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Identification of the Adapter Molecule MTSS1 as a Potential Oncogene-Specific Tumor Suppressor in Acute Myeloid Leukemia.

Schemionek M, Kharabi Masouleh B, Klaile Y, Krug U, Hebestreit K, Schubert C, Dugas M, Büchner T, Wörmann B, Hiddemann W, Berdel WE, Brümmendorf TH, Müller-Tidow C, Koschmieder S - PLoS ONE (2015)

Bottom Line: High expression of MTSS1 predicted better clinical outcome of patients with normal-karyotype AML.Pharmacological treatment of AML cell lines carrying the FLT3-ITD mutation with the specific FLT3 inhibitor PKC-412 caused upregulation of MTSS1.Moreover, treatment of acute promyelocytic cells (APL) with all-trans retinoic acid (ATRA) increased MTSS1 mRNA levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany.

ABSTRACT
The adapter protein metastasis suppressor 1 (MTSS1) is implicated as a tumor suppressor or tumor promoter, depending on the type of solid cancer. Here, we identified Mtss1 expression to be increased in AML subsets with favorable outcome, while suppressed in high risk AML patients. High expression of MTSS1 predicted better clinical outcome of patients with normal-karyotype AML. Mechanistically, MTSS1 expression was negatively regulated by FLT3-ITD signaling but enhanced by the AML1-ETO fusion protein. DNMT3B, a negative regulator of MTSS1, showed strong binding to the MTSS1 promoter in PML-RARA positive but not AML1-ETO positive cells, suggesting that AML1-ETO leads to derepression of MTSS1. Pharmacological treatment of AML cell lines carrying the FLT3-ITD mutation with the specific FLT3 inhibitor PKC-412 caused upregulation of MTSS1. Moreover, treatment of acute promyelocytic cells (APL) with all-trans retinoic acid (ATRA) increased MTSS1 mRNA levels. Taken together, our findings suggest that MTSS1 might have a context-dependent function and could act as a tumor suppressor, which is pharmacologically targetable in AML patients.

No MeSH data available.


Related in: MedlinePlus

MTSS1 expression correlates with better clinical outcome of normal karyotype-AML patients.MTSS1 mRNA levels were measured in patient-derived AML samples (n = 66) with either a Normal Karyotype (NK, n = 38), t(15;17) (n = 15), t(8;21) (n = 5) or inv(16) (n = 8) aberration by qRT-PCR and are expressed as % of GAPDH (A). Using the Leukemia Gene Atlas [37] and data from the German AMLCG 1999 clinical trial [19], two groups of patients (above vs. below the median expression of MTSS1) were analyzed for overall survival (OS) in NK AML patients (n = 163, logrank test P = 0.03; GEO accession number GSE12417; B). Similarly, in the same trial, patients with a DNMT3B expression above vs. below the median were analyzed for survival (n = 163, logrank test P = 0.04; C). We then segregated patients into two groups based on their MTSS1 and DNMT3B expression levels according to high MTSS1 and low DNMT3B or low MTSS1 and high DNMT3B expression and assessed overall survival (AMLCG 1999, n = 163, logrank test P = 0.006; D). Binding of DNMT3B to the MTSS1 promoter was analyzed by chromatin immunoprecipitation experiments using NB4 and Kasumi-1 cells. Precipitated ChIP-DNA was quantified using real-time PCR and SYBR Green for MTSS1 promoter region -864/-645 (E).
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pone.0125783.g002: MTSS1 expression correlates with better clinical outcome of normal karyotype-AML patients.MTSS1 mRNA levels were measured in patient-derived AML samples (n = 66) with either a Normal Karyotype (NK, n = 38), t(15;17) (n = 15), t(8;21) (n = 5) or inv(16) (n = 8) aberration by qRT-PCR and are expressed as % of GAPDH (A). Using the Leukemia Gene Atlas [37] and data from the German AMLCG 1999 clinical trial [19], two groups of patients (above vs. below the median expression of MTSS1) were analyzed for overall survival (OS) in NK AML patients (n = 163, logrank test P = 0.03; GEO accession number GSE12417; B). Similarly, in the same trial, patients with a DNMT3B expression above vs. below the median were analyzed for survival (n = 163, logrank test P = 0.04; C). We then segregated patients into two groups based on their MTSS1 and DNMT3B expression levels according to high MTSS1 and low DNMT3B or low MTSS1 and high DNMT3B expression and assessed overall survival (AMLCG 1999, n = 163, logrank test P = 0.006; D). Binding of DNMT3B to the MTSS1 promoter was analyzed by chromatin immunoprecipitation experiments using NB4 and Kasumi-1 cells. Precipitated ChIP-DNA was quantified using real-time PCR and SYBR Green for MTSS1 promoter region -864/-645 (E).

Mentions: In our cohort of patients with AML, MTSS1 expression was confirmed by qRT-PCR to be increased in t(8;21) or inv(16) AML samples as compared to t(15;17) (Fig 2A). Interestingly, when we analyzed samples from patients with normal karyotype AML, we found MTSS1 expression to be highly variable (Fig 2A). We reasoned that since high MTSS1 expression was found in prognostically favorable AML subgroups, the prognosis of patients with normal karyotypes may differ according to their level of MTSS1 expression. Indeed, when 163 patients with normal karyotype-AML in the German AMLCG 1999 clinical trial [19] were separated into two groups depending on whether their expression of MTSS1 is below or above the MTSS1-probeset, median across all samples, patients with high expression levels of MTSS1 (MTSS1HI) showed a significantly improved overall survival suggesting that MTSS1 is a predictor of favorable outcome (Fig 2B). In the same clinical trial, high expression of DNMT3B, which has been shown to impede MTSS1 expression, showed reversed patterns and correlated with decreased overall survival (Fig 2C). To study if there is a link between MTSS1 and DNMT3B, we segregated the patients in two groups based either on their high MTSS1 and low DNMT3B (MTSS1HI—DNMT3BLO) or low MTSS1 and high DNMT3B expression (MTSS1LO—DNMT3BHI). Strikingly, patients with high MTSS1 and low DNMT3B expression showed a highly significant increased overall survival rate compared to those with low MTSS1 and high DNMT3B expression (Fig 2D). DNMT3B binding to the MTSS1 promoter has previously been described to occur at -864/-645 bp upstream of the transcriptional start site and acts as a mechanism that allows for transcriptional suppression of MTSS1 in hepatocellular carcinoma [6]. To analyze for a direct binding of DNMT3B to the respective MTSS1 promoter region in AML, we performed chromatin immunoprecipitation experiments. We previously detected increased MTSS1 expression in t(8;21) positive cells but not in t(15;17) positive AML (Fig 1B and 1C) and therefore compared DNMT3B binding to the MTSS1 promoter using these two cell types (Fig 2E). We confirmed binding of DNMT3B to the MTSS1 promoter region (-864/-645) in t(15;17) positive cells. Interestingly, this interaction was not detectable in t(8;21) MTSS1 highly expressing AML cells. These data confirm a direct link between DNMT3B activity and decreased MTSS1 expression also in a subset of AML.


Identification of the Adapter Molecule MTSS1 as a Potential Oncogene-Specific Tumor Suppressor in Acute Myeloid Leukemia.

Schemionek M, Kharabi Masouleh B, Klaile Y, Krug U, Hebestreit K, Schubert C, Dugas M, Büchner T, Wörmann B, Hiddemann W, Berdel WE, Brümmendorf TH, Müller-Tidow C, Koschmieder S - PLoS ONE (2015)

MTSS1 expression correlates with better clinical outcome of normal karyotype-AML patients.MTSS1 mRNA levels were measured in patient-derived AML samples (n = 66) with either a Normal Karyotype (NK, n = 38), t(15;17) (n = 15), t(8;21) (n = 5) or inv(16) (n = 8) aberration by qRT-PCR and are expressed as % of GAPDH (A). Using the Leukemia Gene Atlas [37] and data from the German AMLCG 1999 clinical trial [19], two groups of patients (above vs. below the median expression of MTSS1) were analyzed for overall survival (OS) in NK AML patients (n = 163, logrank test P = 0.03; GEO accession number GSE12417; B). Similarly, in the same trial, patients with a DNMT3B expression above vs. below the median were analyzed for survival (n = 163, logrank test P = 0.04; C). We then segregated patients into two groups based on their MTSS1 and DNMT3B expression levels according to high MTSS1 and low DNMT3B or low MTSS1 and high DNMT3B expression and assessed overall survival (AMLCG 1999, n = 163, logrank test P = 0.006; D). Binding of DNMT3B to the MTSS1 promoter was analyzed by chromatin immunoprecipitation experiments using NB4 and Kasumi-1 cells. Precipitated ChIP-DNA was quantified using real-time PCR and SYBR Green for MTSS1 promoter region -864/-645 (E).
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pone.0125783.g002: MTSS1 expression correlates with better clinical outcome of normal karyotype-AML patients.MTSS1 mRNA levels were measured in patient-derived AML samples (n = 66) with either a Normal Karyotype (NK, n = 38), t(15;17) (n = 15), t(8;21) (n = 5) or inv(16) (n = 8) aberration by qRT-PCR and are expressed as % of GAPDH (A). Using the Leukemia Gene Atlas [37] and data from the German AMLCG 1999 clinical trial [19], two groups of patients (above vs. below the median expression of MTSS1) were analyzed for overall survival (OS) in NK AML patients (n = 163, logrank test P = 0.03; GEO accession number GSE12417; B). Similarly, in the same trial, patients with a DNMT3B expression above vs. below the median were analyzed for survival (n = 163, logrank test P = 0.04; C). We then segregated patients into two groups based on their MTSS1 and DNMT3B expression levels according to high MTSS1 and low DNMT3B or low MTSS1 and high DNMT3B expression and assessed overall survival (AMLCG 1999, n = 163, logrank test P = 0.006; D). Binding of DNMT3B to the MTSS1 promoter was analyzed by chromatin immunoprecipitation experiments using NB4 and Kasumi-1 cells. Precipitated ChIP-DNA was quantified using real-time PCR and SYBR Green for MTSS1 promoter region -864/-645 (E).
Mentions: In our cohort of patients with AML, MTSS1 expression was confirmed by qRT-PCR to be increased in t(8;21) or inv(16) AML samples as compared to t(15;17) (Fig 2A). Interestingly, when we analyzed samples from patients with normal karyotype AML, we found MTSS1 expression to be highly variable (Fig 2A). We reasoned that since high MTSS1 expression was found in prognostically favorable AML subgroups, the prognosis of patients with normal karyotypes may differ according to their level of MTSS1 expression. Indeed, when 163 patients with normal karyotype-AML in the German AMLCG 1999 clinical trial [19] were separated into two groups depending on whether their expression of MTSS1 is below or above the MTSS1-probeset, median across all samples, patients with high expression levels of MTSS1 (MTSS1HI) showed a significantly improved overall survival suggesting that MTSS1 is a predictor of favorable outcome (Fig 2B). In the same clinical trial, high expression of DNMT3B, which has been shown to impede MTSS1 expression, showed reversed patterns and correlated with decreased overall survival (Fig 2C). To study if there is a link between MTSS1 and DNMT3B, we segregated the patients in two groups based either on their high MTSS1 and low DNMT3B (MTSS1HI—DNMT3BLO) or low MTSS1 and high DNMT3B expression (MTSS1LO—DNMT3BHI). Strikingly, patients with high MTSS1 and low DNMT3B expression showed a highly significant increased overall survival rate compared to those with low MTSS1 and high DNMT3B expression (Fig 2D). DNMT3B binding to the MTSS1 promoter has previously been described to occur at -864/-645 bp upstream of the transcriptional start site and acts as a mechanism that allows for transcriptional suppression of MTSS1 in hepatocellular carcinoma [6]. To analyze for a direct binding of DNMT3B to the respective MTSS1 promoter region in AML, we performed chromatin immunoprecipitation experiments. We previously detected increased MTSS1 expression in t(8;21) positive cells but not in t(15;17) positive AML (Fig 1B and 1C) and therefore compared DNMT3B binding to the MTSS1 promoter using these two cell types (Fig 2E). We confirmed binding of DNMT3B to the MTSS1 promoter region (-864/-645) in t(15;17) positive cells. Interestingly, this interaction was not detectable in t(8;21) MTSS1 highly expressing AML cells. These data confirm a direct link between DNMT3B activity and decreased MTSS1 expression also in a subset of AML.

Bottom Line: High expression of MTSS1 predicted better clinical outcome of patients with normal-karyotype AML.Pharmacological treatment of AML cell lines carrying the FLT3-ITD mutation with the specific FLT3 inhibitor PKC-412 caused upregulation of MTSS1.Moreover, treatment of acute promyelocytic cells (APL) with all-trans retinoic acid (ATRA) increased MTSS1 mRNA levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany.

ABSTRACT
The adapter protein metastasis suppressor 1 (MTSS1) is implicated as a tumor suppressor or tumor promoter, depending on the type of solid cancer. Here, we identified Mtss1 expression to be increased in AML subsets with favorable outcome, while suppressed in high risk AML patients. High expression of MTSS1 predicted better clinical outcome of patients with normal-karyotype AML. Mechanistically, MTSS1 expression was negatively regulated by FLT3-ITD signaling but enhanced by the AML1-ETO fusion protein. DNMT3B, a negative regulator of MTSS1, showed strong binding to the MTSS1 promoter in PML-RARA positive but not AML1-ETO positive cells, suggesting that AML1-ETO leads to derepression of MTSS1. Pharmacological treatment of AML cell lines carrying the FLT3-ITD mutation with the specific FLT3 inhibitor PKC-412 caused upregulation of MTSS1. Moreover, treatment of acute promyelocytic cells (APL) with all-trans retinoic acid (ATRA) increased MTSS1 mRNA levels. Taken together, our findings suggest that MTSS1 might have a context-dependent function and could act as a tumor suppressor, which is pharmacologically targetable in AML patients.

No MeSH data available.


Related in: MedlinePlus