Targeting FSP1 triggers ferroptosis in lung cancer - PubMed (original) (raw)

[Preprint]. 2025 Aug 23:2025.08.07.668766.

doi: 10.1101/2025.08.07.668766.

Alec J Vaughan 1 2, Jozef P Bossowski 1 3, Yuan Hao 1 3, Aikaterini Ziogou 1 3, Seon Min Kim 4, Tae Ha Kim 4, Mari N Nakamura 1 3, Ray Pillai 1 5 6, Mariana Mancini 1 3, Sahith Rajalingam 1 3, Mingqi Han 7 8, Toshitaka Nakamura 9, Lidong Wang 10, Suckwoo Chung 1 3, Diane Simeone 10, David Shackelford 7 8, Yun Pyo Kang 4, Marcus Conrad 9 11, Thales Papagiannakopoulos 1 3

Affiliations

Targeting FSP1 triggers ferroptosis in lung cancer

Katherine Wu et al. bioRxiv. 2025.

Update in

Abstract

Pre-clinical and clinical studies have demonstrated how dietary antioxidants or mutations activating antioxidant metabolism promote cancer, highlighting a central role oxidative stress in tumorigenesis. However, it is unclear if oxidative stress ultimately increases to a point of cell death. Emerging evidence indicates that cancer cells are susceptible to ferroptosis, a form of cell death triggered by uncontrolled lipid peroxidation1-3. Despite broad enthusiasm about harnessing ferroptosis as a novel anti-cancer strategy, whether ferroptosis is a barrier to tumorigenesis and if it can be leveraged therapeutically remains unknown4,5. Using genetically-engineered mouse models (GEMMs) of lung adenocarcinoma (LUAD), we performed tumor specific loss-of-function studies of the two key ferroptosis suppressors, glutathione peroxidase 4 (Gpx4)6,7 and ferroptosis suppressor protein 1 (Fsp1)8,9, and observed increased lipid peroxidation and robust suppression of tumorigenesis, suggesting that lung tumors are highly sensitive to ferroptosis. Furthermore, across multiple pre-clinical models, we found that FSP1 was required for ferroptosis protection in vivo, but not in vitro, underscoring a heightened need to buffer lipid peroxidation under physiological conditions. Lipidomic analyses revealed that Fsp1-knockout (Fsp1KO) tumors had an accumulation of lipid peroxides, and inhibition of ferroptosis with genetic, dietary, or pharmacological approaches effectively restored the growth of Fsp1KO tumors in vivo. Unlike GPX4, FSP1 expression was prognostic for disease progression and poorer survival in LUAD patients, highlighting its potential as a viable therapeutic target. To this end, we demonstrated that pharmacologic inhibition of FSP1 had significant therapeutic benefit in pre-clinical lung cancer models. Our studies highlight the importance of ferroptosis suppression in vivo and pave the way for FSP1 inhibition as a therapeutic strategy in lung cancer patients.

Keywords: FSP1; GPX4; ferroptosis; lung cancer.

PubMed Disclaimer

Conflict of interest statement

T.P. received funding from Pfizer Medical Education Group, Dracen Pharmaceuticals, Kymera Therapeutics, Bristol Myers Squibb, and Agios Pharmaceuticals not related to the submitted work. M.C. is a co-founder and shareholder of ROSCUE Therapeutics GmbH. M.C. and T.N. have filed a patent (WO2024115673A1) for some of the FSP1 inhibitor compounds described herein. All other authors declare no competing interests.

Figures

Extended Data Fig. 1:

Extended Data Fig. 1:. Gpx4 is required by lung cancer cells

(a) Quantification and representative images of Gpx4 IHC in KP LUAD GEMM tumors with knockout of either control (Neo, n=11) or Gpx4 (n=12). Scale bars: 200μm. (b) Representative 4-hydroxy-2-noneal (4-HNE) IHC of liver tissue from conditional _Gpx4_-knockout mice. Scale bars: 100μm. (c) Top: Western blot of KP LUAD cells with CRISPR/Cas9-mediated genetic deletion of Gpx4 with either two individual or duplexed sgRNAs. Bottom: Representative images of crystal violet clonogenic assay of KP LUAD cells with knockout of either control (Neo) or Gpx4. Cells were treated with 100nM LIP1. (d) Heatmap of LC-MS detection of oxidized phospholipids in KP LUAD cells treated with DMSO control, RSL3 (0.5μM), and RSL3 (0.5μM) + LIP1 (100nM) for 8 hours. (e) Schematic of Gpx4 ectopic overexpression (OE) method in KP LUAD cells. Western blot and representative images of crystal violet clonogenic assay of KP LUAD cells with wildtype (WT) or OE of Gpx4. (f) CellTiter-Glo Luminescence viability assay of KP, Gpx4WT or Gpx4OE cells upon increasing concentrations of RSL3 (n=5 per group). (g) Western blot of KP LUAD cells treated with 20nM Na2SeO3 or DMSO. CellTiter-Glo Luminescence viability assay of KP LUAD cells treated with 20nM Na2SeO3 or DMSO with increasing RSL3 addition (n=5 per group). (h) GPX4 expression in _KRAS_-mutant primary LUAD tumors from TCGA, divided into early and late tumor stages (normal lung, n=54; stage I/II, n=354; stage III/IV, n=98). (i) Overall survival of _KRAS_-mutant LUAD patients (n=464) from TCGA, stratified by high vs low tumor GPX4 expression. (j) Tumor Gpx4 IHC quantification of KP LUAD GEMM tumors at 10 weeks (n=4) vs 14 weeks (n=4) post-tumor initiation. Box plots indicate median (middle line), 25th, 75th percentile (box) and 5th and 95th percentile (whiskers). Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panel h), two-sided student’s t-test (panel j) or Kaplan-Meier simple survival analysis (panel i). For gel source data, see Supplementary Data 1.

Extended Data Fig. 2:

Extended Data Fig. 2:. FSP1’s anti-ferroptotic function is not dependent on oncogenic signaling

(a) FSP1 (AIFM2) expression of _KRAS_-mutant primary LUAD tumors from TCGA, divided into early and late tumor stages (normal lung, n=54; stage I/II, n=354; III/IV, n=98). (b) FSP1 (AIFM2) expression of _KRAS_-mutant primary LUAD tumors from TCGA, separated by the KRAS mutation (G12C n=51; G12V n=32; G12D n=17; G12A n=16; other n=20). (c) FSP1 (AIFM2) expression of primary LUAD tumors from TCGA, separated by oncogenic driver mutation (normal lung, n=59; EGFR, n=71; KRAS, n=135; other n=307). (d) Western blot of KP LUAD human cell lines treated with 50nM RMC-042 for indicated durations. (e) Representative multi-IF images of KP tumors for markers indicated. Panels are 10X, scale bars: 50μm; insets are 20X, scale bars: 20μm. (f) FSP1 (AIFM2) expression of _KRAS_-mutant primary LUAD tumors from TCGA, separated by tumor co-mutation status (KEAP1/STK11 WT n=88; STK11 mutant n=20; KEAP1 mutant n=16; KEAP1/STK11 mutant n=11). (g) FSP1 (AIFM2) expression in KP LUAD human cell lines treated with an Nrf2 activator, KI696, for 5 days (n=3 per group). Box plots indicate median (middle line), 25th, 75th percentile (box) and 5th and 95th percentile (whiskers). Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panels a, b, c, f, g). For gel source data, see Supplementary Data 1.

Extended Data Fig. 3:

Extended Data Fig. 3:. Neither Fsp1 nor Gpx4 have an impact on tumor initiation, proliferation, or apoptosis

(a) Quantification and representative images of tumor Fsp1 IHC in KP LUAD GEMMs with knockout of either control (Neo, n=11) or Fsp1 (n=14). Scale bars: 100μm. (b) Tumor number quantification in KP LUAD GEMMs with tumor-specific knockout of either control (Neo, n=4), Fsp1 (n=7), or Gpx4 (n=6). (c) Individual area of KP LUAD GEMM tumors with knockout of either control (Neo, n=465), Fsp1 (n=453), or Gpx4 (n=384). (d) Quantification of Ki67 IHC of KP LUAD GEMM tumors with knockout of either control (Neo, n=355), Fsp1 (n=268), or Gpx4 (n=291). (e) Quantification of cleaved caspase-3 IHC of KP LUAD GEMM tumors with knockout of either control (Neo, n=46), Fsp1 (n=22), or Gpx4 (n=52). Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panels b-e).

Extended Data Fig. 4:

Extended Data Fig. 4:. Fsp1 is required for cell-autonomous tumor growth in vivo

(a) Top: Western blot of KP LUAD cells with CRISPR/Cas9-mediated genetic deletion of Fsp1 with either two individual or duplexed sgRNAs. Bottom: Representative images of crystal violet clonogenic assay of KP LUAD cells with knockout of either control (Neo) or Fsp1. Cells were treated with 100nM LIP1. (b) Schematic depicting all transplantation models performed using isogenic KP, Fsp1KO and Fsp1WT. (c, d) Longitudinal growth and endpoint tumor weights of KP sg_Fsp1_ (n=8) versus control (sg_Neo_ n=10) subcutaneous (subQ) xenograft tumor transplanted into C57BL/6J male mice. (e) Longitudinal lung tumor growth (measured via bioluminescence) in C57BL/6J male mice orthotopically transplanted with KP LUAD cells with CRISPR/Cas9-mediated knockout of Fsp1 (n=8) or control (n=7). (f) Longitudinal lung tumor growth (measured via bioluminescence normalized to first timepoint (day 7)) in C57BL/6J male mice orthotopically transplanted with isogenic KP, Fsp1KO (n=6) and Fsp1WT (n=7) cells. (g) Longitudinal lung tumor growth (measured via absolute bioluminescence) in NU/J immunocompromised mice orthotopically transplanted with isogenic KP, Fsp1KO (n=6) and Fsp1WT (n=7) cells. (h) Longitudinal lung tumor growth (measured via absolute bioluminescence) in C57BL/6J female mice with orthotopic transplantation of isogenic KP, Fsp1KO (n=8) and Fsp1WT (n=8) cells. (i) Longitudinal tumor growth and endpoint tumor weights of either isogenic KP, Fsp1KO or Fsp1WT subcutaneous (subQ) xenograft tumors transplanted in either C57BL/6J WT (Fsp1KO, male n=10 female n=10; Fsp1WT, male n=8, female n=10) or C57BL/6J _Fsp1_-knockout mice (Fsp1KO, males n=12, females n=12; Fsp1WT, male n=10, female n=10). (j) Western blot of KP LUAD cells with wildtype (WT) or overexpression (OE) of Fsp1. Representative images of crystal violet clonogenic assay of KP LUAD cells with Fsp1WT or Fsp1OE. (k) CellTiter-Glo Luminescence viability assay of KP, Fsp1WT or Fsp1OE cells (5 biological replicates per group) with increasing concentrations of RSL3. (l, m) Longitudinal growth and endpoint weights of KP Fsp1WT (n=10) or Fsp1OE (n=10) subcutaneous (subQ) xenograft tumor transplanted into C57BL/6J male mice. Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panel i), two-way ANOVA with Tukey’s multiple comparisons test (panels c, e, f, g, h, l), or two-sided student’s t-test (panels d, m). For gel source data, see Supplementary Data 1.

Extended Data Fig. 5:

Extended Data Fig. 5:. FSP1 loss in vitro does not induce ferroptosis without RSL3

(a) Top: Western blot; Bottom: baseline growth (normalized to control); and Right: CellTiter-Glo Luminescence viability assay of H2009 cells with the lentiviral addition of Cas9 and guide RNAs targeting FSP1 or a non-targeting control (TOM) (at least 4 biological replicates per group). Viability assay is upon increasing concentrations of RSL3. (b) As in (a), but with H1299 cells. (c) As in (a), but with PC9 cells. (d) As in (a), but with H1975 cells. (e) As in (a), but with A549 cells. (f) As in (a), but with 16645 cells. (g) As in (a), but with KPC7 cells. Data are represented as mean values, error bars represent SEM, significance determined via two-sided student’s t-test (panels a-g). For gel source data, see Supplementary Data 1.

Extended Data Fig. 6:

Extended Data Fig. 6:. Fsp1 loss promotes tumor cell ferroptosis

(a) Western blot of KP, Gpx4KO LUAD cells with either empty vector (—) or Fsp1OE. (b) Western blot of KP, Fsp1KO LUAD cells with re-expression of either empty vector (—), Fsp1WT, Gpx4OE. (c) CellTiter-Glo Luminescence viability assay of KP, Fsp1KO LUAD cells in (b) upon increasing concentrations of RSL3. (d) Endpoint tumor weights of subcutaneous (subQ) xenograft tumors from indicated cell lines transplanted in C57BL/6J female mice (control n=10; Fsp1WT, n=9; Gpx4OE; n=8). (e) Top: Western blot of KP, Fsp1KO LUAD cells with re-expression of either empty vector (—), wildtype (WT) Fsp1, or mutant (mut) Fsp1. Bottom: Representative images of crystal violet clonogenic assay. (f) CellTiter-Glo Luminescence viability assay of KP, Fsp1KO LUAD cells with re-expression of either empty vector (Fsp1KO), Fsp1WT, or Fsp1mut upon increasing concentrations of RSL3. LIP1 used at 100nM. (g) Longitudinal tumor growth (measured via bioluminescence normalized to initial imaging timepoint (day14)) in C57BL/6J male mice orthotopically transplanted with isogenic KP, Fsp1KO LUAD cells with re-expression of either empty control vector (Fsp1KO, n=7), Fsp1WT (n=7), or Fsp1mut (n=7). (h) Representative images of crystal violet clonogenic assay of KP, Fsp1WT or Fsp1KO LUAD cells with CRISPR/Cas9- mediated knockout of control (Neo) or Acsl4. (i) Longitudinal tumor growth (measured by bioluminescence) of C57BL/6J male mice with orthotopic transplantation of isogenic KP, Fsp1KO and Fsp1WT cells with CRISPR/Cas9-mediated knockout of control (Neo) or Acsl4 (WT, n=5; Acsl4KO, n=4; Fsp1KO, n=7; Fsp1KOAcsl4KO, n=7). (j) Representative H&E of experiment in (i). Scale bars: 1000μm. (k) Overall survival of C57BL/6J male mice with orthotopic transplantation of isogenic KP, Fsp1KO and Fsp1WT cells with CRISPR/Cas9-mediated knockout of control (Neo) or Acsl4 (n=5 per group). (l) Longitudinal tumor growth (measured via bioluminescence normalized to first imaging timepoint (day7)) of C57BL/6J mice orthotopically transplanted with KP, Fsp1WT cells and receiving high (n=6) or low (n=5) VitE diets ad libitum 5 days pre-tumor initiation. (m) Tumor bioluminescence signal normalized to first imaging timepoint (day7) of C57BL/6J mice orthotopically transplanted with KP, Fsp1KO cells and receiving Liproxstatin-1 (LIP1, n=7) or Vehicle (Veh, n=5) daily after tumor establishment. (n) Schematic of KP LUAD GEMMs intratracheally infected with pUSEC lentiviruses containing dual sgRNAs targeting Fsp1. Mice were dosed with LIP1 (n=6) or Vehicle (Veh, n=6) every other day starting from tumor initiation. (o) Tumor burden of KP LUAD GEMMs with CRISPR/Cas9-mediated knockout of Fsp1 and treated with Veh or LIP1. (p) Representative H&E and 4-HNE IHC of KP LUAD GEMM tumors with knockout of Fsp1 and treated with Veh or LIP1. Scale bars: 1000μm. Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA (panel d), two-way ANOVA with Tukey’s multiple comparisons (panels g, i, l, m), two-sided student’s t-test (panel o), or Kaplain-Meier simple survival analysis (panel k). For gel source data, see Supplementary Data 1.

Extended Data Fig. 7:

Extended Data Fig. 7:. icFSP1 exerts on-target tumor-suppressive effect

(a) Top: Western blot of KP, Fsp1KO LUAD cells with re-expression of either empty vector (—), wildtype (WT) hFSP1, or Q319K-mutant hFSP1. Bottom: Representative images of crystal violet clonogenic assay. (b) Cell-TiterGlo Luminescence viability assay of KP, Fsp1KO LUAD cells with re-expression of either empty vector (—), WT hFSP1, or Q319K-mutant hFSP1. (c) CellTiter-Glo Luminescence viability assay of human or mouse Fsp1WT-expressing cells upon RSL3 (0.2μM) plus increasing concentrations of iFSP1. (d) CellTiter-Glo Luminescence viability assay as in (c) but with FSEN1. (e) CellTiter-Glo Luminescence viability assay of human or mouse Fsp1WT-expressing cells upon increasing concentrations of icFSP1. (f) CellTiter-Glo Luminescence viability assay of WT or Q319K-mutant hFSP1-expressing cells upon RSL3 (0.2μM) plus increasing concentrations of icFSP1. (g) Longitudinal tumor growth (measured via bioluminescence normalized to first imaging timepoint (day7)) in C57BL/6J mice orthotopically transplanted with isogenic KP, Fsp1KO cells with re-expression of either human FSP1 (hFSP1, n=5), mouse Fsp1 (mFsp1, n=6), or control (n=6) vector. (h) Tumor burden and representative H&E of KP, Fsp1KO (n=4) and hFSP1WT (n=4) orthotopic lung tumors. Scale bars: 1000μm. (i) Representative H&E of KP, hFSP1WT orthotopic lung tumors from indicated treatment groups. Scale bar: 2000μm. (j) Percent of T cells, alveolar macrophages, interstitial macrophages, and neutrophils of total CD45+ cells in tumor bearing lungs from mice treated with the indicated compounds for two weeks (Veh, n=7; LIP1, n=7; icFSP1, n=9; icFSP1+LIP1, n=6). (k) Graphical summary depicting ferroptosis as a barrier to lung cancer, as loss of either Fsp1 or Gpx4 induces tumor cell ferroptosis and restricts disease progression. (l) Summary table of cell lines, their respective tumor lineage, and mutations where Fsp1 loss resulted in tumor suppression in vivo. For all Cell-TiterGlo assays, n=5 biological replicates per group. Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panels g, j) or student’s t test (panel h). For gel source data, see Supplementary Data 1.

Fig. 1:

Fig. 1:. Gpx4 loss triggers ferroptosis in lung tumors

(a) Schematic of KP LUAD GEMMs intratracheally infected with pUSEC lentiviruses containing dual sgRNAs targeting Neo (control; n=13) or Gpx4 (n=13). Mice were dosed with Liproxstatin-1 (LIP1; sg_Neo_ n=6, sg_Gpx4_ n=7) or Vehicle (Veh; sg_Neo_ n=6, sg_Gpx4_ n=7) every other day starting from tumor initiation to experiment endpoint. (b) Tumor burden of KP LUAD tumors with control (sg_Neo_) or Gpx4 (sg_Gpx4_) knockout, treated with either Veh or LIP1. (c) Representative MRI of KP LUAD tumors with control (sg_Neo_) or Gpx4 (sg_Gpx4_) knockout, treated with either Veh or LIP1. (d) Representative H&E of KP LUAD tumors with control (sg_Neo_) or Gpx4 (sg_Gpx4_) knockout, treated with either Veh or LIP1. Scale bars: 2000μm. (e) Tumor 4-HNE expression by IHC of KP LUAD tumors with control (sg_Neo_) or Gpx4 (sg_Gpx4_) knockout, treated with either Veh or LIP1. (f) Representative 4-HNE IHC staining from (e). Scale bars: 20μm. Data are represented as mean values, error bars represent SEM, significance determined via one-way ANOVA with multiple comparisons (panels b, e).

Fig. 2:

Fig. 2:. Fsp1 knockout robustly restricts lung tumorigenesis

(a) Schematic depicting mechanisms of Fsp1- and Gpx4-mediated buffering against cellular lipid peroxidation and ferroptosis. (b) Overall survival of _KRAS_-mutant LUAD patients (n=280) from TCGA, stratified by high versus low primary tumor FSP1 (AIFM2) expression. Median survival for FSP1 high versus low was 1215 days and 1498 days, respectively (hazard ratio 1.51 [1.01–2.25]). (c) Representative IHC for Fsp1 and Gpx4 in KP LUAD GEMM adenomas versus adenocarcinomas. Scale bars: 100μm. (d) Tumor Fsp1 expression by IHC of KP LUAD GEMM tumors at 10- versus 14-weeks post tumor initiation. (e) Schematic of KP LUAD GEMMs intratracheally infected with pUSEC lentiviruses containing double sgRNAs targeting control (Neo; n=4), Fsp1 (n=6), or Gpx4 (n=6). (f) Tumor burden of KP LUAD tumors with knockout of either control (Neo), Fsp1, or Gpx4. (g) Representative H&E of KP LUAD tumors with knockout of either control (Neo), Fsp1, or Gpx4. Scale bars: 2000μm. (h) Relative proportion of KP adenomas versus adenocarcinomas with knockout of either control (Neo), Fsp1, or Gpx4. (i) Representative TUNEL staining of KP, Fsp1KO and Fsp1WT orthotopic lung tumors. Scale bars: 50μm (j) Schematic depicting generation of isogenic Fsp1-wildtype (Fsp1WT) versus Fsp1-knockout (Fsp1KO) cells for paralleled in vitro assays and in vivo syngeneic orthotopic transplantation studies. (k) Representative images of crystal violet clonogenic growth assay in isogenic KP, Fsp1KO and Fsp1WT cells treated with RSL3 (0.5μM) ± LIP1 (100nM). (l) Tumor burden of KP, Fsp1KO (n=8) and Fsp1WT (n=7) orthotopic lung tumors. (m) Representative H&E of KP, Fsp1KO and Fsp1WT orthotopic lung tumors. Scale bars: 2000μm. Data are represented as mean values, error bars represent SEM, significance determined via two-sided student’s t-test (panels d and l), one-way ANOVA with multiple comparisons (panel f) or Kaplan-Meier simple survival analysis (panel b).

Fig. 3:

Fig. 3:. Sensitivity to FSP1 loss is not dependent on tumor mutations or lineage

Longitudinal tumor growth and endpoint tumor weights from indicated cell lines with CRISPR/Cas9-mediated knockout of FSP1 (sg_FSP1_) or control (sg_TOM_, sg_Neo_), transplanted as subcutaneous (subQ) xenograft tumors into NSG mice: (a) H2009, sg_FSP1_ (n=8) or control (n=8). (b) H1299, sg_FSP1_ (n=9) or control (n=9). (c) PC9, sg_FSP1_ (n=9) or control (n=9). (d) H1975, sg_FSP1_ (n=8) or control (n=9). (e) A549, sg_FSP1_ (n=9) or control (n=10). (f) 16645, sg_Fsp1_(n=7) or control (n=9). (g) KPC7 (murine PDAC), sg_Fsp1_ (n=9) or control (n=10). Data are represented as mean values, error bars represent SEM, significance determined via two-sided student’s t test (panels a-g)

Fig. 4:

Fig. 4:. FSP1 is required for the suppression of ferroptosis in vivo

(a) Heatmap of indicated oxidized PE/PC lipid species detected via LC/MS from KP, Fsp1KO and Fsp1WT orthotopic lung tumors. (b) Longitudinal growth of KP, Gpx4KO subQ xenograft tumors with Fsp1OE (n=10) versus control (emptyOE, n=9) in C57BL/6J mice. (c) Longitudinal growth of KP, Fsp1KO subQ xenograft tumors with either Fsp1 restoration (Fsp1WT, n=9), Gpx4 overexpression (Gpx4OE, n=8), or controls (emptyOE, n=9; Fsp1KO n=8) in C57BL/6J mice. (d) Ratio of CoQ9H2/CoQ9 detected via LC/MS in KP, Fsp1KO and Fsp1WT orthotopic lung tumors versus cells treated with DMSO or RSL3 (0.5 μM) for 8 hours. (e) Schematic depicting Acsl4’s pro-ferroptotic function. (f) Tumor burden in C57BL/6J mice with KP, Fsp1KO and Fsp1WT orthotopic lung tumors with CRISPR/Cas9-mediated Acsl4 or control (Neo) deletion (WT n=5, Acsl4KO n=4, Fsp1KO n=7, Fsp1KOAcsl4KO n=7). (g) Schematic depicting dietary Vitamin E (VitE) manipulation studies in (h) and (i). (h) Longitudinal lung tumor growth (measured via bioluminescence normalized to first timepoint) in C57BL/6J mice orthotopically transplanted with KP, Fsp1KO cells and receiving high (n=8) or low (n=7) VitE diets ad libitum 5 days pre-tumor initiation. (i) Longitudinal lung tumor growth (measured via bioluminescence) in C57BL/6J mice orthotopically transplanted with KP, Fsp1KO cells and placed on high (n=6) or low (n=6) VitE diets ad libitum after tumor establishment (day18). (j) Schematic of KP LUAD GEMMs intratracheally infected with pUSEC lentiviruses containing double sgRNAs targeting Neo (control, n=12) or Fsp1 (n=12). Mice were dosed with Liproxstatin-1 (LIP1, n=6 per genotype) or Vehicle (Veh, n=6 per genotype) daily starting 5 weeks post-tumor initiation. (k) Tumor burden of experiment in (j). (l) Representative H&E of tumors of experiment in (j). Scale bars: 100μm. Data are represented as mean values, error bars represent SEM, significance determined via two-sided student’s t test (panel b, d, f, h, i, k) or two-way ANOVA with Tukey’s multiple comparison test (panel c).

Fig. 5:

Fig. 5:. FSP1 is a viable therapeutic target for _KRAS_-mutant lung adenocarcinoma

(a) Overall survival of C57BL/6J mice with orthotopic lung tumors with Fsp1KO (n=7) or hFSP1 re-expression, treated with either icFSP1 (n=8) or Vehicle (n=7). (b) Longitudinal lung tumor growth (measured via bioluminescence) in C57BL/6J mice orthotopically transplanted with hFSP1WT cells and treated with either Vehicle (n=8), LIP1 (n=7), icFSP1 (n=8), or icFSP1+LIP1 (n=8). (c) Overall survival of C57BL/6J mice with Fsp1KO (n=5), hFSP1WT, or hFSP1Q319K-expressing tumors, treated with icFSP1 (hFSP1WT, n=5; hFSP1Q319K, n=7) or Vehicle (hFSP1WT, n=7; hFSP1Q319K, n=7). (d) Longitudinal growth of PDX LX465 tumors treated with icFSP1 (n=10) or Vehicle (n=10) in NSG mice. Data are represented as mean values, error bars represent SEM, significance determined via two-way ANOVA with Tukey’s multiple comparisons test (panel b), two-sided student’s t-test (panel d), or Kaplan-Meier simple survival analysis (panels a, c).

References

    1. Dixon S.J., et al. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell 149, 1060–1072 (2012). -PMC -PubMed
    1. Viswanathan V.S., et al. Dependency of a therapy-resistant state of cancer cells on a lipid peroxidase pathway. Nature 547, 453–457 (2017). -PMC -PubMed
    1. Hangauer M.J., et al. Drug-tolerant persister cancer cells are vulnerable to GPX4 inhibition. Nature 551, 247–250 (2017). -PMC -PubMed
    1. Berndt C., et al. Ferroptosis in health and disease. Redox Biol 75, 103211 (2024). -PMC -PubMed
    1. Nakamura T. & Conrad M. Exploiting ferroptosis vulnerabilities in cancer. Nat Cell Biol (2024). -PubMed

Methods References

    1. Winslow M.M., et al. Suppression of lung adenocarcinoma progression by Nkx2–1. Nature 473, 101–104 (2011). -PMC -PubMed
    1. Criscuolo A., et al. Analytical and computational workflow for in-depth analysis of oxidized complex lipids in blood plasma. Nature Communications 13, 6547 (2022). -PMC -PubMed
    1. Goracci L., et al. Lipostar, a Comprehensive Platform-Neutral Cheminformatics Tool for Lipidomics. Analytical Chemistry 89, 6257–6264 (2017). -PubMed
    1. Adams K.J., et al. Skyline for Small Molecules: A Unifying Software Package for Quantitative Metabolomics. Journal of Proteome Research 19, 1447–1458 (2020). -PMC -PubMed
    1. Kang Y.P., Kim T.H., Ngoc Nguyen C.T., Kim S.M. & Kwon S.W. Robust Determination of Coenzyme Q10 Redox Status Using Two Isotope-Labeled Internal Standards. Available at SSRN 4982487.

Publication types

Grants and funding

LinkOut - more resources