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The combination of Cl-IB-MECA with paclitaxel: a new anti-metastatic therapeutic strategy for melanoma

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Abstract

Purpose

Metastatic melanoma is considered one of the most aggressive malignant tumours, representing the deadliest form of skin cancer. Melanoma progression is associated with the abrogation of normal controls that limit cell proliferation, migration, and invasion, eventually leading to metastasis. Based on the variety of cellular mechanisms involved in metastatic progression, we aimed to evaluate the effect of inosine (50 μM) and of the combination of Cl-IB-MECA (10 μM) with paclitaxel (10 ng/mL) on several stages of melanoma progression.

Methods

Proliferation, migration, adhesion, invasion, and colony formation assays were performed on human C32 and A375 metastatic melanoma cells. Levels of ERK1/2 were also determined using an ELISA kit. Moreover, mouse aortic rings were treated with vascular endothelial growth factor in order to assess the microvessel sprouting (an indicator of angiogenesis) in the presence of the referred compounds.

Results

We demonstrate that inosine induced, through A3 adenosine receptor activation, proliferation, migration, adhesion, and invasion on C32 and A375 melanoma cells, although with dissimilar importance on the two melanoma cell lines. Inosine also increased colony formation on A375 cells. Levels of ERK1/2 were increased after inosine exposure and that increase was dependent on A3 adenosine receptor activation in both cell lines. Moreover, microvessel sprouting stimulated by inosine was decreased by the combination of Cl-IB-MECA with paclitaxel.

Conclusions

Cl-IB-MECA combined with paclitaxel was able to impair almost all of the referred metastatic related mechanisms induced by inosine, making this approach a valuable tool for combinatory therapy against metastatic melanoma.

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Abbreviations

AR:

Adenosine receptor

BME:

Basement membrane extract

BSA:

Bovine serum albumin

Cl-IB-MECA:

2-Chloro-N(6)-(3-iodobenzyl)-adenosine-5′-N-methyl-uronamide

CTR:

Control

DMSO:

Dimethyl sulfoxide

DMEM-HG:

Dulbecco’s modified Eagle’s medium–high glucose

ECM:

Extracellular matrix

ERK:

Extracellular signal-regulated kinase

FBS:

Foetal bovine serum

INO:

Inosine

MAPK:

Mitogen-activated protein kinase

MRE3008F20:

N-[2-(2-furanyl)-8-propyl-8H-pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidine-5-yl]-N′-(4-methoxyphenyl)urea

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide

Opti-MEM:

Opti-minimal essential medium

PBS:

Phosphate-buffered saline

PI3K:

Phosphatidylinositol 3-kinase

PXT:

Paclitaxel

ROS:

Reactive oxygen species

VEGF:

Vascular endothelial growth factor

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Acknowledgments

Work was funded by FEDER through the Program of Operational Competitiveness Factors—COMPETE and National Funds through FCT—Foundation for Science and Technology. ASS and VMC thank FCT for their PhD grant (SFRH/BD/64911/2009) and Post-doc grant (SFRH/BPD/63746/2009), respectively.

Conflict of interest

We declare that we have no conflict of interest regarding this article.

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Authors and Affiliations

  1. REQUIMTE/Laboratório de Farmacologia, Departamento de Ciências do Medicamento, Faculdade de Farmácia, Universidade do Porto, Porto, Portugal

    Ana S. Soares, Carmen Diniz & Paula Fresco

  2. MedInUP – Centro de Investigação Farmacológica e Inovação Medicamentosa, Universidade do Porto, Porto, Portugal

    Ana S. Soares, Carmen Diniz & Paula Fresco

  3. REQUIMTE/Laboratório de Toxicologia, Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, Porto, Portugal

    Vera M. Costa

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  1. Ana S. Soares
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  2. Vera M. Costa
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  4. Paula Fresco
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Correspondence to Paula Fresco.

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Soares, A.S., Costa, V.M., Diniz, C. et al. The combination of Cl-IB-MECA with paclitaxel: a new anti-metastatic therapeutic strategy for melanoma. Cancer Chemother Pharmacol 74, 847–860 (2014). https://doi.org/10.1007/s00280-014-2557-y

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  • DOI: https://doi.org/10.1007/s00280-014-2557-y

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