CAMSAP1 Research Update - Landon's League Foundation

As previously reported, we have discovered that a portion of CAMSAP1 must exist for any organism to survive embryogenesis. This proposition is simply due to the results of the initial attempt at creating a mouse model that proved to be embryonic lethal (aka conditional lethal, see Figure 1) as the CRISPR/cas9 cut was made prior to the CH location along the gene (Figure 1, brown stars). However, further analysis showed that there were viable, heterozygous mice that carried the mutation but were unaffected – this proved that only one functioning copy of CAMSAP1 is necessary for normal development.

Figure 1 – Brown stars: where CRISPR/cas9 cuts were made during first attempt to create CAMSAP1 knockout (KO) mouse model. Blue star: Landon’s paternally inherited mutation location. Pink star: Landon’s maternally inherited mutation location.

This initial attempt also left other questions to be answered:

Does the paternal or maternal mutation create partial protein function throughout Landon’s cells?
If partial protein function does exist,
1. Is it from the paternal or maternal genetic mutation?
2. Can a mouse model be created from either the paternal or maternal mutation, depending on which mutation is responsible for the partial protein function?
If a successful mouse model can be created, can the mutated mouse safely be rescued?
What types of cells express the highest amount of CAMSAP1 normally, and are Landon’s mutated cells devoid of the appropriate expression levels?
Can Landon’s cells be rescued via gene therapy?
Can we analyze Landon’s cells safely to answer these questions?

How Do We Find Out The Answers To These Questions?

Since CAMSAP1 is highly expressed in the nervous system, a brain biopsy would give us some answers but would be far too risky! However, we can create “Induced Pluripotent Stem Cells” (iPSC or iPS). What are iPSCs? Simply put, they are cells that are created from an adult cell (ie. blood cells, skin cells, etc.) that are genetically modified by adding four genes to them (OCT-4, SOX-2, KLF-4, and MYC, also known as “Yamanaka Factors”); however, they are not embryonic stem cells. These genes are specific genetic instructions used to transform or induce adult cells into pluripotent cells (see Video 1). By adding these genes, the adult cells will reverse from their differentiated (specialized) state back to an undifferentiated state. Like embryonic stem cells though, these cells can also be modified to redifferentiate into any cell type in the human body, including cells from the nervous system, but eliminate the ethical dilemma surrounding embryonic stem cell usage.

Video 1

WE ARE EXCITED TO REPORT THAT WE WILL BE WORKING WITH A TEAM OF RESEARCHERS AND PHYSICIANS TO ACHIEVE THIS FEAT!

The plan is to create iPS cells from a simple blood sample from Landon and test the levels of CAMSAP1 in the iPSC samples, as well as an unrelated control sample, to find out whether CAMSAP1 mutations in Landon’s cells lead to partial protein function or gene decay.

Here is an example (Video 2) of the process rescuing of a condition known as Sickle Cell Anemia via gene therapy by way of transforming differentiated skin cells into undifferentiated iPS cells then redifferentiating them into normal red blood cells after the gene was corrected:

Video 2

If partial protein is recognized, then the hope is to proceed with the creation of the mouse model of the same specific mutation that is responsible for the partial protein function recognized in Landon’s cells. The hope is to create a gene therapy to “rescue/cure” the CAMSAP1 mutation by reintroducing the gene into the various cell types to “rescue/cure” them while in the lab.

Gene Therapy

How are these cells fixed? A process in a highly dynamic, new, and constantly evolving field of medicine known as gene therapy, can fix, alter, and/or reintroduce a normal functioning copy of a gene into a cell. This process is completed by altering a virus, eliminating its harmful effects, and engineering it to deliver a message to the cell or deliver the corrected copy of the gene to the cell. Afterwards, the cell will respond appropriately and if everything works as designed, the condition is rescued.

Complex Version of the Potential CAMSAP1 Vector

Other Potential For iPSC Creation

While we are simply scratching the proverbial surface of what this could potentially mean for individuals with CAMSAP1 mutations with respect to the creation/development of a curative gene therapy, these iPS cells could also be used to test different drugs to see if there is an existing medication that could mimic the role of CAMSAP1 throughout the body. Furthermore, we can also see if these drugs have any harmful effects on other tissue types (Figure 2).