Jorge Fernández-Retana

MicroRNAs in cervical cancer: evidences for a miRNA profile deregulated by HPV and its impact on radio-resistance.

Cervical carcinoma (CC) is one of the most common cancers and a leading cause of mortality in women worldwide. Epidemiologic and experimental data have clearly demonstrated a causal role of high-risk Human Papillomavirus (HR-HPV) types in CC initiation and progression, affecting the cellular processes by targeting and inactivating p53 and pRB host proteins. HR-HPV E5, E6 and E7 oncoproteins have the ability to deregulate several cellular processes, mostly apoptosis, cell cycle control, migration, immune evasion, and induction of genetic instability, which promote the accumulation of mutations and aneuploidy. In this scenario, genomic profiles have shown that aberrant expression of cellular oncogenic and tumor suppressive miRNAs have an important role in CC carcinogenesis. It has been stated that HPV infection and E6/E7 expression are essential but not sufficient to lead to CC development; hence other genetic and epigenetic factors have to be involved in this complex disease. Recent evidence suggests an important level of interaction among E6/E7 viral proteins and cellular miRNA, and other noncoding RNAs. The aim of the current review is to analyze recent data that mainly describe the interaction between HR-HPV established infections and specific cellular miRNAs; moreover, to understand how those interactions could affect radio-therapeutic response in tumor cells.

A microRNA expression signature for clinical response in locally advanced cervical cancer

OBJECTIVE Nearly 50% of patients who are diagnosed with locally advanced cervical cancer have an unfavorable pathological response to conventional treatment. MicroRNAs (miRNAs) are potential biomarkers in cervical cancer; however, their role in identifying patients who do not respond to conventional treatment remains poorly investigated. Here, we identify a set of miRNAs that can be used as molecular markers to predict the pathological response in locally advanced cervical cancer patients receiving radiation and chemotherapy treatment. METHODS Forty-one patients diagnosed with locally advanced cervical cancer were invited to participate in this study and enrolled after they signed an informed consent. Two patient cohorts were randomized for miRNA expression profiling, a discovery cohort (n=10) and a validation cohort (n=31); profiling was performed by means of a miScript miRNA PCR Array. After a median clinical follow-up of 45months, statistical analysis was performed to identify miRNAs that could discriminate non-responders from complete pathological responders to conventional treatment. RESULTS miRNA expression profiling identified 101 miRNAs that showed significant differences between non-responders and complete pathological responders (p<0.05). Seven differentially expressed miRNAs were selected, and their expression patterns were confirmed in the validation phase; thus, miR-31-3p, -3676, -125a-5p, -100-5p, -125b-5p, and -200a-5p and miR-342 were significantly associated with clinical response. Expression of this miRNA signature above the median level was a significant predictor of non-response to standard treatment (p<0.001). CONCLUSIONS These seven validated miRNA signatures could be used as molecular biomarkers of chemo- and radio-resistance in locally advanced cervical cancer patients.

The 25 kDa subunit of cleavage factor Im Is a RNA-binding protein that interacts with the poly(A) polymerase in Entamoeba histolytica.

In eukaryotes, polyadenylation of pre-mRNA 3´ end is essential for mRNA export, stability and translation. Taking advantage of the knowledge of genomic sequences of Entamoeba histolytica, the protozoan responsible for human amoebiasis, we previously reported the putative polyadenylation machinery of this parasite. Here, we focused on the predicted protein that has the molecular features of the 25 kDa subunit of the Cleavage Factor Im (CFIm25) from other organisms, including the Nudix (nucleoside diphosphate linked to another moiety X) domain, as well as the RNA binding domain and the PAP/PAB interacting region. The recombinant EhCFIm25 protein (rEhCFIm25) was expressed in bacteria and used to generate specific antibodies in rabbit. Subcellular localization assays showed the presence of the endogenous protein in nuclear and cytoplasmic fractions. In RNA electrophoretic mobility shift assays, rEhCFIm25 was able to form specific RNA-protein complexes with the EhPgp5 mRNA 3´ UTR used as probe. In addition, Pull-Down and LC/ESI-MS/MS tandem mass spectrometry assays evidenced that the putative EhCFIm25 was able to interact with the poly(A) polymerase (EhPAP) that is responsible for the synthesis of the poly(A) tail in other eukaryotic cells. By Far-Western experiments, we confirmed the interaction between the putative EhCFIm25 and EhPAP in E. histolytica. Taken altogether, our results showed that the putative EhCFIm25 is a conserved RNA binding protein that interacts with the poly(A) polymerase, another member of the pre-mRNA 3´ end processing machinery in this protozoan parasite.

Transcript Profiling Distinguishes Complete Treatment Responders With Locally Advanced Cervical Cancer

Cervical cancer (CC) mortality is a major public health concern since it is the second cause of cancer-related deaths among women. Patients diagnosed with locally advanced CC (LACC) have an important rate of recurrence and treatment failure. Conventional treatment for LACC is based on chemotherapy and radiotherapy; however, up to 40% of patients will not respond to conventional treatment; hence, we searched for a prognostic gene signature able to discriminate patients who do not respond to the conventional treatment employed to treat LACC. Tumor biopsies were profiled with genome-wide high-density expression microarrays. Class prediction was performed in tumor tissues and the resultant gene signature was validated by quantitative reverse transcription–polymerase chain reaction. A 27-predictive gene profile was identified through its association with pathologic response. The 27-gene profile was validated in an independent set of patients and was able to distinguish between patients diagnosed as no response versus complete response. Gene expression analysis revealed two distinct groups of tumors diagnosed as LACC. Our findings could provide a strategy to select patients who would benefit from neoadjuvant radiochemotherapy-based treatment.

Transcriptomic Profiling of Adipose Tissue in Obese Women in Response to Acupuncture Catgut Embedding Therapy with Moxibustion

OBJECTIVE Complementary and alternative medicine, such as Traditional Chinese Medicine, represents an efficient therapeutic option for obesity control. It was previously reported that acupuncture catgut embedding therapy (ACET) with moxibustion reduces body weight and reverts insulin resistance in obese women. This study aimed to evidence changes in adipokines and gene expression in adipose tissue that could explain the effects of ACET with moxibustion. DESIGN Overweight/obese women were treated with ACET with moxibustion or sham acupuncture as control. Peripheral blood samples and fat biopsies were taken before and after intervention. Circulating adipokines (leptin, adiponectin, tumor necrosis factor alpha, and resistin) were quantified by enzyme-linked immunosorbent assay. Gene expression in adipose tissue was determined by cDNA microarray assays and assessed by quantitative reverse transcription real-time polymerase chain reaction. RESULTS ACET with moxibustion did not modify circulating adipokines levels. However, correlations with anthropometric and biochemical parameters were affected. Interestingly, transcriptional changes in adipose tissue revealed the modulation of genes participating in homeostasis control, lipid metabolism, olfactory transduction, and gamma-aminobutyric acid signaling pathway. CONCLUSIONS The effects of ACET with moxibustion on body weight and insulin resistance were associated with the regulation of biochemical events that are altered in obesity.

Gene signature based on degradomerelated genes can predict distal metastasis in cervical cancer patients

Cervical cancer is one of the leading causes of death in women worldwide, which mainly affects developing countries. The patients who suffer a recurrence and/or progression disease have a higher risk of developing distal metastases. Proteases comprising the degradome given its ability to promote cell growth, migration, and invasion of tissues play an important role during tumor development and progression. In this study, we used high-density microarrays and quantitative reverse transcriptase polymerase chain reaction to evaluate the degradome profile and their inhibitors in 112 samples of patients diagnosed with locally advanced cervical cancer. Clinical follow-up was done during a period of 3 years. Using a correlation analysis between the response to treatment and the development of metastasis, we established a molecular signature comprising eight degradome-related genes (FAM111B, FAM111A, CFB, PSMB8, PSMB9, CASP7, PRSS16, and CD74) with the ability to discriminate patients at risk of distal metastases. In conclusion, present results show that molecular signature obtained from degradome genes can predict the possibility of metastasis in patients with locally advanced cervical cancer.

PAX8 is transcribed aberrantly in cervical tumors and derived cell lines due to complex gene rearrangements

The transcription factor PAX8, a member of the paired box-containing gene family with an important role in embryogenesis of the kidney, thyroid gland and nervous system, has been described as a biomarker in tumors of the thyroid, parathyroid, kidney and thymus. The PAX8 gene gives rise to four isoforms, through alternative mRNA splicing, but the splicing pattern in tumors is not yet established. Cervical cancer has a positive expression of PAX8; however, there is no available data determining which PAX8 isoform or isoforms are present in cervical cancer tissues as well as in cervical carcinoma-derived cell lines. Instead of a differential pattern of splicing isoforms, we found numerous previously unreported PAX8 aberrant transcripts ranging from 378 to 542 bases and present in both cervical carcinoma-derived cell lines and tumor samples. This is the first report of PAX8 aberrant transcript production in cervical cancer. Reported PAX8 isoforms possess differential transactivation properties; therefore, besides being a helpful marker for detection of cancer, PAX8 isoforms can plausibly exert differential regulation properties during carcinogenesis.

Transcriptome Studies Reveal Altered Signaling Pathways in Cervical Cancer

Transcriptome analysis provides a global idea of the molecular mechanisms affected in different pathologies. Characterization of over- or under-expressed genes constitutes an initial step in this type of analysis. The integration of the information acquired by these global expression profiles regarding signaling pathways or organized modules that work according to specific cellular responses has made it possible for us to understand the development and progression of almost every type of neoplasia. In the case of cervical cancer, transcriptome studies have allowed us to comprehend the viral-mediated carcinogenic process, i.e., human papillomavirus (HPV). In spite of the great progress that has been accomplished regarding radiotherapy and chemotherapy, its impact in cervical cancer in limited; approximately 40% of the patients develop resistance to the conventional treatment schemes, and the disease will eventually recur, leading to the patient’s death. For this reason, knowing which signaling pathways present altered expression in this neoplasia opens a window of opportunity for those patients whose tumors display certain resistance to conventional treatment, further progression or even recurrence. In this chapter, we will summarize the main signaling pathways that are found to be altered in this neoplasia, pathways that have been described in various works in which the cervical cancer transcriptome was analyzed.