Cell-based and gene-based therapies are increasingly being tested and used to treat diseases that have been considered virtually untreatable or uncurable. Now, ongoing research at Cincinnati Children’s is investigating novel approaches to treat bronchopulmonary dysplasia (BPD)—and even rare and lethal alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV).

Vladimir Kalinichenko, MD, PhD, and his team at the Cincinnati Children’s Perinatal Institute’s Center for Lung Regenerative Medicine have published extensively about their research to increase the density of alveolar capillaries in both BPD and ACDMPV. Much of that research has focused on forkhead box (FOX) proteins and their role in regulating cell signaling pathways required for cellular proliferation, differentiation, motility and survival. Their research has uncovered much about how mutations in the FOXF1 gene can disrupt the alveolar microvasculature, affecting lung development or causing neonatal lung disease. Their long-term goal: new therapies to fight lung diseases, including BPD and possibly ACDMPV.

BPD: Fighting a Disease of Prematurity

BPD is a common complication in premature infants with underdeveloped lungs and can lead to long-term breathing problems and poor growth and development. Premature infants often depend on mechanical breathing assistance, further hindering lung development in these babies.

Kalinichenko’s laboratory uses neonatal mice that have been exposed to hyperoxia (over-oxygenation) to model human BPD. One way the team is attacking BPD is through pulmonary c-KIT endothelial progenitor cells, which are common in embryonic and neonatal lungs and help form capillaries and air sacs in the lungs called alveoli. Through extensive single-cell RNA sequencing of mouse and donated human neonatal lung tissue, the team showed that pulmonary c-KIT endothelial progenitor cells require the c-KIT and FOXF1 proteins to stimulate the development of blood vessels and alveoli. High oxygen concentrations damage the cells, impeding lung development in infants on mechanical oxygen assistance. Vascular endothelial growth factor (VEGF) has been shown to improve lung structure and function in rodent BPD models, but severe side effects prevent its use in patients with BPD. However, Kalinichenko’s team found that nanoparticle delivery of FOXM1 or FOXF1—both downstream targets of VEGF—improved elastin fiber organization, decreased alveolar simplification, and preserved long-term lung function in the murine BPD model.

Encouragingly, the team suggests that using c-KIT-positive endothelial cells from donors—or generating them with pluripotent stem cells, which can differentiate into virtually any cell in the body—might be a way to treat BPD and perhaps other pediatric lung disorders associated with loss of alveoli and pulmonary blood vessels. They found that infusion of pulmonary c-KIT endothelial progenitor cells in peripheral blood increased the formation of pulmonary blood vessels and air sacs in murine BPD models.

ACDMPV: Hope on the Horizon?

Infants with ACDMPV cannot grow enough alveolar capillaries to support healthy gas exchange after birth. Without a lung transplant, this condition is usually fatal during the first month of life.

Through extensive analysis of thousands of lung cells from mouse ACDMPV models, the team determined that ACDMPV is linked with changes in a critical signaling pathway involving the proteins BMP9, ACVRL1 and SMAD1. This pathway is critical for healthy blood vessel formation in the lungs. When the FOXF1 protein goes missing or contains certain mutations, the expression of ACVRL1 is reduced, which reduces expression of downstream target genes critical for lung vascular development.

Kalinichenko’s team found that adding synthetic bone morphogenetic protein BMP9 to mice that lacked a copy of the functional FOXF1 gene helped re-create the signaling pathway, telling the lungs to keep making capillaries. The researchers confirmed this through tests in lab cell cultures and in mice.

The results are dramatic, but so far, no drug that simulates BMP9 activity has been approved for human use. If and when a safe synthetic BMP9 molecule is developed and approved, it could potentially change the prognosis for infants with ACDMPV. It could also benefit infants with BPD or congenital diaphragmatic hernia (CDH), a birth defect that strikes about 1,000 newborns a year in the US. 

The Future

Though it may be years before the lab’s work is translated into better treatment options, the difference it could make for infants and families keeps the team committed and focused. Team members are already in communication with the Food and Drug Administration regarding compassionate use of the FOXF1 gene therapy for ACDMPV, as no approved treatment options exist if no transplant is available.

“There are many steps to the process, but within two or three years, some of our research could lead to human trials,” Kalinichenko says. “All our work is for one purpose—to find new and better ways to care for patients with these severe pediatric respiratory diseases and to improve patient lives.”