The Van der Woude syndrome (VWS, MIM 119300), is an autosomal dominant disorder characterized by cleft lip with or without cleft palate, or isolated cleft palate, due to loss-of-function mutations in IRF6 (Interferon Regulatory Factor-6) or GRHL3 (Grainyhead-Like Transcription Factor 3). Most patients show pits on their lower lips. In VWS, most IRF6 alterations are premature stop codons or missense mutations. Pathogenic upstream open reading frame (uORF) mutations are characterized by an out-of-frame upstream start codon (uAUG) located in the 5’UTR and leading to a premature stop codon. uORFs overlap the usual ATG start codon and have a minimum length of 9 nucleotides. We assessed the main features of six uORF-creating mutations identified in VWS patients (two in the lab and four previously described). We also determined all the theoretical SNVs located in IRF6 5’UTR (NM_006147.4) that could create an uORFs and we assessed their potential pathogenicity based on Kozak site in silico prediction. Only four uORF-creating mutations in IRF6 have been previously associated with VWS to date. We report here two novel mutations creating out-of-frame uAUGs (c.-141C>T p.? and c.-162C>T p.?) that probably reduce IRF6 expression. In our lab, IRF6 mutations are found in about 80% of families with VWS and uORFs-creating mutations represent of 3.2% of them (2/63). Previous studies identifying uORFs-creating mutations did not provide detailed phenotypic data. In our group, in the 6 heterozygotes for c.-141C>T, three had a cleft lip with or without cleft palate and three had only a bifid uvula. The patient heterozygote for c.-162C>T had a posterior cleft with an ankyloblepharon. Most genes naturally have uORFs in their 5’UTR region, nonetheless we observed that there were no physiological uORF in IRF6. All 6 uORFs identified in VWS had the same termination codon that occurs in exon 3 (56 nucleotides after the usual ATG). To go further, we searched for all potential uAUG-creating SNV in IRF6 5’UTR. We identified 41 of them, including the 6 identified in patients. Except one, none of them is present in the gnomAD control population. Using a machine-learning-based tool (TIS Predictor, biorxiv.org/content/10.1101/2021.08.17.456657v1, 0 to 1 score range), we then assessed each Kozak similarity score. All 6 uORFs identified in patients had high scores comprised between 0.71 and 0.83, compared to 0.81 for the usual ATG. Among the 35 theoretical SNVs creating an uAUG, 10 may create a potential but weak Kozak site (score range: 0.50-0.71). In conclusion, uORF-creating mutations can thus be responsible for typical VWS. As untranslated regions are not included in most captured regions in high throughput sequencing strategies, this category of variants may be underdiagnosed in VWS and in human pathology in general.