Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase

Elina Mattila, Heidi Marttila, Niko Sahlberg, Pekka Kohonen, Siri Tähtinen, Pasi Halonen, Merja Perälä, Johanna Ivaska

Research output: Contribution to journalArticleScientificpeer-review

22 Citations (Scopus)

Abstract

Background

T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds.

Methods

We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays.

Results

From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells.

Conclusions

In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening to identify small molecule PTP activators that could function as RTK antagonists in cells.

Original languageEnglish
Article number7
Number of pages12
JournalBMC Cancer
Volume10
DOIs
Publication statusPublished - 2010
MoE publication typeA1 Journal article-refereed

Fingerprint

Non-Receptor Type 2 Protein Tyrosine Phosphatase
Receptor Protein-Tyrosine Kinases
Vascular Endothelial Growth Factor A
Non-Receptor Protein Tyrosine Phosphatases
Phosphorylation
Activator Appliances
Spermidine
Integrins
Tail
Neoplasms
Endothelial Cells
Fibroblasts
Molecular Weight
Western Blotting

Cite this

Mattila, E., Marttila, H., Sahlberg, N., Kohonen, P., Tähtinen, S., Halonen, P., ... Ivaska, J. (2010). Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase. BMC Cancer, 10, [7]. https://doi.org/10.1186/1471-2407-10-7
Mattila, Elina ; Marttila, Heidi ; Sahlberg, Niko ; Kohonen, Pekka ; Tähtinen, Siri ; Halonen, Pasi ; Perälä, Merja ; Ivaska, Johanna. / Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase. In: BMC Cancer. 2010 ; Vol. 10.
@article{894e74488d254eb9882bc57aed82fbdf,
title = "Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase",
abstract = "Background T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. Methods We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. Results From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. Conclusions In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening to identify small molecule PTP activators that could function as RTK antagonists in cells.",
author = "Elina Mattila and Heidi Marttila and Niko Sahlberg and Pekka Kohonen and Siri T{\"a}htinen and Pasi Halonen and Merja Per{\"a}l{\"a} and Johanna Ivaska",
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Mattila, E, Marttila, H, Sahlberg, N, Kohonen, P, Tähtinen, S, Halonen, P, Perälä, M & Ivaska, J 2010, 'Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase', BMC Cancer, vol. 10, 7. https://doi.org/10.1186/1471-2407-10-7

Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase. / Mattila, Elina; Marttila, Heidi; Sahlberg, Niko; Kohonen, Pekka; Tähtinen, Siri; Halonen, Pasi; Perälä, Merja; Ivaska, Johanna.

In: BMC Cancer, Vol. 10, 7, 2010.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Inhibition of receptor tyrosine kinase signalling by small molecule agonist of T-cell protein tyrosine phosphatase

AU - Mattila, Elina

AU - Marttila, Heidi

AU - Sahlberg, Niko

AU - Kohonen, Pekka

AU - Tähtinen, Siri

AU - Halonen, Pasi

AU - Perälä, Merja

AU - Ivaska, Johanna

PY - 2010

Y1 - 2010

N2 - Background T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. Methods We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. Results From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. Conclusions In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening to identify small molecule PTP activators that could function as RTK antagonists in cells.

AB - Background T-cell protein tyrosine phosphatase (TCPTP/TC45) is a ubiquitously expressed intra-cellular non-receptor protein tyrosine phosphatase involved in the negative regulation of several cancer relevant cellular signalling pathways. We have previously shown that interaction between the α-cytoplasmic tail of α1β1 integrin and TCPTP activates TCPTP by disrupting an inhibitory intra-molecular bond in TCPTP. Thus, inhibition of the regulatory interaction in TCPTP is a desirable strategy for TCPTP activation and attenuation of oncogenic RTK signalling. However, this is challenging with low molecular weight compounds. Methods We developed a high-throughput compatible assay to analyse activity of recombinant TCPTP in vitro. Using this assay we have screened 64280 small molecules to identify novel agonists for TCPTP. Dose-dependent response to TCPTP agonist was performed using the in vitro assay. Inhibition effects and specificity of TCPTP agonists were evaluated using TCPTP expressing and null mouse embryonic fibroblasts. Western blot analysis was used to evaluate attenuation of PDGFRβ and EGFR phosphorylation. Inhibition of VEGF signalling was analysed with VEGF-induced endothelial cell sprouting assays. Results From the screen we identified six TCPTP agonists. Two compounds competed with α1-cytoplasmic domain for binding to TCPTP, suggesting that they activate TCPTP similar to α1-cyt by disrupting the intra-molecular bond in TCPTP. Importantly, one of the compounds (spermidine) displayed specificity towards TCPTP in cells, since TCPTP -/- cells were 43-fold more resistant to the compound than TCPTP expressing cells. This compound attenuates PDGFRβ and VEGFR2 signalling in cells in a TCPTP-dependent manner and functions as a negative regulator of EGFR phosphorylation in cancer cells. Conclusions In this study we showed that small molecules mimicking TCPTP-α1 interaction can be used as TCPTP agonists. These data provide the first proof-of-concept description of the use of high-throughput screening to identify small molecule PTP activators that could function as RTK antagonists in cells.

U2 - 10.1186/1471-2407-10-7

DO - 10.1186/1471-2407-10-7

M3 - Article

VL - 10

JO - BMC Cancer

JF - BMC Cancer

SN - 1471-2407

M1 - 7

ER -