Why did the Dinosaur Trust choose to fund this study?

Following the success of the BMPR2 research projects that The Dinosaur Trust funded, we became aware of CRISPR/Cas9. We could see that this could be significant in finding a cure for pulmonary hypertension and it was decided that exploring CRISPR/Cas9 would be the next research project The Dinosaur Trust would fund.

We reached out to The PVRI to ask if they could assist with finding a laboratory or team that were working with CRISPR/Cas9 who might be able to conduct research into the possibility of CRISPR/Cas9 providing a cure for Pulmonary hypertension.

Through The PVRI and under the guidance of their Medical Advisory Committee, The Dinosaur Trust awarded a 3 year Grant to Soni Savai-Pullamsetti and Werner Seeger, Justus-Liebig of University, Giessen, Germany.

What is Crispr/cas9?

CRISPR is short for:
Clustered
Regular
Interspaced
Short
Palindromic
Repeats

Crisper consists of two components; the Cas9 protein that can cut DNA and a guide RNA that can recognise the sequence of DNA to be edited. To use CRISPR/Cas9 scientist first identify the sequence of the human genome which is causing a health problem. 

They then create a specific guide RNA to recognise that particular stretch of A’s, T’s, G’s & C’s in the DNA. The guide RNA is attached to the DNA cutting enzyme, Cas9, and then this complex is introduced to the target cells. 

It locates the target letter sequence and cuts the DNA. At this point scientists can then edit the existing genome by either modifying, deleting or inserting new sequences. 

It effectively makes CRISPR/Cas9 a cut and paste tool for DNA editing. 

It is hoped that in the future scientist’s could use CRISPR/Cas9 to develop critical advances in patient care or even cure lifelong inherited diseases. 

The Research Project

Abstract
80% of the cases of familial pulmonary arterial hypertension (PAH) are connected to heterozygous mutations of the BMPR2 gene but most mutation carriers will never develop PAH in their lives. Although PAH pathogenesis has been addressed in many studies the reason for the low penetrance of the mutation is unknown and molecular mechanisms underlying the devastating disease hallmarked by severe remodelling of the pulmonary arterial vasculature are yet to be deciphered.

The aim of our project is to decipher the functional consequences of the BMPR2 mutations found in patients on the regulation of the whole set of genes that concur to the physiological control of the vascular cells, in particular those that are suspected to participate in the pathological process. It is hypothesized that environmental impacts that lead to epigenetic rearrangements in pulmonary arterial cell types, so called second hits, alter the regulation of whole gene sets, which then, in combination with a defect in the BMPR2 gene leads to vascular remodelling and PH.

Thus, to investigate molecular mechanisms underlying PAH pathogenesis we would like to work on four levels and employ different in vitro and in vivo models to dissect the molecular basis of PAH and design a patient-specific approach for gene therapy.

  1. In vitro differentiation of patient PBMC derived iPSCs to endothelial-like cells (iPSC-ECs) to study functional consequences of the BMPR2 mutation in vitro in comparison to non-affected (heathy) BMPR2 mutation carriers and healthy family members without BMPR2 mutation.
  2. Correction of the BMPR2 mutation in patient derived iPSC-ECs with the CRISPR/Cas9 system for in vitroassessment of the molecular consequences of a “therapeutic” gene editing approach.
  3. Employing the personalized iPSC-EC in vitro system for drug screening and validation of the molecular phenotype by RNA sequencing.
  4. Development of CRISPR/Cas9-based gene therapy approaches. Different strategies such as engagement of in situ CRISPR effects by aerosol or intravascular application of adenoviral vectors or transplantation of vascular cell types with a “corrected” version of BMPR2 will be established using a BMPR2 deficient mouse model.

The Dinosaur Trust has awarded 3 years of funding to the University of Giessen, Germany  for its research: The Correction of a BMPR2 Mutation using CRISPR/Cas9.