Effects of Local Acidification on Benthic Communities at Shallow Hydrothermal Vents of the Aeolian Islands (Southern Tyrrhenian, Mediterranean Sea).

The Aeolian Islands (Mediterranean Sea) host a unique hydrothermal system called the "Smoking Land" due to the presence of over 200 volcanic CO2-vents, resulting in water acidification phenomena and the creation of an acidified benthic environment. Here, we report the results of a study conducted at three sites located at ca. 16, 40, and 80 m of depth, and characterized by CO2 emissions to assess the effects of acidification on meio- and macrobenthic assemblages. Acidification caused significant changes in both meio- and macrofaunal assemblages, with a clear decrease in terms of abundance and a shift in community composition. A noticeable reduction in biomass was observed only for macrofauna. The most sensitive meiofaunal taxa were kinorhynchs and turbellarians that disappeared at the CO2 sites, while the abundance of halacarids and ostracods increased, possibly as a result of the larger food availability and the lower predatory pressures by the sensitive meiofaunal and macrofaunal taxa. Sediment acidification also causes the disappearance of more sensitive macrofaunal taxa, such as gastropods, and the increase in tolerant taxa such as oligochaetes. We conclude that the effects of shallow CO2-vents result in the progressive simplification of community structure and biodiversity loss due to the disappearance of the most sensitive meio- and macrofaunal taxa.

Apoptosis inhibition restrains primary malignant traits in different Drosophila cancer models.

Tumor cells exploit multiple mechanisms to evade apoptosis, hence the strategies aimed at reactivating cell death in cancer. However, recent studies are revealing that dying cells play remarkable pro-oncogenic roles. Among the mechanisms promoting cell death, cell competition, elicited by disparities in MYC activity in confronting cells, plays the primary role of assuring tissue robustness during development from Drosophila to mammals: cells with high MYC levels (winners) overproliferate while killing suboptimal neighbors (losers), whose death is essential to process completion. This mechanism is coopted by tumor cells in cancer initiation, where host cells succumb to high-MYC-expressing precancerous neighbors. Also in this case, inhibition of cell death restrains aberrant cell competition and rescues tissue structure. Inhibition of apoptosis may thus emerge as a good strategy to counteract cancer progression in competitive contexts; of note, we recently found a positive correlation between cell death amount at the tumor/stroma interface and MYC levels in human cancers. Here we used Drosophila to investigate the functional role of competition-dependent apoptosis in advanced cancers, observing dramatic changes in mass dimensions and composition following a boost in cell competition, rescued by apoptosis inhibition. This suggests the role of competition-dependent apoptosis be not confined to the early stages of tumorigenesis. We also show that apoptosis inhibition, beside restricting cancer mass, is sufficient to rescue tissue architecture and counteract cell migration in various cancer contexts, suggesting that a strong activation of the apoptotic pathways intensifies cancer burden by affecting distinct phenotypic traits at different stages of the disease.

Restoration of the molecular clock is tumor suppressive in neuroblastoma.

MYCN activation is a hallmark of advanced neuroblastoma (NB) and a known master regulator of metabolic reprogramming, favoring NB adaptation to its microenvironment. We found that the expression of the main regulators of the molecular clock loops is profoundly disrupted in MYCN-amplified NB patients, and this disruption independently predicts poor clinical outcome. MYCN induces the expression of clock repressors and downregulates the one of clock activators by directly binding to their promoters. Ultimately, MYCN attenuates the molecular clock by suppressing BMAL1 expression and oscillation, thereby promoting cell survival. Reestablishment of the activity of the clock activator RORα via its genetic overexpression and its stimulation through the agonist SR1078, restores BMAL1 expression and oscillation, effectively blocks MYCN-mediated tumor growth and de novo lipogenesis, and sensitizes NB tumors to conventional chemotherapy. In conclusion, reactivation of RORα could serve as a therapeutic strategy for MYCN-amplified NBs by blocking the dysregulation of molecular clock and cell metabolism mediated by MYCN.

Preliminary report on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike mutation T478K.

Several severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have emerged, posing a renewed threat to coronavirus disease 2019 containment and to vaccine and drug efficacy. In this study, we analyzed more than 1,000,000 SARS-CoV-2 genomic sequences deposited up to April 27, 2021, on the GISAID public repository, and identified a novel T478K mutation located on the SARS-CoV-2 Spike protein. The mutation is structurally located in the region of interaction with human receptor ACE2 and was detected in 11,435 distinct cases. We show that T478K has appeared and risen in frequency since January 2021, predominantly in Mexico and the United States, but we could also detect it in several European countries.

A G316A Polymorphism in the Ornithine Decarboxylase Gene Promoter Modulates MYCN-Driven Childhood Neuroblastoma.

Ornithine decarboxylase (ODC1), a critical regulatory enzyme in polyamine biosynthesis, is a direct transcriptional target of MYCN, amplification of which is a powerful marker of aggressive neuroblastoma. A single nucleotide polymorphism (SNP), G316A, within the first intron of ODC1, results in genotypes wildtype GG, and variants AG/AA. CRISPR-cas9 technology was used to investigate the effects of AG clones from wildtype MYCN-amplified SK-N-BE(2)-C cells and the effect of the SNP on MYCN binding, and promoter activity was investigated using EMSA and luciferase assays. AG clones exhibited decreased ODC1 expression, growth rates, and histone acetylation and increased sensitivity to ODC1 inhibition. MYCN was a stronger transcriptional regulator of the ODC1 promoter containing the G allele, and preferentially bound the G allele over the A. Two neuroblastoma cohorts were used to investigate the clinical impact of the SNP. In the study cohort, the minor AA genotype was associated with improved survival, while poor prognosis was associated with the GG genotype and AG/GG genotypes in MYCN-amplified and non-amplified patients, respectively. These effects were lost in the GWAS cohort. We have demonstrated that the ODC1 G316A polymorphism has functional significance in neuroblastoma and is subject to allele-specific regulation by the MYCN oncoprotein.

Exploring MYC relevance to cancer biology from the perspective of cell competition.

Cancer has long been regarded and treated as a foreign body appearing by mistake inside a living organism. However, now we know that cancer cells communicate with neighbours, thereby creating modified environments able to support their unusual need for nutrients and space. Understanding the molecular basis of these bi-directional interactions is thus mandatory to approach the complex nature of cancer. Since their discovery, MYC proteins have been showing to regulate a steadily increasing number of processes impacting cell fitness, and are consistently found upregulated in almost all human tumours. Of interest, MYC takes part in cell competition, an evolutionarily conserved fitness comparison strategy aimed at detecting weakened cells, which are then committed to death, removed from the tissue and replaced by fitter neighbours. During physiological development, MYC-mediated cell competition is engaged to eliminate cells with suboptimal MYC levels, so as to guarantee selective growth of the fittest and proper homeostasis, while transformed cells expressing high levels of MYC coopt cell competition to subvert tissue constraints, ultimately disrupting homeostasis. Therefore, the interplay between cells with different MYC levels may result in opposite functional outcomes, depending on the nature of the players. In the present review, we describe the most recent findings on the role of MYC-mediated cell competition in different contexts, with a special emphasis on its impact on cancer initiation and progression. We also discuss the relevance of competition-associated cell death to cancer disease.

Inhibition of polyamine synthesis and uptake reduces tumor progression and prolongs survival in mouse models of neuroblastoma.

Amplification of the MYCN oncogene is associated with an aggressive phenotype and poor outcome in childhood neuroblastoma. Polyamines are highly regulated essential cations that are frequently elevated in cancer cells, and the rate-limiting enzyme in polyamine synthesis, ornithine decarboxylase 1 (ODC1), is a direct transcriptional target of MYCN. Treatment of neuroblastoma cells with the ODC1 inhibitor difluoromethylornithine (DFMO), although a promising therapeutic strategy, is only partially effective at impeding neuroblastoma cell growth due to activation of compensatory mechanisms resulting in increased polyamine uptake from the surrounding microenvironment. In this study, we identified solute carrier family 3 member 2 (SLC3A2) as the key transporter involved in polyamine uptake in neuroblastoma. Knockdown of SLC3A2 in neuroblastoma cells reduced the uptake of the radiolabeled polyamine spermidine, and DFMO treatment increased SLC3A2 protein. In addition, MYCN directly increased polyamine synthesis and promoted neuroblastoma cell proliferation by regulating SLC3A2 and other regulatory components of the polyamine pathway. Inhibiting polyamine uptake with the small-molecule drug AMXT 1501, in combination with DFMO, prevented or delayed tumor development in neuroblastoma-prone mice and extended survival in rodent models of established tumors. Our findings suggest that combining AMXT 1501 and DFMO with standard chemotherapy might be an effective strategy for treating neuroblastoma.

MAX to MYCN intracellular ratio drives the aggressive phenotype and clinical outcome of high risk neuroblastoma.

Childhood neuroblastoma, a disease of the sympathetic nervous system, is the most common solid tumour of infancy, remarkably refractory to therapeutic treatments. One of the most powerful independent prognostic indicators for this disease is the amplification of the MYCN oncogene, which occurs at high levels in approximately 25% of neuroblastomas. Interestingly, amplification and not just expression of MYCN has a strong prognostic value, although this fact appears quite surprising as MYCN is a transcription factor that requires dimerising with its partner MAX, to exert its function. This observation greatly suggests that the role of MYCN in neuroblastoma should be examined in the context of MAX expression. In this report, we show that, in contrast to what is found in normal cells, MAX expression is significantly different among primary NBs, and that its level appears to correlate with the clinical outcome of the disease. Importantly, controlled modulation of MAX expression in neuroblastoma cells with different extents of MYCN amplification, demonstrates that MAX can instruct gene transcription programs that either reinforce or weaken the oncogenic process enacted by MYCN. In general, our work illustrates that it is the MAX to MYCN ratio that can account for tumour progression and clinical outcome in neuroblastoma and proposes that such a ratio should be considered as an important criterion to the design and development of anti-MYCN therapies.

High MYC Levels Favour Multifocal Carcinogenesis.

The term "field cancerisation" describes the formation of tissue sub-areas highly susceptible to multifocal tumourigenesis. In the earlier stages of cancer, cells may indeed display a series of molecular alterations that allow them to proliferate faster, eventually occupying discrete tissue regions with irrelevant morphological anomalies. This behaviour recalls cell competition, a process based on a reciprocal fitness comparison: when cells with a growth advantage arise in a tissue, they are able to commit wild-type neighbours to death and to proliferate at their expense. It is known that cells expressing high MYC levels behave as super-competitors, able to kill and replace less performant adjacent cells; given MYC upregulation in most human cancers, MYC-mediated cell competition is likely to pioneer field cancerisation. Here we show that MYC overexpression in a sub-territory of the larval wing epithelium of Drosophila is sufficient to trigger a number of cellular responses specific to mammalian pre-malignant tissues. Moreover, following induction of different second mutations, high MYC-expressing epithelia were found to be susceptible to multifocal growth, a hallmark of mammalian pre-cancerous fields. In summary, our study identified an early molecular alteration implicated in field cancerisation and established a genetically amenable model which may help study the molecular basis of early carcinogenesis.

Long-term survivors of pancreatic adenocarcinoma show low rates of genetic alterations in KRAS, TP53 and SMAD4.

BACKGROUND: Pancreatic adenocarcinoma (PDAC) is one of the deadliest human malignancies. Although surgery is currently the only effective treatment for PDAC, most patients survive less than 20 months after tumor resection. OBJECTIVE: The primary goal was to investigate alterations in KRAS, TP53, SMAD4 and CDKN2A/p16 in tumors from patients with exceptionally long survival after surgery. METHODS: Tumors from 15 patients with PDAC that survived more than 55 months after surgery ("LS") were analyzed for KRAS, TP53, IDH1, NRAS and BRAF using next-generation sequencing. SMAD4 and CDKN2A/p16 was tested using immunohistochemistry. MGMT promoter methylation was investigated. RESULTS: Tumors from "LS" have a lower prevalence of KRAS and TP53 mutations and had more frequently SMAD4 retained expression, if compared with that of patients died within 24 months from surgery. The survival of patients with wild-type KRAS and TP53 tumors was more than twice longer than that of patients bearing KRAS and TP53 mutations (90.2 vs. 41.1 months). Patients with KRAS wild-type tumors and that retained SMAD4 expression had a survival twice longer than cases with alterations in both genes (83.8 vs. 36.7 months). Eleven tumors (39.3%) showed MGMT methylation. CONCLUSIONS: Our data indicate that absence of KRAS, TP53 and SMAD4 genetic alterations may identify a subset of pancreatic carcinomas with better outcome.

Human Cancer Cells Signal Their Competitive Fitness Through MYC Activity.

MYC-mediated cell competition is a cell-cell interaction mechanism known to play an evolutionary role during development from Drosophila to mammals. Cells expressing low levels of MYC, called losers, are committed to die by nearby cells with high MYC activity, called winners, that overproliferate to compensate for cell loss, so that the fittest cells be selected for organ formation. Given MYC's consolidated role in oncogenesis, cell competition is supposed to be relevant to cancer, but its significance in human malignant contexts is largely uncharacterised. Here we show stereotypical patterns of MYC-mediated cell competition in human cancers: MYC-upregulating cells and apoptotic cells were indeed repeatedly found at the tumour-stroma interface and within the tumour parenchyma. Cell death amount in the stromal compartment and MYC protein level in the tumour were highly correlated regardless of tumour type and stage. Moreover, we show that MYC modulation in heterotypic co-cultures of human cancer cells is sufficient as to subvert their competitive state, regardless of genetic heterogeneity. Altogether, our findings suggest that the innate role of MYC-mediated cell competition in development is conserved in human cancer, with malignant cells using MYC activity to colonise the organ at the expense of less performant neighbours.

MYC, Cell Competition, and Cell Death in Cancer: The Inseparable Triad.

Deregulation of MYC family proteins in cancer is associated with a global reprogramming of gene expression, ultimately promoting glycolytic pathways, cell growth, and proliferation. It is well known that MYC upregulation triggers cell-autonomous apoptosis in normal tissues, while frankly malignant cells develop resistance to apoptotic stimuli, partly resulting from MYC addiction. As well as inducing cell-autonomous apoptosis, MYC upregulation is able to trigger non cell-autonomous apoptotic death through an evolutionarily conserved mechanism known as "cell competition". With regard to this intimate and dual relationship between MYC and cell death, recent evidence obtained in Drosophila models of cancer has revealed that, in early tumourigenesis, MYC upregulation guides the clonal expansion of mutant cells, while the surrounding tissue undergoes non-cell autonomous death. Apoptosis inhibition in this context was shown to restrain tumour growth and to restore a wild-type phenotype. This suggests that cell-autonomous and non cell-autonomous apoptosis dependent on MYC upregulation may shape tumour growth in different ways, soliciting the need to reconsider the role of cell death in cancer in the light of this new level of complexity. Here we review recent literature about MYC and cell competition obtained in Drosophila, with a particular emphasis on the relevance of cell death to cell competition and, more generally, to cancer. Possible implications of these findings for the understanding of mammalian cancers are also discussed.

Failure of the PTEN/aPKC/Lgl Axis Primes Formation of Adult Brain Tumours in Drosophila.

Different regions in the mammalian adult brain contain immature precursors, reinforcing the concept that brain cancers, such as glioblastoma multiforme (GBM), may originate from cells endowed with stem-like properties. Alterations of the tumour suppressor gene PTEN are very common in primary GBMs. Very recently, PTEN loss was shown to undermine a specific molecular axis, whose failure is associated with the maintenance of the GBM stem cells in mammals. This axis is composed of PTEN, aPKC, and the polarity determinant Lethal giant larvae (Lgl): PTEN loss promotes aPKC activation through the PI3K pathway, which in turn leads to Lgl inhibition, ultimately preventing stem cell differentiation. To find the neural precursors responding to perturbations of this molecular axis, we targeted different neurogenic regions of the Drosophila brain. Here we show that PTEN mutation impacts aPKC and Lgl protein levels also in Drosophila. Moreover, we demonstrate that PI3K activation is not sufficient to trigger tumourigenesis, while aPKC promotes hyperplastic growth of the neuroepithelium and a noticeable expansion of the type II neuroblasts. Finally, we show that these neuroblasts form invasive tumours that persist and keep growing in the adult, leading the affected animals to untimely death, thus displaying frankly malignant behaviours.