Limits...
Novel LIMK2 Inhibitor Blocks Panc-1 Tumor Growth in a mouse xenograft model.

Rak R, Haklai R, Elad-Tzfadia G, Wolfson HJ, Carmeli S, Kloog Y - Oncoscience (2014)

Bottom Line: We recently reported the development and validation of a novel LIMK inhibitor, referred to here as T56-LIMKi, using a combination of computational methods and classical biochemistry techniques.We found that T56-LIMKi decreases phosphorylated cofilin (p-cofilin) levels and thus inhibits growth of several cancerous cell lines, including those of pancreatic cancer, glioma and schwannoma.Because the most promising in-vitro effect of T56-LIMKi was observed in the pancreatic cancer cell line Panc-1, we tested the inhibitor on a nude mouse Panc-1 xenograft model.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, The George S. Wise Faculty of Life Sciences, Tel-Aviv University, Tel Aviv, Israel.

ABSTRACT
LIM kinases (LIMKs) are important cell cytoskeleton regulators that play a prominent role in cancer manifestation and neuronal diseases. The LIMK family consists of two homologues, LIMK1 and LIMK2, which differ from one another in expression profile, intercellular localization, and function. The main substrate of LIMK is cofilin, a member of the actin-depolymerizing factor (ADF) protein family. When phosphorylated by LIMK, cofilin is inactive. LIMKs play a contributory role in several neurodevelopmental disorders and in cancer growth and metastasis. We recently reported the development and validation of a novel LIMK inhibitor, referred to here as T56-LIMKi, using a combination of computational methods and classical biochemistry techniques. Here we report that T56-LIMKi inhibits LIMK2 with high specificity, and shows little or no cross-reactivity with LIMK1. We found that T56-LIMKi decreases phosphorylated cofilin (p-cofilin) levels and thus inhibits growth of several cancerous cell lines, including those of pancreatic cancer, glioma and schwannoma. Because the most promising in-vitro effect of T56-LIMKi was observed in the pancreatic cancer cell line Panc-1, we tested the inhibitor on a nude mouse Panc-1 xenograft model. T56-LIMKi reduced tumor size and p-cofilin levels in the Panc-1 tumors, leading us to propose T56-LIMKi as a candidate drug for cancer therapy.

No MeSH data available.


Related in: MedlinePlus

Levels of p-cofilin levels are reduced in a cell-specific manner manner by T56-LIMKiPanc-1, A549, U87, and ST88-14 cells were plated in 10-cm dishes, 5 × 105 cells per plate. The cells were serum-starved for 24 h and then treated with 50 μM T56-LIMKi, 5 μM BMS-5, or vehicle(0.1% DMSO) for 2 h at the indicated concentrations. The cells were then homogenized and their proteins were immunoblotted with specific antibody, quantified, and normalized to p-cofilin, as described in Material and Methods. Average inhibition was calculated from three sets of independent experiments. Results for each cell line are presented as a percentage of the untreated control (mean ± SD, n = 3) All changes observed between controls and treatments, except for T56-LIMKi treatment of A549 cells, were significant (P < 0.01, Student's t-test).
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Figure 5: Levels of p-cofilin levels are reduced in a cell-specific manner manner by T56-LIMKiPanc-1, A549, U87, and ST88-14 cells were plated in 10-cm dishes, 5 × 105 cells per plate. The cells were serum-starved for 24 h and then treated with 50 μM T56-LIMKi, 5 μM BMS-5, or vehicle(0.1% DMSO) for 2 h at the indicated concentrations. The cells were then homogenized and their proteins were immunoblotted with specific antibody, quantified, and normalized to p-cofilin, as described in Material and Methods. Average inhibition was calculated from three sets of independent experiments. Results for each cell line are presented as a percentage of the untreated control (mean ± SD, n = 3) All changes observed between controls and treatments, except for T56-LIMKi treatment of A549 cells, were significant (P < 0.01, Student's t-test).

Mentions: We further tested the ability of T56-LIMKi to inhibit phosphorylation of cofilin in the various cell lines by comparing its inhibitory effect to that of BMS-5, known to inhibit both LIMK1 and LIMK2. The most remarkable T56-LIMKi-induced decrease in p-cofilin among all the cell lines tested was found in the pancreatic cancer cell line Panc-1 (46% ±10%; Fig. 5). Cofilin phosphorylation in Panc-1 was also inhibited by BMS-5 (37% ± 5%); however, unlike in the case of BMS-5 treatment of NF1−depleted MEFs [36], BMS-5 was not more effective in reducing p-cofilin levels in Panc-1 cells, although it inhibited both LIMKs. This finding is consistent with the results of a recent study in which double knockout of LIMK1 and LIMK2 had no additional effect on tumor growth or metastasis of the Panc-1 cell line in a zebrafish xenograft model [27]. In a similar manner, U87 cell growth inhibition was relatively high with both treatments, namely, T56-LIMKi and BMS-5 decreased p-cofilin by 24% ± 10% and 38% ± 12%, respectively, with no significant difference between the two treatments. In contrast A549 and ST88-14 cells reacted differently to the two inhibitors: inhibition of their p-cofilin levels by BMS-5 treatment was significantly higher than that observed with T56-LIMKi (75% ± 20% and 65% ± 55, in ST88-14 and A549, respectively). T56-LIMKi decreased p-cofilin only by 20% ± 8% in ST88-14 cells, but hardly decreased it at all (4% ± 4%) in A549 cells (Fig. 5).


Novel LIMK2 Inhibitor Blocks Panc-1 Tumor Growth in a mouse xenograft model.

Rak R, Haklai R, Elad-Tzfadia G, Wolfson HJ, Carmeli S, Kloog Y - Oncoscience (2014)

Levels of p-cofilin levels are reduced in a cell-specific manner manner by T56-LIMKiPanc-1, A549, U87, and ST88-14 cells were plated in 10-cm dishes, 5 × 105 cells per plate. The cells were serum-starved for 24 h and then treated with 50 μM T56-LIMKi, 5 μM BMS-5, or vehicle(0.1% DMSO) for 2 h at the indicated concentrations. The cells were then homogenized and their proteins were immunoblotted with specific antibody, quantified, and normalized to p-cofilin, as described in Material and Methods. Average inhibition was calculated from three sets of independent experiments. Results for each cell line are presented as a percentage of the untreated control (mean ± SD, n = 3) All changes observed between controls and treatments, except for T56-LIMKi treatment of A549 cells, were significant (P < 0.01, Student's t-test).
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Related In: Results  -  Collection

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Figure 5: Levels of p-cofilin levels are reduced in a cell-specific manner manner by T56-LIMKiPanc-1, A549, U87, and ST88-14 cells were plated in 10-cm dishes, 5 × 105 cells per plate. The cells were serum-starved for 24 h and then treated with 50 μM T56-LIMKi, 5 μM BMS-5, or vehicle(0.1% DMSO) for 2 h at the indicated concentrations. The cells were then homogenized and their proteins were immunoblotted with specific antibody, quantified, and normalized to p-cofilin, as described in Material and Methods. Average inhibition was calculated from three sets of independent experiments. Results for each cell line are presented as a percentage of the untreated control (mean ± SD, n = 3) All changes observed between controls and treatments, except for T56-LIMKi treatment of A549 cells, were significant (P < 0.01, Student's t-test).
Mentions: We further tested the ability of T56-LIMKi to inhibit phosphorylation of cofilin in the various cell lines by comparing its inhibitory effect to that of BMS-5, known to inhibit both LIMK1 and LIMK2. The most remarkable T56-LIMKi-induced decrease in p-cofilin among all the cell lines tested was found in the pancreatic cancer cell line Panc-1 (46% ±10%; Fig. 5). Cofilin phosphorylation in Panc-1 was also inhibited by BMS-5 (37% ± 5%); however, unlike in the case of BMS-5 treatment of NF1−depleted MEFs [36], BMS-5 was not more effective in reducing p-cofilin levels in Panc-1 cells, although it inhibited both LIMKs. This finding is consistent with the results of a recent study in which double knockout of LIMK1 and LIMK2 had no additional effect on tumor growth or metastasis of the Panc-1 cell line in a zebrafish xenograft model [27]. In a similar manner, U87 cell growth inhibition was relatively high with both treatments, namely, T56-LIMKi and BMS-5 decreased p-cofilin by 24% ± 10% and 38% ± 12%, respectively, with no significant difference between the two treatments. In contrast A549 and ST88-14 cells reacted differently to the two inhibitors: inhibition of their p-cofilin levels by BMS-5 treatment was significantly higher than that observed with T56-LIMKi (75% ± 20% and 65% ± 55, in ST88-14 and A549, respectively). T56-LIMKi decreased p-cofilin only by 20% ± 8% in ST88-14 cells, but hardly decreased it at all (4% ± 4%) in A549 cells (Fig. 5).

Bottom Line: We recently reported the development and validation of a novel LIMK inhibitor, referred to here as T56-LIMKi, using a combination of computational methods and classical biochemistry techniques.We found that T56-LIMKi decreases phosphorylated cofilin (p-cofilin) levels and thus inhibits growth of several cancerous cell lines, including those of pancreatic cancer, glioma and schwannoma.Because the most promising in-vitro effect of T56-LIMKi was observed in the pancreatic cancer cell line Panc-1, we tested the inhibitor on a nude mouse Panc-1 xenograft model.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, The George S. Wise Faculty of Life Sciences, Tel-Aviv University, Tel Aviv, Israel.

ABSTRACT
LIM kinases (LIMKs) are important cell cytoskeleton regulators that play a prominent role in cancer manifestation and neuronal diseases. The LIMK family consists of two homologues, LIMK1 and LIMK2, which differ from one another in expression profile, intercellular localization, and function. The main substrate of LIMK is cofilin, a member of the actin-depolymerizing factor (ADF) protein family. When phosphorylated by LIMK, cofilin is inactive. LIMKs play a contributory role in several neurodevelopmental disorders and in cancer growth and metastasis. We recently reported the development and validation of a novel LIMK inhibitor, referred to here as T56-LIMKi, using a combination of computational methods and classical biochemistry techniques. Here we report that T56-LIMKi inhibits LIMK2 with high specificity, and shows little or no cross-reactivity with LIMK1. We found that T56-LIMKi decreases phosphorylated cofilin (p-cofilin) levels and thus inhibits growth of several cancerous cell lines, including those of pancreatic cancer, glioma and schwannoma. Because the most promising in-vitro effect of T56-LIMKi was observed in the pancreatic cancer cell line Panc-1, we tested the inhibitor on a nude mouse Panc-1 xenograft model. T56-LIMKi reduced tumor size and p-cofilin levels in the Panc-1 tumors, leading us to propose T56-LIMKi as a candidate drug for cancer therapy.

No MeSH data available.


Related in: MedlinePlus