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Identification of p38β as a therapeutic target for the treatment of Sézary syndrome.

Bliss-Moreau M, Coarfa C, Gunaratne PH, Guitart J, Krett NL, Rosen ST - J. Invest. Dermatol. (2014)

Bottom Line: Gene set enrichment analysis uncovered candidate genes enriched for an immune-cell signature, specifically the T-cell receptor and mitogen-activated protein kinase signaling pathways.Further analysis identified p38 as a potential therapeutic target that is overexpressed in SS patients and decreased by synergistic-inhibitor treatment.This target was verified through small-molecule inhibition of p38, leading to cell death in both SS cell lines and patient cells.

View Article: PubMed Central - PubMed

Affiliation: Robert H. Lurie Comprehensive Cancer Center, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.

ABSTRACT
Cutaneous T-cell lymphomas (CTCLs) represent a group of hematopoietic malignancies that home to the skin and have no known molecular basis for disease pathogenesis. Sézary syndrome (SS) is the leukemic variant of CTCL. Currently, CTCL is incurable, highlighting the need for new therapeutic modalities. We have previously observed that combined small-molecule inhibition of protein kinase C-β (PKCβ) and glycogen synthase kinase 3 (GSK3) causes synergistic apoptosis in CTCL cell lines and patient cells. Through microarray analysis of a SS cell line, we surveyed global gene expression following combined PKCβ-GSK3 treatment to elucidate therapeutic targets responsible for cell death. Clinically relevant targets were defined as genes differentially expressed in SS patients that were modulated by combination-drug treatment of SS cells. Gene set enrichment analysis uncovered candidate genes enriched for an immune-cell signature, specifically the T-cell receptor and mitogen-activated protein kinase signaling pathways. Further analysis identified p38 as a potential therapeutic target that is overexpressed in SS patients and decreased by synergistic-inhibitor treatment. This target was verified through small-molecule inhibition of p38, leading to cell death in both SS cell lines and patient cells. These data establish p38 as a SS biomarker and a potential therapeutic target for the treatment of CTCL.

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Importance of p38 signaling in primary SS patient samples(a–c) PBMCs were harvested from SS patients (n=3) and healthy volunteers (n=5) and treated with increasing concentrations of either SB202, SB203, or BIRB. Cell death was assessed by flow cytometry. Annexin V staining, positive stained cells quantified as percentage total PBMCs (gating examples: Supplemental Figure S7, online). (a) SB202 (b) SB203 (c) BIRB.
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Figure 5: Importance of p38 signaling in primary SS patient samples(a–c) PBMCs were harvested from SS patients (n=3) and healthy volunteers (n=5) and treated with increasing concentrations of either SB202, SB203, or BIRB. Cell death was assessed by flow cytometry. Annexin V staining, positive stained cells quantified as percentage total PBMCs (gating examples: Supplemental Figure S7, online). (a) SB202 (b) SB203 (c) BIRB.

Mentions: To validate our in vitro observations and determine the clinical relevance of our small-molecule findings, we treated SS patient cells with p38 inhibitors ex vivo. Literature suggests that isolation procedures for blood cells can affect the activation status of the p38-MAPK pathway (Branger et al. 2002). Therefore, we used peripheral blood mononuclear cells (PBMC) from both patients and healthy volunteers rather than enriching for T-cells. After five days in culture, vehicle treated SS patient PBMCs showed a modest, but not statistically significant increase in baseline cell death (Figure 5 and Supplemental Figure S7, online). Apoptosis measured by Annexin V staining was significantly enhanced following low dose (10–25µM) SB202 treatment, in patient samples compared to healthy volunteers (Figure 5a). SB203 treatment had significant effects on patient PBMCs cell death across the full dose range tested (Figure 5b). However, treatment with BIRB, the pan p38 inhibitor, only caused a significant increase of apoptosis in SS patient cells at the lowest dose assayed (10µM; Figure 5c). Apoptotic cell death was confirmed by the measurement of cleaved caspase staining (Supplemental Figures S7 and S8, online). Overall, these data parallel our observations made in cell lines (Figure 4a). We conclude that p38 may be a therapeutic target in SS and the inhibition of p38 using targeted small molecules may provide clinical benefit for SS patients.


Identification of p38β as a therapeutic target for the treatment of Sézary syndrome.

Bliss-Moreau M, Coarfa C, Gunaratne PH, Guitart J, Krett NL, Rosen ST - J. Invest. Dermatol. (2014)

Importance of p38 signaling in primary SS patient samples(a–c) PBMCs were harvested from SS patients (n=3) and healthy volunteers (n=5) and treated with increasing concentrations of either SB202, SB203, or BIRB. Cell death was assessed by flow cytometry. Annexin V staining, positive stained cells quantified as percentage total PBMCs (gating examples: Supplemental Figure S7, online). (a) SB202 (b) SB203 (c) BIRB.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4289446&req=5

Figure 5: Importance of p38 signaling in primary SS patient samples(a–c) PBMCs were harvested from SS patients (n=3) and healthy volunteers (n=5) and treated with increasing concentrations of either SB202, SB203, or BIRB. Cell death was assessed by flow cytometry. Annexin V staining, positive stained cells quantified as percentage total PBMCs (gating examples: Supplemental Figure S7, online). (a) SB202 (b) SB203 (c) BIRB.
Mentions: To validate our in vitro observations and determine the clinical relevance of our small-molecule findings, we treated SS patient cells with p38 inhibitors ex vivo. Literature suggests that isolation procedures for blood cells can affect the activation status of the p38-MAPK pathway (Branger et al. 2002). Therefore, we used peripheral blood mononuclear cells (PBMC) from both patients and healthy volunteers rather than enriching for T-cells. After five days in culture, vehicle treated SS patient PBMCs showed a modest, but not statistically significant increase in baseline cell death (Figure 5 and Supplemental Figure S7, online). Apoptosis measured by Annexin V staining was significantly enhanced following low dose (10–25µM) SB202 treatment, in patient samples compared to healthy volunteers (Figure 5a). SB203 treatment had significant effects on patient PBMCs cell death across the full dose range tested (Figure 5b). However, treatment with BIRB, the pan p38 inhibitor, only caused a significant increase of apoptosis in SS patient cells at the lowest dose assayed (10µM; Figure 5c). Apoptotic cell death was confirmed by the measurement of cleaved caspase staining (Supplemental Figures S7 and S8, online). Overall, these data parallel our observations made in cell lines (Figure 4a). We conclude that p38 may be a therapeutic target in SS and the inhibition of p38 using targeted small molecules may provide clinical benefit for SS patients.

Bottom Line: Gene set enrichment analysis uncovered candidate genes enriched for an immune-cell signature, specifically the T-cell receptor and mitogen-activated protein kinase signaling pathways.Further analysis identified p38 as a potential therapeutic target that is overexpressed in SS patients and decreased by synergistic-inhibitor treatment.This target was verified through small-molecule inhibition of p38, leading to cell death in both SS cell lines and patient cells.

View Article: PubMed Central - PubMed

Affiliation: Robert H. Lurie Comprehensive Cancer Center, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.

ABSTRACT
Cutaneous T-cell lymphomas (CTCLs) represent a group of hematopoietic malignancies that home to the skin and have no known molecular basis for disease pathogenesis. Sézary syndrome (SS) is the leukemic variant of CTCL. Currently, CTCL is incurable, highlighting the need for new therapeutic modalities. We have previously observed that combined small-molecule inhibition of protein kinase C-β (PKCβ) and glycogen synthase kinase 3 (GSK3) causes synergistic apoptosis in CTCL cell lines and patient cells. Through microarray analysis of a SS cell line, we surveyed global gene expression following combined PKCβ-GSK3 treatment to elucidate therapeutic targets responsible for cell death. Clinically relevant targets were defined as genes differentially expressed in SS patients that were modulated by combination-drug treatment of SS cells. Gene set enrichment analysis uncovered candidate genes enriched for an immune-cell signature, specifically the T-cell receptor and mitogen-activated protein kinase signaling pathways. Further analysis identified p38 as a potential therapeutic target that is overexpressed in SS patients and decreased by synergistic-inhibitor treatment. This target was verified through small-molecule inhibition of p38, leading to cell death in both SS cell lines and patient cells. These data establish p38 as a SS biomarker and a potential therapeutic target for the treatment of CTCL.

Show MeSH
Related in: MedlinePlus