Life scientists are all probably well aware of synthetic oligonucleotides which have been manufactured for use in research, for example in PCR or in DNA-based diagnostic kits. More recently, companies have begun to develop oligonucleotide-based therapeutic products too.
During a recent oligonucleotide panel discussion at the Biologics 2024 event in London, it was reported that there are over 100 oligonucleotide drugs in development. The panel also discussed the most considerable challenges for this industry segment, such as patent restrictions, discovery of chemical modifications that increase stability and more ‘off-the-shelf’ target delivery mechanisms needed.
In this blog we ask ‘What are Oligonucleotides?’ and discuss their therapeutic uses and how BioPharmaSpec characterizes these drugs in their global laboratories.
Oligonucleotide therapeutics are modified single- or double-stranded nucleotide polymers that interact with target mRNA to decrease, increase, or restore protein expression. They can be classified as one of three main types:
Aptamers are also single-stranded RNA; however, they function by binding target proteins to affect activity.
Amazingly, all currently approved oligonucleotide therapeutics have been generated through only a handful of modifications. These include modifications at the 2’-carbon of the ribose which is the location of the ribose hydroxyl in RNA and modifications to the phosphodiester backbone which connects adjacent nucleotides. In the latter, replacement of a non-bridging oxygen with a sulfur atom increases nuclease resistance and the lifetime of the drug in vivo. Modifications to the ribose increase overall stability as well as binding affinity to the mRNA targets.
Another successful class of structural analogues is the morpholinos, which contain a neutral backbone completely resistant to nucleases and a morpholine ring in place of the ribose. Conjugation of N-acetyl galactosamine (GalNAc) to siRNA therapeutics has also been successful in targeting drugs to receptors located on liver cells.
Currently there are seventeen FDA approved oligonucleotide therapeutics. These include three vaccines (HEPLISAV-B, COMIRNATY, and SPIKEVAX), three ASOs that cause degradation of target mRNA (VITRAVENE, KYNAMRO, and TEGSEDI), five SSOs that modulate splicing to exclude, or include, specific exons (EXONDYS 51, VYONDYS 53, VILTEPSO, AMONDYS 45, and SPINRAZA), five siRNA that also result in degradation of target mRNA (ONPATTRO, GIVLAARI, OXLUMO, LEQVIO, and AMVUTTRA), and one aptamer (MACUGEN) that binds a target protein affecting its activity.
In 1998 VITRAVENE was the first oligonucleotide therapeutic approved by the FDA. It was used to treat viral retinitis in immunocompromised patients, however, it was subsequently discontinued in 2004 due to lack of clinical need.
The approved vaccines result in the production of antibodies to battle Hepatitis B or Covid-19, respectively. The remaining therapeutics, except for VITRAVENE, are used in the treatment of genetic disorders including familial hypercholesterolemia, transthyretin amyloidosis, Duchenne muscular dystrophy, spinal muscular atrophy, acute hepatic porphyria, and primary hyperoxaluria.
Oligonucleotides are considered to be more akin to small molecules than biologics due to being manufactured synthetically. However, the chemical modifications necessary to impart functionality and the final formulations add a level of complexity which increases the analytical challenge. BioPhamaSpec scientists utilize a range of chromatographic, electrophoretic and mass spectrometry techniques to support identification and purity assessment including LC-ESI-MS, nano-ESI-MS, IP-LC-ESI-MS and MALDI-TOF MS.
Challenges still exist concerning the delivery of oligonucleotide therapeutics but the idea of drug targeting is commonplace and chemistries continue to be developed to bind small molecule drugs to antibody delivery systems (e.g. Antibody Drug Conjugates (ADCs)). This type of approach has led to the development of antibody-oligonucleotide conjugates (AOCs), where different oligonucleotide products are conjugated to antibodies which recognize different protein targets such as the HIV gp160 protein, CD71, CD44 and the epidermal growth factor receptor. These types of systems, whilst still in their infancy, provide an excellent mechanism for targeting oligonucleotides to where they are required.
Oligonucleotides are clearly an area of high importance that will continue to develop for the treatment of rare or currently untreatable diseases. Their outlook is promising as there are currently dozens of oligonucleotide candidates in late-stage trials. Of course, a robust analytical characterization strategy must be used to interrogate the structure of the oligonucleotide product and the high-end chromatographic, spectroscopic and mass spectrometric instrumentation and expertise available at BioPharmaSpec is ideally placed to support this requirement.
Contact us to discuss how our scientists can use their expert knowledge to help start your project.
