Tag: genetic diseases

Quantum Leap for Genetic Disease Therapy with Baculovirus DNA Repair Kit

DNA repair
Source: Pixabay/CC0

Genetic mutations behind a genetic kidney disease affecting children and young adults have been fixed in patient-derived kidney cells with a high-capacity DNA ‘repair kit’. The advance, developed by University of Bristol scientists, is published in Nucleic Acids Research.

In this new study, the international team describe how they created a DNA repair vehicle to genetically fix faulty podocin, a common genetic cause of inheritable Steroid Resistant Nephrotic Syndrome (SRNS).

Podocin is a protein normally located on the surface of specialised kidney cells and is essential for kidney function. Faulty podocin, however, remains stuck inside the cell and never makes it to the surface, terminally damaging the podocytes. Since the disease cannot be cured with medications, gene therapy which repairs the genetic mutations causing the faulty podocin offers hope for patients.

Typically, human viruses have been utilised in gene therapy applications to carry out genetic repairs. These are used as a ‘Trojan Horse’ to enter cells carrying the errors. Currently dominating systems include lentivirus (LV), adenovirus (AV) and adeno-associated virus (AAV), which are all relatively harmless viruses that readily infect humans. Their viral shells however restrict the amount of cargo they can carry and deliver, namely the DNA kit necessary for efficient genetic repair. This limits the scope of their application in gene therapy.

By applying synthetic biology techniques, the team led by Dr Francesco Aulicino and Professor Imre Berger, re-engineered baculovirus, a insect virus which has a nearly unlimited cargo capacity.

“What sets apart baculovirus from LV, AV, and AAV is the lack of a rigid shell encapsulating the cargo space.” said Dr Francesco Aulicino, who led the study. The shell of baculovirus resembles a hollow stick, simply lengthening when the cargo increases. This allows a much more sophisticated tool-kit can be delivered by the baculovirus.

First, baculovirus had to be equipped to penetrate human cells which it normally would not do. “We decorated the baculovirus with proteins that enabled it to enter human cells very efficiently.” explained Dr Aulicino. The scientists then used their engineered baculovirus to deliver much larger DNA pieces than was previously possible, and build these into the genomes of a whole range of human cells.

The DNA in the human genome comprises 3 billion base-pairs making up ~25,000 genes, which encode for the proteins that are essential for cellular functions. If faulty base-pairs occur in our genes, faulty proteins are made which can make us ill, resulting in hereditary disease. Gene therapy promises repair of hereditary disease at its very root, by rectifying such errors in our genomes. Gene editing approaches, in particular CRISPR/Cas-based methods, have greatly advanced the field by enabling genetic repair with base-pair precision.

The team used patient-derived podocytes carrying the disease-causing error in the genome to demonstrate the aptitude of their technology. By creating a DNA repair kit, comprising protein-based scissors and the nucleic acid molecules that guide them – and the DNA sequences to replace the faulty gene, the team delivered with a single engineered baculovirus a healthy copy of the podocin gene concomitant with the CRISPR/Cas machinery to insert it with base-pair precision into the genome. This was able to reverse the disease-causing phenotype and restore podocin to the cell surface.

Professor Imre Berger explained: “We had previously used baculovirus to infect cultured insect cells to produce recombinant proteins for studying their structure and function.” This method, called MultiBac, has been highly successful to make very large multiprotein complexes with many subunits, in laboratories world-wide. “MultiBac already exploited the flexibility of the baculovirus shell to deliver large pieces of DNA into the cultured insect cells, instructing them to assemble the proteins we were interested in.” When the scientists realised that the same property could potentially transform gene therapy in human cells, they created this new DNA repair kit.

Dr Aulicino added: “There are many avenues to utilise our system. In addition to podocin repair, we could show that we can simultaneously correct many errors in very different places in the genome efficiently, by using our single baculovirus delivery system and the most recent editing techniques available.”

Source: University of Bristol

Sex-differentiation Genes Also Contribute to Disease Risks

Man and woman about to sprint
Source: Andrea Piacquadio on Pexels

Some physical traits that differ between sexes are known to be linked to certain single nucleotide polymorphisms (SNPs) outside the X and Y chromosomes. New research now suggests that many of these ‘sex-heterogenous’ SNPs also contribute to a person’s risk for a variety of diseases. Michela Traglia and colleagues at the University of California San Francisco presented their findings in PLOS Genetics.

Millions of SNPs are in each genome, with each SNP representing a difference in a certain DNA building block in a particular stretch of DNA. Many associations have been uncovered between certain SNPs and people’s distinct traits. Understanding SNPs has a number of applications, such as predicting individual treatment effectiveness or disease risks.

Traglia and colleagues previously found that SNPs associated with certain differences in physical traits between men and women, such as waist-hip ratio and basal metabolic rate, may also affect the biology of autism spectrum disorder and other complex diseases. Building on this work with two large genomic datasets, the identified an updated list of 2320 sex-heterogeneous SNPs.

Analysis of these SNPs revealed that they are also associated with a variety of health-related traits and diseases, some with strong sex bias and some without, including schizophrenia, type 2 diabetes, anorexia, heart failure, and ADHD.

These SNPs are located in stretches of DNA that are either within or near genes involved in skeletal and muscle development in a growing embryo. In addition, these SNPs appear to play a role in regulating gene expression and DNA methylation, which are fundamental processes by which a person’s DNA is translated into their distinct biology and traits.

Overall, the researchers conclude that the identified SNPs play a role in early-life biological processes shaping sex-distinct traits and which also affect health and disease risk later in life. More work is needed to understand the mechanisms behind these sex-heterogeneous SNPs.

“We found that genetic alleles with differing effects on measured physical traits in men and women also play an outsized role in health risks,” remarked study co-author Lauren Weiss. “We hope this work helps us to understand the genetic underpinnings of sexual dimorphism and its relationship with both early development and later disease risk.”

Source: EurekAlert!

Whole Genome Sequencing Yields Diagnoses for Rare Diseases

With the integration of whole genome sequencing in Swedish healthcare, some 1200 individuals with rare diseases have received a diagnosis, with novel disease genes discovered in the process.

“We’ve established a way of working where hospital and university collaborate on sequencing each patients’ entire genome in order to find genetic explanations for different diseases,” said first author Henrik Stranneheim, researcher at the Department of Molecular Medicine and Surgery, Karolinska Institutet. “This is an example of how precision medicine can be used to make diagnoses and tailor treatments to individual patients.”

The technology of large-scale whole genome sequencing to yield a person’s entire DNA, is not yet widely used in hospitals despite the technology becoming much more accessible over the last ten years. Whole genome sequencing has uncovered a great genetic variety among different populations, such as in South Africa, where a pilot study uncovered a high rate of novel variants in African populations.

Karolinska University Laboratory and the Clinical Genomics facility at SciLifeLab launched the Genomic Medicine Centre Karolinska-Rare Diseases (GMCK-RD) five years ago. Since then, the centre has sequenced the genomes of 3219 patients, which led to molecular diagnoses for 40% of them with rare diseases.

In addition to these, the researchers found pathogenic mutations in more than 750 genes and discovered 17 novel disease genes. 

“Clinical whole genome sequencing has had huge implications for the area of rare diseases,” explained corresponding author Anna Wedell, professor at the Department of Molecular Medicine and Surgery, Karolinska Institutet. “Used in the right way, targeted at each patient’s specific clinical situation, new groups of patients can receive the right diagnosis and treatment in a way that hasn’t been possible before.”

Whole genome sequencing is challenging in part due to having to manage and interpret the millions of variations that exist within each invidual’s genome. In order to overcome this difficulty, the centre came up with a model that directs the initial analysis to pathogenic variants in genes relevant for that patient’s clinical symptomsIn this way, doctors play an important role in choosing which genetic analyses to run first.
Should the first assessment fail, the analyses are then broadened to more gene panels, which has uncovered new disease genes.

“For us to succeed with precision medicine, a multidisciplinary collaboration between health care and academia is essential,” said second corresponding author Anna Lindstrand, professor at the Department of Molecular Medicine and Surgery, Karolinska Institutet and consultant at Karolinska University Hospital’s Department of Clinical Genetics. “Through these initiatives we combine clinical expertise with bioinformatic tools and together deliver accurate diagnoses and individualised treatments.”

Source: Medical Xpress

Journal information: “Integration of whole genome sequencing into a health care setting: High diagnostic rates across multiple clinical entities in 3219 rare disease patients,” Genome Medicine (2021). DOI: 10.1186/s13073-021-00855-5