Journal: bioRxiv

Article Title: Pitavastatin counteracts venetoclax resistance mechanisms in acute myeloid leukemia by depleting geranylgeranyl pyrophosphate

doi: 10.1101/2025.10.27.684888

Figure Lengend Snippet: PUMA upregulation contributes to pitavastatin cytotoxicity, is dependent on GGPP depletion, and is recapitulated by inhibition of GGTase-1. (A) The cytotoxic effect of pitavastatin +/-30nM venetoclax was measured in two clones of OCI-AML3 cells with PUMA knockout and one clone each of MOLM14 and MOLM13. Two-way ANOVA with Tukey’s post hoc test, n = 3. (B) PUMA expression in a statin-sensitive AML cell line (OCI-AML3) was assessed by western blot. An antibody to vinculin was used as a loading control. Detection of unprenylated RAP1A was used to determine which treatments suppress protein geranylgeranylation. Note rescue of PIT effect on PUMA by mevalonate (MEV) and GGPP more than by FPP. (C) Quantitation of four replicates of the experiment in panel B, with PUMA expression in vehicle (Veh) treated cells set at 1. Data are expressed as mean +/-SEM ** p < 0.01, ***p < 0.001, one-way ANOVA with Tukey’s post hoc test, n = 2-4. (D-E) Different GGTase-1 inhibitors have distinct effects on PUMA expression and viability. (D) Representative western blot of PUMA expression in OCI-AML3 and MOLM13 treated with pitavastatin (0.3 or 1 µM), GGTI298 (10 µM) or GGTI2133 (10 µM). (E) Graphs showing percentage of viable OCI-AML3 and MOLM13 measured by Annexin V and PI staining after 48h of treatment with pitavastatin (1 µM+, GGTI298 (10 µM) or GGTI2133 (10 µM) in combination with venetoclax. Data are expressed as mean +/-SEM ***p < 0.001, ****p < 0.0001. Two-way ANOVA with Tukey’s post hoc test, n ≥ 3.

Article Snippet: The filters were blocked in 5% milk for 1h at room temperature and then incubated overnight at 4° with specific antibodies: PUMA (1:1000 Cell Signaling #12450), unprenylated RAP1A (1:50 Santa Cruz sc-373968), p44/42 MAPK (Erk1/2) (1:1000 Cell Signaling #4696), c-Myc (1:1000 Cell Signaling #18583) Vinculin (1:1000 Cell Signaling #13901), GILZ (1:1000 ThermoFisher Scientific # 12352-1-AP), Mcl1 (1:1000 Cell Signaling #39224).

Techniques: Inhibition, Clone Assay, Knock-Out, Expressing, Western Blot, Control, Quantitation Assay, Staining

Journal: bioRxiv

Article Title: Pitavastatin counteracts venetoclax resistance mechanisms in acute myeloid leukemia by depleting geranylgeranyl pyrophosphate

doi: 10.1101/2025.10.27.684888

Figure Lengend Snippet: Pitavastatin treatment downregulates cMYC expression and increases PUMA and GILZ in AML cell lines and primary cells. (A) Two PIT-sensitive cell lines, one PIT-resistant cell line, and two PIT-sensitive primary AML samples were treated for 16hr as indicated and lysates subjected to western blotting with antibodies to cMYC, PUMA, GILZ, unprenylated RAP1A, and total ERK. (B) Graph of quantitation of western blots from 3-4 experiments for cell lines in panel A, or single experiment for two primary cell samples UCI-01 and UCI-06. Data are expressed as mean +/- SEM *p < 0.05, ***p < 0.001, ****p < 0.0001 two-way ANOVA with Dunnett’s test. (C) PIT and GGTI-298 cause G1 arrest in OCI-AML3 cells that are statin sensitive and show consistent MYC downregulation, but not in HNT-34 cells where MYC downregulation is less prominent. Data are expressed as mean +/- SEM *p < 0.05, ** p < 0.01, ****p < 0.0001 two-way ANOVA with Tukey’s test, n = 3.

Article Snippet: The filters were blocked in 5% milk for 1h at room temperature and then incubated overnight at 4° with specific antibodies: PUMA (1:1000 Cell Signaling #12450), unprenylated RAP1A (1:50 Santa Cruz sc-373968), p44/42 MAPK (Erk1/2) (1:1000 Cell Signaling #4696), c-Myc (1:1000 Cell Signaling #18583) Vinculin (1:1000 Cell Signaling #13901), GILZ (1:1000 ThermoFisher Scientific # 12352-1-AP), Mcl1 (1:1000 Cell Signaling #39224).

Techniques: Expressing, Western Blot, Quantitation Assay