Targeting Mitochondrial Complex I: How Homoharringtonine Overcomes T315I Resistance in Chronic Myeloid Leukemia

Chronic myeloid leukemia (CML) has long been treated through targeted inhibition of the BCR-ABL tyrosine kinase, yet drug resistance—particularly driven by the T315I mutation—continues to challenge therapeutic success. Recent research has uncovered a novel metabolic mechanism by which homoharringtonine (HHT), an FDA-approved natural compound, exerts potent anti-leukemic activity independent of kinase inhibition. Using proteomic and functional analyses, investigators identified oxidative phosphorylation (OXPHOS) as the most significantly disrupted pathway following HHT treatment, with mitochondrial complex I emerging as a major target. HHT-induced downregulation of complex I proteins, coupled with reductions in ATP production, oxygen consumption, and enzymatic activity, reveals a metabolic vulnerability in resistant CML cells that HHT can effectively exploit. This newly defined mechanism highlights the therapeutic potential of targeting mitochondrial metabolism as a strategy to overcome T315I-associated resistance and offers a promising direction for innovation in CML treatment beyond traditional TKI approaches.

Introduction: The Persistent Challenge of Drug Resistance in CML

Chronic myeloid leukemia (CML) is a hematologic malignancy defined by the presence of the BCR-ABL fusion gene, a constitutively active tyrosine kinase that drives uncontrolled proliferation of myeloid cells. The introduction of tyrosine kinase inhibitors (TKIs) revolutionized CML treatment, transforming what was once a fatal disease into a manageable chronic condition for many patients. First-line therapies such as imatinib, and later second- and third-generation TKIs, have significantly improved long-term survival and disease control. However, the clinical landscape is far from resolved. A major and persistent barrier in CML therapy is the emergence of drug resistance, particularly in patients who harbor mutations that reduce TKI binding efficiency.

Among these resistance-associated mutations, the T315I mutation in the BCR-ABL kinase domain stands out as the most challenging. Often referred to as the “gatekeeper mutation,” T315I alters a critical residue within the ATP-binding pocket, preventing most TKIs from achieving adequate inhibition. As a result, patients with T315I-positive CML frequently experience therapeutic failure, disease progression, and limited treatment options. Although ponatinib, a third-generation TKI, was specifically designed to overcome this mutation, concerns regarding cardiovascular toxicity and variable patient tolerance underscore the ongoing need for alternative strategies.

In this context, homoharringtonine (HHT)—a naturally derived, FDA-approved therapeutic—has resurfaced as a promising candidate for resistant CML. Historically used for hematologic malignancies, HHT has demonstrated potent anti-leukemic activity, including efficacy in cases where TKIs fall short. Yet despite decades of therapeutic application, the precise mechanisms driving its success have remained incompletely defined. Unraveling these mechanisms is crucial, not only for optimizing HHT’s clinical utility but also for guiding the development of next-generation treatments for resistant CML.

The growing body of research on metabolic vulnerabilities in cancer has opened new avenues for therapeutic intervention. As recent findings begin to illuminate unexpected mitochondrial targets of HHT, particularly in drug-resistant CML models, the possibility of using metabolic disruption as a complementary or alternative strategy to kinase inhibition is gaining momentum. Understanding the foundational challenges of resistance and the limitations of current therapies sets the stage for exploring how HHT provides a novel—and much needed—mechanistic advantage in overcoming T315I-driven CML.

Homoharringtonine: A Rediscovered Therapeutic With Unresolved Mechanisms

Homoharringtonine (HHT), a cephalotaxine ester derived from the Cephalotaxus genus of evergreen shrubs, has long held a place in the treatment landscape for hematologic malignancies. First isolated in the 1960s and later synthesized as omacetaxine mepesuccinate, HHT has demonstrated potent anti-leukemic properties across multiple clinical settings. Its FDA approval for chronic myeloid leukemia (CML)—particularly in patients resistant or intolerant to multiple tyrosine kinase inhibitors (TKIs)—highlighted its value as a non–BCR-ABL–targeting therapeutic option. Unlike TKIs, which directly inhibit the catalytic activity of the BCR-ABL fusion protein, HHT exerts its effects through a distinct biological mechanism, primarily involving inhibition of protein synthesis via the ribosome. This unique mode of action accounts for its ability to bypass resistance pathways that commonly undermine TKI efficacy.

Despite its clinical success, the precise molecular mechanisms underlying HHT’s anti-leukemic activity have remained incompletely understood. Early studies emphasized its capacity to block peptide chain elongation, leading to reduced synthesis of short-lived oncoproteins such as MCL-1. However, these findings did not fully explain HHT’s selective efficacy against malignant cells or its ability to overcome resistance mutations like T315I, which severely limit the activity of most TKIs. As the complexity of cancer biology has become increasingly apparent, it is now recognized that effective therapies often act through multiple interconnected pathways rather than a single molecular target.

This realization has sparked renewed interest in using modern analytical tools—including proteomics, metabolomics, and systems biology—to uncover previously overlooked aspects of HHT’s activity. Recent investigations have identified potential mitochondrial involvement, metabolic stress responses, and alterations in oxidative phosphorylation (OXPHOS) associated with HHT treatment. These insights suggest that HHT may exert a broader impact on cellular homeostasis than previously appreciated, contributing to its ability to suppress highly resistant CML subtypes.

Understanding the full spectrum of HHT’s actions is more than an academic exercise—it is essential for refining therapeutic strategies, identifying biomarkers of response, and guiding rational combination therapies. As research continues to unveil new layers of HHT’s biological activity, the drug stands as a compelling example of how revisiting established therapeutics with modern technology can reveal transformative insights into cancer treatment mechanisms.

New Mechanistic Insight: HHT Disrupts Oxidative Phosphorylation in CML Cells

As research tools evolve, so does our understanding of how cancer therapies exert their effects beyond traditional signaling pathways. In the case of homoharringtonine (HHT), emerging studies have revealed an unexpected connection between its anti-leukemic activity and the disruption of oxidative phosphorylation (OXPHOS)—a central metabolic process that fuels cellular energy production. Using advanced proteomic profiling, investigators have identified OXPHOS as the most significantly altered pathway in BCR-ABL T315I–bearing CML cells treated with HHT. This discovery represents a major step forward in deciphering the drug’s multifaceted mechanism of action and opens new possibilities for targeting metabolic vulnerabilities in treatment-resistant CML.

OXPHOS, located within the inner mitochondrial membrane, is essential for generating ATP through electron transport and oxidative metabolism. Cancer cells, including leukemic cells, often rely on a flexible metabolic network that allows them to adapt to therapeutic pressure. Although the Warburg effect historically emphasized glycolysis as the dominant metabolic pathway in cancer, recent research has demonstrated that many malignancies—including subsets of leukemia—retain a strong dependence on mitochondrial respiration. This is particularly true for drug-resistant cancer stem cells, which use elevated OXPHOS activity to sustain survival and evade therapy.

The proteomic analysis conducted in this study revealed substantial downregulation of multiple OXPHOS-associated proteins following HHT treatment, highlighting a global impairment of mitochondrial respiratory function. Notably, proteins localized to mitochondrial complex I, the first and largest enzyme complex of the electron transport chain, showed pronounced decreases. Since complex I serves as the entry point for electron transfer and plays a pivotal role in ATP production, its disruption can severely compromise cellular metabolism.

These findings point to a novel explanation for HHT’s potency against T315I-resistant CML cells: rather than solely blocking protein synthesis, HHT appears to induce metabolic collapse by impairing mitochondrial respiration. This is particularly significant because mitochondrial targeting represents an emerging therapeutic strategy for overcoming TKI resistance, as resistant leukemic cells often upregulate OXPHOS to compensate for inhibited oncogenic signaling.

Fig.1 HHT Targets Mitochondrial Complex I to Halt Energy Production in CML

By uncovering OXPHOS dysregulation as a central consequence of HHT treatment, this research reframes the drug as not only a protein synthesis inhibitor but also a mitochondrial disruptor, expanding our understanding of its therapeutic reach and highlighting new metabolic avenues for intervention in resistant CML.

Mitochondrial Complex I Inhibition: The Key to HHT’s Anti-Leukemic Power

The emerging evidence that homoharringtonine (HHT) disrupts oxidative phosphorylation (OXPHOS) has shifted attention toward a more specific mitochondrial target: mitochondrial complex I (MCI). As the first and largest enzyme of the electron transport chain, complex I is responsible for initiating electron transfer from NADH to ubiquinone, driving proton pumping and ultimately powering ATP synthesis. Its central role in cellular energy production makes it a critical determinant of metabolic fitness, particularly in cancer cells that depend on mitochondrial respiration for survival under therapeutic stress. The discovery that HHT significantly downregulates complex I proteins and impairs its activity offers a compelling mechanistic explanation for the drug’s powerful anti-leukemic effects.

Proteomic analyses revealed a marked decline in several complex I subunits following HHT treatment, indicating that the drug may interfere with complex I assembly, stability, or expression. This disruption was further supported by Western blot validation, confirming the loss of key structural components associated with complex I function. Given that T315I mutant CML cells often adopt a high-OXPHOS metabolic state to compensate for inhibited BCR-ABL signaling, the impairment of complex I places direct pressure on their metabolic resilience.

Functionally, inhibition of complex I by HHT leads to sharp reductions in ATP levels and oxygen consumption rate (OCR)—two essential readouts of mitochondrial health. ATP depletion compromises energy-intensive cellular processes, while decreased OCR signals a breakdown in electron transport chain activity. Together, these effects generate profound metabolic stress that leukemic cells are poorly equipped to withstand, especially those already burdened by oncogenic and therapeutic pressures. The resulting energy crisis contributes to reduced proliferation, increased apoptosis, and overall suppression of CML cell viability.

The significance of this mechanism becomes even clearer when viewed in the context of drug-resistant disease. T315I mutant cells, which evade most TKIs through structural changes in the BCR-ABL kinase, remain vulnerable to mitochondrial disruption because this metabolic dependency is unaffected by their mutation status. By bypassing the kinase altogether and targeting a universal energy-producing system, HHT exploits a critical weakness that resistant cells cannot easily escape.

Altogether, the identification of mitochondrial complex I as a major functional target of HHT underscores the drug’s potential as a metabolic therapy for CML. This mechanistic insight not only enhances our understanding of HHT’s therapeutic action but also points toward combination strategies that synergize with mitochondrial inhibition to overcome resistance more effectively.

Implications for Future CML Treatment: A Novel Strategy Against T315I Resistance

The identification of mitochondrial complex I inhibition as a central mechanism of homoharringtonine (HHT) action marks a significant advancement in the ongoing effort to overcome therapy resistance in chronic myeloid leukemia (CML). As T315I and other resistant mutations continue to challenge the effectiveness of tyrosine kinase inhibitors (TKIs), the need for alternative strategies that bypass the BCR-ABL kinase altogether has become increasingly urgent. HHT’s ability to induce metabolic collapse in leukemic cells, regardless of their mutation status, positions it as a valuable therapeutic tool in the evolving landscape of CML treatment.

One of the most promising implications of this discovery is the potential integration of metabolic targeting into CML therapy. For years, treatment development has focused primarily on kinase inhibition, leaving mitochondrial vulnerabilities largely unexplored. The new insight that T315I mutant cells may rely heavily on oxidative phosphorylation (OXPHOS) for survival suggests that disrupting energy production could serve as a complementary—or in some cases alternative—approach to traditional TKI therapy. HHT’s dual ability to inhibit protein synthesis and impair mitochondrial complex I provides a multifaceted attack on leukemic cell viability, widening the therapeutic window for resistant disease.

Another important consideration is the expansion of combination therapy strategies. Metabolic inhibitors, when paired with TKIs or other targeted agents, may enhance treatment efficacy by simultaneously suppressing oncogenic signaling and energy metabolism. Preclinical studies have shown that mitochondrial inhibitors can sensitize resistant leukemia cells to existing therapies, raising the possibility that HHT could serve as a backbone for combination regimens designed to prevent or overcome resistance.

Moreover, these findings may guide future research aimed at developing next-generation metabolic therapies specifically tailored for hematologic malignancies. Understanding how CML cells adapt their metabolism under drug pressure can help identify biomarkers that predict responsiveness to mitochondrial inhibitors, enabling more personalized treatment approaches.

Ultimately, the discovery of HHT’s impact on mitochondrial complex I not only reveals a previously overlooked therapeutic mechanism but also redefines how resistant CML might be treated. By shifting the focus toward metabolic dependency, this research paves the way for innovative strategies that transcend traditional kinase-centric therapies and offer renewed hope for patients facing T315I-driven resistance.