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Purpose The role of non-genetic factors as modifiers of TP53-related hereditary breast cancer (BC) risk is debated. In this regard, little is known about the impact of germline TP53 mutations on BC in sub-Saharan Africa, where the disease often presents in non-contraceptive multiparous premenopausal women with extended history of breastfeeding. Herein, we report the germline TP53 mutations found in a series of 92 Sudanese premenopausal BC patients characterized for reproductive history. Methods The entire TP53 coding sequence, including intron–exon boundaries and UTRs, was analyzed via DHPLC and direct sequencing, and the association of TP53 genotypes with BC risk and with individual lifetime exposures to reproductive factors was investigated with statistical tools. Results The germline TP53 mutation spectrum comprised 20 variants, 15 in the non-coding and 5 in the coding region. The latter included a deleterious missense mutation, c.817C>T (p.Arg273Cys), in a unique patient, and the common and functionally relevant coding polymorphism at amino acid 72 [Pro72Arg (rs1042522)]. The non-coding mutations included c.919+1G>A, a known deleterious splice site mutation, also in a unique patient. Notably, the 2 carriers of deleterious TP53 mutations clustered in the subset of cases with stronger reproductive history relative to childbearing age. When analyzed in comparison to population controls, the codon 72 polymorphism did not reveal associations with BC. Conclusions Our study suggests that the codon 72 Arg>Pro polymorphism is not implicated in premenopausal BC susceptibility, whereas multiparity and breastfeeding might be BC risk factors for carriers of deleterious TP53 mutations. Electronic supplementary material The online version of this article (10.1007/s10549-019-05168-1) contains supplementary material, which is available to authorized users.

The clinical progression of B cell chronic lymphocytic leukemia (CLL) is associated with immune cell dysfunction and a strong decrease of miR-181b-5p (miR-181b), promoting the death of CLL cells. Here we investigated whether the reduction of miR-181b impairs the immune response in CLL. We demonstrate that activated CD4+ T cells increase miR-181b expression in CLL through CD40&ndash, CD40L signaling, which enhances the maturation and activity of cytotoxic T cells and, consequently, the apoptotic response of CLL cells. The cytotoxic response is facilitated by a depletion of the anti-inflammatory cytokine interleukin 10, targeted by miR-181b. In vivo experiments in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice confirmed that miR-181b promotes the apoptotic death of CLL cells only when functional T cells are restored. Overall, our findings suggest that the reinstatement of miR-181b in CLL cells could be an exploitable adjuvant therapeutic option for the treatment of CLL.

The crypt-villus axis represents the essential unit of the small intestine, which integrity and functions are fundamental to assure tissue and whole-body homeostasis. Disruption of pathways regulating the fine balance between proliferation and differentiation results in diseases development. Nowadays, it is well established that microRNAs (miRNAs) play a crucial role in the homeostasis maintenance and perturbation of their levels may promote tumor development. Here, by using microarray technology, we analysed the miRNAs differentially expressed between the crypt and the villus in mice ileum. The emerged miRNAs were further validated by Real Time qPCR in mouse model (ApcMin/+), human cell lines and human tissue samples (FAP) of colorectal cancer (CRC). Our results indicated that miRNAs more expressed in the villi compartment are negatively regulated in tumor specimens, thus suggesting a close association between these microRNAs and the differentiation process. Particularly, from our analysis let-7e appeared to be a promising target for possible future therapies and a valuable marker for tumor staging, being upregulated in differentiated cells and downregulated in early-stage colonic adenoma samples.

Simple Summary Low expression of miR-181b in chronic lymphocytic leukemia (CLL) is linked to progression and cell death resistance. Patients with the progression of the disease show immune dysfunction. We aim at studying whether miR-181b is also involved in this process. We demonstrate that miR-181b can be increased in CLL cells by crosstalk with CD40L+ T cells, enhancing the maturation of CD8+ T cells in cytotoxic T lymphocytes (CTL) and, in turn, the anti-tumor cytotoxic T cell response in in vitro and in vivo models. These results demonstrate a role of the miR-181b in the immune response against tumor and envisage a therapeutic action of miR-181b in a specific milieu surrounding CLL cells. Abstract The clinical progression of B cell chronic lymphocytic leukemia (CLL) is associated with immune cell dysfunction and a strong decrease of miR-181b-5p (miR-181b), promoting the death of CLL cells. Here we investigated whether the reduction of miR-181b impairs the immune response in CLL. We demonstrate that activated CD4+ T cells increase miR-181b expression in CLL through CD40–CD40L signaling, which enhances the maturation and activity of cytotoxic T cells and, consequently, the apoptotic response of CLL cells. The cytotoxic response is facilitated by a depletion of the anti-inflammatory cytokine interleukin 10, targeted by miR-181b. In vivo experiments in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice confirmed that miR-181b promotes the apoptotic death of CLL cells only when functional T cells are restored. Overall, our findings suggest that the reinstatement of miR-181b in CLL cells could be an exploitable adjuvant therapeutic option for the treatment of CLL.

Helicobacter pylori (Hp) is the major recognized risk factor for non-cardia gastric cancer (GC), but only a fraction of infected subjects develop GC, thus GC risk might reflect other genetic/environmental cofactors and/or differences in virulence among infectious Hp strains. Focusing on a high GC risk area of Northern Italy (Cremona, Lombardy) and using archived paraffin-embedded biopsies, we investigated the associations between the Hp vacA and cagA genotype variants and gastric intraepithelial neoplasia (GIN, 33 cases) versus non-neoplastic gastroduodenal lesions (NNGDLs, 37 cases). The glmM gene and the cagA and vacA (s and m) genotypes were determined by polymerase chain reaction (PCR) and sequencing. Hp was confirmed in 37/37 (100%) NNGDLs and detected in 9/33 GINs (27%), consistently with the well-known Hp loss in GC. CagA was detected in 4/9 Hp-positive GINs and in 29/37 NNGDLs. The vacA s1a and m1 subtypes were more common in GINs than in NNGDLs (6/7 vs. 12/34, p=0.014, for s1a; 7/7 vs. 18/34, p=0.020 for m1), with significant vacA s genotype-specific variance. The GIN-associated vacA s1a sequences clustered together, suggesting that aggressive Hp strains from a unique founder contribute to GC in the high-risk area studied.

Helicobacter pylori (Hp) is the major recognized risk factor for non-cardia gastric cancer (GC), but only a fraction of infected subjects develop GC, thus GC risk might reflect other genetic/environmental cofactors and/or differences in virulence among infectious Hp strains. Focusing on a high GC risk area of Northern Italy (Cremona, Lombardy) and using archived paraffin-embedded biopsies, we investigated the associations between the Hp vacA and cagA genotype variants and gastric intraepithelial neoplasia (GIN, 33 cases) versus non-neoplastic gastroduodenal lesions (NNGDLs, 37 cases). The glmM gene and the cagA and vacA (s and m) genotypes were determined by polymerase chain reaction (PCR) and sequencing. Hp was confirmed in 37/37 (100%) NNGDLs and detected in 9/33 GINs (27%), consistently with the well-known Hp loss in GC. CagA was detected in 4/9 Hp-positive GINs and in 29/37 NNGDLs. The vacA s1a and m1 subtypes were more common in GINs than in NNGDLs (6/7 vs. 12/34, p=0.014, for s1a, 7/7 vs. 18/34, p=0.020 for m1), with significant vacA s genotype-specific variance. The GIN-associated vacA s1a sequences clustered together, suggesting that aggressive Hp strains from a unique founder contribute to GC in the high-risk area studied.

The interaction between NPs and immune system has been demonstrated, however, the data available are limited. Among all traits, i.s. hydrophilicity, lipophilicity, catalytic activity, composition, electronic structure, capacity to bind or coat surface species and solubility, the dimension, and consequently the surface area, seems to be the main factor that contribute to the interactions of NPs with biological tissues and immune system in particular. Certain NPs accumulate to regional lymph nodes, where they can be taken up and processed by dendritic cells, interact with self-proteins and, hence, modify their antigenicity and elicit altered immune responses and even autoimmunity. Other NPs may induce allergic sensitization, i.e. allergic contact dermatitis to Pd. In vitro studies demonstrated that NPs can modulate cytokine production toward Th1 (Pl, Pd, Ni, Co) or Th2 (Ti, mw and sw Carbon) production patterns. Some NPs have been linked to allergic sensitization, however, It is unlikely that NPs can act as a hapten inducing a specific IgE production, likely they can act as adjuvant and induce a specific pattern of cytokines, antibody and cells that favor allergic sensitization to environmental allergens. Furthermore, NPs demonstrated pro-inflammatory effects in the lung in experimental animal with increased expression on IL-1beta, MIP-1alpha, MCP-1, MIP-2, keratinocyte chemoattractant, TARC, GM-CSF, MIP-1alpha and activation of the stress-activated MAPKs p38 and JNKs. All considered, the available data suggest that through the elicitation of an oxidative stress mechanism, engineered NPs may contribute to pro-inflammatory disease processes in the lung, particularly allergy.

Tumours can be viewed as aberrant tissues or organs sustained by tumorigenic stem-like cells that engage into dysregulated histo/organogenetic processes. Paragangliomas, prototypical organoid tumours constituted by dysmorphic variants of the vascular and neural tissues found in normal paraganglia, provide a model to test this hypothesis. To understand the origin of paragangliomas, we built a biobank comprising 77 cases, 18 primary cultures, 4 derived cell lines, 80 patient-derived xenografts and 11 cell-derived xenografts. We comparatively investigated these unique complementary materials using morphofunctional, ultrastructural and flow cytometric assays accompanied by microRNA studies. We found that paragangliomas contain stem-like cells with hybrid mesenchymal/vasculoneural phenotype, stabilized and expanded in the derived cultures. The viability and growth of such cultures depended on the downregulation of the miR-200 and miR-34 families, which allowed high PDGFRA and ZEB1 protein expression levels. Both tumour tissue- and cell culture-derived xenografts recapitulated the vasculoneural paraganglioma structure and arose from mesenchymal-like cells through a fixed developmental sequence. First, vasculoangiogenesis organized the microenvironment, building a perivascular niche which in turn supported neurogenesis. Neuroepithelial differentiation was associated with severe mitochondrial dysfunction, not present in cultured paraganglioma cells, but acquired in vivo during xenograft formation. Vasculogenesis was the Achilles’ heel of xenograft development. In fact, imatinib, that targets endothelial-mural signalling, blocked paraganglioma xenograft formation (11 xenografts from 12 cell transplants in the control group versus 2 out of 10 in the treated group, P = 0.0015). Overall our key results were unaffected by the SDHx gene carrier status of the patient, characterized for 70 out of 77 cases. In conclusion, we explain the biphasic vasculoneural structure of paragangliomas and identify an early and pharmacologically actionable phase of paraganglioma organization. Electronic supplementary material The online version of this article (10.1007/s00401-017-1799-2) contains supplementary material, which is available to authorized users.

Clonal evolution of chronic lymphocytic leukemia (CLL) often follows chemotherapy and is associated with adverse outcome, but also occurs in untreated patients, in which case its predictive role is debated. We investigated whether the selection and expansion of CLL clone(s) precede an aggressive disease shift. We found that clonal evolution occurs in all CLL patients, irrespective of the clinical outcome, but is faster during disease progression. In particular, changes in the frequency of nucleotide variants (NVs) in specific CLL-related genes may represent an indicator of poor clinical outcome.

Carotid body and vagal paragangliomas, although considered indolent tumors, represent a challenge for the treating physician. This is mainly because of their peculiar localization, in close proximity with important anatomical structures. In addition, there is no chemotherapy available for these tumors, the role of radiation therapy is debated, and the only successful therapy is surgery. However, to achieve the best treatment goals, it is fundamental for the professional caregiver to master not only the clinical and surgical procedures but also the genetic backgrounds and the histopathological features of these tumors. We provide in this chapter a comprehensive review of the above mentioned aspects, with the aim to address the complexity of these tumors with a multidisciplinary approach.

Paragangliomas and pheochromocytomas are rare tumors of the autonomic nervous system that represent a diagnostic and therapeutic challenge. While the classic sites of origin are the adrenal medulla and the autonomic branches of the lower cranial nerves, they may arise from any site in connection with the autonomic neural branches in the thoracic, abdominal, and head and neck regions. The uncontrolled release of catecholamines by these tumors can be life-threatening. Paragangliomas and pheochromocytomas are the tumor types in which the impact of genetic predisposition factors is the highest. The treatment of paraganglioma and pheochromocytoma requires specialized knowledge and solid experience, which, given the rarity of these diseases, is available only in highly specialized centers. This book includes specific chapters written by experts from several countries that deal with relevant aspects of paraganglioma and pheochromocytoma diagnosis and therapy. Chapter 1 addresses the genetic findings that are most relevant to diagnosis and management, chapter 2 deals with the biochemical diagnosis, chapter 3 with imaging, chapter 4 with the surgical and pharmacological management, mainly of trunk paraganglioma and pheochromocytoma, chapters 5 and 6 with the histopathological, genetic, and clinical characteristics of carotid body, and vagal and tympano-jugular paragangliomas.

Aim: To investigate the genome-wide methylation of genetically characterized colorectal cancer stem cell (CR-CSC) lines. Materials & methods: Eight CR-CSC lines were isolated from primary colorectal cancer (CRC) tissues, cultured and characterized for aneuploidy, mutational status of CRC-related genes and microsatellite instability (MSI). Genome-wide DNA methylation was assessed by MethylationEPIC microarray. Results: We describe a distinctive methylation pattern that is maintained following in vivo passages in immune-compromised mice. We identified an epigenetic CR-CSC signature associated with MSI. We noticed that the preponderance of the differentially methylated positions do not reside at CpG islands, but spread to shelf and open sea regions. Conclusion: Given that CRCs with MSI-high status have a lower metastatic potential, the identification of a MSI-related methylation signature could provide new insights and possible targets into metastatic CRC.

Aneuploidy and overexpression of hsa-miR-155-5p (miR-155) characterize most solid and hematological malignancies. We recently demonstrated that miR-155 sustains aneuploidy at early stages of in vitro cellular transformation. During in vitro transformation of normal human fibroblast, upregulation of miR-155 downregulates spindle checkpoint proteins as the mitotic checkpoint serine/threonine kinase budding uninhibited by benzimidazoles 1 (BUB1), the centromere protein F (CENPF) and the zw10 kinetochore protein (ZW10), compromising the chromosome alignment at the metaphase plate and leading to aneuploidy in daughter cells. Here we show that the heterogeneous nuclear ribonucleoprotein L (HNRNPL) binds to the polymorphic marker D2S1888 at the 3&prime, UTR of BUB1 gene, impairs the miR-155 targeting, and restores BUB1 expression in chronic lymphocytic leukemia. This mechanism occurs at advanced passages of cell transformation and allows the expansion of more favorable clones. Our findings have revealed, at least in part, the molecular mechanisms behind the chromosomal stabilization of cell lines and the concept that, to survive, tumor cells cannot continuously change their genetic heritage but need to stabilize the most suitable karyotype.

In this review, we propose that paraganglioma is a fundamentally organized, albeit aberrant, tissue composed of neoplastic vascular and neural cell types that share a common origin from a multipotent mesenchymal-like stem/progenitor cell. This view is consistent with the pseudohypoxic footprint implicated in the molecular pathogenesis of the disease, is in harmony with the neural crest origin of the paraganglia, and is strongly supported by the physiological model of carotid body hyperplasia. Our immunomorphological and molecular studies of head and neck paragangliomas demonstrate in all cases relationships between the vascular and the neural tumor compartments, that share mesenchymal and immature vasculo-neural markers, conserved in derived cell cultures. This immature, multipotent phenotype is supported by constitutive amplification of NOTCH signaling genes and by loss of the microRNA-200s and -34s, which control NOTCH1, ZEB1, and PDGFRA in head and neck paraganglioma cells. Importantly, the neuroepithelial component is distinguished by extreme mitochondrial alterations, associated with collapse of the ΔΨm. Finally, our xenograft models of head and neck paraganglioma demonstrate that mesenchymal-like cells first give rise to a vasculo-angiogenic network, and then self-organize into neuroepithelial-like clusters, a process inhibited by treatment with imatinib.

Background Current approaches aimed at inducing immunogenic cell death (ICD) to incite an immune response against cancer neoantigens are based on the use of chemotherapeutics and other agents. Results are hampered by issues of efficacy, combinatorial approaches, dosing and toxicity. Here, we adopted a strategy based on the use of an immunomolecule that overcomes pharmachemical limitations. Methods Cytofluorometry, electron microscopy, RT-PCR, western blotting, apotome immunofluorescence, MLR and xenografts. Results We report that an ICD process can be activated without the use of pharmacological compounds. We show that in Kras-mut/TP53-mut colorectal cancer cells the 15 kDa βGBP cytokine, a T cell effector with onco-suppressor properties and a potential role in cancer immunosurveillance, induces key canonical events required for ICD induction. We document ER stress, autophagy that extends from cancer cells to the corresponding xenograft tumours, CRT cell surface shifting, ATP release and evidence of dendritic cell activation, a process required for priming cytotoxic T cells into a specific anticancer immunogenic response. Conclusions Our findings provide experimental evidence for a rationale to explore a strategy based on the use of an immunomolecule that as a single agent couples oncosuppression with the activation of procedures necessary for the induction of long term response to cancer.

Extracellular vesicles act as shuttle vectors or signal transducers that can deliver specific biological information and have progressively emerged as key regulators of organized communities of cells within multicellular organisms in health and disease. Here, we survey the evolutionary origin, general characteristics, and biological significance of extracellular vesicles as mediators of intercellular signaling, discuss the various subtypes of extracellular vesicles thus far described and the principal methodological approaches to their study, and review the role of extracellular vesicles in tumorigenesis, immunity, non-synaptic neural communication, vascular-neural communication through the blood-brain barrier, renal pathophysiology, and embryo-fetal/maternal communication through the placenta.

Paragangliomas (PGLs) are infiltrating autonomic nervous system tumors that cause important morbidity. At present, surgery is the only effective therapeutic option for this rare tumor. Thus, new agents for PGL treatment should be identified. Using unique PGL cell models established in our laboratory, we evaluated the effect of dichloroacetate (DCA) as single agent or in a novel combination with other metabolic drugs, including GW6471 and metformin. DCA and metformin had not been tested before in PGL. DCA reduced PGL cell viability and growth through mechanisms involving reactivation of PDH complex leading to promotion of oxidative metabolism, with lowering of lactate and enhanced ROS production. This resulted in cell cycle inhibition and induction of apoptosis in PGL cells, as shown by flow cytometry and immunoblot analyses. Moreover, DCA drastically impaired clonogenic activity and migration of PGL cells. Also metformin reduced PGL cell viability as single agent and the combinations of DCA, GW6471 and metformin had strong effects on cell viability. Furthermore, combined treatments had drastic and synergistic effects on clonogenic ability. In conclusion, DCA, GW6471 and metformin as single agents and in combination appear to have promising antitumor effects in unique cell models of PGL.

Head and neck paragangliomas (HNPGLs) are rare tumors that may cause important morbidity, because of their tendency to infiltrate the skull base. At present, surgery is the only therapeutic option, but radical removal may be difficult or impossible. Thus, effective targets and molecules for HNPGL treatment need to be identified. However, the lack of cellular models for this rare tumor hampers this task. PPARα receptor activation was reported in several tumors and this receptor appears to be a promising therapeutic target in different malignancies. Considering that the role of PPARα in HNPGLs was never studied before, we analyzed the potential of modulating PPARα in a unique model of HNPGL cells. We observed an intense immunoreactivity for PPARα in HNPGL tumors, suggesting that this receptor has an important role in HNPGL. A pronounced nuclear expression of PPARα was also confirmed in HNPGL-derived cells. The specific PPARα agonist WY14643 had no effect on HNPGL cell viability, whereas the specific PPARα antagonist GW6471 reduced HNPGL cell viability and growth by inducing cell cycle arrest and caspase-dependent apoptosis. GW6471 treatment was associated with a marked decrease of CDK4, cyclin D3 and cyclin B1 protein expression, along with an increased expression of p21 in HNPGL cells. Moreover, GW6471 drastically impaired clonogenic activity of HNPGL cells, with a less marked effect on cell migration. Notably, the effects of GW6471 on HNPGL cells were associated with the inhibition of the PI3K/GSK3β/β-catenin signaling pathway. In conclusion, the PPARα antagonist GW6471 reduces HNPGL cell viability, interfering with cell cycle and inducing apoptosis. The mechanisms affecting HNPGL cell viability involve repression of the PI3K/GSK3β/β-catenin pathway. Therefore, PPARα could represent a novel therapeutic target for HNPGL.

*Department of Otology and Skull Base Surgery, Gruppo Otologico, Piacenza-Rome; ÞDepartment of Clinicaland Experimental Medicine, Otolaryngology Unit, University-Hospital of Parma, Parma; þUnit of GeneralPathology, Aging Research Center (Ce.S.I.), G. d’Annunzio University Foundation, Chieti; §Department ofMedical, Oral and Biotechnological Sciences, G. d’Annunzio University, Chieti; and kDepartment ofOtolaryngology, University of Chieti, ItalyObjective: To define the role of wait-and-scan and radiotherapy(RT) in the treatment of temporal bone paragangliomas andto review the literature on the outcomes of RT as a treatmentmodality.Materials and Methods: This was a retrospective and literaturereview.Therecordsof381patientswith382tumorsbetween1988and 2012 were analyzed. Patients who were subjected to initialwait-and-scan or primary RT were analyzed for tumor response,facial, and lower cranial nerve status on follow-up (FU).Results and Observations: None of the 145 patients (38%)with Class A and B tumors were subjected to wait-and-scan orRT and all were operated on. Of the 237 patients (62%) withClass C and D tumors, 47 (19.8%) were subjected to an initialwait-and-scan policy and 1 (0.4%) underwent primary RT. In24 patients subjected to wait-and-scan with a FU of less than3 years, tumor remained stable in 22 patients (92%). In 12patients with a FU between 3 and 5 years, the tumor remainedstable or regressed in 10 patients (83%). In 11 patients with aFU of more than 5 years, the tumor remained stable or regressedin 5 patients (45%). A literature review of the results of RT didnot conclusively prove that it was effective as a primary mo-dality of treatment for temporal bone paragangliomas.Conclusion: In elderly patients with advanced tumors, a wait-and-scan approach is recommended, and RT is initiated onlywhen fast-growing tumors are detected. There is insufficientevidence in literature to suggest that RT could be an effectivealternative to surgery in Class C and D tumors. Key Words: Lowercranial nerve palsyVRadiotherapyVStereotactic radiosurgeryVTemporal bone paragangliomas VTympanojugular paragangliomas VTympanomastoid paragangliomasVWait-and-scan.Otol Neurotol 00:00Y00, 2014.

Premenopausal breast cancer (BC) is one of the most common cancers of women in rural Africa and part of the disease load may be related to hereditary predisposition, including mutations in the BRCA1 gene. However, the BRCA1 mutations associated with BC in Africa are scarcely characterized. We report here 33 BRCA1 point mutations, among which 2 novel missense variants, found in 59 Central Sudanese premenopausal BC patients. The high fractions of mutations with intercontinental and uniquely African distribution (17/33, 51.5 % and 14/33, 42.4 %, respectively) are in agreement with the high genetic diversity expected in an African population. Overall 24/33 variants (72.7 %) resulted neutral; 8/33 of unknown significance (24.3 %, including the 2 novel missense mutations); 1 (3.0 %) overtly deleterious. Notably, in silico studies predict that the novel C-terminal missense variant c.5090G>A (p.Cys1697Tyr) affects phosphopeptide recognition by the BRCA1 BRCT1 domain and may have a pathogenic impact. Genetic variation and frequency of unique or rare mutations of uncertain clinical relevance pose significant challenges to BRCA1 testing in Sudan, as it might happen in other low-resource rural African contexts.

Ki-67 labeling index has been linked to patient outcome in breast cancer patients. However, very few published reports have examined Ki-67 labeling index in African breast cancer patients. Sixty-two Sudanese breast cancer patients with primary invasive tumors were immunostained for Ki-67, ER, PR, Her-2/neu, CK5/6, and CK17. Ki-67 labeling index ranged from 0% to 50%, with a median of 5% (interquartile range 0–10). Low Ki-67 labeling index (immunostaining < 10%) was detected in 43/62 (69.4%) with a median of 0 (interquartile range 0–5), whereas high Ki-67 labeling index (immunostaining ≥ 10%) was revealed in 19/62 (30.6%) with a median of 20 (interquartile range 12–26). Ki-67 labeling index was significantly associated with tumor grade (P=0.022, Mann-Whitney U Test). There were no significant group differences between Ki-67 labeling index and ER (P=0.43), PR (P=0.7), Her-2/neu (P=0.45), CK5/6 (P=0.29), CK17 (P=0.55), pathologic stage (P=0.4), tumor histology (P=0.99), breast cancer subtypes (P=0.47), tumor size (P=0.16), and age at diagnosis (P=0.6). These results suggested that Ki-67 labeling index correlates with tumor differentiation and not with the tumor size or any other tested marker in Sudanese breast cancers. Thus, Ki-67 labeling index could be considered as a reliable measure of tumor proliferative fraction in Sudan.

The present day distribution of Y chromosomes bearing the haplogroup J1 M267(*)G variant has been associated with different episodes of human demographic history, the main one being the diffusion of Islam since the Early Middle Ages. To better understand the modes and timing of J1 dispersals, we reconstructed the genealogical relationships among 282 M267(*)G chromosomes from 29 populations typed at 20 YSTRs and 6 SNPs. Phylogenetic analyses depicted a new genetic background consistent with climate-driven demographic dynamics occurring during two key phases of human pre-history: (1) the spatial expansion of hunter gatherers in response to the end of the late Pleistocene cooling phases and (2) the displacement of groups of foragers/herders following the mid-Holocene rainfall retreats across the Sahara and Arabia. Furthermore, J1 STR motifs previously used to trace Arab or Jewish ancestries were shown unsuitable as diagnostic markers for ethnicity.

// Laura Lupini 1, * , Felice Pepe 2, 8, * , Manuela Ferracin 3 , Chiara Braconi 4 , Elisa Callegari 1 , Sara Pagotto 2, 8 , Riccardo Spizzo 5 , Barbara Zagatti 1 , Paola Lanuti 6 , Francesca Fornari 7 , Reza Ghasemi 2 , Renato Mariani-Costantini 2, 8 , Luigi Bolondi 7 , Laura Gramantieri 7 , George A. Calin 9 , Silvia Sabbioni 10 , Rosa Visone 2, 8 , Angelo Veronese 2, 8 , Massimo Negrini 1 1 Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy 2 Unit of General Pathology, Aging Research Center (Ce.S.I.), G. d’Annunzio University Foundation, Chieti, Italy 3 Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy 4 Division of Cancer Therapeutics, Institute of Cancer Research, London, UK 5 Division of Experimental Oncology 2 CRO, Aviano, Italy 6 Department of Medicine and Aging Science, G. d’Annunzio University, Chieti, Italy 7 S.Orsola-Malpighi University Hospital, Bologna, Italy 8 Department of Medical, Oral and Biotechnological Sciences, G. d’Annunzio University, Chieti, Italy 9 Department of Experimental Therapeutics, MD Anderson Medical Centre, Houston, TX, USA 10 Department of Life Sciences and Biotechnology, University of Ferrara, Ferrara, Italy * These authors have contributed equally to this work Correspondence to: Angelo Veronese, email: a.veronese@unich.it Massimo Negrini, email: ngm@unife.it Keywords: TP53, hsa-miR-145-5p, hsa-miR-483-3p, HCC, PUMA Received: October 15, 2015 Accepted: April 10, 2016 Published: April 22, 2016 ABSTRACT The miR-145-5p, which induces TP53-dependent apoptosis, is down-regulated in several tumors, including hepatocellular carcinomas (HCCs), but some HCCs show physiological expression of this miR. Here we demonstrate that in HCC cells carrying wild-type TP53 the steady activation of the miR-145 signaling selects clones resistant to apoptosis via up-regulation of the oncogenic miR-483-3p. Expression of the miR-145-5p and of the miR-483-3p correlated negatively in non-neoplastic liver (n=41; ρ=-0.342, P=0.028), but positively in HCCs (n=21; ρ=0.791, P

The Sudan, the largest and most diverse country in Africa, is experiencing a growing cancer problem, but little is presently known on tumor patterns, cancer epidemiology and ethnic or environmental cancer risk factors. We review here the current status of knowledge, summarizing data from local and international publications as well as primary information from the only two cancer hospitals of the country, both located in Central Sudan (Khartoum and Wad Medani). We provide frequencies reported for cancers detected in adults and children, and summarize studies on specific cancer types, as well as information on risk factors that most likely impact on tumor patterns.

Metabolic syndrome (MS) is a cluster of different diseases, namely central obesity, hypertension, hyperglycemia, and dyslipidemia, together with a pro-thrombotic and pro-inflammatory state. These metabolic abnormalities are often associated with an increased risk for cardiovascular disease (CVD) and cancer. Dietary and lifestyle modifications are currently believed more effective than pharmacological therapies in the management of MS patients. Nevertheless, the relatively low grade of compliance of patients to these recommendations, as well as the failure of current therapies, highlights the need for the discovery of new pharmacological and nutraceutic approaches. A deeper knowledge of the patho-physiological events that initiate and support the MS is mandatory. Lipid-sensing nuclear receptors (NRs) are the master transcriptional regulators of lipid and carbohydrate metabolism and inflammatory responses, thus standing as suitable targets. This review focuses on the physiological relevance of the NRs (peroxisome proliferator-activated receptors, liver X receptors, and farnesoid X receptor) in the control of whole-body homeostasis, with a special emphasis on lipid and glucose metabolism, and on the relationships between metabolic unbalances, systemic inflammation, and the onset of CVD. Future perspectives and possible clinical applications are also presented.

Adjuvant therapy is a systemic treatment administered after primary tumor resection to reduce the risk of relapse and death in colorectal cancer (CRC) patients. Generally, adjuvant treatment is recommended for stage III and ‘high-risk' stage II CRC (NCCN guidelines). Nonetheless, adjuvant treatment is still debated for stage IIA (T3N0) patients, whose estimated recurrence rate is 15-20% in the absence of any further therapy after resection of the primary tumor. With the aim to find additional prognostic biomarkers that could improve current patient selection, we analyzed the global methylation profile of a discovery group of 10 relapsing/non-relapsing stage IIA colon cancer samples, without any other known risk factor (low-risk), by Illumina Epic 850K array. We identified a panel of 25 Illumina probes that were able to predict recurrence in chemotherapy-naïve patients. Then, we assessed the methylation of the most predictive Cytosine-Guanine (CpG) dinucleotides in 107 stage IIA colon cancer patients from two different cohorts, by EpiTYPER technology. We validated several CpGs as differentially methylated in relapsing vs. non-relapsing CRCs (p Citation Format: Sara Pagotto, Elisa Porcellini, Maria G. Bacalini, Diletta Sarti, Ingrid Garajova, Elisabetta Broseghini, Noemi Laprovitera, Francesco Vasuri, Paolo Garagnani, Renato Mariani-Costantini, Michele Basso, Rosa Visone, Luigi M. Larocca, Manuela Ferracin, Angelo Veronese. Epigenetic biomarkers of prognosis in stage IIA colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3313.

The given and family names of two co-authors were incorrect in the published article. The correct spelling should read as: Sampath Chandra Prasad and Vinagolu K Rajasekhar.

Juvenile breast cancer is rare and poorly known. We studied a series of five breast cancer patients diagnosed within 25 years of age that included two adolescents, 12- and 15-years-old, and 3 young women, 21-, 21-, and 25-years-old, respectively. All cases were scanned for germline mutations along the entire BRCA1/2 coding sequences and TP53 exons 4-10, using protein truncation test, denaturing high performance liquid chromatography and direct sequencing. Paraffin-embedded primary tumors (available for 4/5 cases), and a distant metastasis (from the 15-years-old) were characterized for histological and molecular tumor subtype, human papilloma virus (HPV) types 16/18 E6 sequences and tumor-associated mutations in TP53 exons 5-8. A BRCA2 germline mutation (p.Ile2490Thr), previously reported in breast cancer and, as compound heterozygote, in Fanconi anemia, was identified in the 21-year-old patient diagnosed after pregnancy, negative for cancer family history. The tumor was not available for study. Only germline polymorphisms in BRCA1/2 and/or TP53 were detected in the other cases. The tumors of the 15- and 12-years-old were, respectively, classified as glycogen-rich carcinoma with triple negative subtype and as secretory carcinoma with basal subtype. The tumors of the 25-year-old and of the other 21-year-old were, respectively, diagnosed as infiltrating ductal carcinoma with luminal A subtype and as lobular carcinoma with luminal B subtype. No somatic TP53 mutations were found, but tumor-associated HPV 16 E6 sequences were retrieved from the 12- and 25-year-old, while both HPV 16 and HPV 18 E6 sequences were found in the tumor of the 15-year-old and in its associated metastasis. Blood from the 15- and 25-year-old, diagnosed with high-stage disease, resulted positive for HPV 16 E6. All the HPV-positive cases were homozygous for arginine at TP53 codon 72, a genotype associated with HPV-related cancer risk, and the tumors showed p16(INK4A) immunostaining, a marker of HPV-associated cancers. Notably menarche at 11 years was reported for the two adolescents, while the 25-year-old was diagnosed after pregnancy and breast-feeding. Our data suggest that high-risk HPV infection is involved in a subset of histopathologically heterogeneous juvenile breast carcinomas associated with menarche or pregnancy and breast-feeding. Furthermore we implicate BRCA2 in a juvenile breast carcinoma diagnosed at 21 years of age, 4 years after an early full-term pregnancy, in absence of cancer family history. Fil: Aceto, Gitana Maria. University of G. D'annunzio Chieti And Pescara; Italia Fil: Solano, Angela Rosario. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Neuman, Maria Isabel. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Bioquímica; Argentina Fil: Veschi, Serena. University of G. D'annunzio Chieti And Pescara; Italia Fil: Morgano, Annalisa. University of G. D'annunzio Chieti And Pescara; Italia Fil: Malatesta, Sara. University of G. D'annunzio Chieti And Pescara; Italia Fil: Chacon, Reinaldo Daniel. Instituto Alexander Fleming; Argentina Fil: Pupareli, Carmen. Instituto Alexander Fleming; Argentina Fil: Lombardi, Mercedes. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina Fil: Battista, Pasquale. University of G. D'annunzio Chieti And Pescara; Italia Fil: Marchetti, Antonio. University of G. D'annunzio Chieti And Pescara; Italia Fil: Mariani Costantini, Renato. University of G. D'annunzio Chieti And Pescara; Italia Fil: Podesta, Ernesto Jorge. Universidad de Buenos Aires. Facultad de Medicina. Departamento de Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Background: Altered germline expression of genes may represent a powerful marker of genetic or epigenetic predisposition to cancer or other diseases.Methods: We developed and validated a method of nonfluorescent primer extension that uses a single dideoxynucleotide and denaturing HPLC (DHPLC) to analyze the relative allele expression. We devised 5 independent assays for measuring allele-specific expression (ASE) to exploit different markers of mismatch repair genes MLH1 [mutL homolog 1, colon cancer, nonpolyposis type 2 (E. coli)] and MSH2 [mutS homolog 2, colon cancer, nonpolyposis type 1 (E. coli)]. We initially confirmed method reproducibility with genomic DNA (gDNA) from individuals heterozygous for a frequent single-nucleotide polymorphism in the MLH1 gene. After this preliminary validation with gDNA, we confirmed assay reproducibility with cDNA templates from control individuals. Relative allele expression was estimated by comparing the heights of the peaks corresponding to the 2 alleles. Results obtained with gDNA templates were used to normalize cDNA results.Results: With these DHPLC-based primer-extension assays, we detected and confirmed a 5-fold imbalance in MLH1 allele expression in a mutation-negative patient with hereditary nonpolyposis colorectal cancer and in another patient with a modest degree of imbalance in MLH1 expression. Among control individuals, the relative expression of MLH1 alleles displayed a narrow range of variation.Conclusions: Independent DHPLC-based primer-extension assays for measuring and confirming ASE can be developed for different sequence variants of interest. This DHPLC application provides a cost-effective method for detecting ASE in cases for which conventional screening fails to detect pathogenic mutations in candidate genes and may be applicable for confirming ASE revealed by other methods, such as those used for transcriptome-wide analyses. .

Background and aims: Self-renewal and differentiation of intestinal epithelium is a tightly regulated process, whose perturbations are implicated in human colorectal tumourigenesis. The insulin/insulin-like growth factor (IGF) signalling pathway may play an important role in intestinal epithelium homeostasis. Insulin receptor substrate 2 (IRS2) is a poorly characterised component in this pathway. Methods: Using complementary in vitro and in vivo human and murine models, expression (mRNA and protein levels), localisation (immunohistochemistry) and regulation of IRS2 were investigated in the normal intestine and colorectal tumours. In silico analysis of the human IRS2 promoter was performed together with reporter and chromatin immunoprecipitation assays. Results: Significant IRS2 expression was detected in the intestine, with specific protein localisation in the villus region of the ileum and in the surface epithelium of the colon. In human HT29 and Caco2 cells, IRS2 mRNA levels increased with spontaneous and induced differentiation, together with CDX2 (caudal-related homeobox protein 2), P21 and KLF4 (Kruppel-like factor 4). Adenoviral infection with human CDX2 induced IRS2 expression in APC- (adenomatous polyposis coli) and β-catenin-mutated cells. On the other hand, IRS2 downregulation was observed in differentiated enterocytes after adenoviral infection with short hairpin CDX2 (shCDX2), in the intestine of CDX2 heterozygous mice and in colorectal tumours of Apc Min/+ and patients with familial adenomatous polyposis (FAP). The human IRS2 promoter region presents several CDX2-binding sites where CDX2 immunoprecipitated in vivo. IRS2 reporters were functionally activated via CDX2 and blocked via a dominant-negative CDX2 protein. Conclusions: Combining gain- and loss-of-function approaches, an intriguing scenario is presented whereby IRS2 is significantly expressed in the apical intestinal compartment and is directly controlled by CDX2 in normal intestine and tumours.

Bio-cultural adaptations to new foods played a key role in human evolution. The fossil record and sequence differences between human and chimpanzee genes point to a major dietary shift at the stem of human evolution. The earliest representatives of the human lineage diverged from the ancestors of chimpanzees because of their better adaptation to hard and abrasive foods. Bipedalism and modifications of the hand, which allowed tool manufacture and use, impacted on dietary flexibility, facilitating access to foods of animal origin. This promoted major anatomic, physiologic and metabolic adaptations. Encephalization, which requires high-quality diet, characterizes the evolutionary sequence that, through the Homo ergaster/erectus stages, led to our species, Homo sapiens, which originated in Africa about 200,000 years ago. At the end of the Ice Age, climatic changes and human impact determined a major food crisis, which triggered the agricultural revolution. This affected nutrition and health, with rapid evolutionary adaptations through the selection of genetic variants that allowed better utilization of new foods, different in relation to geography and culture. Today population growth, globalization and economic pressure powerfully affect diets worldwide. We must take into account our evolutionary past to meet the present nutritional challenges.

Africa is the ultimate source of modern humans and as such harbors more genetic variation than any other continent. For this reason, studies of the patterns of genetic variation in African populations are crucial to understanding how genes affect phenotypic variation, including disease predisposition. In addition, the patterns of extant genetic variation in Africa are important for understanding how genetic variation affects infectious diseases that are a major problem in Africa, such as malaria, tuberculosis, schistosomiasis, and HIV/AIDS. Therefore, elucidating the role that genetic susceptibility to infectious diseases plays is critical to improving the health of people in Africa. It is also of note that recent and ongoing social and cultural changes in sub-Saharan Africa have increased the prevalence of non-communicable diseases that will also require genetic analyses to improve disease prevention and treatment. In this review we give special attention to many of the past and ongoing studies, emphasizing those in Sub-Saharan Africans that address the role of genetic variation in human disease.

CRC-associated P53 mutations have not been studied extensively in non-Western countries at relatively low CRC risk. We examined, for the first time, 196 paraffin-embedded CRC cases from Northern Iran for mutations in P53 exons 5-8 using PCR-direct sequencing. P53 status and mutation site/type were correlated with nuclear protein accumulation, clinicopathologic variables and data on K-ras mutations and high-level microsatellite instability (MSI-H). We detected 96 P53 mutations in 87 (44.4%) cases and protein accumulation in 84 cases (42.8%). P53 mutations correlated directly with stage and inversely with MSI-H. Distal CRCs were more frequently mutated at major CpG hotspot codons [248 (8/66, 12.1%), 175 (7/66, 10.6%), and 245 (7/66, 10.6%)], while in proximal tumors codon 213, emerged as most frequently mutated (5/28, 17.9% vs. 3/66, 4.5%, P = 0.048). Transitions at CpGs, the most common mutation type, were more frequent in non-mucinous (25% vs. 10.4% in mucinous, P = 0.032), and distal CRC (27% vs. 12.5% in proximal, P = 0.02), and correlated with K-ras transversions. Transitions at non-CpGs, second most common P53 mutation, were more frequent in proximal tumors (15.6% vs. 4.7% in distal, P = 0.01), and correlated with K-ras transitions and MSI-H. Overall frequency and types of mutations and correlations with P53 accumulation, stage and MSI-H were as reported for non-Iranian patients. However P53 mutation site/type and correlations between P53 and K-ras mutation types differed between proximal and distal CRC. The codon 213 P53 mutation that recurred in proximal CRC was previously reported as frequent in esophageal cancer from Northern Iran.