p>Overall, our findings indicate that XP and TTD patient-derived primary skin cells, sharing mutations in the same gene but exhibiting opposite skin cancer susceptibilities, provide an exceptional framework for uncovering the molecular mechanisms underlying skin carcinogenesis and identifying novel targets for therapeutic intervention.</p><p>Ubiquitylation plays a crucial role in Nucleotide Excision Repair (NER). In the subpathway Global Genome NER (GG-NER), XPC detects lesions genome wide, with its activity enhanced after ubiquitylation by CRL4^DDB2. Conversely, Transcription-Coupled NER (TC-NER) is restricted to the transcribed strand of active genes, initiated by recognition of the lesion-stalled RNA Polymerase II (RNAPII) by CSB and subsequent recruitment of CSA, the substrate recognition subunit of CRL4^CSA. Ubiquitylation of RNAPII by CRL4^CSA at lysine 1268 (K1268) of its RPB1 subunit was previously shown to be important for TC-NER. Moreover, RPB1 K1268 ubiquitylation suppresses global gene expression by inducing RPB1 degradation, lowering RNAPII levels.</p><p>Intriguingly, however, cells defective in TC-NER still display robust RPB1 ubiquitylation after DNA damage but do not degrade RPB1. This led us to speculate that ubiquitylated RPB1 (ubi-RPB1) might have alternative, non-degradative roles. As ubi-RPB1 is only a small fraction of total RPB1 in the cell, we developed a strategy based on sequential enrichment steps followed by mass-spectrometry to study its interactome. For these experiments, we used an RPB1-K1268R mutant as a control to identify factors that specifically associate with ubiquitylated RPB1. This approach revealed a number of unexpected, high-confidence interactors, that led us to uncover a novel signaling pathway with major implications for Nucleotide Excision Repair.</p><p>Debora Ferri^a,*, Giulia Branca^a,*, Manuela Lanzafame^a, Erica Gandolfi^a, Valentina Riva^a, Giovanni Maga^a, Claudia Landi^c, Luca Bini^c, Lavinia Arseni^a, Fiorenzo A. Peverali^a, Emmanuel Compe^b and Donata Orioli^a</p><p>^aIstituto di Genetica Molecolare (IGM) L.L. Cavalli Sforza, CNR, 27100 Pavia, Italy;</p><p>^bInstitut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Illkirch Cedex 67404, Strasbourg, France;</p><p>^cDepartment of Life Sciences, University of Siena, 53100 Siena, Italy.</p><p>*Equal contribution</p><p><b>Abstract</b></p><p>The transcription factor IIH (TFIIH) is a ten-subunits complex organized in two sub-complexes, the ternary CDK activating kinase (CAK) and the hexameric core-TFIIH, bridged together by the XPD subunit. TFIIH is involved in different cellular mechanisms such as basal transcription, gene expression regulation and nucleotide excision repair (NER), the DNA repair pathway that specifically removes the bulky DNA adducts induced by ultraviolet (UV) light or other genotoxic agents. Mutations in the <i>ERCC2/XPD</i> gene are responsible for different clinical conditions, including the cancer-prone xeroderma pigmentosum (XP) and the photosensitive form of trichothiodystrophy (PS-TTD), a cancer-free neurodevelopmental disorder. At the cellular level, all <i>XPD</i> mutations result in NER defects and accumulation of unrepaired DNA photolesions. Furthermore, mutations specifically associated to PS-TTD clinical features also lead to reduced TFIIH cellular amount and transcription deregulations. Recently, our laboratory has demonstrated that PS-TTD more than XP primary dermal fibroblasts suffer of wide transcriptional impairments that, in some cases, result in altered protein content contributing to PS-TTD clinical features. To accomplish its function in DNA repair and transcription, TFIIH requires a direct interaction with the chromatin and relies on the presence of both the CAK and core sub-complexes. In the present study, we investigate the impact of <i>XPD</i> mutations on the association/dissociation of the two sub-complexes and their interaction with the chromatin.</p><p>Immunoprecipitation of the chromatin-bound CAK followed by mass spectrometry analysis identifies TFIIH as part of a large multiprotein assembly that assists RNA polymerase II during mRNA synthesis. PS-TTD causing mutations alter the relationships of TFIIH with other components of the protein assembly and give rise to transcriptional stress. Overall, this study sheds light on a still unknown function of TFIIH during transcription, whose impairment contributes to the wide transcriptional defects underlying PS-TTD pathological condition.</p><p>Alexandra Paolino¹, Ruth Keogh², Sinéad M. Langan², Alan R. Lehmann³, Isabel Garrood¹, Hiva Fassihi¹</p><p>¹National Xeroderma Pigmentosum Service, Department of Photodermatology, St John's Institute of Dermatology, Guy's and St Thomas’ Foundation Trust, London SE1 7EH, United Kingdom.</p><p>² Department of Non-communicable Disease Epidemiology, London School of Hygiene and Tropical Medicine, London, WC1E 7HT, United Kingdom.</p><p>³ Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton BN1 9RQ, United Kingdom.</p><p><b>Introduction</b>: Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder of DNA repair, characterized by extreme sensitivity to ultraviolet radiation. Patients are at risk of skin cancers, ocular surface disease, and approximately one-third of patients experience progressive neurodegeneration. Life expectancy varies significantly based on complementation group, disease severity, and access to preventive measures. Obtaining reliable survival estimates for this rare and genetically heterogeneous disease can be challenging, leading to variability in reported survival rates. A National Institutes of Health study of 106 patients (1971 to 2009) reported a median age at death of 29 years for XP patients with neurodegeneration and 37 years for those without (1). These statistics are frequently cited in academic literature and widely reported on publicly accessible websites.</p><p><b>Aim</b>: To assess the life expectancy of XP patients receiving multidisciplinary care at the UK National XP Service and assess patients and families understanding of their prognosis.</p><p><b>Methods</b>: We conducted a longitudinal study of 89 patients examined and treated annually in the UK National XP Service from 2015 to 2024. Mortality events were documented and categorised between those with and without neurodegeneration. To evaluate patient perceptions on life expectancy, 24 clinic attendees (16 neurologically unaffected patients and eight relatives) completed a short survey prior between February and July 2023.</p><p><b>Results</b>: Over a 9-year period, we recorded 15 deaths (17%), comprising 11 cases with neurodegeneration (groups A, B, D, F, and G) and 4 cases without neurodegeneration (groups A, C and V). Notably, there were no deaths attributable to skin cancer. Four deaths were linked to internal malignancies [median age: 52.5 years (IQR 41.8-58.75). The median survival age is 50 years (95% CI [34, Inf]) for those with neurological involvement, and 81 years (95% CI [81, Inf]) for those without. The latter is comparable to the general UK population (78.6 years for males, 82.6 years for females, 2020–2022) (2). The estimated probability of survival beyond age 40 is 0.68 (95% CI [0.47,0.98]) for individuals with neurological involvement and 0.90 (95% CI [0.73,1.00]) for those without.</p><p>Only 11 of 24 survey respondents (46%) believed that life expectancy in neurologically unaffected XP patients was normal. One respondent (4%) estimated life expectancy at 20-29 years, 8 (31%) at 30-39 years, 1 (4%) at 50-59 years, and 3 reported uncertainty.</p><p><b>Conclusion</b>: We present evidence of improved survival outcomes in patients with XP in the UK, both with and without neurological involvement, compared to previously reported literature. While life expectancy for individuals with XP can be limited, particularly without appropriate care, advances in medical management and strict adherence to preventive measures have improved outcomes. It is important that outdated and potentially harmful misinformation online is updated and to convey to through discussions with patients and families that regular medical follow-ups, diligent photoprotection, and timely intervention for skin cancers, can enhance both longevity and quality of life for those affected by XP.</p><p><b>REFERENCES</b>:</p><p>Bradford PT, Goldstein AM, Tamura D, Khan SG, Ueda T, Boyle J, Oh KS, Imoto K, Inui H, Moriwaki S, Emmert S, Pike KM, Raziuddin A, Plona TM, DiGiovanna JJ, Tucker MA, Kraemer KH. Cancer and neurologic degeneration in xeroderma pigmentosum: long term follow-up characterises the role of DNA repair. J Med Genet. 2011 Mar;48(3):168-76. doi: 10.1136/jmg.2010.083022. Epub 2010 Nov 19. PMID: 21097776; PMCID: PMC3235003.</p><p>Office for National Statistics (ONS), released 11 January 2024, ONS website, statistical bulletin, National life tables – life expectancy in the UK: 2020 to 2022</p><p>Alexandra Paolino¹, Sally Turner¹, Tanya Henshaw¹, Joanne Palfrey¹, Karla Balgos¹, Paola Giunti, Ana M. S. Morley¹, Shehla Mohammed¹, Adesoji Abiona¹, Alan R. Lehmann², Hiva Fassihi¹</p><p>¹National Xeroderma Pigmentosum Service, Department of Photodermatology, St John's Institute of Dermatology, Guy's and St Thomas’ Foundation Trust, London SE1 7EH, United Kingdom;</p><p>²Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton BN1 9RQ, United Kingdom.</p><p><b>Introduction</b>: There is marked heterogeneity in the presenting clinical features of Xeroderma Pigmentosum (XP), both between and within complementation groups. Effective management relies on early diagnosis, stringent photoprotection to mitigate skin cancer-related morbidity and mortality, and regular dermatological and ophthalmological examinations to detect and treat malignancies promptly. Despite these measures, delays in diagnosis remain a challenge, emphasizing the need for increased awareness among healthcare providers.</p><p><b>Aim</b>: To evaluate presenting clinical features of XP and time from first symptom onset to diagnosis.</p><p><b>Methods</b>: A retrospective review was conducted on 133 XP patients (67 females, 66 males; age range 0–87 years) receiving specialist multidisciplinary care at a national centre. Of these, 100 patients were included in the study, excluding 29 diagnosed through older siblings and 4 with incomplete histories. Data collected included complementation group, presenting clinical features categorised into dermatological, ophthalmological, or neurological signs, age of symptom onset, age at diagnosis, and referring specialty.</p><p><b>Results</b>: The complementation groups represented included XP-A (n = 20), XP-B (n = 2), XP-C (n = 29), XP-D (n = 14), XP-E (n = 6), XP-F (n = 6), XP-G (n = 7), XP-V (n = 16). By age 3, 84% of patients had developed clinical signs of the condition (excluding those presenting with skin cancers), yet the mean age at diagnosis was 12.2 years (median 6; range 0.5–64 years).</p><p>Initial clinical signs varied significantly. Exposed-site pigmentary changes appeared at a mean age of 5 years (range 0.5–44 years) and were predominantly observed in XP-C (83%, mean age 2.3 years, range 0.75–6 years) and XP-V (56%, mean age 14.4, range 2–44) patients.</p><p>Photosensitivity, characterised by severe prolonged sunburn, was the primary symptom in XP-D (100%), XP-F (100%), XP-G (86%), and XP-A (55%) patients, excluding those with the milder XP-A variant. Median diagnostic delays were shorter in these complementation groups (range 3.2-11 years).</p><p>Skin cancers at presentation were most common in XP-E (66%, mean age 26 years, range 15–40), XP-V (44%, mean age 35 years, range 16–46) and XP-C (7%, mean age 16 years, range 4–28 years).</p><p>Ocular signs, including conjunctivitis and photophobia were the first presenting features in two XP-C patients aged 0.5 and 1 year. Neurological symptoms were the initial presenting feature in two XP-A patients who exhibited developmental delays by age two.</p><p>Diagnostic delays were particularly pronounced in XP-E and XP-V patients, with median delays of 28.5 and 21.5 years, respectively. Dermatologists were the most frequent referrers, accounting for 73% of referrals, followed by geneticists (7%), general practitioners (10%), paediatricians (5%), neurologists (3%) and ophthalmologists (2%).</p><p><b>Conclusion</b>: Most XP patients exhibit clinical signs early in life, but significant diagnostic delays exist, particularly in groups with milder cutaneous manifestations where severe sunburn reactions or pigmentary changes are lacking. Dermatologists are the primary referrers, underscoring their critical role in early recognition. Increased awareness across all specialties is essential to improve diagnostic timelines and outcomes for XP patients.</p><p>Guzzon Diletta^1*, Paccosi Elena^1*, Filippi Silvia¹, Valeri Emma¹, De Lanerolle Primal² and Proietti-De-Santis Luca^1#</p><p>¹Unit of Molecular Genetics of Aging, Department of Ecology and Biology (DEB), University of Tuscia, 01100 Viterbo, Italy.</p><p>²Department of Physiology and Biophysics, College of Medicine, University of Illinois at Chicago, 835 S. Wolcott, Chicago, IL 60612.</p><p>Cockayne Syndrome group A (CSA) is an ubiquitin E3 ligase belonging to the family of WD-40 repeat proteins. CSA was initially characterized, together with Cockayne syndrome group B (CSB) protein, as playing a role in Transcription Coupled Repair even if, in the last years, a growing body of evidence shows how this protein, together with CSB, exerts a key role in the regulation by ubiquitination and proteasomal degradation of a plethora of different proteins involved in the most disparate cellular processes. Mutations in CSA, as well as in CSB, result in Cockayne syndrome (CS), a human autosomal recessive disorder characterized by a variety of clinical features, including growth deficiency and severe neurological and developmental manifestations.</p><p>Actin, a major component of the cytoplasm, has been recently described as abundant in its filamentous form (F-Actin) also in the nucleus, where it is involved in a variety of nuclear processes including transcription and chromatin remodeling. The nuclear export of Actin is mediated by Chromosomal Maintenance 1 or Exportin 1 (CRM1). Here, we demonstrate that CSA may regulate the nuclear localization of Actin by ubiquitinating its nuclear exporter CRM1, in such a manner to avoid the passage of CRM1/Actin complexes through the nuclear pore complex. We also demonstrate that CS-A patients derived fibroblasts display a hampered Actin nuclear retention, in favour of its nuclear export, as a consequence of a defective CRM1 ubiquitination. This unbalance in Actin localization results in both an altered and stiff cytoskeletal shape and in a drastic reduction of nuclear Actin foci. Furthermore, while normal cells display a preferential localization of CRM1 on the nuclear membrane, CS-A mutant cells show, instead, an aberrant presence of CRM1 as cytoplasmic foci, further confirming an abnormal rate of CRM1-mediated export. How to reconcile this defective export of nuclear Actin with Cockayne syndrome features? It is well known that nuclear Actin localizes at the promoters of certain genes, where it helps the recruitment or RNA polymerase II (RNA polII). Interestingly, in CS-A mutated patient's derived fibroblasts, we found a hampered recruitment of Actin on promoters of some genes, such as BDNF and BRD7 ones, corresponding to a defective recruitment of RNA polII on the same promoters and to a defective transcription of these genes. These results may potentially justify the transcriptional defects shown by CS-A patients, especially regarding the neurodevelopmental manifestations.</p><p>It is still to determine if also CSB is someway involved in this new regulatory pathway, maybe being responsible of the degradation of the nuclear exporter protein CRM1. Further studies will be required to better define both the eventual role of CSB and the consequences of the impairment of Actin shuttling on the clinical features of CS patients.</p><p>Silvia Filippi, Emma Valeri, Elena Paccosi, Diletta Guzzon and Luca Proietti De Santis</p><p>Unit of Molecular Genetics of Aging, Department of Ecology and Biology (DEB), University of Tuscia, 01100 Viterbo, Italy.</p><p>Cockayne syndrome (CS), defined as Nucleotide excision repair (NER) syndrome, is a rare autosomal recessive disorder linked to mutations in ERCC8 and ERCC6 genes, which encode for CS group A (CSA) and group B (CSB) proteins respectively, both of which play a role in Transcription Couples repair (TCR), a sub-pathway of NER, devoted to removal of lesions on transcribed genes. Although CS patients exhibit hypersensitivity to UV irradiation, they do not experience an increased risk of skin cancer occurrence in contrast to another NER syndrome defined as Xeroderma pigmentosum. While a loss-of-function mutation in the ERCC8 and ERCC6 genes causes a variety of senescence- and cell death-related abnormalities, increased expression of CSA and CSB proteins has been reported in cancer cells from different tissues often associated with increased proliferation and cell robustness. Recently, we demonstrated that CSA protein is over-expressed in breast cancer cells and its down-regulation by ASO technology dramatically reduces not only the tumorigenicity but also enhances the sensitization of BC cells to both oxaliplatin and paclitaxel drugs, two of the major chemotherapy agents used for triple-negative breast subtype. In this line, we decided to analyze the CSA expression in primary (WM115) and metastatic (WM266-4) melanoma cells. Melanoma is the most aggressive form of skin cancer. Research has made a great stride, in the last years, trough the introduction of innovative and effective therapies: immunotherapy and target therapy, unfortunately, some types of melanomas are refractory to therapeutic treatments. In this case, conventional chemotherapy with methylating agents: Dacarbazine (DTIC) and Temozolomide (TMZ) is considered as a last-line option. Both DTIC and TMZ are associated with unclear survival benefits and treatment-related toxicities. Our studies demonstrated that both WM115 and WM266-4 cells display an overexpression of CSA. Furthermore, CSA suppression greatly sensitizes both cell lines, in particular the metastatic ones, to both Dacarbazine (DTIC) and Temozolomide (TMZ) drugs, even at very low doses which are not harmful to normal cells, in term of cell proliferation, cell survival and apoptotic response. Further, studies are mandatory to evaluate whether CSA may be considered a very attractive target for the development of more effective antimelanoma therapies.</p><p>Julie Soutourina</p><p>Institute for Integrative Biology of the Cell (I2BC), CEA - CNRS - University Paris-Saclay, 91191 Gif-sur-Yvette France</p><p>Transcription and DNA repair are fundamental functions of the cell. Their dysfunctions lead to cell death, mutagenesis and pathologies. In NER, transcription-coupled repair removes DNA lesions interfering with RNA polymerase II progression. Inherited NER defects lead to xeroderma pigmentosum, Cockayne syndrome and triochothiodystrophy with complex syndromes including sunlight sensitivity, skin cancer or premature aging and neurological symptoms. Mediator is an essential and conserved multisubunit coactivator complex. In human, neurodevelopmental diseases mapped to Mediator mutations were regrouped as Mediatorpathies. New Mediator variants were recently uncovered in patients with transcription and TCR defects. However, many questions remain unanswered on mechanisms of complex diseases related to transcription and NER deficiencies.</p><p>The yeast <i>Saccharomyces cerevisiae</i> offers a unique opportunity to unveil the fundamental eukaryotic mechanisms thanks to its powerful genetic and genomic tools. Our work contributed to understanding of Mediator transcription function. Moreover, we have discovered its novel role by connecting transcription and NER via Rad2, the yeast homolog of human XP/CS-related XPG. Taking advantage of yeast, we transposed pathological Mediator mutations associated with CS-like symptoms and showed that they led to growth and UV-sensitivity phenotypes, and genetic interactions with TCR components. We are characterising their impact on physical interactions with Pol II and Rad2, transcription and DNA repair.</p><p>Transcription also affects mutagenesis and DNA repair prevents mutations responsible for aging and diseases. However, mechanisms involved in mutagenesis associated with transcription and DNA repair remain to be fully understood. Recently, we developed an innovative microfluidic-based system, coupled with high-throughput sequencing, for mutation accumulation in yeast. We transposed XP, CS or TTD-associated mutations and are analysing their impact on mutagenesis using our microfluidic-based and reporter-gene approaches combined with mutagen treatments.</p><p>In conclusion, we propose how the yeast model can give important insights into our understanding of the molecular mechanisms at the origin of complex human diseases related to transcription and NER deficiencies.</p><p>Philipp-Kjell Ficht¹, Anna Staffeld¹, Wilhelm Sponholz¹, Rüdiger Panzer¹, Anneke Lemken¹, Nataliya DiDonato², Joseph Porrmann², Mensuda Hasanhodzic³, Ales Maver⁴, Tasja Scholz⁵, Maja Hempel⁵, Oleksandra Kuzmich⁶, Steffen Emmert¹, Lars Boeckmann¹</p><p>¹Clinic and Policlinic for Dermatology and Venereology, University Medical Center Rostock, Germany</p><p>²Institute for Clinical Genetics, University Hospital Carl Gustav Carus at the Technische Universität Dresden, Germany</p><p>³Department of endocrinology, metabolic diseases and genetics University Clinical Centre Tuzla, Bosnia and Herzegovina</p><p>⁴Centre for Mendelian Genomics, Clinical Institute of Medical Genetics, UMC Ljubljana, Slovenia</p><p>⁵Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany</p><p>⁶DWI - Leibniz Institute for Interactive Materials e. V., Aachen, Germany</p><p>Xeroderma pimentosum (XP) is an autosomal recessive disorder characterized by increased photosensitivity, actinic skin damage, neurological abnormalities, and an elevated risk of skin and mucosal cancers. This condition is caused by defects in genes encoding for components of the nucleotide excision repair (NER) pathway. In this study, genetic and functional analyses were performed on patients with a suspected clinical diagnosis of XP or those identified with mutations in an XP-related gene. For patients with an unknown genetic cause, DNA sequencing was conducted to determine the underlying genetic defect. Additionally, functional analyses of patient cells were performed to assess their repair capacity and survival rates following UV-C irradiation. Exome or Sanger sequencing revealed identical compound heterozygote mutations in ERCC2 (XPD) in two patients, one patient with a homozygous missense variant in DDB2 (XPE), one with a homozygous single nucleotide variant in XPA and one with a homozygous mutation in XPC. Sequencing of DNA from two further patients is currently conducted. The presents of a heterozygous variant in one of the parents was confirmed for the patients with a homozygous variant in XPA or XPC. The exposure of patient derived fibroblasts to UV-C irradiation showed no reduced post-UV-survival compared to wild type cells for cells from patients with genetic alteration in ERCC2 or DDB2, a slightly reduced post-UV-survival for XPC and no reduced post-UV-survival for cells with alterations in XPA. Preliminary results of a host cell reactivation assay (HCR) showed that the reduced DNA repair capacity of cells with a defect in DDB2 could be compensated by the transfection of the cells with wild type DDB2. No compensation was observed for cells with alterations in ERCC2. Hair analyses for patients with compound heterozygous mutations in ERCC2 showed neither a reduced cysteine content nor the typical tiger-tail pattern as seen in a patient with trichothiodystrophy (TTD) under polarized light microscopy. Overall this ongoing study characterizes and correlates the genotypes and phenotypes of seven patients with varied clinical symptoms.</p><p>Lars Boeckmann¹, Philipp Ficht¹, Anna Staffeld¹, Rüdiger Panzer¹, Steffen Emmert¹</p><p>¹Clinic and Policlinic for Dermatology and Venereology, University Medical Center Rostock, Germany</p><p>The nucleotide excision repair (NER) is essential for the repair of ultraviolet (UV)-induced DNA damage, such as cyclobutane pyrimidine dimers (CPDs) and 6,4-pyrimidine-pyrimidone dimers (6,4-PPs). Alterations in genes of the NER can lead to NER-defective syndromes. The main NER-defective syndromes comprise xeroderma pigmentosum (XP), Cockayne syndrome (CS), and trichothiodystrophy (TTD).</p><p>Clinical and molecular-genetic assessment of XP-C patients from Germany revealed absence of sun-sensitive as well as neurological symptoms. The mean age of XP diagnosis was 9.4 years, and the median age of the first skin cancer was 7 years. We identified five new mutations including an amino acid deletion (c.2538_2540delATC; p.Ile812del) resulting in repair deficiency but no XPC message decay.</p><p>Assessment of XPG-defective patients revealed that the type and location of mutations determine the clinical phenotype. We identified 3 missense mutations and showed by molecular means that these missense mutations in the I-region of the XPG protein impaired both repair and transcription and delayed the recruitment of other XP proteins to UV photodamage as well as their redistribution thereafter. The patients exhibited a XP/CS complex phenotype.</p><p>We also identified two XPG and XPF splice variants with residual repair capabilities in NER. Almost all variants are severely C-terminally truncated and lack important protein-protein interaction domains. Interestingly, XPF-202, differing to XPF-003 in the first 12 amino acids only, had no repair capability at all, suggesting an important role of this region during DNA repair.</p><p>German XPD-deficient patients exhibited a XP phenotype in accordance with established XP-causing mutations (c.2079G&gt;A, p.R683Q; c.2078G&gt;T, p.R683W; c.1833G&gt;T, p.R601L; c.1878G&gt;C, p.R616P; c.1878G&gt;A, p.R616Q). One TTD patient was homozygous for the known TTD-causing mutation p.R722W (c.2195C&gt;T). Two patients were compound heterozygous for a TTD-causing mutation (c.366G&gt;A, p.R112H) and p.D681H (c.2072G&gt;C) amino acid exchange, but exhibited different TTD and XP/CS complex phenotypes. Interestingly, the XP/CS patient's cells exhibited a reduced but well detectable mutated XPD protein expression compared with hardly detectable XPD expression of the TTD patient's cells.</p><p>Further genotype-phenotype studies could be performed within the European Reference Networks for Rare Diseases (ERN-Skin).</p><p>Francesca Brevi¹, Arjan Theil², Alan Lehmann³, Sebastian Iben⁴, Donata Orioli¹ and Elena Botta¹</p><p>¹Istituto di Genetica Molecolare (IGM) CNR, Pavia, Italy</p><p>²Department of Molecular Genetics, Erasmus MC, University Erasmus Medical Center, Rotterdam, The Netherlands</p><p>³Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton, UK</p><p>⁴Department of Dermatology and Allergic Diseases, Ulm University, Ulm, Germany</p><p>Trichothiodystrophy (TTD) is a multi-system disease characterized by skin, neurological and growth abnormalities. The presence of photosensitivity defines the two main forms of TTD – the photosensitive (PS) and non-photosensitive (NPS) forms. Mutations in a variety of genes have been associated to the disease. They include three PS-TTD related genes that encode distinct subunits of the transcription/DNA repair factor TFIIH and seven NPS-TTD related genes encoding the beta subunit of transcription factor IIE -TFIIEβ, the splicing factors TTDN1 and RNF113A and four aminoacyl-tRNA synthetases which operate in translation. All these TTD-related factors participate in gene expression, thus raising the notion of TTD as a gene expression syndrome. It has been demonstrated that TTD-causing mutations in TFIIH subunits or TFIIEβ as well as knockout/knockdown of TTDN1 and RNF113A impact on transcription by RNA polymerase I. Consequently, ribosomal biogenesis is affected with concomitant error-prone translation and loss of protein homeostasis (proteostasis). Now, by taking advantage of our recently identified TTD cases mutated in methionyl-, threonyl- or alanyl-tRNA synthetase, we have investigated the quality of translation in tRNA synthetase-defective TTD cases. By using a luciferase-based assay, we found a high translation error rate in all tested TTD cells, indicating translation infidelity. In addition, survival assays in the presence of elevated concentrations of single specific amino acids suggested impaired accuracy of tRNA charging. These alterations are accompanied by accumulation of misfolded proteins, indicating a loss of proteostasis. Overall, translation infidelity and loss of proteostasis appear as a common underlying pathomechanism for the different forms of TTD, which may contribute to impaired development and neurodegeneration in patients.</p><p>Jordana McLoone^1,2, Kyra Webb², Kathy Tucker³, Antoinette Anazodo², Denise Wilson⁵, Linda Martin^1,4,5</p><p>¹UNSW Sydney, School of Clinical Medicine</p><p>²Kids Cancer Centre, Sydney Children's Hospital</p><p>³Hereditary Cancer Centre, Sydney Children's Hospital, UNSW</p><p>⁴Dept Dermatology, Sydney Children's Hospital</p><p>⁵Melanoma Institute Australia</p><p><b>Background</b>: In Australia there are no dedicated diagnostic or management services for patients with xeroderma pigmentosum (XP).</p><p><b>Aim</b>: To understand the supportive care needs of families who have a child diagnosed with XP.</p><p><b>Methods</b>: Australian parents and carers of a child with XP, as well as children with XP 5–18 years with no intellectual disability were invited to participate. Participants were identified via clinical networks and social media patients support groups. A semi-structured interview was conducted in-person or via Zoom and focused on the diagnostic journey, current care, preferred models of care, psychosocial impact, and information needs. Interview data were transcribed verbatim and coded line-by-line using QSR NVivo Pro. Inductive thematic analysis was used to organize nodes and themes.</p><p><b>Results</b>: Eight adult carers and 3 children with XP where interviewed, including one family with XP-A, one family with XP-C, and two families with XP-D. Five out of the seven families identified participated.</p><p>Most families had long delays before the diagnosis with misinterpretation of presenting symptoms. Wait times for genetic testing were 6–12 months. All families reported highly unmet informational needs, including how to sun protect, prognosis and management. Parents reported unmet psychological needs, including guilt, fear, grief, isolation and hypervigilance. Adaptions and sun protective requirements were costly.</p><p>The preferred model of care for patients was a multidisciplinary team that included dermatology, ophthalmology, neurology, and allied health. Continuity of care was emphasized.</p><p><b>Conclusion</b>: The development of comprehensive clinical services to address the diagnostic, preventative, surveillance, treatment and supportive care needs of Australian XP patients is urgent.</p><p>Riccardo Paolini¹, Yvette Walker², T. Xiong¹, Konstantina Vasilakopoulou,¹ Linda Martin^3,4</p><p>¹UNSW School of Built Environment, Faculty of Art, Design &amp; Architecture.</p><p>²UNSW Rare Diseases</p><p>³UNSW School of Clinical Medicine, Faculty of Medicine &amp; Health.</p><p>⁴Sydney Children's Hospital</p><p>⁵Melanoma Institute Australia.</p><p><b>Background</b>: Complete protection against all ultraviolent (UV) radiation requires individuals with xeroderma pigmentosum (XP) to wear highly protective garments, including a hood with a visor, full clothing, and gloves. Commercially available garments have been designed and tested for UV protection only, but may lead to substantial solar heat gains, resulting in human thermal discomfort, de facto restricting outdoor activity and impacting quality of life.</p><p><b>Objective</b>: To test the effect on thermal properties with of the addition of solar control film to the XP visor (PS90 by 3M).</p><p><b>Methods</b>: Surface temperature was measured with a thermal camera (T540 by FLIR). The optical properties of materials were characterised with a UV-Vis-NIR spectrophotometer with a 150 mm integrating sphere (Lambda 1050+ by Perkin Elmer), and the broadband properties (solar, uv, vis, and nir) were computed with the solar irradiance distribution of global horizontal radiation with air mass 1, following ASTM E903.</p><p><b>Results</b>: The UV transmittance of the plain visor is zero in the 250-380 nm wavelength range, but there is some light transmission in the 380-400 nm range (from 0.02 at 385 nm to 0.64 at 400 nm), resulting in a total UV transmittance of 0.08, and solar transmittance of 0.83.</p><p>With solar control film (PS90), the total UV transmittance reduced to 0.02, and solar transmittance decreased to 0.59. The PS90 film added to the visor reduces visible transmittance by 0.11, without compromising a clear vision, and reduces near-infrared transmission by 0.45, therefore almost halving the solar heat gains in the non-visible portion of the solar spectrum. Further, the clear film displays low solar absorbance and, therefore, limits surface overheating. When these combinations were tested outdoors (clear sky conditions 28.4°C, solar radiation of 365 W/m2 and UV radiation of 13 W/m2), the addition of PS90 film on top of the was 2.9 °C cooler than the plane visor 0.6 m/s, solar radiation of 365 W/m2 and UV radiation of 13 W/m2.</p><p><b>Conclusion</b>: XP patients do not have easy access to protective garments with are both UV safe and thermally safe, particularly in warm climates. Addition of a readily available solar film to the UV visor halved solar heat gains.</p><p>Vuong-Brender Thanh^1,2 and Schumacher Björn^1,2</p><p><i>¹Institute for Genome Stability in Aging and Disease, Medical Faculty, University and University Hospital of Cologne, Cologne, Germany</i>.</p><p><i>²Excellence Cluster for Cellular Stress Responses in Aging-Associated Diseases (CECAD), Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany</i>.</p><p>Thanks to a simple body architecture, the nematode <i>C. elegans</i> can be used to study DNA damage response both at cellular and organismal levels. We used <i>C. elegans xpc-1</i> mutants, ortholog of the mammalian sensor of DNA damage XPC, to understand alternative repair in mutants defective for the Global Genome Nucleotide Excision Repair (GG-NER) pathway. XPC-1 is strongly expressed in the germline of <i>C. elegans</i>. When first instant larvae of <i>xpc-1</i> mutants are UV treated, the primordial germ cells (PGCs) fail to develop a germline during the subsequent larval growth to adulthood. We discovered that mutations in the endonuclease <i>gen-1</i>, ortholog of mammalian resolvase of Holiday junctions GEN1, strongly enhanced the UV-induced germline developmental defects of <i>xpc-1</i> mutants. This increase in UV-sensitivity requires the catalytic activity of GEN-1 as well as its C-terminal region. Our data shows that GEN-1 meditates germ cell arrest in <i>xpc-1</i> mutants in response to UV, suggesting GEN-1 regulates DNA damage checkpoint in PGCs when the lesions are not repair by the canonical GG-NER pathway. The arrest seems to allow an alternative repair pathway independent of the double strand break repair mediated by <i>brc-1</i>/BRCA1 and <i>brd-1</i>/BARD1, and the Fanconi Anaemia pathway mediated by <i>fcd-2</i>/FANCD2. The absence of GEN-1 – dependent alternative repair results in replication and chromosomal segregation defects during subsequent S-phase and mitosis respectively, that can be rescued partially by activating the spindle assembly checkpoint. Our results point to a potentially important regulator of DNA repair in XP patients.</p><p>Teodora Svilenska¹, Chiara Cimmaruta², Claudia Bogner³, Vincent Laugel⁴, Wolfram Gronwald³, Miria Ricchetti², York Kamenisch¹* and Mark Berneburg¹*</p><p>¹Department of Dermatology, University Hospital Regensburg, 93042 Regensburg, Germany, Tel.: 0941 944-9601,</p><p>²U5 Molecular Mechanisms of Pathological and Physiological Ageing, Institut Pasteur, 25, rue du Dr. Roux, 75724 Paris Cedex 15, France</p><p>³Institute of Functional Genomics, Department Functional Genomics, Am BioPark 9, 93053 Regensburg, Germany, Tel.: 0941 943 5015</p><p>⁴University Hospital of Strasbourg, Neuromuscular Centre at Hautepierre Hospital, Hautepierre Hospital, Avenue Molière, 67000 Strasbourg, France</p><p>*: corresponding authors</p><p>email: <span>[email protected]</span>, <span>[email protected]</span></p><p>Cockayne syndrome (CS), a rare genetic disease with progeroid symptoms, like, progressive severe neurological defects and UV sensitivity, has no treatment up to now. The investigation of metabolic changes, which are associated with aging processes or stressors, is necessary to increase the knowledge of the complex processes leading to aging of cells and organisms.</p><p>It was shown in previous work, that exposure of human skin cells to stressors like UVA irradiation or reactive oxygen species (ROS) leads to significant changes in the cell metabolism, especially in the glucose metabolism. The high levels of UVA induced consumption of glucose and pyruvate could be involved in the ROS detoxification strategies of these UVA exposed cells. In addition to this it is known that CS cells can exhibit higher levels of cellular ROS damage than WT cells.</p><p>In this project, we investigated the impact of stressors (ROS) on the metabolism and oxygen consumption in primary human skin fibroblasts derived from CS patients with the premature aging syndrome or from healthy individuals (WT). These cells were treated with repetitive low dose UVA irradiation (inducing ROS) with subsequent measurement of cellular oxygen consumption using a Clark type electrode and metabolic changes in the supernatant of the cells, using nuclear magnetic resonance spectroscopy (NMR).</p><p>UVA irradiation induced significant changes in many metabolites (glucose, lactate, pyruvate, acetate, glutamine, glutamate, choline, alanine, betaine) in the cellular supernatant. Similar to WT cells, CS cells showed higher glucose and pyruvate consumption, as well as higher lactate and alanine secretion upon UVA treatment. Interestingly, metabolic differences between WT and CS cells are already present without external stressors (UVA irradiation) and many of these differences increase upon UVA treatment. Concerning cellular respiration, differences in the oxygen consumption rate, between CS and WT cells were visible without external stressors. WT cells show a higher oxygen consumption rate than CS cells. The application of stressors (UVA irradiation) enhanced these differences between WT cells and CS cells.</p><p>This study revealed differences in metabolism and respiration between CS cells of patients with premature aging symptoms and WT cells. It is known, that, under specific conditions, processes of cellular respiration can generate high levels of ROS. Therefore, it can be speculated, that CS cells try to exploit metabolic ROS detoxification strategies associated to glycolysis (higher glucose and pyruvate consumption than WT cells) in order to reduce ROS production during respiration (lower respiration rate than WT cells).</p><p><b>Introduction</b>: Cutaneous malignancies are the leading cause of premature mortality among patients with Xeroderma Pigmentosum. Cutaneous Squamous Cell Carcinoma (cSCC) is the most prevalent skin cancer seen among XP patients in our setting. Rapid local and distant metastases due to their impaired DNA function make it difficult to treat with surgery. Cemiplimab has been approved for treating locally advanced and metastatic cSCC. It is however not available in developing countries like Tanzania.</p><p><b>Case Presentation</b>: A 9 year old male presented to us in January 2023 with two large ulcerated tumours on the scalp (temporal region), a fungating mass on the left mandibular region. Prior to attending our facility he had multiple excisions done at peripheral hospitals but the tumours always recurred. He had also received 4 cycles of chemotherapy at the national hospital. Multidisciplinary team effort with ENT and general surgeons was crucial from the beginning of treatment. Biopsies from the national hospital revealed SCC on both scalp tumours invading bone, CT scan of the mandibular tumour revealed SCC with invasion of the parotid gland, pushing the left external auditory meatus and causing severe pain. Paracetamol and ibuprofen did not relief the pain and we requested to start him on morphine. He had anaemia of 7g/dl since the parotid mass bled profusely during wound dressing and was transfused with 3 units of blood. ENT and general surgeons did debulking surgery; underlying muscles were infiltrated with the mass hence complete excision was not possible. A week later the scalp tumours were excised and he was planned for palliative care on discharge. In September 2023 he came for follow up with recurrent tumours on the same sites and even bigger than previously. After several discussions in the tumour board and after asking for donations, we were able to acquire the monoclonal antibody Cemiplimab. He was started on cycles of Cemiplimab to be given after every 2 weeks and he has received 21 cycles. Significant shrinkage of the tumour was noted from cycle 15 where he only had shallow ulcerations on the scalp. There is no new or recurrent cancer growth seen and all the wounds have closed up.</p><p><b>Conclusion</b>: This case is an example of the hurdles, unsuccessful as they may be, that had to be undertaken to treat advanced cSCC in XP due to lack of availability of novel treatments like Cemiplimab which through research has proven to be successful in treating aggressive cSCC. XP patients are perfect candidates for treatment with immunotherapy and this should be available in all settings.</p><p>Andrey A. Yurchenko</p><p>INSERM U981, Gustave Roussy Cancer Campus, Université Paris Saclay, Villejuif, France</p><p><span>[email protected]</span></p><p>Fatemeh Rajabi</p><p>INSERM U981, Gustave Roussy Cancer Campus, Université Paris Saclay, Villejuif, France</p><p><span>[email protected]</span></p><p>Tirzah B. P. Lajus</p><p><i>Departamento de Biologia Celular e Genética, Universidade Federal do Rio Grande do Norte, Av. Senador Salgado Filho, s/n, Natal, 59078–970, Brazil</i></p><p><span>[email protected]</span></p><p>Hiva Fassihi</p><p>National Xeroderma Pigmentosum Service, Department of Photodermatology, St John's Institute of Dermatology, Guy's and St Thomas’ Foundation Trust, London SE1 7EH, United Kingdom</p><p><span>[email protected]</span></p><p>Chikako Nishigori</p><p>Division of Dermatology, Department of Internal Related, Kobe University Graduate School of Medicine, Kobe, Japan</p><p><span>[email protected]</span></p><p>Carlos F. M. Menck</p><p>Department of Microbiology, Institute of Biomedical Sciences, University of Sao Paulo, Sao Paulo, SP, Brazil</p><p><span>[email protected]</span></p><p>Alain Sarasin</p><p>CNRS UMR9019 Genome Integrity and Cancers, Institut Gustave Roussy, Villejuif, France</p><p><span>[email protected]</span></p><p>Sergey Nikolaev</p><p>INSERM U981, Gustave Roussy Cancer Campus, Université Paris Saclay, Villejuif, France</p><p><span>[email protected]</span></p><p>Xeroderma Pigmentosum (XP) is characterized by a 1000-fold increased risk of skin cancer due to deficiencies in the NER or TLS pathways. The genomic mechanisms of somatic mutagenesis responsible for this increase are not yet fully understood.</p><p>We sequenced genomes from a unique collection of skin tumors (n = 38) originating from cancer-prone XP subgroups (XP-A/C/D/E/V) and developed in vitro models (XP-C/V) to elucidate the mechanisms of UV-induced mutagenesis in XP using detailed bioinformatic analyses.</p><p>The ultra-mutated tumor phenotype was observed in skin tumors with impaired GG-NER (XP-E = 350 mut/MB, XP-C = 162 mut/MB) and translesion synthesis (XP-V = 248 mut/MB). Mutational profiles of XP skin tumors with NER defects were dominated by C&gt;T mutations, with each group exhibiting specific trinucleotide context features. XP-C and XP-D tumors showed a notably large percentage of CC&gt;TT double-base substitutions (17–20%) compared to sporadic skin cancers (4.7%). Mutations in XP-A and XP-D tumors, with impaired GG-NER and TC-NER, exhibited a uniform distribution along chromatin genomic compartments, implying that NER activity is a major modulator of genome-wide mutation rate heterogeneity.</p><p>XP-V skin tumors with dysfunctional TLS polymerase eta were characterized by a specific mutational profile, including an excess of TG&gt;TT substitutions (28%) absent in sporadic skin cancers. These mutations likely result from error-prone bypass of rare, atypical photolesions in TpA and TpG contexts in the absence of polymerase eta. The mutational signature of canonical UV-induced C&gt;T mutations in pyrimidine dimers was distinct in XP-V tumors, primarily defined by incorrect nucleotide insertion at the 3’ end of photodimers (40-fold higher compared to sporadic cancers). This pattern likely reflects error-prone insertion and error-free extension steps performed by different TLS polymerases.</p><p>WGS analysis of clones from UV-irradiated XPC-KO and POLH-KO cell lines revealed 3-fold and 11-fold increased mutagenesis rates, respectively, compared to wild-type cells. These findings demonstrate that error-free TLS bypass plays a major, if not greater, role in reducing UV-induced mutations alongside NER. Genomic analysis of cell lines transfected with photolesion-specific photolyases highlighted the primary role of CPD in mutation generation and 6-4PP in the DNA damage response.</p><p>The accumulation of unrepaired DNA lesions on the untranscribed strand, nonspecific TLS activity, and the inability to bypass atypical photolesions error-free collectively explain the increased incidence of skin cancer in XP patients.</p><p>Genetic analyses of large cohorts of cancer patients have revealed a correlation between somatic mutations in DNA repair pathway genes and the responsiveness of patients to DNA-damaging chemotherapeutics and immune checkpoint inhibition. In line with this, several case studies have shown that Xeroderma Pigmentosum (XP) patients with melanoma respond well to anti-PD1 therapy, suggesting that XP melanomas enable an immune-favoured tumor environment. To examine nucleotide excision repair (NER) deficient melanomas in vivo, we focused on the helicase ERCC2/XPD and developed a melanoma mouse model with ERCC2 wildtype (wt) or ERCC2-mutant tumors that harboured the deleterious point mutations K48R or D234N, the latter occurring in XP patients. For this purpose, we engineered murine melanoma cells that were first characterized in vitro and then transferred to C57BL/6J mice. As expected, XPD-mutant cells displayed increased sensitivity to UV irradiation and cisplatin treatment in cell culture. In vivo, XPD-K48R melanomas developed only in a minority of cases. XPD-D234N tumors developed earlier than XPD-wt tumors, consistent with the observed increased proliferation rate of XPD-D234N cells under basal conditions. Despite their rapid onset, 50% of XPD-D234N melanomas underwent spontaneous regression within 2–3 weeks, a phenomenon never observed in XPD-wt tumors. When mice bearing XPD-D234N melanomas were treated with a combination of cisplatin and PD-1 blocking antibody, almost all tumors regressed, while no therapy response was observed for mice with XPD-wt melanomas. These findings suggest that XPD-mutant melanomas are more easily resolved by the immune system of the host, particularly under conditions of NER-associated DNA damage and immune therapy. Future research will focus on the molecular mechanisms underlying this enhanced therapeutic responsiveness, paving the way for potential clinical applications targeting XPD in cancer.</p><p>Tycho E.T. Mevissen^1,2, Maximilian Kümmecke³, Ernst W. Schmid¹, Lucas Farnung³, and Johannes C. Walter^1,2</p><p>¹Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA</p><p>²Howard Hughes Medical Institute</p><p>³Department of Cell Biology, Harvard Medical School, Boston, MA</p><p>In transcription-coupled DNA repair, RNA polymerase II stalling at DNA lesions leads to the efficient activation of nucleotide excision repair (TC-NER). This pathway was discovered almost 40 years ago, yet fundamental questions about its mechanism remain unanswered, in large part due to the lack of a cell-free system that supports this pathway. We have recapitulated vertebrate cell-free TC-NER, and we use this system to study STK19, a candidate TC-NER factor known to promote transcription recovery after cellular exposure to UV light. When a plasmid containing a site-specific cisplatin DNA intrastrand crosslink is transcribed in frog egg extracts, error-free repair is observed that depends on the canonical repair proteins CSB, CRL4^CSA, UVSSA, ELOF1, as well as STK19. Structural prediction and cryo-electron microscopy indicate that STK19 is an integral component of the TC-NER complex that interacts with CSA-DDB1, the RNA polymerase II subunit RPB1, and the XPD helicase subunit of TFIIH. Mutating these interfaces disrupts cell-free TC-NER, and molecular modeling suggests that STK19 positions TFIIH such that the damaged DNA strand is threaded into the XPD helicase for subsequent lesion verification. In conclusion, our novel cell-free system that supports <i>bona fide</i> eukaryotic TC-NER strongly suggests that STK19 couples RNA polymerase II stalling to TFIIH-dependent downstream nucleotide excision repair events.</p><p>Sikandar G. Khan¹, Wenelia Baghoomian¹, Christiane Kuschal¹, Deborah Tamura¹, Maxwell P. Lee¹, John J. DiGiovanna^{1, *}, Rodrigo Cepeda-Valdes², Julio Salas-Alanis³ and Kenneth H. Kraemer¹</p><p>¹Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States</p><p>²Dystrophic Epidermolysis Bullosa Research Association Mexico Foundation, Monterrey, Mexico; ³Instituto Dermatologico de Jalisco, Jalisco, Mexico, *Deceased</p><p>Trichothiodystrophy (TTD) is a rare (1 per million) autosomal recessive multisystem developmental disorder characterized by short, brittle hair with transverse “tiger-tailed banding” on polarized microscopy. TTD has been reported to be caused by mutations several different functional groups of genes: nucleotide excision repair (NER)/ basal transcription factor II H (TFIIH) genes: <i>ERCC2/XPD, ERCC3/XPB</i> and <i>GTF2H5/ TTDA;</i> amino acid charging tRNA genes (<i>TARS, MARS1, AARS1</i>, and <i>CARS</i>); basal transcription factor II E (<i>GTF2E2/TFIIEβ)</i>; X-linked ring finger protein 113A (<i>RNF113A)</i>, and a gene of unknown function (<i>MPLKIP/TTDN1)</i>. We performed whole exome sequencing to identify a candidate gene in Sabinas brittle hair syndrome, a mild form of TTD. We report 5 non-photosensitive adult patients from 3 unrelated families with a homozygous missense mutation in <i>DBR1</i> (p.D262Y) encoding the RNA lariat debranching enzyme DBR1 which catalyzes the removal of introns from pre-mRNA in the nucleus. Post-UV DNA repair was normal, indicating that DBR1 was not involved in NER/TFIIH. The cells had reduced levels of <i>DBR1</i> mRNA and no detectable DBR1 protein. Previous reports described interaction of DRB1 and TTDN1 proteins. We found markedly reduced levels of TTDN1 protein in cells from patients with <i>DBR1</i> mutations and no detectable levels of TTDN1 protein in the cells from patients with <i>TTDN1</i> mutations. The stabilization of either of these proteins is critical for splicing and transcription. Genetic analysis suggests an ancient origin of this mutation. Thus, we identified <i>DBR1</i> as causing Sabinas brittle hair syndrome supporting the hypothesis that TTD can be caused by transcriptional impairment.</p><p>Marc Majora, Rituparna Bhattacharjee, Selina Dangeleit, Andrea Rossi, Jean Krutmann</p><p>IUF – Leibniz-Institut für umwelt- medizinische Forschung GmbH, Düsseldorf, Germay</p><p>Xeroderma pigmentosum type A (XPA), an inherited disease characterized by UV hypersensitivity, high skin cancer risk and premature aging, is thought to be caused by defective repair of nuclear DNA. Recent studies, however, suggest that XPA proteins might have functions beyond nuclear DNA repair. Here we show – by analyzing purified mitochondrial fractions from primary human dermal fibroblasts (HDF) – that XPA proteins are located inside the mitochondria. In particular, mitochondrial XPA content was increased when HDF were either irradiated with UVB or treated with the pro-oxidant menadione suggesting that XPA might be important for repairing damage inside the mitochondria and protecting their integrity. Accordingly, sequencing of mitochondrial DNA (mtDNA) of HDF obtained from XPA patients (XPA HDF) revealed an elevated load of mutations as compared with healthy HDF, which was further increased by UVB irradiation. These data suggest that XPA is not only involved in the repair of nuclear DNA but also participates in the repair of mtDNA. The pivotal role of XPA in maintaining mitochondrial function was also reflected by a RNA-Seq transcriptome analysis showing that “Mitochondrial Gene Expression” and “Mitochondrial Translation” were among the most severely suppressed biological processes in UVB-irradiated XPA HDF. To assess potential consequences for mitochondrial function, we next measured the cellular ATP production rate. We found that unirradiated XPA HDF had a higher total ATP production rate than healthy HDF, which was due to increased mitochondrial but not glycolytic ATP production rate. If cells were stressed by UVB irradiation, mitochondrial ATP production rate in XPA HDF was reduced by more than 50% while it remained stable in normal HDF. Thus, XPA HDF have an increased ATP demand, which in unstressed cells can still be met by their mitochondrial function. Upon irradiation, however, compensation fails and XPA HDF become energy deficient. As the chaperone HSP90 serves as an ATP sensor which stabilizes proteins in an ATP-dependent manner, we next analyzed the abundance of HSP90 client proteins. HSP90 levels remained constant, but a marked loss of ErbB2, EGFR, STAT3 and SIRT1 was detected in irradiated XPA HDF when compared to healthy HDF, reflecting collapse of proteostasis. Our results indicate that XPA proteins are present in mitochondria where they maintain integrity of mtDNA and mitochondrial function to ensure cellular ATP supply and thereby prevent collapse of proteostasis, a well-known driver of aging.</p><p>Paola Giunti, Hector Garcia-Moreno, Douglas R Langbehn, Adesoji Abiona, Isabel Garrood, Zofia Fleszar, Marta Antonia Manes, Ana M Susana Morley, Emma Craythorne, Shehla Mohammed, Tanya Henshaw, Sally Turner, Harsha Naik, Istvan Bodi, Robert P E Sarkany, Hiva Fassihi, Alan R Lehmann</p><p>National Xeroderma Pigmentosum Service, Department of Photodermatology, St John's Institute of Dermatology, Guy's and St Thomas' Foundation Trust, London SE1 7EH, United Kingdom</p><p>and</p><p>Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton BN1 9RQ, United Kingdom</p><p>Xeroderma pigmentosum (XP) results from biallelic mutations in any of eight genes involved in DNA repair systems, thus defining eight different genotypes (XPA, XPB, XPC, XPD, XPE, XPF, XPG and XP variant or XPV). In addition to cutaneous and ophthalmological features, some patients present with XP neurological disease. It is unknown whether the different neurological signs and their progression differ among groups. Therefore, we aim to characterize the XP neurological disease and its evolution in the heterogeneous UK XP cohort.</p><p>Patients with XP were followed in the UK National XP Service, from 2009 to 2021. Age of onset for different events was recorded. Cerebellar ataxia and additional neurological signs and symptoms were rated with the Scale for the Assessment and Rating of Ataxia (SARA), the Inventory of Non-Ataxia Signs (INAS) and the Activities of Daily Living questionnaire (ADL). Patients’ mutations received scores based on their predicted effects. Data from available ancillary tests were collected.</p><p>Ninety-three XP patients were recruited. Thirty-six (38.7%) reported neurological symptoms, especially in the XPA, XPD and XPG groups, with early-onset and late-onset forms, and typically appearing after cutaneous and ophthalmological symptoms. XPA, XPD and XPG patients showed higher SARA scores compared to XPC, XPE and XPV. SARA total scores significantly increased over time in XPD (0.91 points/year, 95% confidence interval: 0.61, 1.21) and XPA (0.63 points/year, 95% confidence interval: 0.38, 0.89). Hyporeflexia, hypopallesthaesia, upper motor neuron signs, chorea, dystonia, oculomotor signs and cognitive impairment were frequent findings in XPA, XPD and XPG. Cerebellar and global brain atrophy, axonal sensory and sensorimotor neuropathies, and sensorineural hearing loss were common findings in patients. Some XPC, XPE and XPV cases presented with abnormalities on examination and/or ancillary tests, suggesting underlying neurological involvement. More severe mutations were associated with a faster progression in SARA total score in XPA (0.40 points/year per 1-unit increase in severity score) and XPD (0.60 points/year per 1-unit increase), and in ADL total score in XPA (0.35 points/year per 1-unit increase).</p><p>Symptomatic and asymptomatic forms of neurological disease are frequent in XP patients, and neurological symptoms can be an important cause of disability. Typically, the neurological disease will be preceded by cutaneous and ophthalmological features, and these should be actively searched in patients with idiopathic late-onset neurological syndromes. Scales assessing cerebellar function, especially walking and speech, and disability can show progression in some of the groups. Mutation severity can be used as a prognostic biomarker for stratification purposes in clinical trials.</p><p>Mark Berneburg</p><p>Department of Dermatology, University Hospital Regensburg, Germany</p><p>The human genome is constantly exposed to various sources of DNA damage. Ineffective protection from this damage leads to genetic instability, which can ultimately give rise to somatic disease, premature aging and cancer. Therefore, our organism commands a number of highly conserved and effective mechanisms responsible for DNA repair. If these repair mechanisms are defective due to germline mutations in relevant genes, rare diseases with DNA repair deficiencies can arise. Today, a limited number of rare hereditary diseases characterized by genetic defects of DNA repair mechanisms is known, comprising ataxia telangiectasia, Nijmegen breakage syndrome, Werner syndrome, Bloom Syndrome, Fanconi anemia, Cockayne syndrome Trichothiodystrophy as well as Xeroderma pigmentosum. Although heterogeneous in respect to selected symptoms, these rare disorders share many clinical features such as growth retardation, neurological disorders, premature aging, skin alterations including abnormal pigmentation, telangectasia, xerosis cutis, pathological wound healing as well as an increased risk of developing different types of cancer. Better understanding of underlying molecular pathology in these diseases, has given rise to potential treatment approaches. In this talk, the different routes and modes of action for therapeutic approaches – ranging from treatments of underlying mechanisms to treatments of the different segmental clinical symptoms of these patients – will be discussed.</p><p>Nihan Erden¹, Björn Schumacher¹</p><p>¹Institute for Genome Stability in Ageing and Disease, Cologne Excellence Cluster for Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne</p><p>DNA damage is a causal factor of both cancer development and the aging process. The tumor suppressor p53 is a central mediator of the DNA damage response (DDR) and the single most frequently mutated gene in human cancer. Many studies showed that cell-cycle and apoptosis functions of p53 are important for preventing tumor development and the activation of p53 was regulated cell-autonomously depending on the type and severity of the DNA damage. It was recently discovered that the p53-mediated DDR in stem cells is not regulated only cell-autonomously but also regulated through signalling via the niche cells. The translation initiation factor IFE-4 in <i>C. elegans</i> is activated in somatic gonad precursor niche cells that surround the primordial germ cells, when the latter carry DNA damage. Moreover, it was demonstrated that the IFE-4 ortholog eIF4E2 in mammals is induced in niche cells upon UV-induced DNA damage and is required for the induction of p53 in hair follicle stem cells (HFSCs). These data thus indicate a highly conserved mechanism of non-cell-autonomous regulation of the p53-mediated DDR in stem cells.</p><p>We are currently employing in vivo and <i>ex vivo</i> experimental systems with eIF4E2 epidermis specific knockout mice to dissect the mechanisms of the interactions between niche and stem cells, and to understand the role of eIF4E2 in skin homeostasis and carcinogenesis. To further assess the clinical relevance of the eIF4E2 function, we will characterize eIF4E2 in human squamous cell carcinoma.</p><p>Dr Hiva Fassihi</p><p>National Xeroderma Pigmentosum Service, Department of Photodermatology, St John's Institute of Dermatology, Guy's and St Thomas’ Foundation Trust, London SE1 7EH, UK</p><p>Prof Alan Lehmann</p><p>Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton BN1 9RQ, UK</p><p>Over 100 patients with Xeroderma Pigmentosum (XP), over 30 with Cockayne Syndrome (CS), and 13 with Trichothiodystrophy (TTD) are under long-term follow-up in the UK National Nucleotide Excision Repair (NER) Multidisciplinary Service. Involvement of the UK patient support groups, Action for XP and Amy and Friends, is essential in the care of these patients.</p><p>The aims of the specialist team of clinicians, nurses and scientists is to improve clinical outcomes and reduce morbidity and mortality. Over the last 14 years, life expectancy of patients with XP in the UK has significantly improved when compared to data on 106 patients from a National Institutes of Health study analysing the worldwide literature from 1971 to 2009. In the UK, XP patients without neurodegeneration now have a life expectancy comparable to the general UK population, because of early diagnosis, meticulous photoprotection and prompt diagnosis and treatment of any skin and ocular cancers.</p><p>Functional studies and molecular analysis to determine the pathogenic variants in the NER genes (part of the specialist service) have enabled the detailed genotype-phenotype examination of these patients. The careful clinical assessment of suspected cases has led to the discovery of a new XP complementation group, XP-J, with mutations in the <i>GTF2H4</i> gene, and contributed to the reporting of <i>MORC2</i>-related disorder with phenotypic similarities to CS.</p><p>Among the patients visiting the multidisciplinary clinic, several groups have relatively mild phenotypes. In most of these cases, in XP-A, D and G, they are associated with mutations at splice donor sites (outside the invariant GT nucleotides), causing abnormal splicing of most of the mRNA, but a residual amount of normally spliced mRNA. The resulting small amount of normal protein is sufficient to ameliorate or delay the onset of the clinical features. In four CS patients, mutation of the G at the fifth base of a splice donor site results in a markedly delayed onset of the clinical features.</p><p>Marvin van Toorn, Jurgen Marteijn</p><p>Department of Molecular Genetics and Oncode Institute, Erasmus University Medical Centre, Rotterdam, the Netherlands</p><p>Nucleotide excision repair (NER) preserves genome stability through a ‘cut-and-patch’-type DNA repair reaction. Whereas the initial NER reaction steps that sequentially recognize, verify and excise helix-distorting DNA lesions through dual incision are well-characterized, mechanistic insight how the generated ssDNA gap is subsequently restored remains limited. Here, we study the mechanisms and factors involved in this late, post-incision NER step. We show that RFC mediates PCNA recruitment and loading at NER-generated ssDNA gaps, after which POLD3 displaces RFC to recruit POLD1 and enable polδ-dependent repair synthesis. Subsequent LIG3-mediated nick ligation promotes PCNA unloading by ATAD5-RLC, which facilitates PCNA recycling to newly NER-generated ssDNA gaps. Interestingly, we found that PCNA and polδ recruitment to sites of DNA damage was severely reduced in actively replicating cells. This could explain the existence of a parallel, non-redundant and PCNA-independent gap restoration pathway involving the alternative clamp loader CTF18-RLC, polε and LIG1. Collectively, our study provides important insights into the intricate mechanisms that preserve genomic integrity by coordinating the productive restoration of NER-generated ssDNA gaps throughout the cell cycle.</p><p>Chikako Nishigori¹, Mariko Tsujimoto¹ and Shinichi Moriwaki²</p><p>¹Division of Dermatology, Department of Internal Related, Graduate School of Medicine, Kobe University, Kobe, Japan</p><p>² Department of Dermatology, Osaka Medical and Pharmaceutical University</p><p>Xeroderma pigmentosum (XP) is an autosomal recessive DNA repair disorder characterized by photosensitivity and progressive central and peripheral nervous system impairment. Currently, there are no effective treatments for XP aside from avoiding UV exposure. It has been shown that XP cells are impaired in scavenging ROS and oxidative DNA damage. Therefore, several antioxidants were examined for reducing cytotoxicity caused by oxidative stress in XP-A cells. Since in vitro experiments showed N-acetyl-5-methoxytryptamine and nicotinamide reduced cytotoxicity of XP-A cells caused by radical inducers, we conducted an in vivo experiment to investigate the therapeutic potential of N-acetyl-5-methoxytryptamine. Treatment with N-acetyl-5-methoxytryptamine mitigated UV-induced inflammation, skin tumorigenesis and hearing deterioration in XP model mice. Our results show that N-acetyl-5-methoxy-tryptamine could alleviate XP symptoms through its anti-inflammatory and antioxidant properties. &lt;https://www.sciencedirect.com/science/article/pii/S0923181125000039&gt;.</p><p>Therefore, we conducted a clinical trial on the efficacy of N-acetyl-5-methoxytryptamine (NPC-15) for patients with XP with exaggerated sunburn-reaction type by a multicenter, double-blinded placebo-controlled, two-group crossover study followed by a 52-weeks open study. Ethics approval was overseen by the Kobe University Institutional Review Board and Osaka Medical and Pharmaceutical University Institutional Review Board, and the study was conducted in accordance with the approved protocol (Japan Registry of Clinical Trials (jRCT) identifier: jRCTs051210181. Registered on February 23, 2022. The clinical trial was conducted from April 2022 to December 2023. Twenty patients (Age; 10.6±6.8, F/M; 12/8). Eighteen patients were XP-A. After key open, all data was analyzed and statistically investigated. In the primary endpoint, the MED, 72 hours (+/-6 hours) after UV irradiation on the 15th day (crossover period I and crossover period II) of investigational drug administration, the treatment effect in NPC-15 administration did not show a statistically significant improvement compared to placebo administration. From the viewpoint of individual patients, data suggesting a usefulness on some of the items including neurological severity scale score on XP were obtained. NPC-15 had no significant safety problem. Brief summary is available through the URL below. &lt;https://jrct.niph.go.jp/latest-detail/jRCT2051210181&gt;.</p><p>Melanie van der Woude, Karen L. Thijssen, Mariangela Sabatella, Jurgen A. Marteijn, Wim Vermeulen, Hannes Lans</p><p>Department of Molecular Genetics, Erasmus MC, Rotterdam, Netherlands</p><p>The versatile nucleotide excision repair (NER) pathway protects organisms against the harmful effects of various types of helix-distorting DNA damage. Hereditary NER deficiency gives rise to several human cancer-prone and/or progeroid disorders, including xeroderma pigmentosum, Cockayne syndrome, trichothiodystrophy, or a combination of these. It is not exactly understood how defects in the same DNA repair pathway cause different disease features and severity.</p><p>To better understand the pathogenesis underlying different NER disorders, we make use of human cellular and <i>C. elegans</i> model systems to investigate mechanisms of DNA repair and their in vivo impact. We previously reported that the core NER machinery is continuously targeted to DNA damage in human cells carrying severe NER disease mutations. Here we show, using real-time imaging, that different types of NER deficiency differently affect the binding of core NER factors to DNA damage and that the prolonged binding of specific NER factors to DNA damage correlates to NER disease severity. Using <i>C. elegans</i>, we show that DNA damage can cause a strong developmental arrest and neuronal dysfunction, which is dependent on transcription-coupled DNA repair and on the persistence of specific DNA repair intermediates in the absence of repair. Together, these results identify stalled transcription-coupled NER intermediates as a pathogenic feature of NER deficiency and show that these adversely affect tissue functionality and organismal development.</p><p>Vilhelm A Bohr</p><p>University of Copenhagen, Center for Healthy Aging, and previously National Institute on Aging, NIH</p><p>We find that some DNA repair defective diseases with severe neurodegeneration have mitochondrial dysfunction. Our studies involve cell lines, the worm (c.elegans), and mouse models and include the premature aging syndromes Xeroderma pigmentosum group A, Cockaynes syndrome, Ataxia telangiectasia and Werner syndrome. It also includes models of Alzheimers Disease (AD). Under these conditions we find a pattern of hyperparylation, deficiency in NAD⁺ and Sirtuin signaling and mitochondrial stress, including deficient mitophagy as a prominent feature. NAD supplementation stimulates mitochondrial functions including mitophagy and stimulates DNA repair pathways. Cockayne syndrome (CS) mice have a hearing deficit that reflects the one seen in the patients. Short term NAD supplementation with NR deminishes the hearing loss. The hearing loss is a of a similar type seen in age related hearing loss. Treatment of mice with NR improves the age related hearing loss. Current studies in CS mice show kidney disease reflecting what is also seen in many CS patients and the mice have deficits in NAD synthesis pathways.</p><p>Clinical trials with NAD intervention have shown benefits in AT patients and a current intervention study in Werner patients also shows some benefits. These will be discussed.</p><p>Giulia Branca^a, Debora Ferri^a, Manuela Lanzafame^a, Erica Gandolfi^a, Gianluca Guida^a, Tiziana Nardo^a, E. Botta^a and Donata Orioli^a</p><p>^aIstitute of Molecular Genetics (IGM) L.L. Cavalli Sforza, CNR, 27100 Pavia, Italy</p><p>The multiprotein complex TFIIH is a key factor in transcription and nucleotide excision repair (NER) pathway. TFIIH malfunctioning results in distinct autosomal recessive disorders, including the cancer-prone xeroderma pigmentosum (XP) and the multisystemic trichothiodystrophy (TTD), the latter one being characterized by physical and mental retardation, signs of premature ageing, but no skin cancer. Both XP and TTD cells are characterized by hypersensitivity to UV irradiation, persistence of NER proteins at the site of damage and accumulation of unrepaired DNA lesions. We have shown that TTD more than XP cells suffer of wide transcription deregulations. Whether this is accountable to TFIIH malfunctioning caused by TTD-specific mutations or to the findings that all TTD-causing mutations affect the stability of the entire complex and result in reduced TFIIH amount, is still an open question. The finding that TFIIH stabilization by low temperature or chemical treatment partially recover TFIIH functionality in primary cells from thermo-sensitive TTD cases (whose clinical features worsen with fever episodes), suggested to us the relevance of preserving TFIIH amount in human cells. We have now identified a novel TFIIH-containing multiprotein assembly bound to the chromatin, whose function is to assists RNA polymerase II during transcription. Reduced levels of wild-type TFIIH or other components of the protein assembly lead to transcription alterations similar to those observed in TTD. Overall, our findings demonstrate that quantitative, as well as qualitative, TFIIH alterations contribute to the wide spectrum of TTD clinical features.</p><p>Kenneth H. Kraemer¹, Deborah Tamura², Sikandar G. Khan, John J. DiGiovanna³</p><p>¹Lab of Cancer Biology and Genetics, NCI, NIH, Bethesda, MD, USA emeritus</p><p>²retired</p><p>³deceased</p><p>Studies of xeroderma pigmentosum (XP) began at NIH in 1971 to gain insights into clinical disease and mechanisms of DNA damage and repair. Detailed clinical and lab exams of skin and eye abnormalities, neurologic degeneration, hearing loss, skin and internal cancers, and aging were performed. We collaborated within NIH and with extra-mural scientists. Findings included discovery of 5 excision repair (ER) deficient XP complementation groups, the ER proficient XP variant, and measuring the &gt; 1,000-fold increase in skin and eye cancer. We developed host cell reactivation assays for UV survival and mutagenesis and found that one DNA dimer inactivated expression in XPA cells. XP patients treated with oral retinoids provided the first demonstration of effective cancer chemoprevention in humans. Audiograms were used to assess the rate of XP neurological degeneration. Autopsies revealed infantile brains in adult XP patients attesting to the massive atrophy caused by XP. Internal cancers include brain and spinal cord neoplasms, thyroid cancer, leukemia, lymphoma and lung cancer (in smokers). Premature menopause, a feature of aging, was observed (median age 29.5 years was &gt; 20 years younger than in the U.S. population). We found XPA patients with severe, intermediate, or mild disease. Trichothiodystrophy (TTD) patients with mutations in the XPD gene had abnormal development including absent myelin in their brain, multiple infections, bone abnormalities, aseptic necrosis of the hips and premature death but no increase in skin cancer. Mothers of TTD children with XPD mutations had many pregnancy abnormalities while mothers of XP children with XPD mutations had normal pregnancies thereby linking XPD function with human fetal development. We found two new RNA related TTD causing genes: TFIIE2 and DBR1. With Dr V. Bohr in 1985 we established the DNA Repair Interest Group with monthly videoconferences (several hundred archived at http://videocast.nih.gov) and initiated the first 3 of this series of international XP meetings (in 2004, 2006, and 2010). These intensive investigations of XP at NIH and worldwide have resulted in advances in understanding disease mechanisms and, most importantly, in increasing patient wellbeing and prolonging their life expectancy.</p><p>Dr. R. Sarkany</p><p>Consultant Dermatologist, UK XP National Service, London</p><p>The most common cause of premature death in XP is metastatic skin cancer (1) and 80% of skin cancers of XP are on the face, head and neck (2). So rigorous and absolute photoprotection from ultraviolet is crucial in the management of patients with XP, particularly of the face.</p><p>We have recently used a new methodology to accurately estimate the daily dose of UV reaching the skin of the face over a prolonged period, and used this to study photoprotection behaviour in detail over a 3 week period in 36 XP patients in the UK. We found a wide range of UV protection (i.e. of daily UV dose to the face), with around 35% of adult XP patients protecting significantly more poorly than the rest (3). We went on to demonstrate that poor photoprotection correlated closely with 9 variables, 7 of which were potentially reversible psychological factors (4).</p><p>We went on to design a personalised behaviour change intervention (‘XPAND’) to target these psychological factors shown to correlate with poor photoprotection (i.e. high daily UV dose to the face). It involves 7 one-to-one sessions delivered by a nurse or a psychologist.</p><p>In this lecture I present the results of our randomised controlled trial of this intervention in 16 XP patients with poor photoprotection. The design was a two-arm parallel group randomised control trial with a delayed intervention control arm who received XPAND the following year. The primary endpoint was the mean daily UV dose to the face during a 3 week period in June or July following XPAND (or no intervention in the control group).</p><p>Of 16 patients randomised, 13 provided sufficient data for primary outcome analysis. The XPAND group (n = 8) had a lower mean daily UV dose to the face than the control group (p&lt;0.001). The delayed intervention control group also had a lower mean daily dose to the face after the intervention than before, but not at a statistically significant level. Secondary endpoints were also analysed. Health economic analysis was also carried out and predicted that XPAND was a cost-effective treatment, by reducing long term treatment costs associated with UV exposure.</p><p>In this talk I will discuss these data, what can and what cannot be concluded from this small randomised controlled trial, and the implications for clinical management of XP.</p><p>Alain Sarasin¹, Andrey A. Yurchenko² and Sergey Nikolaev²</p><p>¹UMR9019 CNRS, Gustave Roussy and Paris-Saclay University, Villejuif, France</p><p>²INSERM U981, Gustave Roussy and Paris-Saclay University, Villejuif, France</p><p>Xeroderma pigmentosum (XP) is a rare, autosomal, recessive genodermatosis caused by Excision Nucleotide Repair (NER) deficiency. These patients are extremely sensitive to sunlight leading to an increased frequency of skin cancers on exposed body sites. Due to constant photoprotection and better therapeutic education, XP patients live longer than in the past. Recently, it was found that some XP patients are at a very high risk of developing internal tumors, including central nervous systems, hematological malignancies, gynecological and thyroid cancers (Nikolaev <i>et al.</i>, Orphanet J. Rare Dis., 2022, <span>17</span>, 104).</p><p>We analyzed four published, international, clinically well-defined XP cohorts (from the States, France, England and Brazil) and calculated the cancer risk of these patients. The Odds Ratio (OR) are very high for young XPs (more than a thousand times higher than for the general population) for the 0-20-year-old French XPs. Very high OR are also observed for English, French, and American XPs for developing brain tumors, hematological malignancies, and thyroid tumors. Among XP patients, the most susceptible to developing internal tumors are the patients from the XP-C group, particularly those who bear the founder <i>XPC</i> mutation from North Africa (Sarasin <i>et al.</i>, Blood, 2019, <span>133</span>,2718; Sarasin, Cancers, 2023, <span>15</span>, 2706).</p><p>We analyzed, by whole genome sequencing, 22 internal tumors from XP patients, mainly hematological malignancies, gynecological and thyroid tumors. Results demonstrated a dramatic increase in mutation rates in XP-C tumors versus sporadic tumors. A strong mutational asymmetry with respect to transcription and level of gene expression demonstrated the existence of transcription-coupled repair in vivo in humans. Mutations appeared to be caused by unrepaired bulky guanine lesions on the untranscribed strands of DNA (Yurchenko <i>et al.</i>, Nature Comm., 2020, <span>11,</span> 5834; Yurchenko <i>et al.</i>, Commun Med, 2023, <span>3,</span> 109). The mutational profiles of internal XP-C tumors are very specific to NER-deficiency and nearly identical between all XP-C internal tumors regardless of the tumor type, and completely different from the tissue-matched sporadic tumors.</p><p>Physicians following XP patients should be aware of the high risk of internal tumors. Prevention and early detection of leukemia and brain, gynecological, and thyroid tumors are needed. Screening should be done every year starting around the age of 10–12 years (for example, early anemia occurs several years before MDS/ leukemia in some XP-C patients).</p><p>Diana van den Heuvel^1,11, Marta Rodríguez-Martínez^2,11, Paula J. van der Meer^1,11, Nicolas Nieto Moreno³, Jiyoung Park⁴, Hyun-Suk Kim⁴, Janne J.M. van Schie¹, Annelotte P. Wondergem¹, Areetha D'Souza⁴, George Yakoub¹, Anna E. Herlihy², Krushanka Kashyap³, Thierry Boissière^2,3, Jane Walker², Richard Mitter⁶, Katja Apelt¹, Klaas de Lint⁷, Idil Kirdök⁷, Mats Ljungman^8,9, Rob M.F. Wolthuis⁷, Patrick Cramer¹⁰, Orlando D. Schärer^4,5, Goran Kokic^10,12, Jesper Q Svejstrup^2,3,12, Martijn S. Luijsterburg^1,12</p><p>¹Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands</p><p>²Mechanisms of Transcription Laboratory.</p><p>³Department of Cellular and Molecular Medicine, University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen, Denmark.</p><p>⁴Center for Genomic Integrity, Institute for Basic Science, Ulsan, Republic of Korea</p><p>⁵Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan, Republic of Korea</p><p>⁶Bioinformatics and Biostatistics, The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK.</p><p>⁷Department of Clinical Genetics, Section Oncogenetics, Cancer Center Amsterdam, Amsterdam University Medical Center, Amsterdam, the Netherlands</p><p>⁸Department of Radiation Oncology, University of Michigan, Ann Arbor, MI, USA</p><p>⁹Department of Environmental Health Sciences, Rogel Cancer Center and Center for RNA Biomedicine, University of Michigan, Ann Arbor, MI, USA</p><p>¹⁰Max Planck Institute for Multidisciplinary Sciences, Department of Molecular Biology, 37077 Göttingen, Germany.</p><p>¹¹ These authors contributed equally</p><p>¹² Senior authors</p><p>Transcription-coupled DNA repair (TCR) removes bulky DNA lesions impeding RNA polymerase II (RNAPII) transcription. Recent studies have outlined the stepwise assembly of TCR factors CSB, CSA, UVSSA, and TFIIH around lesion-stalled RNAPII. However, the mechanism and factors required for the transition to downstream repair steps, including RNAPII removal to provide repair proteins access to the DNA lesion, remain unclear. Here, we identify STK19 as a TCR factor facilitating this transition. Loss of STK19 does not impact initial TCR complex assembly or RNAPII ubiquitylation but delays lesion-stalled RNAPII clearance, thereby interfering with the downstream repair reaction. Cryo-EM and mutational analysis reveal that STK19 associates with the TCR complex, positioning itself between RNAPII, UVSSA, and CSA. The structural insights and molecular modeling suggest that STK19 positions the ATPase subunits of TFIIH onto DNA in front of RNAPII. Together, these findings provide new insights into the factors and mechanisms required for TCR.</p>","PeriodicalId":14758,"journal":{"name":"Journal Der Deutschen Dermatologischen Gesellschaft","volume":"23 S2","pages":"3-20"},"PeriodicalIF":5.5000,"publicationDate":"2025-03-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/ddg.15735_g","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal Der Deutschen Dermatologischen Gesellschaft","FirstCategoryId":"3","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1111/ddg.15735_g","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"DERMATOLOGY","Score":null,"Total":0}