By Gilang Cahya 
In the meticulous world of scientific inquiry, where hypotheses are rigorously tested and predictions are made, the unexpected often serves as a potent catalyst for groundbreaking discovery. Such was the case for a collaborative research effort involving scientists at Memorial Sloan Kettering Cancer Center (MSK) and the Icahn School of Medicine at Mount Sinai. Their laboratory investigations, yielding results contrary to initial expectations, have illuminated a novel pathway to enhance therapeutic strategies that employ small RNAs to silence disease-causing genes, with significant implications for conditions like cancer.
The Serendipitous Discovery
The genesis of this discovery lies in a seemingly straightforward experiment. Developmental biologist Eric Lai, PhD, a principal investigator at MSK, described the core of the research: "Sometimes you do an experiment. You think you’re testing one idea, but when it doesn’t turn out the way you planned, it can lead you to find something else that’s much more interesting." This sentiment perfectly encapsulates the journey of the research team, led by Seungjae Lee, PhD, a postdoctoral fellow in Dr. Lai’s lab.
Their initial objective was to elucidate the role of a protein known as ALAS1 in the biogenesis of microRNAs, a critical class of small regulatory RNAs. The prevailing scientific understanding at the time posited that ALAS1 was integral to the production of heme, a vital molecule involved in numerous biological processes including oxygen transport and energy production. Researchers hypothesized that ALAS1’s involvement in heme synthesis might indirectly influence the generation of microRNAs. Consequently, when the team experimentally removed ALAS1 from cellular models, they anticipated a commensurate decrease in microRNA levels.
However, the experimental outcome defied this prediction. Instead of a decline, the researchers observed a surprising and significant increase in microRNA abundance. "But instead, we were surprised to see them increase," Dr. Lai recounted, emphasizing the counterintuitive nature of the finding. This unexpected result immediately signaled the existence of a previously unrecognized function for ALAS1, extending beyond its well-established role in heme biosynthesis.
Unraveling the "Moonlighting" Enzyme
This anomalous observation prompted a deeper investigation into ALAS1’s cellular activities. Dr. Lee’s subsequent experiments confirmed that the removal of ALAS1 consistently led to a surge in microRNA production. Crucially, when other enzymes involved in the heme biosynthesis pathway were experimentally depleted, microRNA levels remained unaffected. This critical distinction strongly suggested that ALAS1 possessed a distinct, independent function unrelated to its canonical role in heme production.
"This told us that ALAS1 has another job outside of helping to make heme, which no one had realized," stated Dr. Lee, highlighting the novelty of their finding. Dr. Lai further elaborated on this concept, describing it as a "moonlighting" function, a biological phenomenon where a single protein performs multiple, seemingly unrelated tasks. "And here we discovered that ALAS1 has this secret role regulating microRNAs that’s not connected to its normal role in heme synthesis," he added.
The findings were formally published in the prestigious scientific journal Science, underscoring the significance of this discovery within the broader scientific community.
The Power of Small RNAs in Therapeutics
To fully appreciate the implications of this discovery, it is essential to understand the mechanism and therapeutic potential of small RNAs. MicroRNAs, alongside related small interfering RNAs (siRNAs), are diminutive RNA molecules, typically comprising 21 to 22 nucleotides. Their primary function is to bind to specific messenger RNA (mRNA) molecules, effectively silencing them and thereby preventing the translation of genetic information into proteins.
A complex cellular machinery, akin to a "bucket brigade," works in concert to process longer RNA precursors into these potent, short regulatory RNAs. This intricate biological process has been a key focus for therapeutic development. Scientists have successfully harnessed this knowledge to engineer small RNA-based drugs capable of precisely targeting and silencing genes implicated in various diseases.
A landmark achievement in this field was the U.S. Food and Drug Administration (FDA) approval of patisiran in 2018. This groundbreaking siRNA drug was the first of its kind to be authorized for the treatment of hereditary transthyretin amyloidosis, a debilitating genetic disorder. Since then, a growing number of siRNA-based therapies have received regulatory approval, with many more progressing through rigorous clinical trials. The potential applications are vast, ranging from rare genetic disorders to more prevalent conditions. These siRNA drugs, often referred to as RNAi drugs, operate by interfering with the accumulation of messenger RNA.
A Collaborative Endeavor and In Vivo Validation
The unexpected findings from the Lai Lab at MSK prompted a strategic collaboration with researchers at the Icahn School of Medicine at Mount Sinai. This partnership brought together expertise in heme regulation and the intricacies of ALAS genes, specifically from Makiko Yasuda, MD, PhD, Robert Desnick, MD, PhD, and postdoctoral fellow Sangmi Lee, PhD. The Mount Sinai team had been actively developing custom animal models, providing an ideal platform for the MSK researchers to extend their cell-based discoveries into a more complex biological system.
In these custom animal models, specifically in mouse liver cells, the depletion of ALAS mirrored the cellular findings: a global increase in microRNA levels was observed. This in vivo validation solidified the hypothesis that ALAS1 acts as a crucial regulator, potentially a "brake," on microRNA production.
"The emerging picture is that ALAS acts as a brake on the production of microRNAs," Dr. Lai explained. This insight immediately sparked a new line of inquiry: could this newfound understanding be leveraged to enhance the efficacy of existing or future siRNA therapies? The team posited that by understanding how to release this "brake," they might be able to amplify the silencing capabilities of siRNA drugs.
Implications for Cancer Therapy and Beyond
The theoretical implications of this discovery are far-reaching, particularly for the development of more potent and versatile therapeutic agents. The ability to modulate ALAS1 activity could potentially boost the effectiveness of siRNA drugs against any gene that is aberrantly overactive in disease states. This includes oncogenes, the genetic drivers of many cancers, which are often prime targets for therapeutic intervention.
However, Dr. Lai cautioned that significant hurdles remain before this potential can be fully realized. "But we’re not quite there yet," he stated. Current siRNA drugs face limitations in their therapeutic index and systemic delivery. The majority of FDA-approved siRNA drugs, for instance, are designed to target hepatocytes, the primary functional cells of the liver. This concentration in liver-targeting is largely due to the liver’s inherent role as a filtration organ, making it a relatively accessible target for drug delivery.
As a testament to their progress, the team conducted a proof-of-concept study. They successfully demonstrated that not only could they deplete ALAS in mouse liver cells, leading to an increase in microRNAs, but this intervention also significantly enhanced the gene-silencing activity of a model siRNA compound administered to the mice.
A Promising Synergy with Existing Therapies
Adding a layer of serendipity to this discovery is the fact that one of the existing FDA-approved siRNA drugs, givosiran, works by inhibiting ALAS1 itself. Givosiran is used to treat acute hepatic porphyrias, a group of rare genetic disorders. The established safety and efficacy of givosiran in humans provides compelling evidence that targeting ALAS1 with siRNA technology is not only feasible but also well-tolerated.
This convergence suggests a potentially powerful synergistic strategy: combining an siRNA agent designed to inhibit ALAS1 with other siRNA drugs targeting different disease-causing genes. "Since an siRNA against ALAS1 works effectively and safely in humans, this raises the possibility of combining such an agent to enhance other siRNA drugs," Dr. Lai noted. He further speculated that this approach could be broadly applicable across a wide range of siRNA therapies.
The potential benefits of such a combined therapeutic strategy are substantial. Enhanced efficacy could lead to lower required doses of siRNA drugs, thereby reducing the risk of side effects and improving cost-effectiveness. Furthermore, if these therapies can be made more potent, it might open avenues for targeting cell types beyond the liver, expanding the therapeutic reach of RNAi technology.
The Enduring Value of Foundational Research
This remarkable discovery serves as a powerful illustration of the importance of curiosity-driven, fundamental scientific research. In December 2024, the Nobel Prize in Physiology or Medicine was awarded to Gary Ruvkun, PhD, and Victor Ambros, PhD, for their pioneering discovery of microRNAs and their role in gene regulation in the early 1990s. Dr. Lai, who conducted his undergraduate thesis research in Dr. Ruvkun’s lab, credits his mentor with igniting his passion for science.
"I got my first real exposure to how science was actually done and gained lifelong interests in developmental biology and small RNAs," Dr. Lai remarked, emphasizing how his mentor’s recent accolade highlights the profound impact of foundational science. He elaborated on the essence of such research: "Dr. Ruvkun didn’t start out looking for microRNAs. Like Dr. Ambros, he was investigating the development of nematodes, these tiny worms that live in the soil. And not only did this unveil an entirely new paradigm for how genes are controlled, the field they started eventually resulted in a novel class of human therapies."
Dr. Lai strongly advocates for continued investment in basic research, even when its immediate practical applications are not apparent. "When people ask why we’re not spending all of our research dollars directly studying diseases like cancer, why we’re funding research into cells and processes in model organisms like fruit flies, yeast, and bacteria — this is a great example of how discovery science fuels the biggest breakthroughs," he asserted. He underscored the critical need for sustained public and governmental support for scientific endeavors, particularly in an era marked by societal and political debates surrounding research funding priorities. "And I think it is especially critical to keep this conversation active, given how much uncertainty and disagreement there is in society and government about how much to publicly fund scientific research and in what areas. Hopefully, there will be continued support to keep the engine of foundational research strong."
Funding and Intellectual Property
The research underpinning this discovery was supported by grants from the National Institutes of Health (R01DK134783, R01-GM083300, P30-CA008748), a Cooperative Centers of Excellence in Hematology pilot grant (10040500-05S1), and a NYSTEM training award (C32559GG).
The researchers have filed a patent application related to their methods for enhancing the efficacy of RNAi therapy by targeting ALAS1/ALAS2 (WO2024148236A1). Additionally, Drs. Yasuda and Desnick are co-inventors on a patent concerning RNAi therapy for acute hepatic porphyrias and have reported pharmaceutical consulting work.
