Tumor-draining lymph node (TDLN) invasion by metastatic cells in breast cancer correlates with poor prognosis and is associated with local immunosuppression, which can be partly mediated by ...regulatory T cells (Tregs). Here, we study Tregs from matched tumor-invaded and non-invaded TDLNs, and breast tumors. We observe that Treg frequencies increase with nodal invasion, and that Tregs express higher levels of co-inhibitory/stimulatory receptors than effector cells. Also, while Tregs show conserved suppressive function in TDLN and tumor, conventional T cells (Tconvs) in TDLNs proliferate and produce Th1-inflammatory cytokines, but are dysfunctional in the tumor. We describe a common transcriptomic signature shared by Tregs from tumors and nodes, including CD80, which is significantly associated with poor patient survival. TCR RNA-sequencing analysis indicates trafficking between TDLNs and tumors and ongoing Tconv/Treg conversion. Overall, TDLN Tregs are functional and express a distinct pattern of druggable co-receptors, highlighting their potential as targets for cancer immunotherapy.
Anal carcinomas (AC) are associated with human papillomavirus (HPV) DNA sequences, but little is known about the physical state of the viral genome in carcinoma cells. To define the integration ...status and gene(s) targeted by viral insertions in AC, tumor DNAs extracted from 35 tumor specimen samples in patients with HPV16‐associated invasive carcinoma were analyzed using the detection of integrated papillomavirus sequences‐PCR approach. The genomic status at integration sites was assessed using comparative genomic hybridization‐array assay and gene expression using reverse transcription quantitative PCR (RT‐qPCR). HPV16 DNA was found integrated in 25/35 (71%) cases and the integration locus could be determined at the molecular level in 19 cases (29 total integration loci). HPV DNA was inserted on different chromosomes, but 5 cases harbored viral sequences at 19p13.2, within the nuclear factor I X (NFIX) locus. Viral DNA mapped between the most distal and the two proximal alternatively expressed exons of this gene in three cases (CA21, CA04, and CA35) and upstream of this gene (663 kb and 2.3 Mb) in the others. CGH arrays showed genomic gains/amplifications at the NFIX region, associated with HPV within the gene and RT‐qPCR, revealed NFIX mRNA overexpression. Other genes targeted by integration were IL20RB, RPS6KA2, MSRA1, PIP5K1B, SLX4IP, CECR1, BCAR3, ATF6, CSNK1G1, APBA2, AGK, ILF3, PVT1, TRMT1, RAD51B, FASN, CCDC57, DSG3, and ZNF563. We identified recurrent targeting of NFIX by HPV16 insertion in anal carcinomas, supporting a role for this gene in oncogenesis, as reported for non‐HPV tumors.
JunD regulates genes involved in antioxidant defence. We took advantage of the chronic oxidative stress resulting from junD deletion to examine the role of reactive oxygen species (ROS) in tumour ...development. In a model of mammary carcinogenesis, junD inactivation increased tumour incidence and revealed an associated reactive stroma. junD‐inactivation in the stroma was sufficient to shorten tumour‐free survival rate and enhance metastatic spread. ROS promoted conversion of fibroblasts into highly migrating myofibroblasts through accumulation of the hypoxia‐inducible factor (HIF)‐1α transcription factor and the CXCL12 chemokine. Accordingly, treatment with an antioxidant reduced the levels of HIF and CXCL12 and numerous myofibroblast features. CXCL12 accumulated in the stroma of HER2‐human breast adenocarcinomas. Moreover, HER2 tumours exhibited a high proportion of myofibroblasts, which was significantly correlated to nodal metastases. Interestingly, this subset of tumours exhibited a significant nuclear exclusion of JunD and revealed an associated oxido‐reduction signature, further demonstrating the relevance of our findings in human cancers. Collectively, our data uncover a new mechanism by which oxidative stress increases the migratory properties of stromal fibroblasts, which in turn potentiate tumour dissemination.
Specific human papillomavirus genotypes are associated with most ano‐genital carcinomas and a large subset of oro‐pharyngeal carcinomas. Human papillomavirus DNA is thus a tumour marker that can be ...detected in the blood of patients for clinical monitoring. However, data concerning circulating human papillomavirus DNA in cervical cancer patients has provided little clinical value, due to insufficient sensitivity of the assays used for the detection of small sized tumours. Here we took advantage of the sensitive droplet digital PCR method to identify circulating human papillomavirus DNA in patients with human papillomavirus‐associated carcinomas.
A series of 70 serum specimens, taken at the time of diagnosis, between 2002 and 2013, were retrospectively analyzed in patients with human papillomavirus‐16 or human papillomavirus‐18‐associated carcinomas, composed of 47 cases from the uterine cervix, 15 from the anal canal and 8 from the oro‐pharynx. As negative controls, 18 serum samples from women with human papillomavirus‐16‐associated high‐grade cervical intraepithelial neoplasia were also analyzed. Serum samples were stored at −80°C (27 cases) or at −20°C (43 cases). DNA was isolated from 200 µl of serum or plasma and droplet digital PCR was performed using human papillomavirus‐16 E7 and human papillomavirus‐18 E7 specific primers.
Circulating human papillomavirus DNA was detected in 61/70 (87%) serum samples from patients with carcinoma and in no serum from patients with cervical intraepithelial neoplasia. The positivity rate increased to 93% when using only serum stored at −80°C. Importantly, the two patients with microinvasive carcinomas in this series were positive. Quantitative evaluation showed that circulating viral DNA levels in cervical cancer patients were related to the clinical stage and tumour size, ranging from 55 ± 85 copies/ml (stage I) to 1774 ± 3676 copies/ml (stage IV).
Circulating human papillomavirus DNA is present in patients with human papillomavirus‐associated invasive cancers even at sub‐clinical stages and its level is related to tumour dynamics. Droplet digital PCR is a promising method for circulating human papillomavirus DNA detection and quantification. No positivity was found in patients with human papillomavirus‐associated high grade cervical intraepithelial neoplasia.
Mammalian cells possess two isoforms of the histone H3–H4 chaperone anti‐silencing function 1 (Asf1), Asf1a and Asf1b. However to date, whether they have individual physiological roles has remained ...elusive. Here, we aim to elucidate the functional importance of Asf1 isoforms concerning both basic and applied aspects. First, we reveal a specific proliferation‐dependent expression of human Asf1b unparalleled by Asf1a. Strikingly, in cultured cells, both mRNA and protein corresponding to Asf1b decrease upon cell cycle exit. Depletion of Asf1b severely compromises proliferation, leads to aberrant nuclear structures and a distinct transcriptional signature. Second, a major physiological implication is found in the applied context of tissue samples derived from early stage breast tumours in which we examined Asf1a/b levels. We reveal that overexpression of Asf1b mRNA correlate with clinical data and disease outcome. Together, our results highlight a distribution of tasks between the distinct Asf1 isoforms, which emphasizes a specialized function of Asf1b required for proliferation capacity. We discuss the implications of these results for breast cancer diagnosis and prognosis.
Anti‐silencing function 1 (Asf1) is a H3–H4 chaperone involved in both replication‐coupled and replication‐independent nucleosome assembly pathways. Here in mammalian cells Asf1b is the physiologically important isoform for cell proliferation and overexpression of Asf1b correlates with prognosis and metastasis status of breast cancer.
Dendritic cells (DCs) initiate adaptive immune responses in lymph nodes (LNs). In mice, LN DCs can be divided into resident and tissue-derived populations, the latter of which migrate from the ...peripheral tissues. In humans, different subsets of DCs have been identified in the blood, spleen, and skin, but less is known about populations of resident and migratory tissue-derived DCs in LNs. We have analyzed DCs in human LNs and identified two populations of resident DCs that are present in all LNs analyzed, as well as in the spleen and tonsil, and correspond to the two known blood DC subtypes. We also identify three main populations of skin-derived migratory DCs that are present only in skin-draining LNs and correspond to the DC subsets found in the skin. Resident DCs subsets induce both Th1 and Th2 cytokines in naive allogeneic T lymphocytes, whereas the corresponding blood subsets failed to induce efficient Th2 polarization. LN-resident DCs also cross-present antigen without in vitro activation, whereas blood DCs fail to do so. Among migratory DCs, one subset was poor at both CD4(+) and CD8(+) T cell activation, whereas the other subsets induced only Th2 polarization. We conclude that in humans, skin-draining LNs host both resident and migratory DC subsets with distinct functional abilities.
Although there is evidence that redox regulation has an essential role in malignancies, its impact on tumor prognosis remains unclear. Here we show crosstalk between oxidative stress and the miR-200 ...family of microRNAs that affects tumorigenesis and chemosensitivity. miR-141 and miR-200a target p38α and modulate the oxidative stress response. Enhanced expression of these microRNAs mimics p38α deficiency and increases tumor growth in mouse models, but it also improves the response to chemotherapeutic agents. High-grade human ovarian adenocarcinomas that accumulate miR-200a have low concentrations of p38α and an associated oxidative stress signature. The miR200a-dependent stress signature correlates with improved survival of patients in response to treatment. Therefore, the role of miR-200a in stress could be a predictive marker for clinical outcome in ovarian cancer. In addition, although oxidative stress promotes tumor growth, it also sensitizes tumors to treatment, which could account for the limited success of antioxidants in clinical trials.
Objectives
With the perspective of prophylactic vaccination against high‐risk human papillomavirus (HPV), we analyzed the viral epidemiology of cervical neoplasia in Senegal.
Methods
All patients ...were treated at the Institut Joliot Curie du Cancer in Dakar. HPV genotypes were characterized using a real‐time polymerase chain reaction‐based approach and sequencing.
Results
Histologically, there were 224 invasive carcinomas, 17 high‐grade intraepithelial neoplasia (CIN), and five undetermined histologies. Molecular analysis was conclusive in 241 cases. HPV DNA was found in 207/241 (85.9%) cases while 34/241 (14.1%) remained HPV negative. There was one single genotype in 127/207 (61.4%) cases and several in 80/207 (38.6%) corresponding to 308 genotypes identified. Viral genotyping found HPV16 in 175 (56.8%) cases, HPV18 in 45 (14.6%), HPV45 in 40 (13.0%), HPV58 in 35 (11.4%), HPV33 in 6 (2.0%), HPV35 in 3 (1.0%), HPV31 in 2 (0.6%), HPV39 and HPV56 in one (0.3% each).
Conclusion
Our analysis showed that 98.4% of the HPV‐positive cases were associated with viral genotypes covered by the 9‐valent HPV vaccine. However, 14.1% of cases remained HPV negative. Therefore, prophylactic vaccination using a 9‐valent vaccine should dramatically reduce the incidence of HPV‐associated neoplasia but the detection and treatment of CIN remain necessary for the optimal prevention of cervical cancer.
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