2026 Theses Doctoral

A mitochondrial lipid-peroxidation-to-apoptosis signaling cascade centered on complex I

Zhu, Qiulin

Ferroptosis is an iron-dependent form of regulated cell death driven by lipid peroxidation of cellular membranes. While the core machinery governing ferroptosis, including the GPX4/glutathione axis and the FSP1/CoQ10 axis, has been substantially elucidated, how upstream mitochondrial determinants set ferroptosis susceptibility, and how ferroptotic stress interfaces with apoptotic execution, remain poorly understood. Identifying novel molecular nodes that govern ferroptosis sensitivity is essential for expanding the therapeutic landscape of this cell death modality in cancer.

In this thesis, I investigated the mechanism and molecular target of SGC-GAK-1, a small molecule originally used as a putative ADCK3 inhibitor to target CoQ₁₀ biosynthesis in cancer cells. This work originated from two exploratory projects pursued at the outset of my doctoral studies: a high-throughput screen of kinase inhibitors using TfR1 plasma membrane translocation as a ferroptosis-specific readout, and a hypothesis-driven approach targeting ADCK3-dependent CoQ₁₀ biosynthesis in breast cancer. While the kinase inhibitor screen identified EGFR inhibitors as ferroptosis sensitizers, discrepancies between pharmacological and genetic ADCK3 depletion, together with a striking concentration-dependent shift from ferroptosis to apoptosis, revealed that SGC-GAK-1 engages targets beyond ADCK3 and motivated the mechanistic studies that became the core of this thesis.

I discovered that SGC-GAK-1 induces a mitochondria-centered cell death program characterized by cardiolipin oxidation, CoQ₁₀ depletion, and a Ferrostatin-1–sensitive but Liproxstatin-1–resistant lipid peroxidation phenotype, establishing the first systematic demonstration that these two commonly used ferroptosis inhibitors are not functionally interchangeable. At higher concentrations, sustained mitochondrial stress activates a ceramide–sphingosine–VDAC signaling module that drives apoptotic execution downstream of the initial ferroptotic insult, defining a dose-dependent ferroptosis-to-apoptosis cascade.

Using thermal proteome profiling (CETSA), I identified the mitochondrial complex I accessory subunit NDUFA5 as the proximal molecular target engaged by SGC-GAK-1. Regulatory-network similarity analysis revealed that structurally diverse compounds can converge on this same mitochondrial stress state, and I discovered that PIM1-high, FLT3-ITD acute myeloid leukemia (AML) exhibits selective vulnerability to this NDUFA5-dependent death axis, validated in vitro cell line models. Collectively, these findings define a mitochondrial lipid peroxidation–to-apoptosis signaling axis centered on complex I, provide a framework for understanding how stress intensity tunes cell death outcomes, and reveal a therapeutically exploitable vulnerability in mitochondria-dependent malignancies.