1. Cancer Biology
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Loss of functional BAP1 augments sensitivity to TRAIL in cancer cells

  1. Krishna Kalyan Kolluri
  2. Constantine Alifrangis
  3. Neelam Kumar
  4. Yuki Ishii
  5. Stacey Price
  6. Magali Michaut
  7. Steven Williams
  8. Syd Barthorpe
  9. Howard Lightfoot
  10. Sara Busacca
  11. Annabel Sharkey
  12. Zhenqiang Yuan
  13. Elizabeth K Sage
  14. Sabarinath Vallath
  15. John Le Quesne
  16. David A Tice
  17. Doraid Alrifai
  18. Sylvia von Karstedt
  19. Antonella Montinaro
  20. Naomi Guppy
  21. David A Waller
  22. Apostolos Nakas
  23. Robert Good
  24. Alan Holmes
  25. Henning Walczak
  26. Dean A Fennell
  27. Mathew Garnett
  28. Francesco Iorio
  29. Lodewyk Wessels
  30. Ultan McDermott  Is a corresponding author
  31. Samuel M Janes  Is a corresponding author
  1. University College London, United Kingdom
  2. Wellcome Trust Sanger Institute, United Kingdom
  3. The Netherlands Cancer Institute, Netherlands
  4. University of Leicester, United Kingdom
  5. Oncology Research, MedImmune, Inc., United States
  6. UCL Cancer Institute, University College London, United Kingdom
  7. Glenfield Hospital, University Hospitals of Leicester, United Kingdom
  8. European Bioinformatics Institute, United Kingdom
Short Report
Cite this article as: eLife 2018;7:e30224 doi: 10.7554/eLife.30224
3 figures, 2 tables and 7 additional files


Figure 1 with 2 supplements
A chemical screen in mesothelioma cell lines identifies a BAP1-mutant population sensitised to the death receptor ligand rTRAIL.

(A) Area under the curve (AUC) values for 15 malignant mesothelioma (MM) cells treated for 6 days with 94 compounds. Each dot indicates the AUC value for an individual cell line treated. AUC <0.7 is indicated by the red dotted line — only those compounds with ≥2 cell lines below this value were analysed for statistically significant associations with gene mutations. The AUC values for rTRAIL are indicated by the red asterisk. (B) A Welch t-test was used to test for significant pharmacogenomics interactions between the 94 single agents in the screen and the presence of driver mutations in any of 5 MM cancer genes. Each volcano plot circle corresponds to a significant gene–drug interaction whose position on the x-axis indicates the corresponding effect size. Both half-axes are positive; the right side (green circles) indicates the effect sizes of sensitivity associations, whereas the left side (red circles) corresponds with the effect sizes of resistance associations. The position on the y-axis indicates the statistical significance of the identified interaction. The size of a given circle is proportional to the number of samples in which the selected functional event involved in the corresponding interaction occurs. Specific examples of associations are indicated where the effect size is large (rTRAIL and BAP1 mutations) or highly significant (cisplatin and CDKN2A mutations). (C) Cell viability between wild-type BAP1 (wt BAP1) (n = 10) and mutant BAP1 (mt BAP1) (n = 5) MM lines following 6 days of treatment with rTRAIL (t-test; *p=0.015). (D) Cell viability data for 17 MM lines treated for 6 days with a concentration range of rTRAIL (0.4–50 ng/ml). MM lines are coloured according to their sensitivity pattern (green = sensitive (S); orange = partially sensitive (PS); red = resistant (R)). *Indicates cell lines harbouring BAP1 mutations. (E) Immunoblot of BAP1 protein expression in BAP1-mutant versus BAP1-wild-type MM cell lines. Sensitivity to rTRAIL treatment is indicated as font colour: green (S); orange (PS); red (R).

Figure 1—figure supplement 1
Mutation status of 5 candidate tumour driver genes in the 15 MM lines used in the combinatorial chemical inhibitor screen.
Figure 1—figure supplement 2
BAP1 and the response to alternative apoptotic stimuli in MM cells.

72 hour cell viability results for 9 MM cell lines (4 BAP1-mutant - green and five wild-type - red) treated with (A) cisplatin (B) pemetrexed (C) FAS receptor agonistic antibody CH11, (D) TNF-α and 5 μM LCL161 or (E) DR5 agonist MEDI3039 assessed by MTT assay.

Figure 2 with 9 supplements
BAP1-induced TRAIL resistance extends to other cancer subtypes and is dependent upon functional deubiquitinase and ASXL-binding sites.

(A) BAP1-wild-type H2818, MPP-89, H2373 and H2869 MM lines were transduced with BAP1 (shBAP1) or empty vector (EV) shRNA. Immunoblot confirmed BAP1 knockdown in the BAP1 shRNA-transduced cells. Parental and transduced cells were treated with rTRAIL (1000 ng/ml) and cell viability assessed after 72 hr by MTT assay (t-test; ****p<0.0001). (B) The BAP1-wild-type breast cancer line MDAMB-231 and the renal cell carcinoma (RCC) lines Caki-1 and BB65 were transduced with BAP1 (shBAP1) or empty vector (EV) shRNA. Immunoblot confirmed BAP1 knockdown in the BAP1 shRNA transduced cells. Parental and transduced cells were treated with rTRAIL (1000 ng/ml) and cell viability assessed after 72 hr by MTT assay (t-test; ****p<0.0001). (C) The rTRAIL-sensitive H226 MM line, which harbours a homozygous deletion of BAP1, was transduced with either a GFP control, wild-type BAP1 or a mutant BAP1 containing an inactive functional domain: C91A — inactivating mutation of deubiquitinase catalytic site; ΔHBM — deletion of HCF-1-binding motif; T493A — inactivating mutation of FOXK2-binding site; ΔASXL — deletion of ASXL1/2 protein-binding site; ΔCTD — deletion of C-terminal domain containing nuclear localisation signal. These transduced lines were treated with 50 ng/ml rTRAIL and cell death assessed with XTT assay (one-way ANOVA; **p<0.01). (D) The parental and transduced H226 MM lines were treated with a concentration range (1–100 pM) of the small molecule death receptor agonist MEDI3039 and cell viability assessed with XTT assay. (E) The BAP1-wild-type MPP-89 MM line was transduced with ASXL1 (shASXL1), ASXL2 (shASXL2) or empty vector (EV) shRNA. qPCR confirmed a decrease in ASXL1 and ASXL2 mRNA expression in the ASXL1 shRNA and ASXL2 shRNA-transduced cells, respectively (Figure 2—figure supplement 6). Parental and transduced cells were treated with a concentration range (1–100 pM) of MEDI3039 and cell viability assessed with XTT assay. (F) Differential gene expression of apoptosis regulator genes in the catalytically inactive BAP1-mutant (C91A) relative to the wild-type BAP1-transduced (wt BAP1) H226 cells. (G) Immunoblot of apoptosis regulator proteins in the catalytically inactive BAP1-mutant (C91A), inactive ASXL1/2-binding site BAP1-mutant (ΔASXL) or wild-type BAP1-transduced (wt BAP1) H226 cells. (H) Flow cytometry analysis of death receptor 4 (DR4) and 5 (DR5) cell surface expression in H226 cells transduced with the catalytically inactive BAP1-mutant (C91A) or wild-type BAP1 (wt BAP1) and of BAP1-wild-type H2818 MM cells transduced with BAP1 (KD) or empty vector (EV) shRNA. The values represent the median fluorescence intensity (MFI).

Figure 2—figure supplement 1
shRNA knockdown of BAP1 increases sensitivity to rTRAIL in MM cells.

Three BAP1-wild-type MM cell lines (A) MPP-89, (B) H2869 and (C) H2818 were transduced with empty vector (EV) or BAP1 shRNA (shBAP1). Immunoblot confirmed BAP1 knockdown. The parental, EV and shBAP1 cells were treated with rTRAIL for 24 hr and cell death measured by Annexin V/DAPI flow cytometry assay.

Figure 2—figure supplement 2
shRNA knockdown of BAP1 increases sensitivity to DR agonists in breast cancer cells.

The MDAMB-231 breast cancer cell line was transduced with empty vector (EV) or BAP1 shRNA (shBAP1). Immunoblot confirmed BAP1 knockdown. Cells were treated with (A) rTRAIL and (B) MEDI3039 and cell viability measured with MTT assay at 72 hr. (C) Cells were treated with rTRAIL for 24 hr and cell death measured with Annexin V/DAPI flow cytometry assay.

Figure 2—figure supplement 3
shRNA knockdown of BAP1 increases sensitivity to DR agonists in clear cell renal carcinoma cells.

Clear cell renal carcinoma cell lines, Caki-1 and BB65, were transduced either with either empty vector (EV) or BAP1 shRNA (shBAP1). Immunoblot confirmed BAP1 knockdown. Cells were treated with rTRAIL (A and C) or MEDI3039 (B and D) for 72 hr and cell viability measured by MTT assay.

Figure 2—figure supplement 4
Cell viability of non-mesothelioma BAP1-mutant cell lines following rTRAIL treatment.

Bladder (RT4) and breast (HCC1187) cancer cell lines harbouring nonsense mutations in BAP1 show increased sensitivity to rTRAIL compared with renal cell carcinoma or bladder cancer cell lines harbouring missense (769P and RCC10RGB) or wild-type BAP1 (BB65RCC and SW1710). Cell viability was measured after 6 days of treatment with 100 ng/ml rTRAIL.

Figure 2—figure supplement 5
Overexpression of wild-type BAP1 induces resistance to rTRAIL in BAP1 mutant MM cells. 

The rTRAIL-sensitive H2804(A) and H28(B) mesothelioma cell lines, which harbour mutations in BAP1, were transduced with wild-type BAP1 (wt BAP1) or BAP1 with an inactive deubiquitinase catalytic domain (C91A) and treated with a dose range of rTRAIL.

Cell death was assessed with Annexin V/DAPI apoptosis assay.

Figure 2—figure supplement 6
shRNA knockdown of ASXL1 increases sensitivity of MM cells to rTRAIL.

(A) Cell viability of parental, empty vector, ASXL1 and ASXL2 shRNA-transduced MPP-89 cells treated with rTRAIL (0–1000 ng/ml) for 3 days measured with XTT assay. (B) Efficacy of ASXL1 and ASXL2 shRNA knockdown assessed by qPCR.

Figure 2—figure supplement 7
Ubiquitinated histone 2A at K119 (H2AK119Ub) expression and BAP1 function.

(A) Immunoblot analysis of H2AK119Ub levels in the parental, GFP-, wild-type BAP1 (wt BAP1)-, deubiquitinase mutant BAP1 (C91A)- and ASXL-binding mutant BAP1 (ΔASXL)-transduced H226 MM cell lines. (B) Immunofluorescence images of H2AK119Ub staining in the parental, deubiquitinase mutant-transduced (C91A), ASXL-binding mutant-transduced (ΔASXL) and wild-type BAP1-transduced H226 cell lines. (C) Quantification of immunofluorescence staining in 2B (normalised to cell number; one-way ANOVA; ***p<0.001).

Figure 2—figure supplement 8
Differential gene expression data from H226 MM cells expressing C91A-mutant (mt BAP1) or wild-type BAP1 (wt BAP1).

Only genes with logFC ≥2 and adj.p <0.05 are displayed.

Figure 2—figure supplement 9
Signalling pathway impact analysis of gene expression data from H226 MM cells expressing C91A-mutant (mt BAP1) or wild-type BAP1 (wt BAP1).

The proteins in the pathway are highlighted in green if the expression in mt BAP1 is significantly less than wt BAP1 and red if the expression in mt BAP1 is significantly more than wt BAP1.

Figure 3 with 3 supplements
Loss of functional BAP1 leads to TRAIL sensitivity in early passage mesothelioma cell lines, human tumour explants and mouse xenograft models.

(A) Mean cell viability effect between human early passage MM cell lines (positive nuclear BAP1 staining; n = 13 and negative nuclear BAP1 staining; n = 12) as assessed by immunohistochemistry following 3 days of treatment with rTRAIL (50 ng/ml) (t-test, p=0.0067). (B) Immunohistochemical images of tumour explants derived from three MM patients treated with either vehicle or rTRAIL for 24 hr. Explants were stained with anti-BAP1 and anti-cleaved PARP (marker for apoptosis) antibodies. (C) The percentage of cleaved PARP-positive cells in tumour explants derived from three patients and treated with either vehicle or 0, 50, 100 and 200 ng/ml of rTRAIL for 24 hr was scored based on the percentage of cells with nuclear cleaved PARP-positive staining. (D) Weights of tumour xenografts derived from BAP1-wild-type (wt BAP1) versus catalytically inactive BAP1-mutant (C91A mt BAP1) transduced MM cells following treatment with either vehicle or TRAIL (600 μg per mouse) at the time of sacrifice (day 42) (t-test). (E) Serial bioluminescence imaging of BAP1-wild-type (wt BAP1) and catalytically inactive BAP1-mutant (C91A) MM xenografts in mice treated with either vehicle or TRAIL. Mice were imaged on day 0 (after tumour inoculation), day 13 (before TRAIL administration) and day 41 (time of sacrifice). The intensity of luminescence is denoted by colour: red - high luciferase signal (high tumour burden) and blue - low luciferase signal (low tumour burden). (F) A time-course of bioluminescence scores in BAP1-wild-type (wt BAP1) versus catalytically inactive BAP1-mutant (C91A) MM tumour xenografts. Bioluminescence was measured on days 0, 13, 19, 26 and 41, 15 min after injecting the mice with 0.2 ml luciferin intraperitoneally. The number of photons emitted per second indicates the tumour burden (two way ANOVA).

Figure 3—figure supplement 1
BAP1 expression in early passage MM cultures.

(A) Immunohistochemical analysis and (B) immunoblot. Cultures are grouped by sensitivity to rTRAIL.

Figure 3—figure supplement 2
Ex vivo experimental protocol.

Tumour explants were obtained by cutting primary pleural tissue from patients with MM who underwent pleurectomy into fragments of approximately 2 mm3.

The explants were treated with vehicle or rTRAIL (50 ng/ml, 100 ng/ml or 200 ng/ml) for 24 hr, following which time explants were fixed and stained for cleaved-PARP (which is a marker of apoptosis).

Figure 3—figure supplement 3
In vivo experimental protocol.

(A) Schematic of in vivo experimental protocol. Mice were injected with H226 cells transduced with wild-type BAP1 and luciferase or catalytically inactive BAP1-mutant (C91A) and luciferase on the right and left flanks, respectively. Mice were divided into two groups, each of which received 600 μg TRAIL or vehicle 6 days a week (day 14–40). Tumour size was assessed longitudinally with bioluminescence imaging on days 0, 13, 19, 26 and 41. (B) Size of tumours derived from BAP1-wild-type (wt) versus catalytically inactive (C91A) BAP1-mutant (mt) MM cells following treatment with either vehicle or TRAIL (600 μg per mouse) at time of sacrifice (day 42). A centimetre scale is included in the photograph for comparison.



Table 1
BAP1 immunoblot status, nuclear BAP1 staining and rTRAIL sensitivity (50 ng/ml) of the 25 human early passage MM cultures.
Sample nameWestern blotNuclear BAP1-IHCSensitivity
36Low ExpressionSensitive
52Partially Sensitive
2Partially Sensitive
30Low Expression+Partially Sensitive
15Low Expression+Partially Sensitive
35++Partially Sensitive
24++Partially Sensitive
Key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
BRCA associated protein-1 (human)BAP1Entrez Gene NCBIGene ID: 8314
Additional sex combs like 1 (human)ASXL1Entrez Gene NCBIGene ID: 171023
strain, strain background
NOD.CB17-Prkdcscid/NcrCrlNOD SCID miceCharles River Laboratories, UKRRID:IMSR_CRL:394
cell line
Early passage mesotheliomacell cultures7T, 8T, 45, 19, 14T,
23T, 40, 36, 26, 12T, 3T, 52, 2, 30, 15, 35, 24, 43, 34, 50T, 33T, 18, 53T, 38
MesobanK, Mesothelioma UKwww.mesobank.com
Tissue Bank, Papworth Hospital NHS Trust, UK
NCI-H2373H2373Wellcome Trust Sanger Institute, UKRRID:CVCL_A533
NCI-H2803H2803Wellcome Trust Sanger Institute, UKRRID:CVCL_U997
NCI-H2452H2452Wellcome Trust Sanger Institute, UKRRID:CVCL_1553
NCI-H2722H2722Wellcome Trust Sanger Institute, UKRRID:CVCL_U994
NCI-H2369H2369Wellcome Trust Sanger Institute, UKRRID:CVCL_A532
NCI-H2795H2795Wellcome Trust Sanger Institute, UKRRID:CVCL_U996
NCI-H2869H2869Wellcome Trust Sanger Institute, UKRRID:CVCL_V001
NCI-H2591H2591Wellcome Trust Sanger Institute, UKRRID:CVCL_A543
MPP 89MPP-89Wellcome Trust Sanger Institute, UKRRID:CVCL_1427
NCI-H2810H2810Wellcome Trust Sanger Institute, UKRRID:CVCL_U999
NCI-H2818H2818Wellcome Trust Sanger Institute, UKRRID:CVCL_V000
NCI-H513H513Wellcome Trust Sanger Institute, UKRRID:CVCL_A570
NCI-H2595H2595Wellcome Trust Sanger Institute, UKRRID:CVCL_A545
NCI-H2461H2461Wellcome Trust Sanger Institute, UKRRID:CVCL_A536
NCI-H2731H2731Wellcome Trust Sanger Institute, UKRRID:CVCL_U995
NCI-H2804H2804Wellcome Trust Sanger Institute, UKRRID:CVCL_U998
NCI-H28H28Wellcome Trust Sanger Institute, UKRRID:CVCL_1555
NCI-H226H226Szlosarek lab, Barts Cancer Institute, UKRRID:CVCL_1544
MDA-MB-231MDAMB-231Wellcome Trust Sanger Institute, UKRRID:CVCL_0062
Caki-1Caki-1Wellcome Trust Sanger Institute, UKRRID:CVCL_0234
BB65BB65Wellcome Trust Sanger Institute, UKRRID:CVCL_1078
BAP1 (C-4) mouse mAbanti-BAP1Santa Cruz Biotechnology, Santa Cruz, CACat# sc-28383 RRID:AB_6267231:500 in milk; 1:50 for flow cytometry
Caspase-8 (1C12) mouse mAbanti-caspase 8Cell Signaling Technology, Danvers, MACat# 9746 RRID:AB_22751201:1000 in BSA
FLIP (7F10) mouse mAbanti c-FLIPEnzo Life Sciences, Farmingdale, NYCat# ALX-804-961-0100 RRID:AB_27139151:1000 in milk
c-IAP1 (D5G9) rabbit mAbanti-cIAP1Cell Signaling Technology,Danvers, MACat# 7065S RRID:AB_108908621:1000 in BSA
c-IAP2 (58C7) rabbit mAbanti-cIAP2Cell Signaling Technology,
Danvers, MA
Cat# 3130S RRID:AB_106932981:1000 in BSA
FADD rabbit pAbanti-FADDCell Signaling Technology,
Danvers MA
Cat# 2782 RRID:AB_21004841:1000 in BSA
XIAP (3B6) rabbit mAbanti-XIAPCell Signaling Technology,
Danvers, MA
Cat# 2045 RRID:AB_22148661:1000 in milk
survivin rabbit pAbanti-survivinCell Signaling Technology,
Danvers, MA
Cat# 2803 RRID:AB_4908071:1000 in BSA
α-Tubulin (11H10) Rabbit mAbanti-α-tubulinCell Signaling Technology,
Danvers, MA
#21251:2000 in milk
Ubiquityl-Histone H2A (Lys119) (D27C4) XPRabbit mAbanti-H2AK119UbCell Signaling Technology,
Danvers, MA
Cat# 8240P RRID:AB_108916181:2000 in BSA
Histone H2A (D6O3A) Rabbit mAbanti-H2ACell Signaling Technology,
Danvers, MA
Cat# 12349 RRID:AB_26878751:1000 in BSA
Anti-mouse IgG, HRP-linked antibodyanti-mouse HRPCell Signaling Technology,
Danvers, MA
Cat# 7076 RRID:AB_3309241:2000 in milk
Anti-rabbit IgG, HRP-linked antibodyanti-rabbit HRPCell Signaling Technology,
Danvers, MA
Cat# 7074 RRID:AB_20992331:2000 in milk
Donkey anti-Mouse IgG (H + L) Highly Cross-Adsorbed Secondary Antibody, AlexaFluor 488AlexaFluor 488-conjugated anti-mouse antibodyThermo Fisher Scientific, UKCat# A-21202 RRID:AB_1416071:200 for flow cytometry
Annexin V, AlexaFluor 647 conjugateAnnexin V AlexaFluor 647-conjugated antibodyThermo Fisher Scientific, UKCat# A23204 RRID:AB_23411491:100 for flow cytometry
PE anti-human CD261 (DR4, TRAIL-R1) antibodyPE-conjugated antibody to DR4Biolegend, UKCat# 307205 RRID:AB_3146691:100 for flow cytometry
PE anti-human CD262 (DR5, TRAIL-R2) antibodyPE-conjugated antibody to DR5Biolegend, UKCat# 307405 RRID:AB_3146771:100 for flow cytometry
PE Mouse IgG1, κ Isotype Ctrl AntibodyPE isotype control antibodyBiolegend, UKCat# 4001121:100 for flow cytometry
Goat anti-Rabbit IgG (H + L) Secondary Antibody, AlexaFluor 488 conjugateAlexaFluor 488-conjugated anti-rabbit secondary antibodyThermo Fisher Scientific, UKCat# R37116 RRID:AB_25565441:200 for flow cytometry
Anti-Cleaved PARP1 (E51) mAbcleaved PARP primary antibody; anti-cleaved PARPAbcam, UKCat# ab32064 RRID:AB_777102(1:6000) for immunohistochemistry
recombinant DNA reagent
BAP1 (NM_004656) Human cDNA ClonepCMV6-AC BAP1 plasmidOrigene, Rockville, MDCat# SC117256
pHIV-Luc-ZsGreenZS-green luciferase plasmid, pHIV-Luc-ZsGreenBryan Welm Lab, University of Utah, Addgene, Logan, UTCat# 39196
pCMVR8.74pCMV-dR8.74Thrasher lab, UCL, Addgene,
Cat# 22036
pMD2.GpMD2.GThrasher lab, UCL, Addgene,
Cat# 12259
sequence based reagent
BAP1 GIPZ Lentiviral shRNABAP1 shRNAUCL RNAi Library (Dharmacon, Lafayett, CO)V2LHS 41473
ASXL1 GIPZ Lentiviral shRNAASXL1 shRNAUCL RNAi Library (Dharmacon, Lafayett, CO)V2LHS 78829
ASXL2 GIPZ Lentiviral shRNAASXL2 shRNAUCL RNAi Library (Dharmacon, Lafayette, CO)V3LHS_313940
peptide, recombinant protein
Recombinant Human sTRAILrTRAILPeprotech, UKCat# 310–04
commercial assay or kit
Cell Proliferation Kit XTTXTT reagentApplichem, UKA8088
Q5 Site-Directed Mutagenesis KitSite directed mutagenesisNew England Biolabs, Ipswich, MACat# E0554
Rabbit specific HRP/DAB (ABC) Detection IHC Kitrabbit-specific HRP/DAB (ABC) detection IHC kitAbcam, UKCat# ab64261
chemical compound, drug
MEDI3039MEDI3039MedImmune, UK
software, algorithm
GraphPad Prism softwareGraphpad PrismGraphPad Software, CA, USA
CaVEMan algorithmCaVEManhttps://github.com/cancerit/CaVEMan
Pindel algorithmPindelhttps://github.com/genome/pindel
Predicting Integral Copy Numbers In Cancer algorithmPICNIChttp://www.sanger.ac.uk/science/tools/picnic
FlowJo softwareFlowjoFlowJo LLC
RIPA bufferRIPASigma-Aldrich, St. Louis, MOCat# R0278
Syto™ 60 red fluorescent nucleic acid stainSyto 60Thermo Fisher Scientific, UKCat# S11342
Thiazolyl Blue Tetrazolium Bromide (MTT)MTT reagentSigma-Aldrich, St. Louis, MOCat# M2128
jetPEI DNA transfection reagentjetPEISource Bioscience, UKCat# 101–10
PolybrenePolybreneSigma-Aldrich, St Louis, MOCat# 1076898 μg/ml
Hoechst 33342 Solution (20 mM)Hoechst 33342Thermo Fisher Scientific, UKCat# 62249
4’, 6-diamidino-2-phenylindoleDAPISigma-Aldrich, St Louis, MOCat# D9542200 μg/ml

Additional files

Supplementary file 1

List of 94 compounds used either as single agents or in combination with rTRAIL.

Listed are the unique ID number, the compound name and target, the cellular process targeted and the minimum and maximum concentration (micromolar) of the 5-point concentration range used for each compound.

Supplementary file 2

Name and histological subtype (where known) of the 15 mesothelioma cell lines.

Supplementary file 3

1425 area under the curve (AUC) viability scores for 94 experimental agents tested against 15 mesothelioma cell lines after 6 days of treatment.

Supplementary file 4

Results of Welch’s two sample t-test from analysis of 45 single compounds that ≥2 cell lines demonstrated sensitivity to (AUC <0.7) and using the mutation status of eight genes implicated as drivers in mesothelioma in each cell line.

A 6 day viability assay was used to determine cell line sensitivity. False discovery associations < 0.2 are highlighted as red font. Whether a mutation is associated with resistance or sensitivity to that compound is indicated by red or green in the ‘effect’ column, respectively.

Supplementary file 5

Description of BAP1 mutations detected in 15 mesothelioma cell lines and the sensitivity of the cell lines to rTRAIL (as measured by a 6 day viability assay).

The sensitivity of each cell line is indicated in the last column as sensitive (green), partially sensitive (orange) or resistant (red).

Supplementary file 6

Differential gene expression values of apoptotic genes in H226 mesothelioma cells transduced with either the catalytically inactive C91A BAP1 mutant (C91A) or wild-type BAP1 (WT).

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