Implementation
The ultimate goal for the research done for CAR_Ma has always been its eventual use in clinical applications. Modern immunotherapies have had brilliant success, but have striking weaknesses targeting solid tumors, lacking accessibility and affordability, and resulting in some long-term risks (Food and Drug Administration, 2024). With the development of CAR_Ma, we hope to bridge the gap between effectiveness and affordability bringing the most advanced immunotherapies to use globally.
As designed, CAR_Ma is a complete system integrating the vector, functional units and delivery system ready for use as a therapeutic. CAR_Ma was envisioned as an injectable in vivo therapeutic capable of treating cancer without the need and additional cost in equipment and expertise required from existing ex vivo immunotherapies like the CAR-T systems currently on the market or the extreme cost of CRISPR based gene therapies. By using a single multi-modal RNA capable of both sustained expression of CAR for up to 60 days while at the same time conducting gene knockout or gene silencing packaged in a lipid nanoparticle, we are able to create an simple and injectable immunotherapy with the production and logistics of an mRNA vaccine like those developed by Moderna and Pfizer. However, even in comparison to the state-of-the-art mRNA vaccines developed by these companies, CAR_Ma is built on newer nanoparticle technology that can allow storage and transport of our therapeutic at 4°C or even room temperature if lyophilized which is a dramatic improvement compared to Pfizer which requires -80°C storage and Moderna which requires -20°C storage. All in all, CAR_Ma will be easier to produce, dose, transport and apply, with the requirements for sub-zero storage and transport alleviated CAR_Ma will have a much greater global reach.
Another prime key factor affecting the implementation, adoption and accessibility is cost. Existing therapies for cancer are extremely expensive: A study by Nadeem et al. (2016) on a group of chemotherapies revealed 6-month mean cost of chemotherapy was US$26,989, and the median cost was US$9,611. In Hong Kong, Cigna Healthcare estimates the cost of radiotherapy treatment to range from US$3,800 (HK$30,000) USD to US$25,600 (HK$200,000) (Cigna Worldwide Life Insurance Company Limited, n.d.) depending on the type of cancer. The problem is even further exacerbated with more powerful immunotherapies. A review by Cliff et al. (2023) placed the manufacturing cost of a CAR-T therapeutic around US$20,000 and for another CAR-T therapeutic that relied on an automated production system had a cost of approximately US$35,107 per patient. These prices create a therapeutic market that is inaccessible and essentially ineffective to a large proportion of the global population. CAR_Ma on the other hand will have a projected cost to be similar if not slightly greater than that of the mRNA COVID-19 vaccines. Light and Lexchin (2021) found that all capital, personnel and material costs for Moderna’s mRNA vaccine add up to US$2.85 a dose, and Pfizer’s vaccine only US$1.18 a dose. If CAR_Ma is expected to cost about equivalent to 100% more per dose in the worst case, each dose of CAR_Ma will cost around US$5.00 and require a dose every 2 months. Combined with a course of trimethoprim at the price of US$2.23 per 100 mg pill, keeping in mind CAR_Ma would require a dose 45-fold lower in order to have control (Cafferty et al., 2021). A 6-month regime of CAR_Ma would cost about US$280. The low cost would not only reap great benefits to patients, but insurance companies and even governments, ensuring that as many people globally will be able to receive the most advanced therapeutics.
Ultimately, CAR_Ma is to be implemented as a next-generation immunotherapy with superior
effectiveness, streamlined logistics, simple administration and patient compliance, low
discomfort or side-effects and a low cost. All-in-all producing an effective therapeutic
that is accessible and affordable.
Carma Roadmap
The roadmap of CAR_Ma can be broken down into several phases encompassing our past work, current endeavors and future plans.
Phase I: 1st Generation CAR-Ma
The 1st Generation CAR-Ma was designed as a proof-of-concept for the expression of CAR via
self-replicating RNA in macrophages. This was carried out by our team for the iGEM
Competition 2023 (HKU iGEM, 2023)
Phase IIa: 2nd Generation CAR_Ma
The second generation of CAR_Ma began work in this year in Phase IIa, with the initial
design revolving around our multi-modal RNA vector carrying our 5th generation CAR with a
small-molecule variable control system and our first attempt at integrating gene modulation
into our system via a simplex Cas9-mediated knockout targeting SIRPa-CD47 and a multiplex
Cas12a-mediated knockout targeting SIRPa-CD47, PD1-PDL1 and other targets. Designs were
completed in the first quarter of 2024. The design team continued to Phase IIb and this
iteration of CAR_Ma moved on to extensive lab testing.
Phase IIb: 2nd Generation CAR_Ma
Phase IIb proceeded as iterative design improvements enhancing the 2nd generation of CAR_Ma.
This Phase focused significantly on packaging and optimization with our team pioneering
nanostructured lipid carriers and protein-based nanoparticles for design. Our team also
iterated with our second attempt at gene modulating leveraging RNAi rather than CRISPR. This
allowed us to develop a smaller overall package, improving production and reducing cost, but
more importantly allowed us to have a fully transient system where the cell could return to
a native state unlike with CRISPR which led to permanent changes which could result in
long-term complications to the immune system.
Phase IIc: 2nd Generation CAR_Ma
Phase IIc revolves around the extensive testing and characterisation that has happened and
will continue past the iGEM competition. This involves characterizing the effectiveness of
our CAR system, the variable control system, its effectiveness against 3D tumor spheroids,
immune system signaling and more. This stage will work to characterize CAR_Ma and all of its
constituent systems with as much detail as possible.
Phase IId: 2nd Generation CAR_Ma
Phase IId is the culmination and conclusion of Phase II, where all the data collected will
be published. We are expecting to publish by the first half of 2025, where we will also
patent any technologies relevant.
Phase III: Tape-out and Expansion
In Phase III, we will split our workforce with one team finalizing CAR_Ma and preparing for
large scale production and improving production quality, and another team restarting the
cycle adapting CAR_Ma to other cancers by changing the antigen recognising domain. We expect
this work to begin by the second half of 2025.
Future work
Once CAR_Ma is produced in enough quality and quantity, we plan to proceed with pre-clinical
and clinical studies preparing CAR_Ma for use as a therapy. At the same time increasing
production output and preparing for scale-up or scale-out operations. Simultaneously, our
research and development teams will continually adapt and upgrade the fundamental
technologies behind CAR_Ma opening up further applications in cancer therapy, immunotherapy,
gene therapy and further applications in other fields of synthetic biology.
References
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