<p><b>Background</b>: Congenital cataract displays large phenotypic (syndromic and isolated cataracts) and genetic heterogeneity. Mutations in several transcription factors involved in eye development, like <i>PITX3</i>, have been associated with congenital cataracts and anterior segment mesenchymal disorders.</p> <p><b>Materials and methods</b>: Targeted sequencing of 187 genes involved in ocular development was performed in 96 patients with mainly anophthalmia and microphthalmia. Additionally, Sanger sequencing analysis of <i>PITX3</i> was performed on a second cohort of 32 index cases with congenital cataract and Peters anomaly and/or sclereocornea.</p> <p><b>Results</b>: We described five families with four different <i>PITX3</i> mutations, two of which were novel. In Family 1, the heterozygous recurrent c.640_656dup (p.Gly220Profs*95) mutation cosegregated with eye anomalies ranging from congenital cataract to Peters anomaly. In Family 2, the novel c.669del [p.(Leu225Trpfs*84)] mutation cosegregated with dominantly inherited eye anomalies ranging from posterior embryotoxon to congenital cataract in heterozygous carriers and congenital sclereocornea and cataract in a patient homozygous for this mutation. In Family 3, we identified the recurrent heterozygous c.640_656dup (p.Gly220Profs*95) mutation segregating with congenital cataract. In Family 4, the <i>de novo</i> c.582del [p.(Ile194Metfs*115)] mutation was identified in a patient with congenital cataract, microphthalmia, developmental delay and autism. In Family 5, the c.38G>A (p.Ser13Asn) mutation segregated dominantly in a family with Peters anomaly, which is a novel phenotype associated with the c.38G>A variant compared with the previously reported isolated congenital cataract.</p> <p><b>Conclusions</b>: Our study unveils different phenotypes associated with known and novel mutations in <i>PITX3</i>, which will improve the genetic counselling of patients and their families.</p
