By Gilang Cahya 
In the rigorous pursuit of scientific understanding, outcomes that defy initial expectations are not merely anomalies; they are often the fertile ground from which groundbreaking discoveries sprout. This principle has been vividly illustrated by a recent study conducted by researchers at Memorial Sloan Kettering Cancer Center (MSK) and their collaborators at the Icahn School of Medicine at Mount Sinai. Their unexpected laboratory findings have illuminated a novel therapeutic pathway, holding significant promise for enhancing treatments that employ small RNAs to silence disease-causing genes, including those implicated in various cancers.
The serendipitous nature of scientific inquiry was underscored by developmental biologist Eric Lai, PhD, who remarked, "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 undertaken by the research team, led by Seungjae Lee, PhD, a postdoctoral fellow in the Lai Lab at MSK’s Sloan Kettering Institute. Their initial objective was to investigate the role of a protein named ALAS1 in the biogenesis of microRNAs, a class of small regulatory RNA molecules. The prevailing hypothesis was that ALAS1’s involvement in heme production, a critical component in numerous biological processes, would directly correlate with microRNA levels. Specifically, the researchers anticipated that reducing ALAS1 in cellular models would lead to a corresponding decrease in microRNA abundance.
However, the experimental results delivered a profound surprise. Instead of the predicted decline, the team observed a significant and unexpected increase in microRNA levels when ALAS1 was depleted. This counterintuitive outcome catalyzed a deeper investigation, revealing an uncharacterized function for ALAS1 that extends beyond its well-established role in heme synthesis. The implications of this discovery, published in the prestigious journal Science, are far-reaching, potentially revolutionizing the application of RNA-based therapeutics.
The Power of Small RNA Snippets in Gene Silencing
To fully appreciate the significance of this new finding, it is essential to understand the fundamental mechanism by which small RNAs exert their influence. Both microRNAs (miRNAs) and their closely related counterparts, small interfering RNAs (siRNAs), are remarkably short RNA molecules, typically measuring only 21 to 22 nucleotides in length. These molecular entities operate by binding with high specificity to messenger RNAs (mRNAs), the cellular transcripts that carry genetic instructions from DNA to the protein-making machinery of the cell. Upon binding, miRNAs and siRNAs effectively "silence" the target mRNA, preventing the production of the corresponding protein.
The intricate cellular machinery involved in generating these potent silencing agents involves a complex cascade of enzymatic reactions. This process converts longer RNA precursor molecules into the mature, active small RNA products. Crucially, scientists have successfully harnessed this biological pathway to develop a novel class of drugs. These therapeutic siRNAs are designed to target and silence specific genes that are aberrantly expressed and contribute to disease pathogenesis.
The impact of this technological advancement is already evident in clinical practice. The U.S. Food and Drug Administration (FDA) approved the first siRNA drug, patisiran, in 2018. This groundbreaking therapy is used to treat hereditary transthyretin amyloidosis, a debilitating genetic disorder. Since then, several other siRNA drugs have gained regulatory approval, with numerous others progressing through various stages of clinical trials. The therapeutic potential of siRNA medicines is recognized for both rare genetic conditions and more prevalent diseases, with these drugs sometimes referred to as RNAi (RNA interference) drugs due to their mechanism of interfering with mRNA accumulation.
Unveiling a "Moonlighting" Enzyme: ALAS1’s Dual Role
The unexpected surge in microRNA levels observed in the Lai Lab, following the depletion of ALAS1, prompted Dr. Lee to conduct further rigorous experiments. These investigations systematically examined the impact of removing other enzymes involved in the heme biosynthesis pathway. The critical finding was that the depletion of these other enzymes did not elicit any significant change in microRNA levels. This stark contrast strongly suggested that ALAS1 possessed a distinct and independent function, separate from its established 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, articulating the profound nature of their discovery. Dr. Lai further elaborated on this novel function, characterizing it as a "moonlighting" role, a term used in biochemistry to describe proteins that perform multiple, unrelated functions. "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.
This "moonlighting" function of ALAS1 represents a significant departure from previous understanding. ALAS1, or aminolevulinate synthase 1, is the rate-limiting enzyme in the heme biosynthesis pathway. Heme itself is an indispensable cofactor in various physiological processes, including oxygen transport via hemoglobin, energy production through cellular respiration, and, as now revealed, the intricate regulation of gene expression by microRNAs. The dual nature of ALAS1, acting both as a key player in heme synthesis and as a regulator of microRNA biogenesis, opens up entirely new avenues for therapeutic intervention.
A Collaborative Effort to Enhance siRNA Drug Efficacy
Recognizing the far-reaching implications of their cell-based findings, the MSK research team sought to validate their observations in more complex biological systems. This led to a crucial collaboration with experts in heme regulation and ALAS genes at the Icahn School of Medicine at Mount Sinai. This partnership involved Makiko Yasuda, MD, PhD, Robert Desnick MD, PhD, and postdoctoral fellow Sangmi Lee, PhD. Their collective expertise facilitated the translation of the MSK team’s findings from cell cultures into sophisticated custom animal models that the Mount Sinai group had been developing.
The experiments conducted in these mouse models corroborated the initial discoveries. Upon depleting ALAS in liver cells, a global increase in microRNAs was observed, mirroring the results seen in cell culture. This consistency across different experimental platforms solidified the understanding of ALAS’s regulatory role.
"The emerging picture is that ALAS acts as a brake on the production of microRNAs," Dr. Lai explained. This conceptualization of ALAS as a "brake" provided a clear rationale for its potential therapeutic application. "So we thought, now that we know how to remove this brake, maybe we can use that to improve the efficacy of siRNA drugs and their ability to silence their target genes."
The theoretical framework suggests that by modulating ALAS activity, researchers could potentially enhance the effectiveness of siRNA drugs. This strategy could be particularly impactful for targeting genes that are overexpressed and drive disease processes, including oncogenes that fuel cancer development.
Addressing the Limitations of Current siRNA Therapies
Despite the remarkable progress in developing siRNA drugs, significant challenges remain. "Therapeutic siRNA drugs don’t work well enough against all targets and are currently limited in where they can be used in the body," Dr. Lai noted. A prime example of this limitation is the current restriction of all six FDA-approved siRNA drugs to targeting hepatocytes, the primary functional cells of the liver. The liver’s role as the body’s principal filtration organ makes it a relatively accessible target for drug delivery.
To demonstrate the practical feasibility of their hypothesis, the team conducted a proof-of-concept study. They successfully depleted ALAS in mouse liver cells, confirming the expected increase in microRNA levels. More importantly, this intervention led to a significant enhancement in the silencing activity of a model siRNA compound administered to the mice. This experimental success provided tangible evidence that manipulating ALAS levels can indeed boost the efficacy of siRNA therapeutics.
Adding a layer of remarkable coincidence and immediate therapeutic relevance, one of the existing FDA-approved siRNA drugs, givosiran, functions by inhibiting ALAS1. This drug is specifically used to treat acute hepatic porphyrias, a group of rare genetic disorders affecting heme synthesis. The established safety and efficacy of givosiran in humans strongly suggest that combining such an ALAS1-inhibiting agent with other siRNA drugs could be a viable strategy to enhance their therapeutic impact. Dr. Lai posited that this approach could be broadly applicable to any siRNA therapy.
The potential benefits of improving siRNA drug efficacy are multifaceted. Enhanced potency could lead to cost-effectiveness by enabling lower therapeutic doses, thereby potentially reducing the incidence of side effects. Furthermore, a deeper understanding of ALAS regulation might pave the way for expanding the therapeutic reach of siRNA drugs beyond the liver to target other cell types and tissues, broadening their applicability to a wider range of diseases.
The Enduring Value of Discovery Science
The journey from an unexpected experimental result to a potential therapeutic breakthrough underscores the indispensable role of curiosity-driven, foundational research. This is particularly relevant in the context of recent accolades in the field of small RNA biology. In December 2024, Harvard geneticist Gary Ruvkun, PhD, was awarded the Nobel Prize in Physiology or Medicine, alongside Victor Ambros, PhD, for their seminal discovery of microRNAs and their role in gene regulation in the early 1990s. Dr. Lai himself had the privilege of conducting his undergraduate thesis research in Dr. Ruvkun’s laboratory during that formative period.
Reflecting on this profound influence, Dr. Lai stated, "I got my first real exposure to how science was actually done and gained lifelong interests in developmental biology and small RNAs." He emphasized that his mentor’s Nobel Prize serves as a powerful testament to the transformative impact of fundamental, curiosity-driven research.
"Dr. Ruvkun didn’t start out looking for microRNAs," Dr. Lai observed. "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."
This narrative serves as a compelling argument for continued investment in basic science. "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," Dr. Lai asserted. He stressed the critical importance of maintaining robust public discourse and sustained support for scientific research, particularly in an era marked by societal and governmental uncertainty regarding funding priorities. "Hopefully, there will be continued support to keep the engine of foundational research strong," he concluded.
The research detailed in this study 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). Furthermore, the researchers have filed a patent application pertaining to their methods for enhancing the efficacy of RNAi therapy by targeting ALAS1/ALAS2 (WO2024148236A1). Drs. Yasuda and Desnick are also co-inventors on a patent related to RNAi therapy for acute hepatic porphyrias and have reported pharmaceutical consulting work.
