Caracterização da nocicepção em um modelo de dor do câncer de mama e avaliação da participação do receptor TRPA1 neste modelo em camundongos fêmeas

Abstract

Breast cancer can cause bone metastasis, contributing to patients' pain. There is still a portion of patients with breast cancer who have inadequate treatment for pain. The mechanisms associated with cancer pain are still uncertain, and several ion channels seem to be involved in the transduction of noxious stimuli. The transient receptor potential ankyrin 1 (TRPA1) is a sensor for noxious stimuli, such as allyl isothiocyanate (AITC) and hydrogen peroxide (H2O2). This study aimed to characterize nociception in a breast cancer pain model with bone metastasis and to evaluate the TRPA1 receptor involvement in this model. 4T1 cells (104) were inoculated into the fourth mammary gland of female BALB/c mice; then, a time curve was performed to observe the development of mechanical and cold allodynia, and changes in the facial score. To determine the presence of bone metastasis, the clonogenic metastatic test and assessment of serum calcium levels were performed. At 20 days after tumor induction, the antinociceptive effect of different analgesics (paracetamol, naproxen, codeine or morphine), a cannabinoid agonist (WIN 55,212-2), TRPA1 antagonists (acute and repeated oral and intraplantar), of intrathecal administration of an antisense (AS) oligonucleotide for TRPA1, of an antioxidant (α-lipoic acid), were tested. The action of repeated administration of an antagonist was tested. H2O2 and NADPH oxidase and superoxide dismutase (SOD) activity and TRPA1 receptor expression were performed. Bone metastasis was confirmed by clonogenic assay and hypercalcemia was observed 20 days after cell inoculation. The administration of paracetamol, naproxen, codeine, morphine, WIN 55,212-2, TRPA1 antagonists (HC-030031 or A967079), antisense oligonucleotide, α-lipoic acid had an antinociceptive effect. Repeated dose treatment using a TRPA1 antagonist also had an antinociceptive effect. Intraplantar injection of TRPA1 agonist (AITC) caused chemical hyperalgesia 20 days after inoculation of 4T1 cells. However, there was no increase in TRPA1 expression in bone. SOD and NADPH oxidase enzyme activity and H2O2 levels were increased in paw skin, sciatic nerve and bone tissue 20 days after inoculation of 4T1 cells. The cell viability test showed that TRPA1 antagonist (HC-030031) and the antioxidant compound did not decrease cell viability, while paclitaxel significantly decreased. Cells of the 4T1 lineage do not express TRPA1. By the SAR technique, 4T1 cells do not have protein expression at the transcriptional level of TRPA1 and neither do osteoblasts and osteoclasts of the analyzed strains. There is no protein and transcriptional expression of TRPA1 in bone tissue of mice, however there is evidence of the presence of TRPV4 at the transcriptional level. Thus, this cancer pain model can be a reliable way to look at the mechanisms of pain induced by breast cancer with bone metastasis or to look at the efficacy of new analgesic compounds and new analgesic mechanisms. Furthermore, TRPA1 is an important target for the study of cancer pain and the development of new pharmacological treatments for cancer pain.