RNAi, siRNAs, ribozymes and other fun things
RNA interference (RNAi) is an important pathway used for the regulation of gene expression. It used to be called post-transcriptional gene silencing (PTGS) and as the names imply it is a mechanism through which mRNA fragments are selectively silenced (prior to translation).
The two main components of the RNAi pathway are micro RNA (miRNA) and small\short interfering RNA (siRNA). They both form a double stranded RNA fragment with a hairpin loop (called primary pri-miRNA for miRNA, double stranded dsRNA in plants and short hairpin shRNA in humans for siRNA), the hairpin loop is cleaved by an enzyme called dicer, the strands separate and are incorporated into the RNA-induced silencing complex (RISC, miRISC when bound to miRNA and siRISC when bound to siRNA). The RNA strand of the RISC assembly binds to a complimentary mRNA and the mRNA is thus degraded through cleavage by Ago2, a catalytic component of the RISC (to be more specific, given complete complementarity between the miRNA and target mRNA sequence, Ago2 can cleave the mRNA and lead to direct mRNA degradation; in the absence of complementarity, silencing is achieved by preventing translation by simply ‘being in the way’ of translation machinery1).
The two differ in that miRNAs, especially those in animals, typically have incomplete base pairing to a target and inhibit the translation of many different mRNAs with similar sequences. In contrast, siRNAs typically base-pair perfectly and induce mRNA cleavage only in a single, specific target, and are thus more therapeutically useful. The above explanation is a broad overview of the process but the exact details2 will not concern us in this article.
siRNA therapy is simply the administration of exogenous siRNA molecules with the intention of silencing a particular gene. siRNA therapy, by virtue of how it works, does not last permanently - once the siRNA fragments and siRISCs are degraded the target gene will no longer be silenced. Unlike gene therapy and gene editing, which delivers new DNA or makes permanent changes to it respectively, the effect of RNA therapy is more short-lived. On the other hand, RNA therapy has less potential side effects and is easier to carry out.
For example, self reported data from Thermo Fisher states3, “a single transfection of 5 nM Silencer Select [a line of siRNAs produced by Thermo Fisher4] siRNAs achieved >80% knockdown that lasted 5–7 days post-transfection, then progressively diminished. Higher siRNA concentrations did not result in stronger or longer-lasting knockdown, but are likely to cause off-target effects. In some cases, repeated siRNA transfection resulted in improved knockdown during select timepoints post-transfection.” Most commercial approved siRNA therapies need to be re-administered once every few months.
Ribozymes (ribonucleic acid enzymes) are RNA molecules that have the ability to catalyse specific biochemical reactions. They perform an important function both in the conversion of pre-mRNA to mRNA through RNA splicing, as well as mRNA silencing through cleavage (acting similarly to siRNAs). The hammerhead, hairpin and hepatitis delta virus (HDV) ribozyme motifs (and many more!5) have the capacity for self-cleavage of a particular phosphodiester bond. Group I and group II intron ribozymes have the capacity for self-splicing, by cleavage and ligation of phosphodiester bonds6. These molecules have extensive therapeutic and research applications since they can be used to selectively silence genes through cleavage of mRNA (again, similarly to siRNA).
To reiterate, the main difference between ribozyme and siRNA technologies is that siRNAs require the recruitment of endogenous proteins to form the siRISC complex whereas ribozymes act independently.
Angela Christiano and the Hairless gene
Gene research
Angela Christiano et al. conducted inordinate quantities of research into the underlying genetics of hairlessness in the 2000’s7. Two particularly relevant genes are described below.
The hairless (Hr) gene encodes a transcriptional co-repressor highly expressed in the mammalian skin. In the mouse, several null and hypomorphic (mutation that causes a partial loss of gene function) Hr alleles have been identified resulting in hairlessness in homozygous animals, characterised by alopecia developing after a single cycle of relatively normal hair growth8. In short, silencing the gene results in hairlessness.
In the skin, epidermal adhesion is mediated in part by specialised cell-cell junctions known as desmosomes, which are characterised by the presence of desmosomal cadherins (e.g. cadherin 11), known as desmogleins (e.g. desmoglein 4) and desmocollins. There is a type of inherited hair loss, named localised autosomal recessive hypotrichosis (LAH, OMIM 607903) that can be caused by an intragenic deletion in the desmoglein 4 gene (DSG4)9. Desmoglein 4 is a key mediator of keratinocyte cell adhesion in the hair follicle, where it coordinates the transition from proliferation to differentiation10. Without DSG4 the cells separate from each other and become disorganised, so people and mice lacking the gene have thin, sparse hair that is fragile and breaks easily11.
Gene therapy research
Christiano et al. published only one study on hair removal through RNA therapy12. They designed ribozymes to target poGene therapy researchtential cleavage sites which were located on an open loop of the mouse hairless mRNA (determined using RNA secondary structure prediction software), which they then formulated as deoxyribozymes. Commercially available mouse brain polyA-RNA served as template for in vitro cleavage reactions to test the efficiency of the deoxyribozymes:
About 800 ng of RNA template was incubated in the presence of 20 mM Mg2+ and Rnase Out Rnase inhibitor (Life Technologies) at pH 7.5 with 2μg of deoxyribozyme for 1 h. After incubation, aliquots of the reaction were used as template for reverse transcriptase-polymerase chain reaction (RTPCR), amplifying regions including the targeted cleavage sites. The RT-PCR products were visualized on an ethidium bromidecontaining 2% agarose gel under UV light, and the intensity of the products was determined as described above (Fig. 1). Those deoxyribozymes that were capable of cleaving the target mouse hr mRNA with the highest efficiency were used for in vivo experiments.
The deoxyribozymes were either dissolved in an 85% EtOH/15% ethylene glycol vehicle or prepared in Superfect liposomes. For each treatment, they applied a compound containing 2μg of deoxyribozyme on a 1 cm^2 area on the back of the animal. During application and for 15 min after, the mice were placed in a temporary restraint to prevent the removal of the formula. Control animals were treated with vehicle containing the same length oligonucleotides of random sequence.
Treatment must be completed for the full duration of the hair growth cycle since the hairless gene comes into play only at specific points in the cycle. One crucial thing to note is that since the treatment completely destroyed the structure of the hair follicles, the hair removal is PERMANENT and only one course of RNA therapy is required.
8-week-old female littermates were wax depilated and killed after 4 weeks of treatment that began immediately after the depilation. Control animals showed active hair regrowth in the depilated area whereas regrowth was of lesser extent in the treated mice and the hair became more sparse. In the treated portion of skin the hair follicles are not able to enter depilation-induced anagen at all or exhibit much lower growth rates as compared with control skin. These results were seen both the 85% EtOH/15% ethylene glycol and liposome vehicles.
Medical Trial
In 2004 she even got a grant of 100,000$ to start a medical trial13 investigating the use of ribozymes as a permanent hair removal therapeutic:
Gene therapy for the treatment of disease is perhaps the most exciting promise of modern medical science. It is feasible that the simple inhibition of a single gene could potentially achieve therapeutic goals in certain medical settings. Such is the case in excessive hair growth, in which selectively inhibiting genes involved in hair cycling can lead to decreased hair growth, and improvement of the clinical phenotype. As an initial drug target, we selected the hairless (hr) gene, which is mutated in the hairless (hr/hr) mouse model. Our earlier biological and morphological studies have demonstrated that hairless expression is required for normal hair cycling. In the absence of hairless expression, either in mutant mice or human patients with complete hair loss, there is no further hair growth after the first cycle - the hair is shed, the hair follicle is destroyed, and the hair never grows. We reasoned, therefore, that if we could transiently inhibit hairless gene expression using ribozyme technology, (mimicking the hairless phenotype), we could permanently inhibit hair growth. In our Preliminary Studies, we tested the in vitro and in vivo efficacy of a simple preparation of topically applied ribozymes directed against the hairless (hr) gene, and showed evidence for targeted hair follicle disruption. Encouraged by our initial proof-of-concept results, in this Phase I STTR proposal, we will investigate more sophisticated methods for improving efficacy, focused on developing effective delivery strategies to the hair follicle, such as liposomes. We will test the hypothesis that topical application of anti-hairless ribozymes encapsulated in liposomes will result in the arrest of hair growth, and recapitulate the pathological changes and permanent hair loss observed in mutant hairless mice. Our goal is to determine the optimal liposome composition for efficient delivery of ribozymes into the hair follicle, and increase our initial success in recapitulating the hairless phenotype following long-term treatment in mice. It is our plan to apply for Phase II STTR funding for extended pre-clinical studies immediately upon successfu/completion of Phase I STTR funding, and eventually to extend these studies into human clinical trials.
As far as I can tell the trial was abandoned.
Patents
A number of patents were filed on research done by Christiano with applications to hair removal14.
Hair Removal and Hair Growth Elimination with Nucleic Acid Target Gene Interference This technology employs nucleic acid interference of specific target genes implicated in hair growth. The expression of these genes, desmoglein 4, nude and hairless, is inhibited by mRNA cleavage and/or translation repression, in order to stifle hair growth or remove hair. Catalytic nucleic acid constructs, both RNA and DNA based, for such interference-mediated inhibition are described, and compositions of carrier and vector containing these constructs have been prepared for use in humans and other mammals.
Patent Status: Patents Issued (US7423022B2); Patents Pending (US20060270621, US20070269395, WO/2004/093788, US20090004169, US20100221318)
Some of this research was sold to a company called Sirna, about which teddyg had the following insightful comment15:
Hair removal is a multi-billion dollar industry and it supports a lot of jobs around the world. Have you seen the price of razor blades???
Too much money to lose for us plebs to have an easy permanent solution.
One day they won’t be able to stop the breakthroughs but for now they have us by the balls.
Yours in hairy misery.
Teddyg
After some looking around I was able to locate the following patents:
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SIRNA THERAPEUTICS
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https://patents.google.com/patent/AU2004288143A1
https://patents.google.com/patent/US20060160757A1
https://patents.google.com/patent/WO2005045036A2
https://patents.google.com/patent/US20050054598A1
https://patents.google.com/patent/US20050176665A1
https://patents.google.com/patent/US20050233996A1
RNA interference mediated inhibition of hairless (HR) gene expression using short interfering nucleic acid (siNA)
James Mcswiggen, Sirna Therapeutics, 2004
https://patents.google.com/patent/US7404969B2
https://patents.google.com/patent/US7893302B2
Lipid nanoparticle based compositions and methods for the delivery of biologically active molecules
Tongqian ChenChandra VargeeseKurt VagleWeimin WangYe Zhang, Sirna therapeutics, 2006
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ANGELA CHRISTIANO
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https://patents.google.com/patent/EP1385948A2/en
https://patents.google.com/patent/US8008471B2/en
(EU and US patent, they differ slightly)
Nucleic acids for inhibiting hairless protein expression and methods of use thereof
Angela Christiano 2002/2002
https://patents.google.com/patent/WO2004093788A9/
Desmoglein 4 is a novel gene involved in hair growth
Angela Christiano 2004
https://patents.google.com/patent/CA2557532A1/
https://patents.google.com/patent/US8329667B2/
https://patents.google.com/patent/US20070269395A1/
(CA and US patent, they look mostly identical)
Inhibition of hairless protein mRNA
Angela Christiano 2005/2006
https://patents.google.com/patent/WO2006128084A3/
Trps1-mediated modulation of hair growth
Angela Christiano 2006
https://patents.google.com/patent/WO2006128085A2/
Cadherin-11-mediated modulation of hair growth
Angela Christiano 2006An article1617 was published in 2008 about the demise of Sirna. TL;DR Merck shut down the department after buying the company because they weren’t interested in dermatology, and the department was struggling with bringing the therapy to market because they could not find an appropriate method for transdermal delivery of siRNA.
When Sirna Therapeutics acquired privately held dermatology firm Skinetics Biosciences in late 2004 (see RNAi News, 12/10/2004), it predicted that it would move its first RNAi-based dermatology product, a permanent hair removal drug, into the clinic two years later.
Sirna never achieved this goal, in part because the company’s dermatology unit was shut down after Merck acquired the RNAi shop early last year (see RNAi News, 1/4/2007). But in the end, technological hurdles facing topical delivery of siRNAs may have been mostly to blame for the failure, according to a former Skinetics offical.
Skinetics CSO and co-founder Joseph Carroll, who headed up Sirna’s dermatology division, said the company was unable to achieve “knockdown consistently in vivo to move the [hair-removal] program forward.”
Carroll, who is now a consultant, noted that while delivery has always been a major issue for the RNAi therapeutics field, transdermal delivery is particularly difficult since the skin is by design a barrier.
“The first hurdle is just getting [siRNA] through the stratum corneum, [the outermost layer of the epidermis], and into the skin,” he told RNAi News last week. “That’s a challenge because you have this macromolecule that’s … pretty big.” But even if delivery through this layer is achieved, “the second challenge is [getting] cells to take it up,” he noted.
Carroll said that before being acquired by Merck, Sirna had both been working on its own topical delivery approaches and evaluating technologies under development elsewhere, but “we just weren’t finding viable delivery options.
“We tried a range of delivery systems for skin and were looking for high levels of consistent knockdown in vivo,” he said. “It just wasn’t achievable using the systems we looked at.”
Carroll said he remains optimistic about the future of topical RNAi drugs, but suggested that a solution to the transdermal delivery problem may not be immediately forthcoming.
“Every organ is a unique challenge, and there will someday be delivery to the skin [of] a topically delivered siRNA,” he said. Still, “the magic bullet hasn’t been found yet.”
Hair Today, Gone Tomorrow
Although a handful of RNAi drug firms have been eyeing the dermatology field, Sirna became the most high profile to do so when it bought Skinetics.
Under the terms of that deal, Sirna paid an estimated $2.5 million in stock for the company and its intellectual property portfolio, which included rights to hairless, a hair growth-related gene discovered by Columbia University researcher and Skinetics co-founder Angela Christiano.
Following the transaction, Sirna established Sirna Dermatology, a business unit focused exclusively on developing the permanent hair-removal drug and RNAi-based treatments for skin disorders including psoriasis and atopic dermatitis.
Despite the fanfare that accompanied the creation of Sirna Dermatology, the unit was quietly shut down a few months after Merck acquired Sirna, according to Carroll.
Merck was “realigning [Sirna] for a couple of key programs, and dermatology was not one of them,” he said. Dermatology “was clearly just not part of their portfolio and not something they wanted to pursue.”
Indeed, Merck has publicly stated that its main areas of interest are Alzheimer’s disease, atherosclerosis, cardiovascular disease, diabetes, vaccines, obesity, cancer, pain, and sleep disorders.
Carroll noted that he and Skinetics’ other co-founders had an option to re-acquire the Skinetics IP from Merck, “but it wasn’t very feasible at that point – we didn’t have a lab, we didn’t have funding, [and] we didn’t have a company.”
More importantly, “I had some concerns about the status of topical siRNA delivery,” he said. “I had explored the idea of starting another venture around siRNA and dermatology, but I think that’s just not going to happen right now” since it would likely be difficult to secure funding for this kind of effort.
Carroll added that the Skinetics IP has since been returned to Columbia.
Sirna Dermatology isn’t the only company to find the topical delivery nut hard to crack.
TransDerm, which is developing RNAi drugs for skin disorders, is developing a topical delivery technology called gene cream for its lead drug candidate, a pachyonychia congenita therapy called TD101.
Last week, however, the company announced that its recently initiated phase Ib study of TD101 would examine the drug when it is administered as an intradermal injection (see RNAi News, 1/17/2007).
According to TransDerm’s Founder and CEO Roger Kaspar, the company continues to work on getting the gene cream ready for human testing, but is waiting for an indication that the RNAi component of TD101 is effective before stepping up this effort.
Carroll noted that he is “very encouraged” by the gene cream data to which he has been privy, but noted that it isn’t yet apparent whether the delivery technology will be applicable to a variety of skin disorders even if it is successfully developed for TD101.
“I’m wondering how amenable [gene cream] is going to be for a range of dermatologic indications or whether it is really specific for” pachyonychia congenita, he said.
So, they couldn’t get things working in 2008, but has any progress in the field been made since then?
Ribozymes or siRNA for therapy?
The above paper by Christiano et al. (2004) clearly demonstrated the efficacy of ribozymes for permanent hair removal, yet many of the mentioned patents (2002-2006) focus instead on siRNA technology. Which is preferable?
An excellent review on the topic of siRNA drug design has been written by Zhang et al.18 siRNA molecules have low bioavailability due to their large size and anionic charge. At the systemic level, delivery is compromised by rapid clearance, especially via the kidneys. Naked unmodified agents may have a half-life as short as 5 min. This is why it took about two decades since the discovery of RNAi before the first siRNA therapies were introduced to market.
Nanocarrier-encapsulated drugs are often subject to serum protein adsorption. This opsonization results in uptake by the reticuloendothelial system (RES) and clearance by phagocytes. siRNAs are also rapidly degraded by nucleases present in plasma, tissues, and the cytoplasm.
After surviving systemic clearance, the drugs must pass through the capillary endothelium into the tissues, which is particularly challenging due to the abundance of adherence and tight junctions. siRNAs may passively accumulate to fenestrated sites, such as the liver or tumor tissue, but this presents a challenge to delivering these therapeutics to other sites besides these organs that preferentially take up these molecules. Even after successful transport into the target tissue compartment, siRNAs must be taken up into the target cells. However, RNAs do not spontaneously cross cellular membranes, which represents a barrier to cellular internalization. Furthermore, endosomal trapping after internalization can be another limiting factor in targeting siRNA molecules to their molecular sites. siRNA therapeutics may enter a cell through transfection or conjugation to a ligand with a high affinity receptor on the target cell. After internalization, less than 1% of siRNA molecules escape the endosomal compartment. siRNA molecules trapped in endosomes are either degraded or recycled back to the surface for extrusion from the cell.
Finally, siRNA may activate an undesired immunogenic response, as extracellular and intracellular immune mediators may falsely recognize these as viral RNA molecules. For instance, Toll-like receptors may recognize certain sequences as immunostimulant motifs. These reactions result in unfavorable adverse reactions
The two main strategies to address these issues are chemical modification of the siRNA and delivery mechanisms such as encapsulation in lipid nanoparticles. For more information on these refer to Zhang’s paper (since many of them are not relevant to transdermal application).
Like siRNA/shRNA therapeutics, ribozymes can either be delivered to the target cells in RNA form or can be transcribed from therapeutic genes. Due to poor stability of fully-RNA ribozymes, therapies that rely on direct delivery often require chemically stabilised ribozymes, including the following modifications: 5’-PS backbone linkage, 2’-O-Me, 2’-deoxy-2’-C-allyl uridine, and terminal inverted 3’-3’ deoxyabasic nucleotides. All of these modifications were incorporated for Angiozyme (RPI.4610), the first synthetic ribozyme tested in clinical trials. Like siRNAs, ribozymes and face similar challenges of delivery and off-target toxicity.19
Functionally, RNAi requires extremely strict recognition and degrades specific sequences that cannot be initiated with even one mistaken base. Thymine at 30 overhangs makes RNAi of higher stability, reducing the possibility of degradation by ribozyme as well as the dependence on chemical modification. Additionally, siRNA exhibits obvious inhibition of target genes at a thousandth of the concentration of AONs (antisense oligonucleotides) or even less, and the effect increases with an increase in concentration. On the other hand, Ribozymes usually recognise their cleavage sites by base pairing, but small mismatches can be tolerated by many ribozymes. They are also often less stable than siRNAs. Thus, siRNA technology is generally preferred to that of ribozymes.20
Transdermal siRNA therapy administration
I will be focusing on actually viable/affordable approaches to transdermal delivery of siRNAs.
One potentially viable approach is the use of microneedling - very cheap and easy to implement in practice. The downside is that you would need to endure a little bit of blood. It is worth noting however that these microneedle arrays are much smaller than those used for hair regrowth, so I am not sure that something like a dermaroller would work.
One study found that “Treating the ear with microneedles showed permeation of siRNA in the skin and could reduce GAPDH gene expression up to 66% in the skin without accumulation in the major organs” and “reported [microneedle] delivery efficiency varied [in previous research papers] from 10% to 85%. The delivery efficiency of the microneedles did not translate to gene silencing efficiency. Although our study cannot evaluate the percentage of transdermal siRNA to show the delivery efficiency, a significant silencing efficiency (66%) was observed.”21
The other viable approach I have found is the use of ionic liquids and deep eutectic solvents (DES) (aka black magic!). One study also investigated GAPDH suppression by siRNA delivered transdermally in a CAGE-BDOA formulation22. “CAGE-BDOA-siRNA exhibited the most significant effect of reduction of GAPDH expression. When compared to a control of CAGE-BDOA (without siRNA), this formulation had reduced the expression of target protein by 44%”. CAGE is easily synthesised by salt metathesis of choline bicarbonate and geranic acid23 (can also be bought cheaply commercially) while BDOA is essentially medical grade benzalkonium chloride, a very common surfactant.
Another study24 found a mixture of CAGE and CAPA (choline and phenylpropanoic acid) to be most effective for transdermal delivery in the context of siRNAs targeting psoriasis. These results are supported by results from molecular simulation, where it was shown that the DES system (CAGE) improved siRNA stability and provided DES-induced solvating and intercalation effects, aided by addition of phenylpropanoic acid.
Design, formulation and cost
Work in progress :)
Also I’m getting my genome sequenced so I could in theory personalise the siRNA therapy to my own genetics.
Some sort of animal model to ensure safety would be ideal.
Some further things to look into:
potential for side effects?
the importance of hair follicle stem cells for repairing skin damage and wounds (have talked about this in previous posts), could this pose an issue?
Lim, L. P.; Lau, N. C.; Garrett-Engele, P.; Grimson, A.; Schelter, J. M.; Castle, J.; Bartel, D. P.; Linsley, P. S.; Johnson, J. M. Microarray Analysis Shows That Some microRNAs Downregulate Large Numbers of Target mRNAs. Nature 2005, 433 (7027), 769–773. https://doi.org/10.1038/nature03315.
RNA Interference. Wikipedia; 2023. https://en.wikipedia.org/wiki/RNA_interference
https://www.thermofisher.com/uk/en/home/references/ambion-tech-support/rnai-sirna/tech-notes/duration-of-sirna-induced-silencing.html
https://www.thermofisher.com/uk/en/home/life-science/rnai/synthetic-rnai-analysis/ambion-silencer-select-sirnas.html
Peng, H.; Latifi, B.; Müller, S.; Lupták, A.; Chen, I. A. Self-Cleaving Ribozymes: Substrate Specificity and Synthetic Biology Applications. RSC Chem. Biol. 2021, 2 (5), 1370–1383. https://doi.org/10.1039/D0CB00207K.
Phylactou, L. A.; Kilpatrick, M. W.; Wood, M. J. Ribozymes as Therapeutic Tools for Genetic Disease. Hum. Mol. Genet. 1998, 7 (10), 1649–1653. https://doi.org/10.1093/hmg/7.10.1649.
https://pubmed.ncbi.nlm.nih.gov/?term=Christiano+AM&filter=years.2000-2010&sort=pubdate
Benavides, F.; Oberyszyn, T. M.; VanBuskirk, A. M.; Reeve, V. E.; Kusewitt, D. F. The Hairless Mouse in Skin Research. J. Dermatol. Sci. 2009, 53 (1), 10–18. https://doi.org/10.1016/j.jdermsci.2008.08.012.
Bazzi, H.; Martinez-Mir, A.; Kljuic, A.; Christiano, A. M. Desmoglein 4 Mutations Underlie Localized Autosomal Recessive Hypotrichosis in Humans, Mice, and Rats. J. Investig. Dermatol. Symp. Proc. 2005, 10 (3), 222–224. https://doi.org/10.1111/j.1087-0024.2005.10110.x.
Kljuic, A.; Bazzi, H.; Sundberg, J. P.; Martinez-Mir, A.; O’Shaughnessy, R.; Mahoney, M. G.; Levy, M.; Montagutelli, X.; Ahmad, W.; Aita, V. M.; Gordon, D.; Uitto, J.; Whiting, D.; Ott, J.; Fischer, S.; Gilliam, T. C.; Jahoda, C. A. B.; Morris, R. J.; Panteleyev, A. A.; Nguyen, V. T.; Christiano, A. M. Desmoglein 4 in Hair Follicle Differentiation and Epidermal Adhesion: Evidence from Inherited Hypotrichosis and Acquired Pemphigus Vulgaris. Cell 2003, 113 (2), 249–260. https://doi.org/10.1016/s0092-8674(03)00273-3.
Newly Identified Hair Loss Gene May Improve Hair Removal. Columbia University Irving Medical Center. https://www.cuimc.columbia.edu/news/newly-identified-hair-loss-gene-may-improve-hair-removal (accessed 2023-11-12).
Cserhalmi-Friedman, P. B.; Panteleyev, A. A.; Christiano, A. M. Recapitulation of the Hairless Mouse Phenotype Using Catalytic Oligonucleotides: Implications for Permanent Hair Removal. Exp. Dermatol. 2004, 13 (3), 155–162. https://doi.org/10.1111/j.0906-6705.2004.0143.x.
GENE INHIBITION STRATEGIES FOR HAIR REMOVAL | SBIR.gov. Accessed March 19, 2024. https://www.sbir.gov/node/308936
https://web.archive.org/web/20161220230002/http://innovation.columbia.edu/technologies/1084_hair-growth-inhibited-with-nucleic-acid-target-gene-interference
https://hairtell.com/forum/t/dr-angela-christiano-nucleic-acid-target-gene-int/40090/2
http://www.rnainews.com/issues/6_4/features/144620-1.html, dead link, not on the internet archive.
https://hairtell.com/forum/t/merck-axed-sirnas-hair-removal-studies/34061/2
Zhang, M. M.; Bahal, R.; Rasmussen, T. P.; Manautou, J. E.; Zhong, X. The Growth of siRNA-Based Therapeutics: Updated Clinical Studies. Biochem. Pharmacol. 2021, 189, 114432. https://doi.org/10.1016/j.bcp.2021.114432.
Burnett, J. C.; Rossi, J. J. RNA-Based Therapeutics- Current Progress and Future Prospects. Chem. Biol. 2012, 19 (1), 60–71. https://doi.org/10.1016/j.chembiol.2011.12.008.
Zhao, Y.; Shu, R.; Liu, J. The Development and Improvement of Ribonucleic Acid Therapy Strategies. Mol. Ther. - Nucleic Acids 2021, 26, 997–1013. https://doi.org/10.1016/j.omtn.2021.09.002.
Deng, Y.; Chen, J.; Zhao, Y.; Yan, X.; Zhang, L.; Choy, K.; Hu, J.; Sant, H. J.; Gale, B. K.; Tang, T. Transdermal Delivery of siRNA through Microneedle Array. Sci. Rep. 2016, 6 (1), 21422. https://doi.org/10.1038/srep21422.
Dharamdasani, V.; Mandal, A.; Qi, Q. M.; Suzuki, I.; Bentley, M. V. L. B.; Mitragotri, S. Topical Delivery of siRNA into Skin Using Ionic Liquids. J. Controlled Release 2020, 323, 475–482. https://doi.org/10.1016/j.jconrel.2020.04.038.
Zakrewsky, M.; Banerjee, A.; Apte, S.; Kern, T. L.; Jones, M. R.; Sesto, R. E. D.; Koppisch, A. T.; Fox, D. T.; Mitragotri, S. Choline and Geranate Deep Eutectic Solvent as a Broad-Spectrum Antiseptic Agent for Preventive and Therapeutic Applications. Adv. Healthc. Mater. 2016, 5 (11), 1282–1289. https://doi.org/10.1002/adhm.201600086.
Mandal, A.; Kumbhojkar, N.; Reilly, C.; Dharamdasani, V.; Ukidve, A.; Ingber, D. E.; Mitragotri, S. Treatment of Psoriasis with NFKBIZ siRNA Using Topical Ionic Liquid Formulations. Sci. Adv. 2020, 6 (30), eabb6049. https://doi.org/10.1126/sciadv.abb6049.
Very impressed by what you do on this blog, please keep it up!