Classification of tumor types using XGBoost machine learning model: a vector space transformation of genomic alterations.

BACKGROUND: Machine learning (ML) represents a powerful tool to capture relationships between molecular alterations and cancer types and to extract biological information. Here, we developed a plain ML model aimed at distinguishing cancer types based on genetic lesions, providing an additional tool to improve cancer diagnosis, particularly for tumors of unknown origin. METHODS: TCGA data from 9,927 samples spanning 32 different cancer types were downloaded from cBioportal. A vector space model type data transformation technique was designed to build consistently homogeneous new datasets containing, as predictive features, calls for somatic point mutations and copy number variations at chromosome arm-level, thus allowing the use of the XGBoost classifier models. Considering the imbalance in the dataset, due to large difference in the number of cases for each tumor, two preprocessing strategies were considered: i) setting a percentage cut-off threshold to remove less represented cancer types, ii) dividing cancer types into different groups based on biological criteria and training a specific XGBoost model for each of them. The performance of all trained models was mainly assessed by the out-of-sample balanced accuracy (BACC) and the AUC scores. RESULTS: The XGBoost classifier achieved the best performance (BACC 77%; AUC 97%) on a dataset containing the 10 most represented tumor types. Moreover, dividing the 18 most represented cancers into three different groups (endocrine-related carcinomas, other carcinomas and other cancers),such analysis models achieved 78%, 71% and 86% BACC, respectively, with AUC scores greater than 96%. In addition, the model capable of linking each group to a specific cancer type reached 81% BACC and 94% AUC. Overall, the diagnostic potential of our model was comparable/higher with respect to others already described in literature and based on similar molecular data and ML approaches. CONCLUSIONS: A boosted ML approach able to accurately discriminate different cancer types was developed. The methodology builds datasets simpler and more interpretable than the original data, while keeping enough information to accurately train standard ML models without resorting to sophisticated Deep Learning architectures. In combination with histopathological examinations, this approach could improve cancer diagnosis by using specific DNA alterations, processed by a replicable and easy-to-use automated technology. The study encourages new investigations which could further increase the classifier's performance, for example by considering more features and dividing tumors into their main molecular subtypes.

MiR-182-5p Is Upregulated in Hepatic Tissues from a Diet-Induced NAFLD/NASH/HCC C57BL/6J Mouse Model and Modulates Cyld and Foxo1 Expression.

Non-alcoholic fatty liver disease (NAFLD) is considered a relevant liver chronic disease. Variable percentages of NAFLD cases progress from steatosis to steatohepatitis (NASH), cirrhosis and, eventually, hepatocellular carcinoma (HCC). In this study, we aimed to deepen our understanding of expression levels and functional relationships between miR-182-5p and Cyld-Foxo1 in hepatic tissues from C57BL/6J mouse models of diet-induced NAFL/NASH/HCC progression. A miR-182-5p increase was detected early in livers as NAFLD damage progressed, and in tumors compared to peritumor normal tissues. An in vitro assay on HepG2 cells confirmed Cyld and Foxo1, both tumor-suppressor, as miR-182-5p target genes. According to miR-182-5p expression, decreased protein levels were observed in tumors compared to peritumor tissues. Analysis of miR-182-5p, Cyld and Foxo1 expression levels, based on datasets from human HCC samples, showed results consistent with those from our mouse models, and also highlighted the ability of miR-182-5p to distinguish between normal and tumor tissues (AUC 0.83). Overall, this study shows, for the first time, miR-182-5p overexpression and Cyld-Foxo1 downregulation in hepatic tissues and tumors from a diet-induced NAFLD/HCC mouse model. These data were confirmed by the analysis of datasets from human HCC samples, highlighting miR-182-5p diagnostic accuracy and demonstrating the need for further studies to assess its potential role as a biomarker or therapeutic target.

NF-κB and mitochondria cross paths in cancer: mitochondrial metabolism and beyond.

NF-κB plays a pivotal role in oncogenesis. This transcription factor is best known for promoting cancer cell survival and tumour-driving inflammation. However, several lines of evidence support a crucial role for NF-κB in governing energy homeostasis and mediating cancer metabolic reprogramming. Mitochondria are central players in many metabolic processes altered in cancer. Beyond their bioenergetic activity, several facets of mitochondria biology, including mitochondrial dynamics and oxidative stress, promote and sustain malignant transformation. Recent reports revealed an intimate connection between NF-κB pathway and the oncogenic mitochondrial functions. NF-κB can impact mitochondrial respiration and mitochondrial dynamics, and, reciprocally, mitochondria can sense stress signals and convert them into cell biological responses leading to NF-κB activation. In this review we discuss their emerging reciprocal regulation and the significance of this interplay for anticancer therapy.

A natural product, voacamine, sensitizes paclitaxel-resistant human ovarian cancer cells.

Most women with ovarian cancer are treated with chemotherapy before or after surgery. Unfortunately, chemotherapy treatment can cause negative side effects and the onset of multidrug resistance (MDR). The aim of this study is to evaluate the chemosensitizing effect of a natural compound, voacamine (VOA), in ovarian (A2780 DX) and colon (LoVo DX) cancer drug-resistant cell lines which overexpress P-glycoprotein (P-gp), in combination with paclitaxel (PTX), or doxorubicin (DOX) or 5-fluorouracil (5-FU). VOA, a bisindole alkaloid extracted from Peschiera fuchsiaefolia, has already been shown to be effective in enhancing the effect of doxorubicin, because it interferes with the P-gp function. Ovarian cancer cytotoxicity test shows that single treatments with VOA, DOX and PTX do not modify cell viability, while pretreatment with VOA, and then PTX or DOX for 72 h, induces a decrease. In colon cancer, since 5-FU is not a-substrate for P-gp, VOA has no sensitizing effect while in VOA + DOX there is a decrease in viability. Annexin V/PI test, cell cycle analysis, activation of cleaved PARP1 confirm that VOA plus PTX induce apoptotic cell death. Confocal microscopy observations show the different localization of NF-kB after treatment with VOA + PTX, confirming the inhibition of nuclear translocation induced by VOA pretreatment. Our data show the specific effect of VOA which only works on drugs known to be substrates of P-gp.

MicroRNAs Expression in Response to rhNGF in Epithelial Corneal Cells: Focus on Neurotrophin Signaling Pathway.

PURPOSE: Nerve growth factor efficacy was demonstrated for corneal lesions treatment, and recombinant human NGF (rhNGF) was approved for neurotrophic keratitis therapy. However, NGF-induced molecular responses in cornea are still largely unknown. We analyzed microRNAs expression in human epithelial corneal cells after time-dependent rhNGF treatment. METHODS: Nearly 700 microRNAs were analyzed by qRT-PCR. MicroRNAs showing significant expression differences were examined by DIANA-miRpath v.3.0 to identify target genes and pathways. Immunoblots were performed to preliminarily assess the strength of the in silico results. RESULTS: Twenty-one microRNAs (miR-26a-1-3p, miR-30d-3p, miR-27b-5p, miR-146a-5p, miR-362-5p, mir-550a-5p, mir-34a-3p, mir-1227-3p, mir-27a-5p, mir-222-5p, mir-151a-5p, miR-449a, let7c-5p, miR-337-5p, mir-29b-3p, miR-200b-3p, miR-141-3p, miR-671-3p, miR-324-5p, mir-411-3p, and mir-425-3p) were significantly regulated in response to rhNGF. In silico analysis evidenced interesting target genes and pathways, including that of neurotrophin, when analyzed in depth. Almost 80 unique target genes (e.g., PI3K, AKT, MAPK, KRAS, BRAF, RhoA, Cdc42, Rac1, Bax, Bcl2, FasL) were identified as being among those most involved in neurotrophin signaling and in controlling cell proliferation, growth, and apoptosis. AKT and RhoA immunoblots demonstrated congruence with microRNA expression, providing preliminary validation of in silico data. CONCLUSIONS: MicroRNA levels in response to rhNGF were for the first time analyzed in corneal cells. Novel insights about microRNAs, target genes, pathways modulation, and possible biological responses were provided. Importantly, given the putative role of microRNAs as biomarkers or therapeutic targets, our results make available data which might be potentially exploitable for clinical applications.

Cancer secretome and inflammation: The bright and the dark sides of NF-κB.

Tumour promoting inflammation is widely recognized as a hallmark of cancer. The source of this chronic inflammation in cancer has been ascribed to the progressive activation over time of immune cells, mostly of the innate arm of the immune system. However, recent evidence has shown that chronic inflammation may also derive, at least in part, from senescent cells. Hence, due to the prominent role of inflammation in cancer, the cancer secretome definition includes all the secretory factors ensuing from the crosstalk between the cancer cell and the tumour microenvironment. The mechanistic basis underlying the paracrine signalling between the cancer cell and the surrounding tumour microenvironment in malignancy have been widely investigated by using in vivo models of cancers, thus identifying the NF-κB transcription factor as the molecular hub linking inflammation and cancer. In this review, we highlight the roles of NF-κB in regulating the inflammation-derived secretome emanating from immune and senescent cells, with a special focus on the bright and the dark sides of their pro-inflammatory signalling on tumorigenesis.

Next-generation sequencing: recent applications to the analysis of colorectal cancer.

Since the establishment of the Sanger sequencing method, scientists around the world focused their efforts to progress in the field to produce the utmost technology. The introduction of next-generation sequencing (NGS) represents a revolutionary step and promises to lead to massive improvements in our understanding on the role of nucleic acids functions. Cancer research began to use this innovative and highly performing method, and interesting results started to appear in colorectal cancer (CRC) analysis. Several studies produced high-quality data in terms of mutation discovery, especially about actionable or less frequently mutated genes, epigenetics, transcriptomics. Analysis of results is unveiling relevant perspectives aiding to evaluate the response to therapies. Novel evidences have been presented also in other directions such as gut microbiota or CRC circulating tumor cells. However, despite its unquestioned potential, NGS poses some issues calling for additional studies. This review intends to offer a view of the state of the art of NGS applications to CRC through examination of the most important technologies and discussion of recent published results.

Adipose-derived stem cells sustain prolonged angiogenesis through leptin secretion.

Recent studies suggest that adipose-derived stem cells (ASCs) play a role in tissue remodeling through the release of cytokines and growth factors. We compared the secreted cytokine profile of hypoxia-conditioned ASCs (hASCs) with normoxic ASCs (nASCs) and we analyzed the effect of ASCs conditioned medium (CM) on endothelial cells. We found that hypoxia induced a transient upregulation of VEGF in ASCs and a notable and enduring upregulation of leptin mRNA expression 30-fold greater than control after 24 h and up to 60-fold greater than control at day 7. CM from hASC stimulated EC tube formation to a significantly greater extent than CM from nASC. This might be due to leptin-secreted factor. Indeed, exogenous leptin stimulated the expression of HIF2-α, but not HIF1-α, and upregulated the expression of Flt-1 and Tie-1 proangiogenic receptors. In conclusion, hASCs may be particularly efficient in sustaining angiogenesis through the release of leptin.

MicroRNA expression analysis in high fat diet-induced NAFLD-NASH-HCC progression: study on C57BL/6J mice.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common malignant tumor of the liver. Non-alcoholic fatty liver disease (NAFLD) is a frequent chronic liver disorder in developed countries. NAFLD can progress through the more severe non alcoholic steatohepatitis (NASH), cirrhosis and, lastly, HCC. Genetic and epigenetic alterations of coding genes as well as deregulation of microRNAs (miRNAs) activity play a role in HCC development. In this study, the C57BL/6J mouse model was long term high-fat (HF) or low-fat (LF) diet fed, in order to analyze molecular mechanisms responsible for the hepatic damage progression. METHODS: Mice were HF or LF diet fed for different time points, then plasma and hepatic tissues were collected. Histological and clinical chemistry assays were performed to assess the progression of liver disease. MicroRNAs' differential expression was evaluated on pooled RNAs from tissues, and some miRNAs showing dysregulation were further analyzed at the individual level. RESULTS: Cholesterol, low and high density lipoproteins, triglycerides and alanine aminotransferase increase was detected in HF mice. Gross anatomical examination revealed hepatomegaly in HF livers, and histological analysis highlighted different degrees and levels of steatosis, inflammatory infiltrate and fibrosis in HF and LF animals, demonstrating the progression from NAFLD through NASH. Macroscopic nodules, showing typical neoplastic features, were observed in 20% of HF diet fed mice. Fifteen miRNAs differentially expressed in HF with respect to LF hepatic tissues during the progression of liver damage, and in tumors with respect to HF non tumor liver specimens were identified. Among them, miR-340-5p, miR-484, miR-574-3p, miR-720, whose expression was never described in NAFLD, NASH and HCC tissues, and miR-125a-5p and miR-182, which showed early and significant dysregulation in the sequential hepatic damage process. CONCLUSIONS: In this study, fifteen microRNAs which were modulated in hepatic tissues and in tumors during the transition NAFLD-NASH-HCC are reported. Besides some already described, new and early dysregulated miRNAs were identified. Functional analyses are needed to validate the results here obtained, and to better define the role of these molecules in the progression of the hepatic disease.

Bioinformatics approach to predict target genes for dysregulated microRNAs in hepatocellular carcinoma: study on a chemically-induced HCC mouse model.

BACKGROUND: Hepatocellular carcinoma (HCC) is an aggressive epithelial tumor which shows very poor prognosis and high rate of recurrence, representing an urgent problem for public healthcare. MicroRNAs (miRNAs/miRs) are a class of small, non-coding RNAs that attract great attention because of their role in regulation of processes such as cellular growth, proliferation, apoptosis. Because of the thousands of potential interactions between a single miR and target mRNAs, bioinformatics prediction tools are very useful to facilitate the task for individuating and selecting putative target genes. In this study, we present a chemically-induced HCC mouse model to identify differential expression of miRNAs during the progression of the hepatic injury up to HCC onset. In addition, we describe an established bioinformatics approach to highlight putative target genes and protein interaction networks where they are involved. RESULTS: We describe four miRs (miR-125a-5p, miR-27a, miR-182, miR-193b) which showed to be differentially expressed in the chemically-induced HCC mouse model. The miRs were subjected to four of the most used predictions tools and 15 predicted target genes were identified. The expression of one (ANK3) among the 15 predicted targets was further validated by immunoblotting. Then, enrichment annotation analysis was performed revealing significant clusters, including some playing a role in ion transporter activity, regulation of receptor protein serine/threonine kinase signaling pathway, protein import into nucleus, regulation of intracellular protein transport, regulation of cell adhesion, growth factor binding, and regulation of TGF-beta/SMAD signaling pathway. A network construction was created and links between the selected miRs, the predicted targets as well as the possible interactions among them and other proteins were built up. CONCLUSIONS: In this study, we combined miRNA expression analysis, obtained by an in vivo HCC mouse model, with a bioinformatics-based workflow. New genes, pathways and protein interactions, putatively involved in HCC initiation and progression, were identified and explored.

The role of IL-1ß in the bone loss during rheumatic diseases.

Several inflammatory diseases have been associated with increased bone resorption and fracture rates and different studies supported the relation between inflammatory cytokines and osteoclast activity. The main factor required for osteoclast activation is the stimulation by receptor activator of nuclear factor kappa-B ligand (RANKL) expressed on osteoblasts. In this context, interleukin- (IL-) 1ß, one of the most powerful proinflammatory cytokines, is a strong stimulator of in vitro and in vivo bone resorption via upregulation of RANKL that stimulates the osteoclastogenesis. The resulting effects lead to an imbalance in bone metabolism favouring bone resorption and osteoporosis. In this paper, we review the available literature on the role of IL-1ß in the pathogenesis of bone loss. Furthermore, we analysed the role of IL-1ß in bone resorption during rheumatic diseases and, when available, we reported the efficacy of anti-IL-1ß therapy in this field.

NF-κB: Governing Macrophages in Cancer.

Tumor-associated macrophages (TAMs) are the major component of the tumor microenvironment (TME), where they sustain tumor progression and or-tumor immunity. Due to their plasticity, macrophages can exhibit anti- or pro-tumor functions through the expression of different gene sets leading to distinct macrophage phenotypes: M1-like or pro-inflammatory and M2-like or anti-inflammatory. NF-κB transcription factors are central regulators of TAMs in cancers, where they often drive macrophage polarization toward an M2-like phenotype. Therefore, the NF-κB pathway is an attractive therapeutic target for cancer immunotherapy in a wide range of human tumors. Hence, targeting NF-κB pathway in the myeloid compartment is a potential clinical strategy to overcome microenvironment-induced immunosuppression and increase anti-tumor immunity. In this review, we discuss the role of NF-κB as a key driver of macrophage functions in tumors as well as the principal strategies to overcome tumor immunosuppression by targeting the NF-κB pathway.

A method to comprehensively identify germline SNVs, INDELs and CNVs from whole exome sequencing data of BRCA1/2 negative breast cancer patients.

In the rapidly evolving field of genomics, understanding the genetic basis of complex diseases like breast cancer, particularly its familial/hereditary forms, is crucial. Current methods often examine genomic variants-such as Single Nucleotide Variants (SNVs), insertions/deletions (Indels), and Copy Number Variations (CNVs)-separately, lacking an integrated approach. Here, we introduced a robust, flexible methodology for a comprehensive variants' analysis using Whole Exome Sequencing (WES) data. Our approach uniquely combines meticulous validation with an effective variant filtering strategy. By reanalyzing two germline WES datasets from BRCA1/2 negative breast cancer patients, we demonstrated our tool's efficiency and adaptability, uncovering both known and novel variants. This contributed new insights for potential diagnostic, preventive, and therapeutic strategies. Our method stands out for its comprehensive inclusion of key genomic variants in a unified analysis, and its practical resolution of technical challenges, offering a pioneering solution in genomic research. This tool presents a breakthrough in providing detailed insights into the genetic alterations in genomes, with significant implications for understanding and managing hereditary breast cancer.

Reverse-phase protein microarray highlights HER2 signaling activation in immunohistochemistry/FISH/HER2-negative breast cancers.

EVALUATION OF: Wulfkuhle JD, Berg D, Wolff C et al. Molecular analysis of HER2 signaling in human breast cancer by functional protein pathway activation mapping. Clin. Cancer Res. 18(23), 6426-6435 (2012). Exhaustive characterization and mapping of pivotal molecules and downstream effectors deregulated in breast cancer is of fundamental clinical value to define the most effective therapy. Wulfkuhle et al. applied reverse-phase protein microarray, a highly sensitive immunoassay able to perform quantitative and multiplexed analysis of total and/or modified cellular proteins, to assess protein levels and activation/phosphorylation status of the HER family (EGFR, HER2, HER3) and downstream signaling molecules in HER2(+) and HER2(-) breast cancers. The research was performed using laser capture microdissected tumor epithelial cells from frozen samples and formalin-fixed paraffin embedded specimens, which were also analyzed by immunohistochemistry (IHC) and FISH. This study identified a subgroup of IHC/FISH/HER2(-) patients with HER2 activation/phosphorylation levels comparable with those obtained from IHC/FISH/HER2(+) tumors. HER2 signaling activation was independent from total HER2 expression and involved HER3 and EGFR activation. These findings indicate that molecular characterization by reverse-phase protein microarray of HER2 and its partners/effectors in the signaling cascade enables the identification of a subgroup of IHC/FISH/HER2(-) patients showing HER2 signaling activation. These patients, currently excluded from targeted therapy administration, could potentially benefit from this and it could improve prognosis and survival.

Comparative Analysis of Dasatinib Effect between 2D and 3D Tumor Cell Cultures.

Three-dimensional cell culture methods are able to confer new predictive relevance to in vitro tumor models. In particular, the 3D multicellular tumor spheroids model is considered to better resemble tumor complexity associated with drug resistance compared to the 2D monolayer model. Recent advances in 3D printing techniques and suitable biomaterials have offered new promises in developing 3D tissue cultures at increased reproducibility and with high-throughput characteristics. In our study, we compared the sensitivity to dasatinib treatment in two different cancer cell lines, prostate cancer cells DU145 and glioblastoma cells U87, cultured in the 3D spheroids model and in the 3D bioprinting model. DU145 and U87 cells were able to proliferate in 3D alginate/gelatin bioprinted structures for two weeks, forming spheroid aggregates. The treatment with dasatinib demonstrated that bioprinted cells were considerably more resistant to drug toxicity than corresponding cells cultured in monolayer, in a way that was comparable to behavior observed in the 3D spheroids model. Recovery and analysis of cells from 3D bioprinted structures led us to hypothesize that dasatinib resistance was dependent on a scarce penetrance of the drug, a phenomenon commonly reported also in spheroids. In conclusion, the 3D bioprinted model utilizing alginate/gelatin hydrogel was demonstrated to be a suitable model in drug screening when spheroid growth is required, offering advantages in feasibility, reproducibility, and scalability compared to the classical 3D spheroids model.

Intranasal delivery of NGF rescues hearing impairment in aged SAMP8 mice.

Hearing loss impacts the quality of life and affects communication resulting in social isolation and reduced well-being. Despite its impact on society and economy, no therapies for age-related hearing loss are available so far. Loss of mechanosensory hair cells of the cochlea is a common event of hearing loss in humans. Studies performed in birds demonstrating that they can be replaced following the proliferation and transdifferentiation of supporting cells, strongly pointed out on HCs regeneration as the main focus of research aimed at hearing regeneration. Neurotrophins are growth factors involved in neuronal survival, development, differentiation, and plasticity. NGF has been involved in the interplay between auditory receptors and efferent innervation in the cochlea during development. During embryo development, both NGF and its receptors are highly expressed in the inner ears. It has been reported that NGF is implicated in the differentiation of auditory gangliar and hair cells. Thus, it has been proposed that NGF administration can decrease neuronal damage and prevent hearing loss. The main obstacle to the development of hearing impairment therapy is that efficient means of delivery for selected drugs to the cochlea are missing. Herein, in this study NGF was administered by the intranasal route. The first part of the study was focused on a biodistribution study, which showed the effective delivery in the cochlea; while the second part was focused on analyzing the potential therapeutic effect of NGF in senescence-accelerated prone strain 8 mice. Interestingly, intranasal administration of NGF resulted protective in counteracting hearing impairment in SAMP8 mice, ameliorating hearing performances (analyzed by auditory brainstem responses and distortion product otoacoustic emission) and hair cells morphology (analyzed by microscopy analysis). The results obtained were encouraging indicating that the neurotrophin NGF was efficiently delivered to the inner ear and that it was effective in counteracting hearing loss.

Molecular Mechanisms Underpinning Immunometabolic Reprogramming: How the Wind Changes during Cancer Progression.

Metabolism and the immunological state are intimately intertwined, as defense responses are bioenergetically expensive. Metabolic homeostasis is a key requirement for the proper function of immune cell subsets, and the perturbation of the immune-metabolic balance is a recurrent event in many human diseases, including cancer, due to nutrient fluctuation, hypoxia and additional metabolic changes occurring in the tumor microenvironment (TME). Although much remains to be understood in the field of immunometabolism, here, we report the current knowledge on both physiological and cancer-associated metabolic profiles of immune cells, and the main molecular circuits involved in their regulation, highlighting similarities and differences, and emphasizing immune metabolic liabilities that could be exploited in cancer therapy to overcome immune resistance.

Targeting costimulatory molecules to improve antitumor immunity.

The full activation of T cells necessitates the concomitant activation of two signals, the engagement of T-cell receptor by peptide/major histocompatibility complex II and an additional signal delivered by costimulatory molecules. The best characterized costimulatory molecules belong to B7/CD28 and TNF/TNFR families and play crucial roles in the modulation of immune response and improvement of antitumor immunity. Unfortunately, tumors often generate an immunosuppressive microenvironment, where T-cell response is attenuated by the lack of costimulatory molecules on the surface of cancer cells. Thus, targeting costimulatory pathways represent an attractive therapeutic strategy to enhance the antitumor immunity in several human cancers. Here, latest therapeutic approaches targeting costimulatory molecules will be described.

Concurrent RAS and RAS/BRAF V600E Variants in Colorectal Cancer: More Frequent Than Expected? A Case Report.

The assessment of RAS and BRAF mutational status is one of the main steps in the diagnostic and therapeutic algorithm of metastatic colorectal cancer (mCRC). Multiple mutations in the BRAF and RAS pathway are described as a rare event, with concurrent variants in KRAS and BRAF genes observed in approximately 0.05% of mCRC cases. Here, we report data from a case series affected by high-risk stage III and stage IV CRC and tested for RAS and BRAF mutation, treated at our Medical Oncology Unit. The analysis of KRAS, NRAS (codons 12, 13, 59, 61, 117, 146), and BRAF (codon 600) hotspot variants was performed in 161 CRC tumors from August 2018 to September 2021 and revealed three (1.8%) patients showing mutations in both KRAS and BRAF (V600E), including two cases with earlier CRC and one with metastatic disease. We also identified one patient (0.6%) with a mutation in both KRAS and NRAS genes and another one (0.6%) with a double KRAS mutation. Notably, the latter was characterized by aggressive behavior and poor clinical outcome. The mutational status, pathological features, and clinical history of these five CRC cases are described. Overall, this study case series adds evidence to the limited available literature concerning both the epidemiological and clinical aspects of CRC cases characterized by the presence of concurrent RAS/BRAF variants. Future multicentric studies will be required to increase the sample size and provide additional value to results observed so far in order to improve clinical management of this subgroup of CRC patients.

EV-Mediated Chemoresistance in the Tumor Microenvironment: Is NF-κB a Player?

Drug resistance is a major impediment to patient survival and remains the primary cause of unsuccessful cancer therapy. Drug resistance occurs in many tumors and is frequently induced by chemotherapy which triggers a defensive response both in cancerous and cancer-associated cells that constitute the tumor microenvironment (TME). Cell to cell communication within the TME is often mediated by extracellular vesicles (EVs) which carry specific tumor-promoting factors able to activate survival pathways and immune escape mechanisms, thus sustaining tumor progression and therapy resistance. NF-κB has been recognized as a crucial player in this context. NF-κB activation is involved in EVs release and EVs, in turn, can trigger NF-κB pathway activation in specific contexts, based on secreting cytotype and their specific delivered cargo. In this review, we discuss the role of NF-κB/EVs interplay that sustain chemoresistance in the TME by focusing on the molecular mechanisms that underlie inflammation, EVs release, and acquired drug resistance.