Bioorthogonal Chemistry

‘Bioorthogonal chemistry’ is a term coined in 2003 by American Chemist & 2022 Nobel Prize Laureate Carolyn Bertozzi. It encompasses a set of chemical reactions which can occur within biological environments, whilst exerting minimal effect on native biomolecules or interference with native biochemical processes of the host organism - these reactions exist ‘orthogonal’ (perpendicular) to biology. Key functional groups in Bioorthogonal Chemistry include the alkynes (carbon-carbon triple bonds) and the azides (⁻N=N⁺=N⁻). The azides are particularly bioorthogonal due to their minute size (which is favourable for cell permeability and avoiding ‘perturbations’ - the alteration of a function of a biological system), metabolic stability, and how, as they don’t naturally exist in cells, they have no competing biological side reactions. Past & present uses of bioorthogonal chemistry include: 

● Vehicle airbags: Modern vehicle airbags contain sodium azide (NaN₃), a shock sensitive, explosive compound. When a vehicle’s crash sensor is triggered, an electrical charge is administered which starts the chemical reaction, inflating the air bag with harmless nitrogen gas (2NaN₃ → 2Na + 3N₂). This reaction can occur in as quickly as 0.03 seconds! 

● Early HIV treatment: Azidothymidine - AZT - (Fig. 1) was the first drug used to treat HIV infection. For viruses to replicate, they use an enzyme called reverse transcriptase to convert their single-stranded RNA genome to double-stranded DNA in a process termed reverse transcription. When this antiretroviral medicine is used, instead of the virus transcribing thymidine, it instead transcribes the AZT, which contains an azide Group, thus stalling DNA synthesis of HIV and producing less viruses. 

Another key feature to consider when discussing uses of Bioorthogonal Chemistry are Click Reactions. Click Reactions occur exclusively between the azides (⁻N=N⁺=N⁻) and alkynes (carbon-carbon triple bond), produce no by-products and therefore have a 100% atom economy. Bioorthogonal ‘Click’ Chemistry has enabled complex chemical reactions to be carried out within living organisms: the reactions do not bring harm to, interfere with or disrupt the biological processes occurring within these systems as they cannot be recognised & used by these systems. ‘Click’ Chemistry is therefore vital in understanding how we may be able to develop Targeted Cancer Therapeutics using Bioorthogonal Chemistry. 

Modern day cancer treatments tend to be delivered intravenously using anthracyclines (notably doxorubicin), a class of antitumour antibiotics used for cancer chemotherapy: they stop the growth of cancerous cells by preventing their enzymatic machinery from engaging in DNA duplication & cell division, causing the cells to die. The long-standing side effect of using such effective drugs is the high likelihood of ‘off-target toxicity’, where non-cancerous cells can also be harmed by the intercalating effects of the anthracyclines. Frequent targets for this ‘off-target toxicity’ tend to be fast growing body cells, like hair & nails, hence why most cancer patients experience some form of hair loss over the course of their chemotherapy treatment. So, scientists began to consider: what if there was a way to develop targeted cancer treatments? Treatments that enabled the activation of these powerful cancer drugs - anthracyclines - at the tumour sites, mitigating the harm of ‘off-target toxicity’? This is where Bioorthogonal ‘Click’ Chemistry comes in. 

‘Click-Activated Protodrugs Against Cancer’ (or ‘CAPAC’) is a platform developed by American Biotechnology Company Shasqi. Through ‘CAPAC’, Shasqi are pioneering the use of Bioorthogonal ‘Click’ Chemistry to target cancer drugs directly to the tumour site, minimising side effects and potentially improving the therapeutic index. They’ve achieved this through exploiting one of the fastest click reactions: a Diels-Alder cycloaddition between a tetrazine (C2H2N4) and a trans-cyclooctene (TCO) - 2 bioorthogonal molecules. The treatment involves two key components: a tetrazine-modified sodium hyaluronate biopolymer & doxorubicin that is connected to a TCO (trans-cyclooctene) unit. Over the course of the treatment (Fig. 2), the patient will undergo multiple stages:

● Local hydrogel injection: The tetrazine-modified sodium hyaluronate biopolymer is injected into a patient’s tumour 

● Protodrug dose: The patient then receives five daily infusions of doxorubicin-TCO

● Concentration: The drug circulates through the body until it meets the tetrazine-modified biopolymer at the tumour site 

● Activation: At the point of meeting, the click reaction brings the tetrazine and TCO together, triggering a rearrangement that frees the doxorubicin right next to the tumour cells 

Compared to prior cancer treatments, this process would not only mitigate the harm of the drug’s ‘off-target toxicity’, limiting the side-effects of the chemotherapy drug, it would also increase the local concentration of doxorubicin far beyond what would normally be possible in a patient, having a greater effect in preventing the growth of cancer cells. 

In the treatment of this life-threatening disease, Shasqi’s research into the ‘CAPAC’ platform, though still ongoing, looks excitingly promising: as recently as March 2023, they’ve proven their platform’s efficacy in humans. During a Phase 1 dose-escalation clinical trial in adult patients with advanced solid tumours, Shasqi were able to demonstrate the activation of their tetrazine-modified sodium hyaluronate biopolymer & doxorubicin-TCO at tumour sites, evidencing it’s safety, systemic pharmacokinetics, and immunological activity. With the continuation of their innovative research, the future treatment of cancer can be significantly aided with the use of Bioorthogonal ‘Click’ Chemistry.

Written by Emmanuella Fernandez

REFERENCES

Acs.org. (2021). Click chemistry sees first use in humans. [online] Available at: https://cen.acs.org/pharmaceuticals/Click-chemistry-sees-first-use/98/web/2020/10. 

Cancer Research UK (2023). Doxorubicin (Adriamycin) | Cancer drugs | Cancer Research UK. [online] www.cancerresearchuk.org. Available at: 

https://www.cancerresearchuk.org/about-cancer/treatment/drugs/doxorubicin. 

Wang, Y., Zhang, C., Wu, H. and Feng, P. (2020). Activation and Delivery of Tetrazine-Responsive Bioorthogonal Prodrugs. Molecules, 25(23), p.5640. doi:https://doi.org/10.3390/molecules25235640. 

Wikipedia Contributors (2019). Reverse transcriptase. [online] Wikipedia. Available at: https://en.wikipedia.org/wiki/Reverse_transcriptase. 

Wikipedia. (2020). Zidovudine. [online] Available at: https://en.wikipedia.org/wiki/Zidovudine. 

Wikipedia. (2022). Bioorthogonal chemistry. [online] Available at: 

https://en.wikipedia.org/wiki/Bioorthogonal_chemistry.