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Metastatic carcinoma of presumed renal origin (rCUP) has recently emerged as a new entity within the heterogeneous entity of Cancers of Unknown Primary (CUP) but their biological features and optimal therapeutic management remain unknown. We report the molecular characteristics and clinical outcome of a series of 25 rCUP prospectively identified within the French National Multidisciplinary Tumor Board for CUP. This cohort strongly suggests that rCUP share similarities with common RCC subtypes and benefit from renal-tailored systemic treatment. This study highlights the importance of integrating clinical and molecular data for optimal diagnosis and management of CUP.
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Tumor immunological characterization includes evaluation of tumor-infiltrating lymphocytes (TILs) and programmed cell death protein ligand-1 (PD-L1) expression. This study investigated TIL distribution, its prognostic value, and PD-L1 expression in metastatic and matched primary tumors (PTs). Specimens from 550 pan-cancer patients of the SHIVA01 trial (NCT01771458) with available metastatic biopsy and 111 matched PTs were evaluated for TILs and PD-L1. Combined positive score (CPS), tumor proportion score (TPS), and immune cell (IC) score were determined. TILs and PD-L1 were assessed according to PT organ of origin, histological subtype, and metastatic biopsy site. We found that TIL distribution in metastases did not vary according to PT organ of origin, histological subtype, or metastatic biopsy site, with a median of 10% (range: 0-70). TILs were decreased in metastases compared to PT (20% [5-60] versus 10% [0-40], p < 0.0001). CPS varied according to histological subtype (p = 0.02) and biopsy site (p < 0.02). TPS varied according to PT organ of origin (p = 0.003), histological subtype (p = 0.0004), and metastatic biopsy site (p = 0.00004). TPS was higher in metastases than in PT (p < 0.0001). TILs in metastases did not correlate with overall survival. In conclusion, metastases harbored fewer TILs than matched PT, regardless of PT organ of origin, histological subtype, and metastatic biopsy site. PD-L1 expression increased with disease progression. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
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BACKGROUND: High tumor mutational burden (TMB) was reported to predict the efficacy of immune checkpoint inhibitors (ICIs). Pembrolizumab, an anti-PD-1, received FDA-approval for the treatment of unresectable/metastatic tumors with high TMB as determined by the FoundationOne®CDx test. It remains to be determined how TMB can also be calculated using other tests. RESULTS: FFPE/frozen tumor samples from various origins were sequenced in the frame of the Institut Curie (IC) Molecular Tumor Board using an in-house next-generation sequencing (NGS) panel. A TMB calculation method was developed at IC (IC algorithm) and compared to the FoundationOne® (FO) algorithm. Using IC algorithm, an optimal 10% variant allele frequency (VAF) cut-off was established for TMB evaluation on FFPE samples, compared to 5% on frozen samples. The median TMB score for MSS/POLE WT tumors was 8.8 mut/Mb versus 45 mut/Mb for MSI/POLE-mutated tumors. When focusing on MSS/POLE WT tumor samples, the highest median TMB scores were observed in lymphoma, lung, endometrial, and cervical cancers. After biological manual curation of these cases, 21% of them could be reclassified as MSI/POLE tumors and considered as "true TMB high." Higher TMB values were obtained using FO algorithm on FFPE samples compared to IC algorithm (40 mut/Mb [10-3927] versus 8.2 mut/Mb [2.5-897], p < 0.001). CONCLUSIONS: We herein propose a TMB calculation method and a bioinformatics tool that is customizable to different NGS panels and sample types. We were not able to retrieve TMB values from FO algorithm using our own algorithm and NGS panel.
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Neoplasias , Humanos , Mutación , Neoplasias/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodosRESUMEN
AIMS: Malignant tumours of the lacrimal apparatus are rare and frequently show a poor prognosis, with no clear therapeutic standards. Characterisation of the genetic landscape of these rare tumours is sparse, and therefore therapeutics generally follow those of their common salivary gland counterparts. To further clarify the pathophysiology and discover potential therapeutic targets, we investigated the genetic landscape of eight tumours of the lacrimal apparatus. METHODS AND RESULTS: DNA and RNA sequencing were performed to identify genetic mutations and gene fusions. Immunohistochemistry, fluorescence in-situ hybridisation and reverse transcription-polymerase chain reaction followed by Sanger sequencing were performed to confirm the identified molecular alterations. Genetic alterations were detected in six tumours. Among five adenoid cystic carcinomas (ACC), four had confirmed alterations of MYB or MYBL1 genes, including a MYB::NFIB fusion, a MYBL1::NFIB fusion, a MYB amplification and a novel NFIB::THSD7B fusion. Mutations in genes encoding epigenetic modifiers, as well as NOTCH1, FGFR2 and ATM mutations, were also identified in ACCs. A carcinoma ex pleomorphic adenoma showed TP53 and CIC mutations and an amplification of ERBB2. A transitional cell carcinoma was associated with HPV16 infection. No genetic alteration was found for one adenocarcinoma, not otherwise specified. CONCLUSIONS: Our study highlights the variety of molecular alterations associated with lacrimal system tumours and emphasises the importance of molecular testing in these tumours, which can reveal potentially targetable mutations. Our results also reinforce the hypothesis of a common physiopathology of all ACCs, regardless of their primary location.
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Adenoma Pleomórfico , Carcinoma Adenoide Quístico , Aparato Lagrimal , Neoplasias de las Glándulas Salivales , Humanos , Aparato Lagrimal/patología , Proteínas de Fusión Oncogénica/genética , Carcinoma Adenoide Quístico/genética , Carcinoma Adenoide Quístico/patología , Adenoma Pleomórfico/genética , Adenoma Pleomórfico/patología , Fusión Génica , Neoplasias de las Glándulas Salivales/genética , Neoplasias de las Glándulas Salivales/patologíaRESUMEN
BACKGROUND: Data on the role of the microbiota in cancer have accumulated in recent years, with particular interest in intratumoral bacteria. Previous results have shown that the composition of intratumoral microbiome is different depending on the type of primary tumour and that bacteria from the primary tumour could migrate to metastatic sites. METHODS: Seventy-nine patients with breast, lung, or colorectal cancer and available biopsy samples from lymph node, lung, or liver site, treated in the SHIVA01 trial were analysed. We performed bacterial 16S rRNA gene sequencing on these samples to characterise the intratumoral microbiome. We assessed the association between microbiome composition, clinicopathological characteristics, and outcomes. RESULTS: Microbial richness (Chao1 index), evenness (Shannon index) and beta-diversity (Bray Curtis distance) were associated with biopsy site (p = 0.0001, p = 0.03 and p < 0.0001, respectively) but not with primary tumour type (p = 0.52, p = 0.54 and p = 0.82, respectively). Furthermore, microbial richness was inversely associated with tumour-infiltrating lymphocytes (TILs, p = 0.02), and PD-L1 expression on immune cells (p = 0.03), or assessed by Tumor Proportion Score (TPS, p = 0.02) or Combined Positive Score (CPS, p = 0.04). Beta-diversity was also associated with these parameters (p < 0.05). Patients with lower intratumoral microbiome richness had shorter overall survival (p = 0.03) and progression-free survival (p = 0.02) in multivariate analysis. CONCLUSION: Biopsy site, rather than primary tumour type, was strongly associated with microbiome diversity. Immune histopathological parameters such as PD-L1 expression and TILs were significantly associated with alpha and beta-diversity supporting the cancer-microbiome-immune axis hypothesis.
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Antígeno B7-H1 , Microbiota , Humanos , ARN Ribosómico 16S/genética , Pulmón , Mama , BacteriasRESUMEN
By 2030, pancreatic cancer will become the second leading cause of cancer-related deaths in the United States and in Europe. The management of patients with advanced pancreatic cancer relies on chemotherapy and poly (ADP-ribose) polymerase inhibitors for patients who carry BRCA1/2 inactivating alterations. Some variants, such as large insertion/deletions (Indels), inactivating BRCA1/2 and therefore of clinical relevance can be hard to detect by next-generation sequencing techniques. Here we report a 47-year-old patient presenting with pancreatic cancer whose tumour harbours a large somatic intra-exonic deletion of BRCA2 of 141 bp. This BRCA2 deletion, located in the C-terminal domain, can be considered as pathogenic and consequently affect tumorigenesis because it is involved in the interaction between the DSS1 protein and DNA. Thanks to the optimized bioinformatics algorithm, this intermediate size deletion in BRCA2 was identified, enabling personalized patient management via the inclusion of the patients in a clinical trial.
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A prevalence of around 26% of human papillomavirus (HPV) in head and neck squamous cell carcinoma (HNSCC) has been previously reported. HPV induced oncogenesis mainly involving E6 and E7 viral oncoproteins. In some cases, HPV viral DNA has been detected to integrate with the host genome and possibly contributes to carcinogenesis by affecting the gene expression. We retrospectively assessed HPV integration sites and signatures in 80 HPV positive patients with HNSCC, by using a double capture-HPV method followed by next-generation Sequencing. We detected HPV16 in 90% of the analyzed cohort and confirmed five previously described mechanistic signatures of HPV integration [episomal (EPI), integrated in a truncated form revealing two HPV-chromosomal junctions colinear (2J-COL) or nonlinear (2J-NL), multiple hybrid junctions clustering in a single chromosomal region (MJ-CL) or scattered over different chromosomal regions (MJ-SC) of the human genome]. Our results suggested that HPV remained episomal in 38.8% of the cases or was integrated/mixed in the remaining 61.2% of patients with HNSCC. We showed a lack of association of HPV genomic signatures to tumour and patient characteristics, as well as patient survival. Similar to other HPV associated cancers, low HPV copy number was associated with worse prognosis. We identified 267 HPV-human junctions scattered on most chromosomes. Remarkably, we observed four recurrent integration regions: PDL1/PDL2/PLGRKT (8.2%), MYC/PVT1 (6.1%), MACROD2 (4.1%) and KLF5/KLF12 regions (4.1%). We detected the overexpression of PDL1 and MYC upon integration by gene expression analysis. In conclusion, we identified recurrent targeting of several cancer genes such as PDL1 and MYC upon HPV integration, suggesting a role of altered gene expression by HPV integration during HNSCC carcinogenesis.
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Alphapapillomavirus , Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Proteínas Oncogénicas Virales , Infecciones por Papillomavirus , Carcinogénesis , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , ADN , Genómica , Neoplasias de Cabeza y Cuello/genética , Humanos , Factores de Transcripción de Tipo Kruppel , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/metabolismo , Papillomaviridae/genética , Infecciones por Papillomavirus/complicaciones , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/patología , Estudios Retrospectivos , Carcinoma de Células Escamosas de Cabeza y Cuello/genéticaRESUMEN
PURPOSE: Almost all cervical cancers are caused by human papillomavirus (HPV) and patients with advanced stage are at high risk for relapse. Circulating HPV DNA (HPV ctDNA) may serve as a residual tumor marker at the end of chemoradiation or to predict relapse during the follow-up period. EXPERIMENTAL DESIGN: We analyzed serum samples from 94 HPV16- or HPV18-related CCs from the BioRAIDs prospective cohort. Samples were collected before and after treatment and during an 18-month follow-up period. Using digital droplet PCR (ddPCR), we assessed the relevance of circulating HPV E7 gene as a marker for residual disease compared to HPV integration site and PIK3CA mutations. Finally, the prognostic impact of circulating HPV E7 gene was assessed with its prediction value of relapse. RESULTS: HPV E7 gene was the most sensitive tumor marker, superior to both HPV integration sites and PIK3CA mutations in serum. Circulating HPV DNA (HPV ctDNA) was detected in 63% (59/94) of patients, before treatment. HPV ctDNA detection in serum sample was associated with high FIGO stage (P = 0.02) and para-aortic lymph node involvement (P = 0.01). The level of HPV ctDNA was positively correlated with HPV copy number in the tumor (R = 0.39, P < 0.001). Complete clearance of HPV ctDNA by the end of treatment was significantly associated with a longer PFS (P < 0.0001). Patients with persistent HPV ctDNA in serum relapsed with a median time of 10 months (range, 2-15) from HPV ctDNA detection. CONCLUSIONS: HPV ctDNA detection is a useful marker to predict relapse in cervical cancer.See related commentary by Wentzensen and Clarke, p. 5733.
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Alphapapillomavirus/genética , Biomarcadores de Tumor/sangre , ADN Viral/sangre , Detección Precoz del Cáncer , Recurrencia Local de Neoplasia/sangre , Recurrencia Local de Neoplasia/virología , Neoplasia Residual/sangre , Neoplasia Residual/virología , Infecciones por Papillomavirus/sangre , Neoplasias del Cuello Uterino/sangre , Neoplasias del Cuello Uterino/virología , Adulto , Anciano , Anciano de 80 o más Años , Quimioradioterapia , Femenino , Humanos , Persona de Mediana Edad , Infecciones por Papillomavirus/complicaciones , Estudios Prospectivos , Neoplasias del Cuello Uterino/terapia , Adulto JovenRESUMEN
Precision oncology is currently based on pairing molecularly targeted agents (MTA) to predefined single driver genes or biomarkers. Each tumor harbors a combination of a large number of potential genetic alterations of multiple driver genes in a complex system that limits the potential of this approach. We have developed an artificial intelligence (AI)-assisted computational method, the digital drug-assignment (DDA) system, to prioritize potential MTAs for each cancer patient based on the complex individual molecular profile of their tumor. We analyzed the clinical benefit of the DDA system on the molecular and clinical outcome data of patients treated in the SHIVA01 precision oncology clinical trial with MTAs matched to individual genetic alterations or biomarkers of their tumor. We found that the DDA score assigned to MTAs was significantly higher in patients experiencing disease control than in patients with progressive disease (1523 versus 580, P = 0.037). The median PFS was also significantly longer in patients receiving MTAs with high (1000+ <) than with low (<0) DDA scores (3.95 versus 1.95 months, P = 0.044). Our results indicate that AI-based systems, like DDA, are promising new tools for oncologists to improve the clinical benefit of precision oncology.
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Development of high-throughput technologies helped to decipher tumor genomic landscapes revealing actionable molecular alterations. We aimed to rank the level of evidence of recurrent actionable molecular alterations in head and neck squamous cell carcinoma (HNSCC) on the basis of the European Society for Medical Oncology (ESMO) Scale for Clinical Actionability of Molecular Targets (ESCAT) to help the clinicians prioritize treatment. We identified actionable alterations in 33 genes. HRAS-activating mutations were ranked in tier IB because of the efficacy of tipifarnib (farnesyltransferase inhibitor) in HRAS-mutated patients with HNSCC (nonrandomized clinical trial). Microsatellite instability (MSI), high tumor mutational burden (TMB), and NTRK fusions were ranked in tier IC because of PD-1 and TRK tyrosine kinase inhibitors tissue-agnostic approvals. CDKN2A-inactivating alterations and EGFR amplification were ranked in tier IIA because of the efficacy of palbociclib (CDK4/6 inhibitor) and afatinib (tyrosine kinase inhibitor) in these respective molecular subgroups in retrospective analyses of clinical trials. Molecular alterations in several genes, including PIK3CA gene, were ranked in tier IIIA because of clinical benefit in other tumor types, whereas molecular alterations in IGF1R and TP53 genes were ranked in tier IVA and tier V, respectively. The most compelling actionable molecular alterations in HNSCC according to ESCAT include HRAS-activating mutations, MSI, high TMB, NTRK fusions, CDKN2A-inactivating alterations, and EGFR amplification.
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Neoplasias de Cabeza y Cuello/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Ciclo Celular/genética , Reparación del ADN/genética , Humanos , Sistema de Señalización de MAP Quinasas/genética , Proteínas Tirosina Quinasas Receptoras/metabolismoRESUMEN
Immunotherapy has made a breakthrough in medical oncology with the approval of several immune checkpoint inhibitors in clinical routine, improving overall survival of advanced cancer patients with refractory disease. However only a minority of patients experience a durable response with these agents, which has led to the development of combination strategies and novel immunotherapy drugs to further counteract tumor immune escape. Epigenetic regulations can be altered in oncogenesis, favoring tumor progression. The development of epidrugs has allowed targeting successfully these altered epigenetic patterns in lymphoma and leukemia patients. It has been recently shown that epigenetic alterations can also play a key role in tumor immune escape. Epidrugs, like HDAC inhibitors, can prime the anti-tumor immune response, therefore constituting interesting partners to develop combination strategies with immunotherapy agents. In this review, we will discuss epigenetic regulations involved in oncogenesis and immune escape and describe the clinical development of combining HDAC inhibitors with immunotherapies.
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High-throughput molecular profiling of solid tumours using core needle biopsies (CNB) allows the identification of actionable molecular alterations, with around 70% success rate. Although several studies have demonstrated the utility of small biopsy specimens for molecular testing, there remains debate as to the sensitivity of the less invasive fine-needle aspiration (FNA) compared to CNB to detect molecular alterations. We aimed to prospectively evaluate the potential of FNA to detect such alterations in various tumour types as compared to CNB in cancer patients included in the SHIVA02 trial. An in-house amplicon-based targeted sequencing panel (Illumina TSCA 99.3 kb panel covering 87 genes) was used to identify pathogenic variants and gene copy number variations (CNV) in concomitant CNB and FNA samples obtained from 61 patients enrolled in the SHIVA02 trial (NCT03084757). The main tumour types analysed were breast (38%), colon (15%), pancreas (11%), followed by cervix and stomach (7% each). We report 123 molecular alterations (85 variants, 23 amplifications and 15 homozygous deletions) among which 98 (80%) were concordant between CNB and FNA. The remaining discordances were mainly related to deletions status, yet undetected alterations were not exclusively specific to FNA. Comparative analysis of molecular alterations in CNB and FNA showed high concordance in terms of variants as well as CNVs identified. We conclude FNA could therefore be used in routine diagnostics workflow and clinical trials for tumour molecular profiling with the advantages of being minimally invasive and preserve tissue material needed for diagnostic, prognostic or theranostic purposes.
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Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Neoplasias/genética , Neoplasias/patología , Medicina de Precisión , Biopsia con Aguja Fina , Biopsia con Aguja Gruesa , Variaciones en el Número de Copia de ADN/genética , ADN de Neoplasias/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Estudios Prospectivos , Reproducibilidad de los ResultadosRESUMEN
BACKGROUND: Cervical cancer (CC) remains a leading cause of gynaecological cancer-related mortality with infection by human papilloma virus (HPV) being the most important risk factor. We analysed the association between different viral integration signatures, clinical parameters and outcome in pre-treated CCs. METHODS: Different integration signatures were identified using HPV double capture followed by next-generation sequencing (NGS) in 272 CC patients from the BioRAIDs study [NCT02428842]. Correlations between HPV integration signatures and clinical, biological and molecular features were assessed. RESULTS: Episomal HPV was much less frequent in CC as compared to anal carcinoma (p < 0.0001). We identified >300 different HPV-chromosomal junctions (inter- or intra-genic). The most frequent integration site in CC was in MACROD2 gene followed by MIPOL1/TTC6 and TP63. HPV integration signatures were not associated with histological subtype, FIGO staging, treatment or PFS. HPVs were more frequently episomal in PIK3CA mutated tumours (p = 0.023). Viral integration type was dependent on HPV genotype (p < 0.0001); HPV18 and HPV45 being always integrated. High HPV copy number was associated with longer PFS (p = 0.011). CONCLUSIONS: This is to our knowledge the first study assessing the prognostic value of HPV integration in a prospectively annotated CC cohort, which detects a hotspot of HPV integration at MACROD2; involved in impaired PARP1 activity and chromosome instability.
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Enzimas Reparadoras del ADN/genética , Hidrolasas/genética , Papillomaviridae/fisiología , Infecciones por Papillomavirus/virología , Neoplasias del Cuello Uterino/virología , Integración Viral/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/virología , Fosfatidilinositol 3-Quinasa Clase I/genética , Femenino , Humanos , Calicreínas/genética , Persona de Mediana Edad , Papillomaviridae/genética , Infecciones por Papillomavirus/genética , Supervivencia sin Progresión , Antígeno Prostático Específico/genética , Neoplasias del Cuello Uterino/genéticaRESUMEN
Recently, we detected a new fusion transcript LMO3-BORCS5 in a patient with Ewing sarcoma within a cohort of relapsed pediatric cancers. LMO3-BORCS5 was as highly expressed as the characteristic fusion oncogene EWS/FLI1. However, the expression level of LMO3-BORCS5 at diagnosis was very low. Sanger sequencing depicted two LMO3-BORCS5 variants leading to loss of the functional domain LIM2 in LMO3 gene, and disruption of BORCS5. In vitro studies showed that LMO3-BORCS5 (i) increases proliferation, (ii) decreases expression of apoptosis-related genes and treatment sensitivity, and (iii) downregulates genes involved in differentiation and upregulates proliferative and extracellular matrix-related pathways. Remarkably, in vivo LMO3-BORCS5 demonstrated its high oncogenic potential by inducing tumors in mouse fibroblastic NIH-3T3 cell line. Moreover, BORCS5 probably acts, in vivo, as a tumor-suppressor gene. In conclusion, functional studies of fusion oncogenes at relapse are of great importance to define mechanisms involved in tumor progression and resistance to conventional treatments.
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Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas con Dominio LIM/genética , Proteínas de Fusión Oncogénica/genética , Sarcoma de Ewing/genética , Animales , Progresión de la Enfermedad , Humanos , Ratones , Células 3T3 NIH , Recurrencia Local de Neoplasia/genética , Fusión de Oncogenes , Sarcoma de Ewing/patologíaRESUMEN
We hypothetized that pediatric cancers would more likely harbor fusion transcripts. To dissect the complexity of the fusions landscape in recurrent solid pediatric cancers, we conducted a study on 48 patients with different relapsing or resistant malignancies. By analyzing RNA sequencing data with a new in-house pipeline for fusions detection named ChimComp, followed by verification by real-time PCR, we identified and classified the most confident fusion transcripts (FTs) according to their potential biological function and druggability. The majority of FTs were predicted to affect key cancer pathways and described to be involved in oncogenesis. Contrary to previous descriptions, we found no significant correlation between the number of fusions and mutations, emphasizing the particularity to study pre-treated pediatric patients. A considerable proportion of FTs containing tumor suppressor genes was detected, reflecting their importance in pediatric cancers. FTs containing non-receptor tyrosine kinases occurred at low incidence and predominantly in brain tumors. Remarkably, more than 30% of patients presented a potentially druggable high-confidence fusion. In conclusion, we detected new oncogenic FTs in relapsing pediatric cancer patients by establishing a robust pipeline that can be applied to other malignancies, to detect and prioritize experimental validation studies leading to the development of new therapeutic options.
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Neoplasias/genética , Medicina de Precisión/métodos , Análisis de Secuencia de ARN/métodos , Adolescente , Adulto , Niño , Preescolar , Femenino , Perfilación de la Expresión Génica/métodos , Humanos , Lactante , Masculino , Recurrencia Local de Neoplasia/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Transcriptoma/genética , Adulto JovenRESUMEN
Pediatric cancers differ from adult tumors, especially by their very low mutational rate. Therefore, their etiology could be explained in part by other oncogenic mechanisms such as chromosomal rearrangements, supporting the possible implication of fusion genes in the development of pediatric cancers. Fusion genes result from chromosomal rearrangements leading to the juxtaposition of two genes. Consequently, an abnormal activation of one or both genes is observed. The detection of fusion genes has generated great interest in basic cancer research and in the clinical setting, since these genes can lead to better comprehension of the biological mechanisms of tumorigenesis and they can also be used as therapeutic targets and diagnostic or prognostic biomarkers. In this review, we discuss the molecular mechanisms of fusion genes and their particularities in pediatric cancers, as well as their relevance in murine models and in the clinical setting. We also point out the difficulties encountered in the discovery of fusion genes. Finally, we discuss future perspectives and priorities for finding new innovative therapies in childhood cancer.
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TTF-1 and PAX-8 are responsible for thyroid organogenesis and for maintenance of differentiation in thyrocytes. Thus, we hypothesized that the induction of these two transcription factors could affect proliferation and tumorigenicity. Moreover, the ability of various pharmacological agents to modulate expression of the TTF-1 and PAX-8 and their effects on apoptosis were also analysed. For this purpose, cell lines derived from papillary (TPC-1 and BHP 10-3) and anaplastic (ARO) thyroid carcinomas were stably transfected with expression vectors containing TTF-1 or PAX-8 genes. Subsequently, the effects on expression at gene and protein levels, as well as on cell growth, cell cycle, migration and in vivo tumorigenicity were studied. Our results showed that: i) TTF-1 reciprocally induces PAX-8 expression; ii) the basal state of TTF-1 or PAX-8 influences proliferation, migration and tumorigenicity; iii) the induction of TTF-1 acts on cell proliferation more than PAX-8 and mainly affects tumorigenicity; and iv) TTF-1 was found to be more sensitive to epigenetic modulators than PAX-8. Therefore, we postulated that both TTF-1 and PAX-8 when co-expressed have anti-proliferative and anti-tumorigenic properties up to a threshold expression level and beyond that, are able to induce pro-tumorigenic effects. Hence in future, it will be quite interesting to systematically take into account the basal state of expression of TTF-1 and PAX-8. It will also be important to study the two thyroid transcription factors as part of a duo. This could open in the long-term, new therapeutic perspectives for thyroid carcinomas.