Studies of rare genetic bone disorders are often limited due to unavailability of tissue specimens and the lack of animal models fully replicating phenotypic features. Craniometaphyseal dysplasia (CMD) is a rare monogenic disorder characterized by hyperostosis of craniofacial bones concurrent with abnormal shape of long bones. Mutations for autosomal dominant CMD have been identified in the ANK gene (ANKH). Here we describe a simple and efficient method to reprogram adherent cells cultured from peripheral blood to human induced pluripotent stem cells (hiPSCs) from eight CMD patients and five healthy controls. Peripheral blood mononuclear cells (PBMCs) were separated from 5–7 mL of whole blood by Ficoll gradient, expanded in the presence of cytokines and transduced with Sendai virus (SeV) vectors encoding OCT3/4, SOX2, KLF4, and c-MYC. SeV vector, a cytoplasmic RNA vector, is lost from host cells after propagation for 10–13 passages. These hiPSCs express stem cell markers, have normal karyotypes, and are capable of forming embryoid bodies in vitro as well as teratomas in vivo. Further differentiation of these patient-specific iPSCs into osteoblasts and osteoclasts can provide a useful tool to study the effects CMD mutations on bone, and this approach can be applied for disease modeling of other rare genetic musculoskeletal disorders.

We identified osteoclast defects in craniometaphyseal dysplasia (CMD) using an easy-to-use protocol for differentiating osteoclasts from human induced pluripotent stem cells (hiPSCs). CMD is a rare genetic bone disorder, characterized by life-long progressive thickening of craniofacial bones and abnormal shape of long bones. hiPSCs from CMD patients with an in-frame deletion of Phe377 or Ser375 in ANKH are more refractory to in vitro osteoclast differentiation than control hiPSCs. To exclude differentiation effects due to genetic variability, we generated isogenic hiPSCs, which have identical genetic background except for the ANKH mutation. Isogenic hiPSCs with ANKH mutations formed fewer osteoclasts, resorbed less bone, expressed lower levels of osteoclast marker genes, and showed decreased protein levels of ANKH and vacuolar proton pump v-ATP6v0d2. This proof-of-concept study demonstrates that efficient and reproducible differentiation of isogenic hiPSCs into osteoclasts is possible and a promising tool for investigating mechanisms of CMD or other osteoclast-related disorders.