Decentralized Medicine Manufacturing on Demand
Next: Decentralised Medicine Research
Decentralized Medicine Manufacturing on Demand
Another trend in decentralizing health on a somewhat longer time frame will be ability to manufacture pharmaceuticals locally.
There are two ways to achieve this:
Through so called flow chemistry that is discussed later in this chapter and more in in upcoming post on concept I call the The Unfactory
Through so called biosimilars
Creation of biosimilars and their local production requires a rather longer explanation with several cutting-edge technologies
PCR for fast duplication of genetic material
DNA sequencing for reading the DNA material
Understanding the function of specific genes and creation of libraries with this information
CRISPR for editing genes and adding new functionalities to cells (like creation of biosimilar medicines by modified microbes)
Locally synthetizing DNA. This is based on transmitting the right genetic information digitally anywhere so that the right medicine can be produced locally when needed.
If this gets too detailed, feel free to zoom past this section.
Polymerase Chain Reaction (PCR)
Polymerase chain reaction (PCR) is a method for fast making billions of copies of a DNA sample. This allows to take a very small sample of DNA and amplify it so it can be studied in detail
PCR works so that first the sample is heated so that DNA separates (denatures) into two pieces of single stranded DNA. Next with the help of Taq polymerase two new full strands of DNA are made. These two strands both contain one old strand from the original DNA and one new strand of DNA. This way the DNA Is copied into two identical pieces. Then this process is repeated to make new copies again. When the cycle of denaturation and synthetizing is repeated enough times, billions of copies can be made. Today the entire process is automated and takes just a few hours.
The large amount of generated DNA allows to do DNA sequencing and analyse the sequence of nucleotide acids of the DNA – essentially what DNA it is. This can be used to fast make diagnostics of infectious diseases caused for example by bacteria and viruses (by detecting their DNA in the sample). It can also be used to detect otherwise hard to analyse slow-growing or hard to grow micro-organisms like anaerobic bacteria or mycobacteria.
PCR test were used during Covid pandemic to detect viral RNA as one of the two main ways to detect it There is a variant of PCR (RT-PCR) where the RNA of a virus is first made into a two stranded DNA and then used the standard PCR process to multiply it for diagnostics.
Antigen tests were the other main way to detect from nasal swabs. They detect certain proteins of the virus, are faster but less accurate. Antigen in general is any molecule - like a protein on the surface of the virus - that causes an immune response in the body.
DNA Sequencing
After copying DNA with PCR, they need to be read. This is done with DNA sequencing. It is used to find out the structure of DNA, i.e., what is the exact sequence of bases (DNA is made of four different types of bases called either A, T, C or G).
Today DNA sequencing is made in automated DNA sequencers that work by electrophoresis. In it, DNA is placed in one end of a gel and electrodes placed at both ends of the slab of gel. When current is applied the DNA, molecules start moving through the gel at different speed based on their size. Bigger molecules move slower and small faster. The DNA needs also to be chopped first to pieces as electrophoresis has limitation of being able to separate about 500 bases.
The sequencers still need to figure out each base during the process. Before sequencing the DNA is copied many times with PCR as described above. At the end of this copying the generated copies are divided into four batches (corresponding to the bases A, T, C and G) and modified base (modified A, T, C or G) is added to each batch that causes the sequences to break during copying at corresponding base. I.e., the base targeted is at the end of the sequence. In one batch all copies end in T in another in A etc.
At the same time different fluorescent dyes of different colors are added. This makes detection easy. Assume we color bases so that A is dyed green, T yellow, C blue and G red. Then a DNA sequence like A T C G G A is broken into segments of:
A green and ATCGGA (green)
AT (yellow)
ATC (blue)
ATCG (red) and ATCGG (red)
When these molecules are put through the gel and a current applied, they come out with smallest first and a detector reads the colour of the fluorescence. As they are ordered by length the detector can directly read the DNA sequence. In our example green (A), yellow (T), blue (C), G (red) etc…
Function of genes
How do we then know what a specific gene sequence does? The simplest is to see what happens if a gene is missing. This is done by the studying organisms that have some gene mutation that results in a physical difference like lack of pigment, curly wings in a fruit fly or lack of a protein causing disease in humans. By comparing normal organisms’ sequence to the organism with the mutation. (BTW: what the gene does in a cell is called ‘gene expression’ in molecular biology).
The results found are stored in large public databases. These DNA libraries can then be used to search what a specific gene sequence does when encountered in some other species.
Gene expression is most easily done with organisms that reproduce rapidly such as bacteria, yeasts etc. This process can be speeded up by artificially increasing mutations with certain chemicals or with sloppy-PCR. Sloppy-PCR is a method where the PCR process works badly causing up to 2% errors but this fault is used to advantage as the mutations are used to study how genes work. After copying the mutant coding sequences must still be cloned into the target cell.
https://www.ncbi.nlm.nih.gov/books/NBK26818/
CRISPR
CRISPR is a gene editing technology based on bacteria’s natural defence mechanism against viruses. When bacterium detects that there is virus DNA present, it starts producing two types of short RNA strands. One of them matches that of the invading virus. These short RNA sequences combine and form a protein called Cas9 (shorthand for CRISPR-associated). Cas9 is an enzyme with the property that it can cut DNA. In bacteria the Cas9 cuts the virus DNA disabling it.
CRISPR technique is based on notion that it can be engineered to cut any DNA at precisely chosen location by changing the guiding RNA to match the target. And this works also in living cells.
When Cas9 cuts the DNA, the cell tries to repair the cut. The repair process is error prone with resulting in mutations that disable the cell. In research this can be used to understand what the function of the gene is.
With CRISPR it is also possible to replace a gene sequence with a new version. This editing can be done also in cultured cells like stem cells that can turn into many different cell types. It can also be done in a fertilized egg. This allows the creation of transgenic animals with desired properties.
CRISP can also target multiple genes at the same time. This is a big bonus as bodily responses are normally controlled by many genes acting together. CRIPRS potential uses are in correcting genetic defects, treating or preventing diseases and improving crops.
CRIPRS can also be used to alter bacteria so that they start producing beneficial molecules for us. For example, bacteria in controlled environment (bio reactor) can produce hormones and other medicines. Or they can be made to produce for example egg white to be used in bakery.
Biosimilar Drugs
Biologic drugs are made with living cells by injecting genes that produce the needed protein. The addition of new genetic instructions is done with the CRISPR technique described above. The cells start making the protein as instructed and finally the molecules are isolated and they become the active ingredient in the biological drug.
The injection of the new generic material can be done either to the DNA in the nucleus of the cell (chromosomal DNA) or to a plasmid in a microbe. Plasmids are small circular DNA molecules freely in the plasm of the microbe (usually a bacteria). Plasmids can replicate independently from cell replication.
This is how a new type of medicines called biosimilars are manufactured. During the manufacturing process there are small differences that result in the final molecule not being identical but closely resembling. These alterations are made to lower the cost making biosimilars an affordable substitution for pricy, large-molecule biologics.
This method can be used to produce either exactly same hormone/enzyme that human body produces or similar that has same effect. The reason for having slight differences are because sometimes small changes and simplifications make either production or extraction more efficient and simpler.
Compared to more common synthetic drugs today, the difference is that synthetics are small molecules while the biologic drugs are large molecules consisting of thousands of molecules.
https://www.arthritis-health.com/treatment/medications/science-behind-biologics
http://www.genomenewsnetwork.org/resources/whats_a_genome/Chp2_2.shtml
Producing Vaccines and Medicine Locally
At the leading edge today is 3D printing medicine. Basic concept is to receive a DNA sequence over the Internet as a file and with that as a “code” be able to produce medicine like insulin, create vaccines or other medicines.
Using the received sequence, a new gene containing the received DNA sequence is synthesised from chemicals in the laboratory and then inserted into a cell using CRISPR. These gene sequences can be natural ones or ones synthesised in software.
The created gene can be inserted either into the nucleus of a microbe or to a plasmid. A genetically modified plasmid is then introduced into a host organism. For example a strain of e.coli (bacteria) or saccharomyces cerevisiae (yeast). These are relatively easy to grow.
The modified host organisms now carries the new gene and begins to produce the desired molecule (begins to express the gene is the common terminology). The created molecule is harvested from the culture and purified. Finally it needs to be packaged (formulated) into the right dosage forms (vials, pens, cartridges etc.)
The idea is for every major hospital, clinic, and corporation in the world to own such a digital-to-biological converter or printer. If a viral outbreak hits, the vaccine could be sent around the world in a digital file in minutes and produced locally, instead of being stockpiled and shipped out.
Today this is upcoming technology and will be heavily patented. Over time patents expire and technology becomes commoditised. Then it becomes possible to decentralise medicine manufacturing to local level. Such facilities could be at the local library, town hall or health center. If the technology can be miniaturised, perhaps personally or at each house.
https://www.theguardian.com/science/2013/oct/13/craig-ventner-mars
https://www.vice.com/en_us/article/59zj9b/craig-venters-digital-to-biological-converter-is-real
Producing other molecules
In principle the above mentioned method can be used to produce other useful molecules as well, not just complex ones needed for health.
Any chemical feedstock or more complex chemical molecule might be possible. This includes simple results like fuels of various sorts or more complex results.