Limits...
A transcriptomic signature mediated by HOXA9 promotes human glioblastoma initiation, aggressiveness and resistance to temozolomide.

Pojo M, Gonçalves CS, Xavier-Magalhães A, Oliveira AI, Gonçalves T, Correia S, Rodrigues AJ, Costa S, Pinto L, Pinto AA, Lopes JM, Reis RM, Rocha M, Sousa N, Costa BM - Oncotarget (2015)

Bottom Line: Additionally, HOXA9 promoted the malignant transformation of human immortalized astrocytes in an orthotopic in vivo model, and caused tumor-associated death.Importantly, the pharmacological inhibition of BCL2 with the BH3 mimetic ABT-737 reverted temozolomide resistance in HOXA9-positive cells.These data establish HOXA9 as a driver of glioma initiation, aggressiveness and resistance to therapy.

View Article: PubMed Central - PubMed

Affiliation: Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Campus de Gualtar 4710-057 Braga, Portugal.

ABSTRACT
Glioblastoma is the most malignant brain tumor, exhibiting remarkable resistance to treatment. Here we investigated the oncogenic potential of HOXA9 in gliomagenesis, the molecular and cellular mechanisms by which HOXA9 renders glioblastoma more aggressive, and how HOXA9 affects response to chemotherapy and survival. The prognostic value of HOXA9 in glioblastoma patients was validated in two large datasets from TCGA and Rembrandt, where high HOXA9 levels were associated with shorter survival. Transcriptomic analyses identified novel HOXA9-target genes with key roles in cancer-related processes, including cell proliferation, DNA repair, and stem cell maintenance. Functional studies with HOXA9-overexpressing and HOXA9-silenced glioblastoma cell models revealed that HOXA9 promotes cell viability, stemness and invasion, and inhibits apoptosis. Additionally, HOXA9 promoted the malignant transformation of human immortalized astrocytes in an orthotopic in vivo model, and caused tumor-associated death. HOXA9 also mediated resistance to temozolomide treatment in vitro and in vivo via upregulation of BCL2. Importantly, the pharmacological inhibition of BCL2 with the BH3 mimetic ABT-737 reverted temozolomide resistance in HOXA9-positive cells. These data establish HOXA9 as a driver of glioma initiation, aggressiveness and resistance to therapy. In the future, the combination of BH3 mimetics with temozolomide should be further explored as an alternative treatment for glioblastoma.

No MeSH data available.


Related in: MedlinePlus

HOXA9 accelerates tumor growth in vivo(A) U87MG cells were subcutaneously xenografted into nude mice; representative photographs of in vivo and ex vivo tumors are shown. (B) Longitudinal assessment of tumor volumes showing increased growth rates of U87MG-HOXA9 tumors as compared to HOXA9-negative tumors. (C) Positive correlation between expression levels of HOXA9 and tumor volume. (D) and (E) Final tumor volumes (D) and weights (E) are significantly higher in HOXA9-positive tumors (n = 10). (F) Hematoxilin-eosin and immunohistochemical staining showing that HOXA9-positive tumors present higher protein levels of Cyclin D1, Ki-67, BCL2, Nestin, and PECAM1. (G) Quantification of the % of vascular area based on PECAM1 staining, showing increased angiogenic potential of U87MG-HOXA9 tumors. Statistical differences were calculated by two-way ANOVA (B) and Spearman correlation (C), or t-tests (D and E) (*p < 0.05; **p < 0.01; ***p < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4480707&req=5

Figure 4: HOXA9 accelerates tumor growth in vivo(A) U87MG cells were subcutaneously xenografted into nude mice; representative photographs of in vivo and ex vivo tumors are shown. (B) Longitudinal assessment of tumor volumes showing increased growth rates of U87MG-HOXA9 tumors as compared to HOXA9-negative tumors. (C) Positive correlation between expression levels of HOXA9 and tumor volume. (D) and (E) Final tumor volumes (D) and weights (E) are significantly higher in HOXA9-positive tumors (n = 10). (F) Hematoxilin-eosin and immunohistochemical staining showing that HOXA9-positive tumors present higher protein levels of Cyclin D1, Ki-67, BCL2, Nestin, and PECAM1. (G) Quantification of the % of vascular area based on PECAM1 staining, showing increased angiogenic potential of U87MG-HOXA9 tumors. Statistical differences were calculated by two-way ANOVA (B) and Spearman correlation (C), or t-tests (D and E) (*p < 0.05; **p < 0.01; ***p < 0.001).

Mentions: Considering that stem cell properties have been linked with tumor malignancy [14, 26, 27], together with our microarray data implicating the HOXA9 transcriptome in typical hallmarks of cancer (Supplementary Figure 3, Supplementary Figure 10 and Supplementary Table 1), we hypothesized that HOXA9 could play a role in tumor growth in vivo. To test this hypothesis, we established subcutaneous GBM xenograft models with U87MG cells. Tumors derived from U87MG-HOXA9 cells presented increased growth rates and were significantly larger than U87MG-MSCV cells (Figure 4A–4E). Interestingly, the final tumor volume was significantly correlated with HOXA9 expression levels (Figure 4C), further strengthening the relevance of HOXA9 on tumor growth kinetics in vivo. Histologically, HOXA9 positive tumors presented a higher proliferative activity, as indicated by increased Ki-67 staining, which includes mostly neoplastic cells but also non-neoplastic cells (e.g. endothelial and stromal cells; Figure 4F). A higher expression of Nestin was also observed, supporting an increased stemness potential of HOXA9-positive cells in vivo. Furthermore, the expression levels of Cyclin D1 and BCL2 were also increased in U87MG-HOXA9 tumors in vivo (Figure 4F). These proteins are critical regulators of cell cycle and cell death, respectively, and were identified as HOXA9 targets in the microarray data. Tumor-associated angiogenesis, another critical hallmark of GBM, as assessed by PECAM1 protein staining, was significantly increased in HOXA9-positive tumors (Figure 4F and 4G). Together, these data establish HOXA9 as a critical mediator of GBM growth and aggressiveness in vivo, and identifies critical molecular mediators sustaining this malignant phenotype.


A transcriptomic signature mediated by HOXA9 promotes human glioblastoma initiation, aggressiveness and resistance to temozolomide.

Pojo M, Gonçalves CS, Xavier-Magalhães A, Oliveira AI, Gonçalves T, Correia S, Rodrigues AJ, Costa S, Pinto L, Pinto AA, Lopes JM, Reis RM, Rocha M, Sousa N, Costa BM - Oncotarget (2015)

HOXA9 accelerates tumor growth in vivo(A) U87MG cells were subcutaneously xenografted into nude mice; representative photographs of in vivo and ex vivo tumors are shown. (B) Longitudinal assessment of tumor volumes showing increased growth rates of U87MG-HOXA9 tumors as compared to HOXA9-negative tumors. (C) Positive correlation between expression levels of HOXA9 and tumor volume. (D) and (E) Final tumor volumes (D) and weights (E) are significantly higher in HOXA9-positive tumors (n = 10). (F) Hematoxilin-eosin and immunohistochemical staining showing that HOXA9-positive tumors present higher protein levels of Cyclin D1, Ki-67, BCL2, Nestin, and PECAM1. (G) Quantification of the % of vascular area based on PECAM1 staining, showing increased angiogenic potential of U87MG-HOXA9 tumors. Statistical differences were calculated by two-way ANOVA (B) and Spearman correlation (C), or t-tests (D and E) (*p < 0.05; **p < 0.01; ***p < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480707&req=5

Figure 4: HOXA9 accelerates tumor growth in vivo(A) U87MG cells were subcutaneously xenografted into nude mice; representative photographs of in vivo and ex vivo tumors are shown. (B) Longitudinal assessment of tumor volumes showing increased growth rates of U87MG-HOXA9 tumors as compared to HOXA9-negative tumors. (C) Positive correlation between expression levels of HOXA9 and tumor volume. (D) and (E) Final tumor volumes (D) and weights (E) are significantly higher in HOXA9-positive tumors (n = 10). (F) Hematoxilin-eosin and immunohistochemical staining showing that HOXA9-positive tumors present higher protein levels of Cyclin D1, Ki-67, BCL2, Nestin, and PECAM1. (G) Quantification of the % of vascular area based on PECAM1 staining, showing increased angiogenic potential of U87MG-HOXA9 tumors. Statistical differences were calculated by two-way ANOVA (B) and Spearman correlation (C), or t-tests (D and E) (*p < 0.05; **p < 0.01; ***p < 0.001).
Mentions: Considering that stem cell properties have been linked with tumor malignancy [14, 26, 27], together with our microarray data implicating the HOXA9 transcriptome in typical hallmarks of cancer (Supplementary Figure 3, Supplementary Figure 10 and Supplementary Table 1), we hypothesized that HOXA9 could play a role in tumor growth in vivo. To test this hypothesis, we established subcutaneous GBM xenograft models with U87MG cells. Tumors derived from U87MG-HOXA9 cells presented increased growth rates and were significantly larger than U87MG-MSCV cells (Figure 4A–4E). Interestingly, the final tumor volume was significantly correlated with HOXA9 expression levels (Figure 4C), further strengthening the relevance of HOXA9 on tumor growth kinetics in vivo. Histologically, HOXA9 positive tumors presented a higher proliferative activity, as indicated by increased Ki-67 staining, which includes mostly neoplastic cells but also non-neoplastic cells (e.g. endothelial and stromal cells; Figure 4F). A higher expression of Nestin was also observed, supporting an increased stemness potential of HOXA9-positive cells in vivo. Furthermore, the expression levels of Cyclin D1 and BCL2 were also increased in U87MG-HOXA9 tumors in vivo (Figure 4F). These proteins are critical regulators of cell cycle and cell death, respectively, and were identified as HOXA9 targets in the microarray data. Tumor-associated angiogenesis, another critical hallmark of GBM, as assessed by PECAM1 protein staining, was significantly increased in HOXA9-positive tumors (Figure 4F and 4G). Together, these data establish HOXA9 as a critical mediator of GBM growth and aggressiveness in vivo, and identifies critical molecular mediators sustaining this malignant phenotype.

Bottom Line: Additionally, HOXA9 promoted the malignant transformation of human immortalized astrocytes in an orthotopic in vivo model, and caused tumor-associated death.Importantly, the pharmacological inhibition of BCL2 with the BH3 mimetic ABT-737 reverted temozolomide resistance in HOXA9-positive cells.These data establish HOXA9 as a driver of glioma initiation, aggressiveness and resistance to therapy.

View Article: PubMed Central - PubMed

Affiliation: Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Campus de Gualtar 4710-057 Braga, Portugal.

ABSTRACT
Glioblastoma is the most malignant brain tumor, exhibiting remarkable resistance to treatment. Here we investigated the oncogenic potential of HOXA9 in gliomagenesis, the molecular and cellular mechanisms by which HOXA9 renders glioblastoma more aggressive, and how HOXA9 affects response to chemotherapy and survival. The prognostic value of HOXA9 in glioblastoma patients was validated in two large datasets from TCGA and Rembrandt, where high HOXA9 levels were associated with shorter survival. Transcriptomic analyses identified novel HOXA9-target genes with key roles in cancer-related processes, including cell proliferation, DNA repair, and stem cell maintenance. Functional studies with HOXA9-overexpressing and HOXA9-silenced glioblastoma cell models revealed that HOXA9 promotes cell viability, stemness and invasion, and inhibits apoptosis. Additionally, HOXA9 promoted the malignant transformation of human immortalized astrocytes in an orthotopic in vivo model, and caused tumor-associated death. HOXA9 also mediated resistance to temozolomide treatment in vitro and in vivo via upregulation of BCL2. Importantly, the pharmacological inhibition of BCL2 with the BH3 mimetic ABT-737 reverted temozolomide resistance in HOXA9-positive cells. These data establish HOXA9 as a driver of glioma initiation, aggressiveness and resistance to therapy. In the future, the combination of BH3 mimetics with temozolomide should be further explored as an alternative treatment for glioblastoma.

No MeSH data available.


Related in: MedlinePlus