The transcriptomic response of cells to a drug combination is more than the sum of the responses to the monotherapies
- Department of Genetics and Genomics Sciences, Icahn School of Medicine at Mount Sinai, United States
- Department of Cell, Developmental, and Regenerative Biology, Icahn School of Medicine at Mount Sinai, United States
- IBM Computational Biology Center, IBM Research, United States
- Department of Business Administration, University of Illinois at Urbana-Champaign, United States
- Department of Systems Biology, Columbia University, United States
- Sulzberger Columbia Genome Center, High Throughput Screening Facility, Columbia University Medical Center, United States
- Department of Biomedical Informatics, Columbia University, United States
- Department of Biochemistry and Molecular Biophysics, Columbia University, United States
- Department of Medicine, Columbia University, United States
- Tisch Cancer Institute, Cancer Immunology, Icahn School of Medicine at Mount Sinai, United States
- Diabetes, Obesity and Metabolism Institute, Icahn School of Medicine at Mount Sinai, United States
- Icahn Institute for Data Science and Genomic Technology, Icahn School of Medicine at Mount Sinai, United States
Figures
Figure 1 with 2 supplements
The transcriptomics of drug combinations mirror their phenotypic characteristics.
(A) Monotherapies and drug combinations used in the study. (B) Workflow of molecular analysis of synergy. Starburst highlights the novel component of RNAseq analysis. Question mark denotes the focus …
Figure 1—figure supplement 1
The response of MCF7 to TM is more synergistic than to TW.
(A) Combination Index for the combinations at the selected doses. Combination Index < 1 indicates synergy. (B) Viability of MCF7 cells treated with increasing doses of T and M performed alongside …
Figure 1—figure supplement 2
Transcriptomic profiles of MFC7 and LNCaP cells with combinations.
(A) Similarity between the gene expression data of two replicates treated with TM at 12 hr. (B) The gene expression of one replicate treated with TM at 12 hr and another replicate treated with TM at …
Figure 2 with 2 supplements
Synergistically expressed genes and correlated monotherapies are associated with synergy.
(A-C) Number of DEGs over time in MCF7. The Venn diagrams correspond to DEGs at 3 hr in (A) T, M, and TM; (B) T, W, and TW; and (C) M, W, and MW. The area represented in each color is in proportion …
Figure 2—figure supplement 1
Gene expression characteristics of differential expression and synergy in MCF7.
(A) Comparison between gene expression in treatment (y-axis) and control (x-axis), for all genes over all treatments from the combination experiments in MCF7. Upregulated and downregulated genes as …
Figure 2—figure supplement 2
Differential expression in LNCaP Number of DEGs over time in LNCaP.
The Venn diagrams correspond to DEGs at 3 hr in (A) T, M, and TM; (B) T, W, and TW; and (C) M, W, and MW. The area represented in each color is in proportion to the number of genes in the …
Figure 3 with 2 supplements
Key biological processes are associated with synergy.
Enrichment of DEGs in T, M, and TM with cancer-relevant gene sets. Only gene sets enriched in at least one condition (time point or treatment) are shown. ‘TUM’ indicates the union of DEGs either in …
Figure 3—figure supplement 1
Biological processes in MCF7 and LNCaP.
(A) Enrichment scores of differentially up and down regulated genes at different time points in W, M, T, TW, and MW, in MCF7 cells with the same cancer-relevant gene sets shown in Figure 3. (B) …
Figure 3—figure supplement 2
Enrichment in gene sets associated with phospholipidosis (PLD) in MCF7 and LNCaP.
Enrichment of DEGs with cellular component gene sets (see Materials and methods) in (A) MCF7 cells for T, M, and TM, (B) MCF7 cells for W, M, T, TW, and MW, and (C) all treatments in LNCaP cells, …
Figure 4
Differentially expressed genes have different time courses.
(A) Mean and standard deviation of gene expression in four clusters identified according to their similarity in expression in T, M, and TM in MCF7 cells. (B) Examples of genes in each cluster with …
Figure 5
New differential splicing emerges in drug combination TM.
Top 100 synergistically spliced exons in combination TM at 12 hr in MCF7 cells.
Figure 6 with 1 supplement
Synergistic splicing is distinct from differential expression and associated with synergy.
(A) Number of synergistically expressed and synergistically spliced genes in TM in MCF7 cells over time; shaded areas correspond to the Venn diagram for 3 hr. (B) Relationship of Excess Over Bliss …
Figure 6—figure supplement 1
Differential and synergistic splicing in MCF7 cells.
(A-C) Number of differentially spliced genes over time with Venn diagrams of differentially spliced genes at 3 hr in (A) T, M, and TM; (B) T, W, and TW; and (C) M, W, and MW. The area represented in …
Figure 7 with 2 supplements
New differentially active transcription factors emerge in combination TM.
Number of DATFs over time with Venn diagrams at 3 hr in (A) T, M, and TM; (B) T, W, and TW; and (C) M, W, and MW in MCF7 cells. Area represented in each color matches the number of genes in the …
Figure 7—figure supplement 1
Possible changes to transcription factor activity.
Four cases of transcription factor activity that were assessed to determine whether a transcription factor was activated or inactivated.
Figure 7—figure supplement 2
Classes of differentially active transcription factors in MCF7 cells.
(A) Differentially active transcription factors for each combination according to the status of the positive and negative effector of each transcription factor. Unique: either positive or negative …
Figure 8 with 1 supplement
Characteristics of differentially active transcription factors.
(A) All instances of DATFs in MCF7 cells according to the differential expression or splicing status of each TF in the corresponding treatment and time point. The top 20 most significant DATFs not …
Figure 8—figure supplement 1
Differentially active transcription factors in W combinations for MCF7 cells.
Heatmap of differentially active transcription factors over time in W, MW, and TW at 3–24 hr. Color intensity reflects the degree and direction of enrichment by Fisher’s Exact test and is shown as …
Figure 9 with 1 supplement
Transcription factors become differentially active in a time-dependent cascade in TM.
The number of DATFs or SEGs at 3–24 hr are shown as bubbles. Blue, red, and white bubbles represent DATFs in T, M, and TM, respectively. TFs (gray bubbles) and SEGs (green bubbles) shown are …
Figure 9—figure supplement 1
Cascade of differential transcription factor activity.
Connections between differentially active transcription factors in TM based on the MCF7 network. Each bubble represents a set of transcription factors that are differentially active in TM at a given …
Figure 10
Correlation of monotherapies is associated with synergy in an independent dataset.
(A) Relationship between Excess Over Bliss (EOB) for 91 drug pairs and the correlation between the gene expression of LY3 DLBCL cells treated with corresponding monotherapies in the DREAM dataset. …
Tables
Table 1
Selection of adjusted p-value cutoff for differentially expressed genes.
The six most differentially expressed genes with respect to DMSO in each treatment at time 0 are shown in ascending order of their Voom score (log10(FDR)). Immediate Early Genes are marked in red. …
| T_0 | TM_0 | M_0 | TW_0 | MW_0 | W_0 | ||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| -6 | BCAN | −17 | ATXN2 | -3 | −54 | −53 | −53 | ||||
| -3 | −17 | -2 | −26 | −28 | −20 | ||||||
| TOB1 | -3 | VIM | −17 | ZNF592 | -1 | −23 | −17 | −19 | |||
| KLF4 | -2 | −15 | ZHX2 | -1 | −17 | PDCD7 | −17 | C17O | −16 | ||
| SGK1 | -2 | MSN | −13 | SCAF4 | -1 | PDCD7 | −16 | −15 | −16 | ||
| PRDM1 | -2 | NCAN | −10 | NAT8L | -1 | C17ORF91 | −16 | −14 | PDCD7 | −15 | |
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Cell line (Homo sapiens) | MCF7 | American Type Culture Collection | Cat No. HTB-22 | RRID:CVCL_0031 |
| Cell line (Homo sapiens) | LNCaP | American Type Culture Collection | Cat No. CRL-1740 | RRID:CVCL_1379 |
| Chemical compound, drug | Withaferin A | Enzo Life Sciences | Cat No. BML-CT104-0010 | |
| Chemical compound, drug | Mefloquine hydrochloride | Sigma-Aldrich | Cat No. M2319-100MG | |
| Chemical compound, drug | Tamoxifen citrate | Tocris Bioscience | Cat No. 0999 |
Additional files
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Source data 1
Log counts per million of MCF7 cell combination treatment experiments.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data1-v2.csv.zip
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Source data 2
Log counts per million of MCF7 cell monotherapy dose experiments.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data2-v2.csv.zip
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Source data 3
Log counts per million of LNCaP cell combination treatment experiments.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data3-v2.csv.zip
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Source data 4
Archive of MCF7 combination experiments differential expression data.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data4-v2.zip
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Source data 5
Archive of MCF7 dose experiments differential expression data.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data5-v2.zip
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Source data 6
Archive of LNCaP differential expression data.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data6-v2.zip
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Source data 7
k-means clusters assigned to genes.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data7-v2.zip
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Source data 8
Archive of differential splicing data.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data8-v2.zip
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Source data 9
Archive of differential transcription factor activity data.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data9-v2.zip
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Source data 10
Archive of transcription factors involved in the transcriptional cascade.
- https://cdn.elifesciences.org/articles/52707/elife-52707-data10-v2.zip
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Supplementary file 1
Viability data and calculated EOB for TM dose matrices at 12, 24, and 48 hr in MCF7.
Actual values of negative inhibition in monotherapies are included in the heatmap at left. Monotherapy inhibition values used to calculate EOB are shown in the table at right (i.e. Drug1_NPI).
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp1-v2.xlsx
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Supplementary file 2
Viability data and calculated EOB for TW dose matrices at 12, 24, and 48 hr in MCF7.
Actual values of negative inhibition in monotherapies are included in the heatmap at left. Monotherapy inhibition values used to calculate EOB are shown in the table at right (i.e. Drug1_NPI).
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp2-v2.xlsx
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Supplementary file 3
Time courses viability data of TM, TW, and MW in MCF7.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp3-v2.xlsx
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Supplementary file 4
Time courses viability data of TM, TW, and MW in LNCaP.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp4-v2.xlsx
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Supplementary file 5
Viability data and calculated EOB for TM, TW, and MW at 48 hr in LNCaP.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp5-v2.xlsx
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Supplementary file 6
Viability data for T and M dose and calculated EOB for sham combinations in MCF7.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp6-v2.csv.zip
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Supplementary file 7
Archive of Raw Fastq IDs.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp7-v2.zip
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Supplementary file 8
Archive of raw expression files.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp8-v2.zip
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Supplementary file 9
Exon counts.
- https://cdn.elifesciences.org/articles/52707/elife-52707-supp9-v2.zip
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Transparent reporting form
- https://cdn.elifesciences.org/articles/52707/elife-52707-transrepform-v2.docx
