Research ArticleCancer

Statins enhance efficacy of venetoclax in blood cancers

See allHide authors and affiliations

Science Translational Medicine  13 Jun 2018:
Vol. 10, Issue 445, eaaq1240
DOI: 10.1126/scitranslmed.aaq1240
  • Fig. 1 Statins selectively enhance the efficacy of venetoclax against blood cancer cells.

    (A) Viability of AML and DLBCL cell lines treated with increasing doses of venetoclax, simvastatin, or the combination for 48 hours. Concentrations for each drug are shown on the x axes for venetoclax (red) and simvastatin (blue). Significance testing was done using one-tailed paired Student’s t test comparing median inhibitory concentration (IC50) values of indicated treatment groups (n = 3). (B) Viability of primary murine lymphoma cells cocultured with irradiated 3T3 stroma and treated for 48 hours with indicated inhibitors ([venetoclax] = 10 nM for 27-L1 and 1000 nM for 27-L2). Significance testing was done using one-tailed paired t test on IC50 values (n = 3). (C) Viability of primary AML cells treated with simvastatin for 16 hours before addition of venetoclax for an additional 8 hours. Cell lines were classified as sensitized if their response to the combination exceeded the response to both single-agent treatments by greater than 10%. n = 1 for each AML sample. (D) Viability of primary CLL cells grown on NK.tert stroma and treated with simvastatin for 16 hours before addition of venetoclax for an additional 8 hours. One-way analysis of variance (ANOVA) with Tukey’s multiple comparisons test. n = 12. Data for individual samples is provided in fig. S4A. (E) Viability of PBMC subsets treated with simvastatin (3 μM), venetoclax (100 nM), or the combination for 48 hours before staining. Significance testing was done by two-tailed paired Student’s t test (n ≥ 3). (A to E) In this and other figures, *P < 0.05, **P < 0.01, ***P < 0.001. ns, not significant.

  • Fig. 2 DBP predicts sensitization to venetoclax by simvastatin.

    Dynamic BH3 profiles for all cells treated with 10 μM simvastatin or 50 nM BEZ235 for 16 hours. Significance testing was performed by two-tailed paired Student’s t test relative to vehicle-treated control samples. n = 4 (OCI-LY8) or 3 (SU-DHL4, OCI-LY1).

  • Fig. 3 Combination of simvastatin and venetoclax is effective in a syngeneic mouse model of lymphoma.

    (A) Percent lymphoma burden in indicated organs of mice treated with venetoclax (75 mg/kg per day, n = 5), simvastatin (50 mg/kg/ per day, n = 6), or the combination (n = 6) for 5 days. (B) Spleen weights for mice treated as in (A). Significance testing was done using unpaired one-tailed t tests. (C) Western blots for pharmacodynamic (PD) effect of simvastatin (accumulation of unprenylated RAP1A) in indicated organs of mice treated as in (A). Note that two spleens from the simvastatin-treated group were lost due to errors in processing. GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

  • Fig. 4 Sensitization to venetoclax requires inhibition of protein geranylgeranylation.

    (A) Viability of cells treated with the combination of 300 nM navitoclax and 10 μM simvastatin supplemented with indicated metabolites for 48 hours. MVA, mevalonate. n = 4 (OCI-LY8 control and GGPP), n = 3 (OCI-LY8 MVA/cholesterol and all SU-DHL4 groups). (B) Viability of cells treated as indicated with 10 μM simvastatin, GGTI-298, or FTI-277 with (black bars) or without (white bars) venetoclax (30 nM for OCI-LY8 and 300 nM for SU-DHL4) for 48 hours. Viability was assessed by flow cytometry using annexin-V and propidium iodide double negativity. n = 3 [Sim and FT inhibitor (FTI)], n = 5 or 6 [GGT inhibitor (GGTI) and dimethyl sulfoxide (DMSO)]. (C) Western blot of cells treated with 10 μM of indicated inhibitors for 16 hours. Statin and FTI-277 but not GGTI-498 caused a mobility shift of HDJ-2 (slower migrating, unprenylated form). Statin and GGTI-498 but not FTI-277 induced the appearance of unprenylated RAP1A. (D) Dynamic BH3 profile of cells treated with 10 μM simvastatin (blue bars) or GGTI-298 (black bars) for 16 hours. n = 5 (OCI-LY8, DMSO, and Sim), n = 2 (OCI-LY8, GGTI), n = 4 (SU-DHL4, all conditions). Significance testing was performed by two-tailed paired Student’s t test relative to vehicle-treated control samples in (A) and (B), one-tailed in (D).

  • Fig. 5 Statins induce transcription of PUMA and HRK in DLBCL and AML cell lines.

    (A and B) Western blot (left) and quantification (right) of whole-cell lysates from DLBCL cells (A) or AML cells (B) treated with 10 μM of indicated inhibitors for 16 hours. (C) Western blot (left) and quantification (right) of mitochondria-enriched fractions from DLBCL cells treated with 10 μM of indicated inhibitors for 16 hours. M, mitochondrial fraction; C, cytoplasmic fraction. COX IV is a mitochondrial marker, and extracellular signal–regulated kinase (ERK) is a cytoplasmic marker. (D) Quantitative polymerase chain reaction of DLBCL cell lines treated with 10 μM of indicated inhibitors for 16 hours. (A to C) Paired one-tailed t tests, n = 3 to 5. (D) One-way ANOVA with Dunnett’s post-test (all samples compared to control).

  • Fig. 6 Response to venetoclax was enhanced among statin users in CLL clinical trials.

    (A) Proportion of CLL subjects in clinical studies of venetoclax monotherapy who achieved any response, depicted as ORR by investigator assessment; results are shown by the presence or absence of statin use in individual studies and when pooled. (B) Proportion of subjects in these studies who achieved CR by investigator assessment; this result reflects the composite of subjects who achieved CR or CRi. (C) Odds ratio for achievement of CR across the pool of studies when adjusted for individual covariates. ECOG, Eastern Cooperative Oncology Group; ALC, absolute lymphocyte count. (D) Adjusted and unadjusted odds ratios for achievement of ORR and CR and hazard ratios for PFS and OS estimated by multivariate analyses across the pool of studies.

Supplementary Materials

  • www.sciencetranslationalmedicine.org/cgi/content/full/10/445/eaaq1240/DC1

    Materials and Methods

    Fig. S1. Simvastatin enhances the effects of venetoclax on a subset of AML and DLBCL cell lines.

    Fig. S2. Statins synergize with venetoclax in blood cancer cells.

    Fig. S3. Simvastatin plus venetoclax induce apoptosis in DLBCL and AML cell lines.

    Fig. S4. Simvastatin enhances killing of primary CLL samples cultured with stimuli from the microenvironment.

    Fig. S5. Statins induce dose-dependent increase in mitochondrial priming but do not sensitize to chemotherapy.

    Fig. S6. The combination of statin with venetoclax extends survival of mice with syngeneic B cell lymphoma.

    Fig. S7. The effect of statins is due to on-target HMGCR inhibition.

    Fig. S8. Mevalonate and GGPP are sufficient to rescue from the effects of simvastatin.

    Fig. S9. Simvastatin inhibits protein geranylgeranylation in a dose-dependent manner in DLBCL.

    Fig. S10. Inhibition of GGT is sufficient to recapitulate the effects of simvastatin in AML cell lines.

    Fig. S11. Simvastatin does not affect expression of many major BCL2 family proteins but does increase PUMA.

    Fig. S12. Simvastatin increases association of BCL2 with PUMA in sensitive OCI-AML3 cells but not in resistant OCI-LY1 cells.

    Fig. S13. PUMA knockdown in OCI-AML3 cells rescues them from sensitization to venetoclax by simvastatin.

    Fig. S14. Statins increase PUMA expression through a mechanism independent of p53 in DLBCL and AML cells.

    Fig. S15. Statin use is associated with longer PFS in CLL patients treated in venetoclax clinical trials.

    Fig. S16. Response to venetoclax was enhanced in CLL clinical trials among patients who received the 400-mg statin dose.

    Fig. S17. Statins do not affect venetoclax pharmacokinetics.

    Table S1. Characteristics of CLL patient samples.

    Table S2. Characteristics of CLL patients in three clinical trials of venetoclax monotherapy grouped by background statin use.

    Table S3. Adverse events in CLL patients in three clinical trials of venetoclax monotherapy grouped by background statin use.

    Table S4. Raw data (in a separate Excel file).

  • Supplementary Material for:

    Statins enhance efficacy of venetoclax in blood cancers

    J. Scott Lee, Andrew Roberts*, Dennis Juarez, Thanh-Trang T. Vo, Shruti Bhatt, Lee-or Herzog, Sharmila Mallya, Richard J. Bellin, Suresh K. Agarwal, Ahmed Hamed Salem, Tu Xu, Jia Jia, Lingxiao Li, John R. Hanna, Matthew S. Davids, Angela G. Fleischman, Susan O'Brien, Lloyd T. Lam, Joel D. Leverson, Anthony Letai, Jonathan H. Schatz, David A. Fruman*

    *Corresponding author. Email: dfruman{at}uci.edu (D.A.F.); awroberts{at}comcast.net (A.R.)

    Published 13 June 2018, Sci. Transl. Med. 10, eaaq1240 (2018)
    DOI: 10.1126/scitranslmed.aaq1240

    This PDF file includes:

    • Materials and Methods
    • Fig. S1. Simvastatin enhances the effects of venetoclax on a subset of AML and DLBCL cell lines.
    • Fig. S2. Statins synergize with venetoclax in blood cancer cells.
    • Fig. S3. Simvastatin plus venetoclax induce apoptosis in DLBCL and AML cell lines.
    • Fig. S4. Simvastatin enhances killing of primary CLL samples cultured with stimuli from the microenvironment.
    • Fig. S5. Statins induce dose-dependent increase in mitochondrial priming but do not sensitize to chemotherapy.
    • Fig. S6. The combination of statin with venetoclax extends survival of mice with syngeneic B cell lymphoma.
    • Fig. S7. The effect of statins is due to on-target HMGCR inhibition.
    • Fig. S8. Mevalonate and GGPP are sufficient to rescue from the effects of simvastatin.
    • Fig. S9. Simvastatin inhibits protein geranylgeranylation in a dose-dependent manner in DLBCL.
    • Fig. S10. Inhibition of GGT is sufficient to recapitulate the effects of simvastatin in AML cell lines.
    • Fig. S11. Simvastatin does not affect expression of many major BCL2 family proteins but does increase PUMA.
    • Fig. S12. Simvastatin increases association of BCL2 with PUMA in sensitive OCI-AML3 cells but not in resistant OCI-LY1 cells.
    • Fig. S13. PUMA knockdown in OCI-AML3 cells rescues them from sensitization to venetoclax by simvastatin.
    • Fig. S14. Statins increase PUMA expression through a mechanism independent of p53 in DLBCL and AML cells.
    • Fig. S15. Statin use is associated with longer PFS in CLL patients treated in venetoclax clinical trials.
    • Fig. S16. Response to venetoclax was enhanced in CLL clinical trials among patients who received the 400-mg statin dose.
    • Fig. S17. Statins do not affect venetoclax pharmacokinetics.
    • Table S1. Characteristics of CLL patient samples.
    • Table S2. Characteristics of CLL patients in three clinical trials of venetoclax monotherapy grouped by background statin use.
    • Table S3. Adverse events in CLL patients in three clinical trials of venetoclax monotherapy grouped by background statin use.

    [Download PDF]

    Other Supplementary Material for this manuscript includes the following:

    • Table S4 Raw data (in a separate Excel file).

Navigate This Article