Ferroptosis, triggered by aberrant polyunsaturated phospholipid peroxidation, contributes to the development of various diseases including neurodegeneration, ischemia/reperfusion-induced damages in the liver, kidney, heart and brain, and hemolysis. Vulnerability to ferroptosis induction is also a common feature of various cancers including clear-cell carcinomas from the kidney and ovary, pancreatic cancer, diffuse large B-cell lymphoma, hepatocellular carcinoma, colorectal cancer, and therapy-resistant cancer cells. Hence, inducing polyunsaturated phospholipid peroxidation and ferroptosis in human tumor cells has emerged as a promising strategy for cancer therapy. This concept is supported by the tumor suppressive effects of ferroptosis-inducing agents including imidazole ketone erastin (IKE) and its combination with other targeted therapies, recombinant cysteinase, and direct administration of polyunsaturated fatty acids. However, rapidly stratifying cancer patients for their likelihood to respond to ferroptosis-inducing therapies remains a major challenge in developing ferroptosis-targeted anti-cancer treatments. This issue is further highlighted by the facts that no specific primary cancer lineage or genotype is predictive of ferroptosis responsiveness, and biomarkers that are indicative of ferroptosis sensitivity in vivo remain largely unavailable.

On November 22, Westlake Laboratory principal investigator Yilong Zou, in collaboration with principal investigator Stuart Schreiber of Broad Institute, presented their work in Cell Chemical Biology entitled “PALP: A rapid imaging technique for stratifying ferroptosis sensitivity in normal and tumor tissues in situ”.

They established and presented a technique termed photochemical activation of membrane lipid peroxidation (PALP) to detect polyunsaturated phospholipids as well as report ferroptosis sensitivity in live cells and tissues in situ (Figure 1). Specifically, they used high-power laser pulses to induce localized lipid peroxidation in membranes, which subsequently resulted in oxidation of a fluorescent BODIPY-C11 probe. They envisioned that the PALP techniques could have broad utilities in basic and clinical research involving polyunsaturated lipid peroxidation and metabolism as well as facilitating the development of ferroptosis-targeted therapies.

Figure 1. Scheme for photochemical activation of membrane lipid peroxidation technique.

PALP technique can report ferroptosis sensitivity in cells using targeted lasers to induce localized polyunsaturated fatty acyl (PUFA)-lipid peroxidation including human renal carcinoma cells, ovarian carcinoma cells, mouse primary neurons and mouse renal epithelial cells etc. PALP-induced fluorescent signals can be suppressed by lipophilic antioxidants and iron chelation, demonstrating that PALP-induced signals depend on PUFA-lipid levels (Figure 2). The polyunsaturated fatty acid content is the major factor in ferroptosis. This team selected four ovarian cancer cell linesthat exhibit a wide range of sensitivity to ferroptosis inducers in vitro. The subcutaneously implanted xenograft ovarian tumor exhibited the same ranking of PALP-induced signals and PUFA-lipid contents using spatial metabolomics approach, supportingthat PALPv1 is likely useful for stratifying ferroptosis sensitivity in tumors in situ in clinically relevant settings (Figure 3). 

Figure 2. PALP-induced signals depend on cellular polyunsaturated fatty acyl lipid levels.

Figure 3. PALP technique enables stratifying ferroptosis sensitivity in tumors in situ.

The PALP technique may be useful for predicting ferroptosis susceptibility in tissue samples to guide patient selection for ferroptosis-targeted therapies, and for characterizing polyunsaturated lipid metabolism during normal development, aging, tumorigenesis, and other relevant disease settings. Given the relative simplicity of the technique, they proposed that PALP might additionally be applicable to the following research settings: 1) to study the mechanisms of ferroptosis at high spatio-temporal resolution and to detect polyunsaturated phospholipids in heterogeneous cell populations; 2) to evaluate how novel chemicals and genetic perturbations affect the dynamics of lipid peroxidation in cells and tissues; 3) to assess the functions and dynamics of polyunsaturated phospholipids in aging-related diseases such as Alzheimer’s Disease and organ degeneration. Overall, the PALP technique series will be widely useful for studying ferroptosis, lipid metabolism, membrane lipid damage and repair, and human diseases associated with aberrant lipid metabolic states.