Association of changes in expression of HDAC and SIRT genes after drug treatment with cancer cell line sensitivity to kinase inhibitors

Figures & data

Table 1. Concerted expression changes of HDAC and SIRT genes after drug treatment in the NCI-TPW dataset.

Gene	Aza	Bor	Cis	Das	Dox	Erl	Gel	Gem	Lap	Pac	Rap	Sor	Sun	Top	Vor
HDAC1	H6↓*, H24↓*, L24↓	H6↓, H24↓, L24↓*	L24↑		H24↓*		H6↓		H6↓			H24↓*, L24↓	H6↓	H6↑	H6↑, H24↑*, L6↑, L24↑*
HDAC2	H24↑, L24↑	L24↓		H6↓, H24↓, L24↓			H6↓, H24↓, L6↓		H24↓		H6↓	H24↓*	H24↓
HDAC3	H6↓, H24↓	H6↓			H6↑, H24↓*	H24↓	H6↓		H24↓			L24↓	H24↓	H2↑, H6↑	H2↑*, H6↑*, H24↑*, L2↑, L6↑, L24↑
HDAC4	H2↓, H6↓*, L6↓*, L24↓*	H24↓***, L24↓**	H6↓*, H24↓**	H24↑*	H2↓*, H6↓**, H24↓***, L6↓*, L24↓	L2↓*,	H6↓*, H24↓**, L6↓*	H24↓*, L24↓*,						H2↓*, H6↓**, H24↓**
HDAC5	H6↓*, H24↑*, L24↑	H24↑*	H24↑	H24↑*, L24↑*	H24↑**, L24↑	H24↑*		H24↑*, L24↑*	H6↑*, H24↑*, L24↑*		H6↑, H24↑*, L6↑, L24↑*	H6↑*, H24↑*, L6↑		H24↑*, L24↑	H6↑*, H24↑**, L6↑*, L24↑*
HDAC6	L24↑				H24↑			H24↓, L24↓		H24↑	H24↑, L24↑		H24↑,		H6↓, L6↓
HDAC7	H24↑, L24↑			H24↓*				H24↓							H6↓*, H24↓*, L6↓, L24↓
HDAC9	H6↑*, H24↑**	L24↑***		H6↓**, H24↓**, L24↓*	H2↓*, H6↓*, L24↑*	H24↑*		H24↑***, L2↓, L24↑***	H6↑*, H24↑*	L6↑		H6↑*, H24↑** L24↑*		H2↓*, H6↓**, H24↑**	H2↓**, H6↓*, L6↓*
HDAC11	L24↑			L24↑					L24↑*
SIRT1	H6↑*, H24↑*, L6↑, L24↑*	H6↑*, H24↑*	H6↑, H24↑*		H2↑, H6↑*, H24↑*		H6↓, L6↓	H24↑*, L24↑*	H6↑*	H24↑	L2↓	L2↓		H2↓, H6↑*, H24↑*	H2↓*, L2↓, L6↓, L24↓*
SIRT2	H24↓*	H24↑*, L24↑*	H24↑*	L24↑*	H6↑, L24↑		H24↑*	H24↑, L24↑*	H24↑			H24↑*, L24↑*		H6↑*, H24↑*, L24↑	H6↑*, H24↑*, L6↑, L24↑*
SIRT3		H6↓, H24↓, L24↓		H24↑, L24↑	H24↑*						H24↑	L24↑	H24↑		H6↑, H24↑, L6↑
SIRT4	L24↓			L24↑	H6↑, H24↑*									H6↑*, H24↑*	H6↑*, H24↑*, L6↑*, L24↑*
SIRT5		H6↓, H24↓*, L6↓, L24↓		L24↑	H24↑	H24↑	H24↓				H24↑, L24↑	H24↑, L24↑	H24↑		H2↓, H6↓, L6↓, L24↓
SIRT6	H24↑*	H24↑*	H24↑		H24↑*		H24↓							H6↑*, H24↑*
SIRT7	H24↑*, L24↑	H24↑*, L6↑*, L24↑*	H6↑*, H24↑		H24↑*		H6↓, L2↓, L6↓	H24↑*, L24↑*		H2↓	H6↓, H24↓, L2↓, L6↓, L24↓	H24↑*	H2↓, H6↑, H24↑	H6↑, H24↑*	H2↑*, H6↑*, H24↑*, L2↑*, L6↑*

Columns represent 15 agents in the NCI-TPW dataset. Aza, azacytidine; Bor, bortezomib; Cis, cisplatin; Das, dasatinib; Dox, doxorubicin; Erl, erlotinib; Gel, geldanamycin; Lap, lapatinib; Gem, gemcitabine; Pac, paclitaxel; Rap, rapamycin; Sor, sorafenib; Sun, sunitinib; Top, topotecan; Vor, vorinostat. Concerted changes in expression (↑, upregulated or ↓, downregulated) are shown for microarray experiments in which nearly all cell lines had a change in the same direction, with no more than 15 cell lines showing a change in the opposite direction. Expression changes are shown for the high (H) or low concentration (L) of each agent. The time when the change was observed is also indicated. Concerted changes for multiple conditions for the same drug-gene combination are separated by commas. High and low concentrations of each agent are listed in the Materials and Methods section.

* Concerted expression change as described above and log₂FC (the difference of log₂ expression values between treated and untreated cells) in that direction in some cell lines was ≥ 1 or ≤ −1.

** Concerted expression change and log₂FC in that direction in some cell lines was ≥ 2.5 or ≤ −2.5.

*** Concerted expression change as described above and log₂FC in that direction in some cell lines was ≥ 4 or ≤ −4.

For example, H↑24***, L↓2, L↑24*** for SIRT9 after treatment with gemcitabine (Gem) indicates that it was upregulated in a concerted manner after treatment with both the high and low concentrations of that agent at 24 hours after treatment, with the change of log₂ expression values in at least some cell lines being ≥ 4 or ≤ −4, and that it was also downregulated at 2 hours after treatment with the low concentration of gemcitabine, but the amplitude of the SIRT9 expression changes under that condition did not reach ≥ 1 or ≤ −1.

Figure 1. Examples of changes in expression of HDAC genes in response to treatment with erlotinib and dasatinib. Shown are transcriptional changes (log₂FC) at 2 (left panel), 6 (middle panel), and 24 hr (right panel) after treatment. Horizontal right bars indicate elevated gene expression, whereas left bars show decreased expression relative to the untreated cell lines. As summarized in Table 1, concerted upregulation at 24 hr (shown on the right most panels) was observed for HDAC5 after treatment with (a) the high (2,000 nM) and (b) the low (100 nM) concentrations of dasatinib and (c) the high (10,000 nM) concentration of erlotinib. (d) expression changes of HDAC2 after treatment with the high concentration of dasatinib showed concerted downregulation at 6 hr (middle panel) and 24 hr (right panel). Colors represent cancer categories. The scale on the bottom represents log₂ difference between expression values of treated and untreated cell lines. The scale for each microarray experiment is specific to that experiment.

Table 2. Spearman correlations between log(GI50) and log₂FC of HDAC and SIRT genes satisfying p_FDR <0.05.

Agent	Gene	Condition	Spearman ρ	p0	pFDR
Dasatinib	HDAC5	High 24	−0.7806	3.74 × 10^−11	6.28 × 10^−8
Dasatinib	HDAC5	Low 24	−0.6871	2.58 × 10^−8	2.17 × 10^−5
Dasatinib	HDAC2	High 24	0.5774	1.40 × 10^−5	0.00336
Dasatinib	SIRT1	High 6	−0.5031	9.05 × 10^−7	0.01382
Dasatinib	SIRT2	High 24	−0.5158	0.00015	0.01925
Dasatinib	HDAC1	Low 6	−0.4628	0.00037	0.04187
Dasatinib	HDAC1	High 6	−0.4551	0.00048	0.04490
Erlotinib	HDAC5	High 24	−0.4625	0.00023	0.02714
Erlotinib	SIRT5	High 24	−0.4429	0.00044	0.04377
Cisplatin	HDAC4	High 24	0.6251	3.37 × 10^−7	0.00019
Cisplatin	SIRT1	High 24	−0.5730	4.83 × 10^−6	0.00190
Cisplatin	HDAC1	High 24	0.4913	0.00014	0.01925
Gemcitabine	HDAC1	Low 24	0.4595	0.00042	0.04377
Topotecan	HDAC1	High 24	0.5429	1.07 × 10^−5	0.00300
Geldanamycin	HDAC4	Low 24	0.5653	5.65 × 10^−6	0.00190
Azacytidine	HDAC9	High 24	−0.6136	3.27 × 10^−5	0.00687
Paclitaxel	HDAC9	Low 24	0.4920	6.53 × 10^−5	0.01219

The results are grouped by agent and sorted by p_FDR.

$\mathit{\rho }$, Spearman correlation coefficient.

Condition indicates concentration (high or low) and the time point (6 or 24 hr after treatment). None of the changes at 2 hr after treatment satisfied p_FDR < 0.05.

p₀, p-value prior to the FDR adjustment.

p_FDR, FDR adjusted p-value.

Figure 2. Experimental validation of transcriptional and protein levels of HDAC5 in the IGROV1, MDA-MB-231, and UACC-257 cell lines after treatment with 2000 nM of dasatinib. (a) Transcriptional changes in mRNA levels using RT-PCR. Results of two independent probes to HDAC5 are normalized to GAPDH. (b) Changes in protein levels using Western blots. Protein results are first normalized to GAPDH and then values divided by normalized untreated cells. Full blots are shown.

Data availability statement

The datasets analysed in the current study are publicly available from the NCI-TPW portal, [https://tpwb.nci.nih.gov] [33] (NCI-TPW drug response data, gene expression changes, and baseline gene expression data and their graphical presentation) and NCBI GEO, [https://www.ncbi.nlm.nih.gov/geo/] (public datasets used for validation of HDAC and SIRT and miRNA expression changes in response to drug treatment). The dataset of mRNA and protein changes of HDAC5 in the IGROV1, MDA-MB-231, and UACC-257 cell lines in response to dasatinib used during the current study is available from the corresponding author on reasonable request.