Identification of key genes and specific pathways potentially involved in androgen-independent, mitoxantrone-resistant prostate cancer

Figures & data

Figure 1 Tumor growth effect of a 6-week treatment with MTX and/or castration.

Notes: (A) Antitumor activities of different concentrations of MTX (control, 0.35, 1, and 3.5 mg/kg) in PCa xenograft mouse models. (B) Combined effects of androgen deprivation and MTX on PCa xenograft growth (P<0.015) (n=5 mice per group).

Abbreviations: MTX, mitoxantrone; PCa, prostate cancer; wk, Week.

Table 1 Top ten up- and downexpressed genes in xenograft of VCaP vs its parent cells

Primary accession	Gene symbol	Log2 ratio	Main function
NM_012445	SPON2	5.524	Antigen binding and lipopolysaccharide binding
NM_001099	ACPP	5.383	Protein homodimerization activity and phosphatase activity
NM_006998	SCGN	5.262	Calcium ion binding and cell proliferation
NM_002982	CCL2	4.853	Chemotactic activity for monocytes and basophils
NM_001618	PARP1	4.632	Poly(A) RNA binding and protein kinase binding
NM_000454	SOD1	4.354	Protein homodimerization activity and enzyme binding
NM_001353	AKR1C1	4.123	Oxidoreductase activity and oxidoreductase activity
NM_020783	SYT4	3.518	Ca(2+)-dependent exocytosis and dendrite formation
NM_000905	NPY	3.473	G-protein-coupled receptor activity and hormone activity
NM_002592	PCNA	3.216	Involved in the RAD6-dependent DNA repair pathway
NM_152703	SAMD9L	−3.074	Endosome fusion, and downregulation of growth factor signaling
NM_030763	HMGN5	−3.132	Preferentially binds to euchromatin and modulates cellular transcription
NM_012242	DKK1	−3.273	Signal transducer activity and low-density lipoprotein particle receptor binding
NM_00100839	GPX8	−3.452	Glutathione peroxidase activity
NM_004929	CALB1	−3.638	Calcium ion binding and vitamin D binding
NM_015163	TRIM9	−4.264	A member of tripartite motif family whose function has not been identified
NM_005978	S100A2	−4.316	Regulation of cellular processes, cell cycle progression, and differentiation
NM_001039492	FHL2	−4.817	Involved in the assembly of extracellular membranes
NM_175887	PRR15	−5.135	May have a role in cell proliferation and/or differentiation
NM_001753	CAV1	−5.237	A negative regulator of the Ras-p42/44 mitogen-activated kinase cascade

Table 2 Top ten up- and downexpressed genes in xenograft of CWR22R vs its parent cells

Primary accession	Gene symbol	Log2 ratio	Main function
NM_002996	CX3CL1	5.721	Receptor binding and chemokine activity
NM_001633	AMBP	5.635	Regulation of inflammatory and pathological processes
NM_000905	NPY	5.274	G-protein-coupled receptor activity and hormone activity
NM_004438	EPHA4	5.126	Identical protein binding and protein kinase activity
NM_001353	AKR1C1	4.823	Oxidoreductase activity and oxidoreductase activity
NM_018014	BCL11A	4.514	Protein homodimerization activity and RNA polymerase II sequence-specific DNA binding
NM_005233	EPHA3	4.465	Transferase activity and protein tyrosine kinase activity
BC005008	CEACAM6	3.917	Cell adhesion affects the sensitivity of tumor cells to adenovirus infection
NM_003295	TPT1	3.764	Poly(A) RNA binding and transcription factor binding
NM_014739	BCLAF1	3.693	BCL2-associated transcription factor and poly(A) RNA binding
NM_004653	KDM5D	−3.136	Oxidoreductase activity and histone demethylase activity
NM_148957	TNFRSF19	−3.417	Tumor necrosis factor-activated receptor activity
NM_004126	GNG11	−3.649	GTPase activity and signal transducer activity
NM_005141	FGB	−3.841	Receptor binding and chaperone binding
NM_004651	USP11	−4.132	Cysteine-type endopeptidase activity and thiol-dependent ubiquitinyl hydrolase activity
NM_001733	C1R	−4.545	Calcium ion binding and serine-type peptidase activity
NM_005613	RGS4	−4.674	GTPase activator activity and G-protein alpha-subunit binding
NM_130902	COX7B2	−5.529	Cytochrome-c oxidase activity
NM_012242	DKK1	−5.537	Signal transducer activity and low-density lipoprotein particle receptor binding
NM_006528	TFPI2	−5.746	Serine-type endopeptidase inhibitor activity and peptidase inhibitor activity

Figure 2 Detection of EPHA4, AKR1C3, HIF1A, and HSD17B12 expression.

Notes: Expression data from VCaPR and CWR22R PCa xenografts from castrated, MTX-treated mice, and parental cell lines were assessed by Western blot (A) and qRT-PCR (B). EPHA4, AKR1C3, and HSD17B12 expression was significantly increased, whereas HIF1A expression was dramatically lower in the MTX-treatment xenografts compared to their parental cell lines (P<0.01).

Abbreviations: qRT-PCR, quantitative reverse transcription PCR; MTX, mitoxantrone; PCa, prostate cancer.

Figure 3 qRT-PCR analysis of DEGs identified in the microarray.

Notes: Comparison of DEGs between PCa xenografts from castrated mice and parental cell lines. (A) CWR22R vs CWR22. (B) VCaPR vs VCaP. Expression data are represented by a log ratio calculated by comparing ΔCq from the xenograft with ΔCq from the parent cells. ΔCq was calculated as the difference between Cq of the targeted genes and Cq of the endogenous control gene, β-actin. Using this method, 16/18 (89%) genes pulled from the microarray analysis were confirmed to be differentially expressed by qRT-PCR.

Abbreviations: DEGs, differentially expressed genes; qRT-PCR, quantitative reverse transcription PCR; PCa, prostate cancer.

Figure 4 Pathway function enrichment and deregulated gene networks.

Notes: Representation of canonical pathways across the entire dataset; y-axis displays the significance. For ratios, taller bars indicate more genes associated with the canonical pathway (A). Deregulated gene networks in MTX-treatment xenografts (B and C). Red and green intensities indicate the degree of upregulation and downregulation, respectively. Genes in uncolored notes were not identified as differentially expressed and were integrated into the computationally generated networks based on evidence stored in the IPA knowledge memory, indicating a relevance to this network.

Abbreviations: IPA, Ingenuity Pathway Analysis; MTX, mitoxantrone.

Data availability

The data of the study have been deposited into the Research Data Deposit (http://www.researchdata.org.cn with the Approval Number asRDDB2018000383).