Efectos de los antiprogestagenos sobre la proliferación del carcinoma de mama canino. Estudios in vivo e in vitro

Los tumores de mama en la especie canina representan casi la mitad de las neoplasias que afectan a la perra. Dada su elevada frecuencia, los tumores de mama caninos son importantes causas de morbilidad y mortalidad. Alrededor del 50% de los tumores mamarios diagnosticados son considerados malignos y por tanto, con una evolución clínica postquirúrgica desfavorable. Muchos de ellos tienen receptores de progesterona (RP), dato que evidencia la importancia de esta hormona en el desarrollo de la enfermedad. Sin embargo, a diferencia de lo que ocurre en la mujer con el receptor de estrógenos, no hay datos sobre los efectos de los antiprogestágenos en el tratamiento sistémico de las perras con cáncer de mama. En este trabajo hemos analizado los efectos in vivo e in vitro de los antiprogestágenos sobre la proliferación del carcinoma de mama canino con expresión de RP. El estudio in vivo se realizó en 27 perras no castradas (22 experimentales y 5 controles) con tumores espontáneos tratadas con 2 dosis de 20 mg/Kg de aglepristona antes de la cirugía y observamos una disminución de la proliferación celular (índice Ki67, MIB-1, método ABC) en los tumores que tenían receptores de progesterona tanto con una técnica inmunohistoquímica (anticuerpo PR10A9, técnica ABC) como la técnica cuantitativa de reacción en cadena de la polimerasa en tiempo real (RTqPCR). Esta última técnica se estandarizó y aplicó por primera vez en la literatura sobre muestras de displasias y tumores de la mama canina procesadas rutinariamente para estudio histopatológico y...Canine mammary tumours account for nearly half of the neoplasms affecting the female dog. Given their high frequency, mammary tumours are important causes of morbidity and mortality. Some 50% of canine mammary tumours are diagnosed as malignant and therefore, are expected to have an unfavourable clinical course after surgery. Many mammary carcinomas have progesterone receptors, evidencing the importance of this hormone in the development of the disease. However, unlike what happens in human breast cancer with respect to oestrogen receptors, there are no data available concerning the role of antiprogestins in the systemic treatment of female dogs with mammary cancer. In this study we have analyzed the in vivo and in vitro effects of antiprogestins on the proliferation of canine mammary carcinomas with progesterone receptors. The in vivo study was performed on 27 no spayed female dogs (22 experimental and 5 controls) with spontaneous tumours treated with 2 doses of 20 mg / Kg aglepristone before surgery. Results showed a decrease in cell proliferation (Ki67 index, MIB- 1 antibody, ABC method) in tumours expressing progesterone receptors as measured by immunohistochemistry (PR10A9 antibody, ABC technique) and by real time quantitative polymerase chain reaction (RTqPCR). The latter technique was standardized and applied for the first time in the literature in the dog by using tissue samples of mammary dysplasias and tumours routinely processed for histopathological examination. This technique allows the study of both the complete progesterone receptor as well as their isoforms..

Sex Steroid Hormones and Tumors in Domestic Animals

Sex steroid hormones play a role in the development and control of animal tumours, particularly in those arising in their target organs. Due to their incidence and prevalence, mammary tumours of female dogs and cats are among the most frequently studied with focus on the role of ovarian oestrogen and progesterone. In these tumours, sex steroid hormones have been shown to act during the three steps of the carcinogenesis cascade: initiation, promotion and progression. Experimental data have shown the mutagenic effect of oestrogens [1] while epidemiologic and clinical studies highlighted the role of ovarian hormones as promoters on mammary tumours in both the dog and the cat [2-9]. Finally, oestrogens and progesterone further act during tumour progression. Their role in the last two steps of carcinogenesis makes it possible to control the evolution of the diseas

Ovarian stimulation with FSH reduces phosphorylation of gonadotrope progesterone receptor and LH secretion in the rat

Administration of human FSH(hFSH) to cyclic rats during the dioestrous phase attenuates progesterone receptor (PR)-dependent events of the preovulatory LH surge in pro-oestrus. The increased bioactivity of the putative ovarian gonadotropin surge inhibiting/attenuating factor induced by hFSH treatment is not associated with a decrease in PR protein expression, and the possibility of its association at a PR posttranslational effect has been raised. The present experiments aimed to analyse PR phosphorylation status in the gonadotrope of ratswith impaired LH secretion induced by in vivo hFSH injection. Two experimental approaches were used. First, incubated pro-oestrous pituitaries from hFSH-injected cycling and oestrogen-treated ovariectomized (OVX) rats were used to analyze the effect of calyculin, an inhibitor of intracellular phosphatases, on PR-dependent LH release, which was measured in the incubation medium by RIA. Second, pituitaries taken from hFSH-injected intact cycling and OVX rats and later incubated with P or GNRH1 were used to assess the phosphorylation rate of gonadotrope. The latter was analysed in formalin-fixed, paraffin-embedded tissue sections by immunohistochemistry using a MAB that recognizes the phosphorylated (p) form of PR at Ser294. Calyculin reduced the ovary-mediated inhibition of hFSH in GNRH1-stimulated LH secretion. In addition, the immunohistochemical expression of pSer294 PR was significantly reduced after ovarian stimulation with hFSH in pituitaries frompro-oestrous rats incubated with PorGNRH1.Altogether, these results suggested that the ovarian-dependent inhibitory effect of FSH injection on the preovulatory LH secretion in the rat may involve an increase in dephosphorylation of PR

Progesterone receptor isoform A may regulate the effects of neoadjuvant aglepristone in canine mammary carcinoma

Background: Progesterone receptors play a key role in the development of canine mammary tumours, and recent research has focussed on their possible value as therapeutic targets using antiprogestins. Cloning and sequencing of the progesterone receptor gene has shown that the receptor has two isoforms, A and B, transcribed from a single gene. Experimental studies in human breast cancer suggest that the differential expression of progesterone receptor isoforms has implications for hormone therapy responsiveness. This study examined the effects of the antiprogestin aglepristone on cell proliferation and mRNA expression of progesterone receptor isoforms A and B in mammary carcinomas in dogs treated with 20 mg/Kg of aglepristone (n = 22) or vehicle (n = 5) twice before surgery. Results: Formalin-fixed, paraffin-embedded tissue samples taken before and after treatment were used to analyse total progesterone receptor and both isoforms by RT-qPCR and Ki67 antigen labelling. Both total progesterone receptor and isoform A mRNA expression levels decreased after treatment with aglepristone. Furthermore, a significant decrease in the proliferation index (percentage of Ki67-labelled cells) was observed in progesterone-receptor positive and isoform-A positive tumours in aglepristone-treated dogs. Conclusions: These findings suggest that the antiproliferative effects of aglepristone in canine mammary carcinomas are mediated by progesterone receptor isoform A

The Combination of Neutrophil–Lymphocyte Ratio and Platelet–Lymphocyte Ratio with Liquid Biopsy Biomarkers Improves Prognosis Prediction in Metastatic Pancreatic Cancer

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with a highly inflammatory microenvironment and liquid biopsy has emerged as a promising tool for the noninvasive analysis of this tumor. In this study, plasma was obtained from 58 metastatic PDAC patients, and neutrophil–lymphocyte ratio (NLR), platelet–lymphocyte ratio (PLR), circulating cell-free DNA (cfDNA) concentration, and circulating RAS mutation were determined. We found that NLR was significantly associated with both overall survival (OS) and progression-free survival. Remarkably, NLR was an independent risk factor for poor OS. Moreover, NLR and PLR positively correlated, and combination of both inflammatory markers significantly improved the prognostic stratification of metastatic PDAC patients. NLR also showed a positive correlation with cfDNA levels and RAS mutant allelic fraction (MAF). Besides, we found that neutrophil activation contributed to cfDNA content in the plasma of metastatic PDAC patients. Finally, a multi-parameter prognosis model was designed by combining NLR, PLR, cfDNA levels, RAS mutation, RAS MAF, and CA19-9, which performs as a promising tool to predict the prognosis of metastatic PDAC patients. In conclusion, our study supports the idea that the use of systemic inflammatory markers along with circulating tumor-specific markers may constitute a valuable tool for the clinical management of metastatic PDAC patients

Circulating Cell-Free DNA-Based Liquid Biopsy Markers for the Non-Invasive Prognosis and Monitoring of Metastatic Pancreatic Cancer

Liquid biopsy may assist in the management of cancer patients, which can be particularly applicable in pancreatic ductal adenocarcinoma (PDAC). In this study, we investigated the utility of circulating cell-free DNA (cfDNA)-based markers as prognostic tools in metastatic PDAC. Plasma was obtained from 61 metastatic PDAC patients, and cfDNA levels and fragmentation were determined. BEAMing technique was used for quantitative determination of RAS mutation allele fraction (MAF) in cfDNA. We found that the prognosis was more accurately predicted by RAS mutation detection in plasma than in tissue. RAS mutation status in plasma was a strong independent prognostic factor for both overall survival (OS) and progression-free survival (PFS). Moreover, RAS MAF in cfDNA was also an independent risk factor for poor OS, and was strongly associated with primary tumours in the body/tail of the pancreas and liver metastases. Higher cfDNA levels and fragmentation were also associated with poorer OS and shorter PFS, body/tail tumors, and hepatic metastases, whereas cfDNA fragmentation positively correlated with RAS MAF. Remarkably, the combination of CA19-9 with MAF, cfDNA levels and fragmentation improved the prognostic stratification of patients. Furthermore, dynamics of RAS MAF better correlated with patients’ outcome than standard CA19-9 marker. In conclusion, our study supports the use of cfDNA-based liquid biopsy markers as clinical tools for the non-invasive prognosis and monitoring of metastatic PDAC patients