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Microvascular inflammation (MVI), defined as the presence of glomerulitis and/or peritubular capillaritis, is the key histological lesion of anti-HLA donor-specific antibodies (DSA)-related antibody mediated rejection, but recently other possible mechanisms of MVI have emerged. However, except for peritubular capillary C4d deposition that is more frequently observed in the presence of anti-HLA-DSA, histological features are similar regardless of MVI origin. Therefore, accurately describing patterns of MVI may help differentiate etiologies and drive therapeutic choices. We describe the case of a kidney transplant recipient (primary nephropathy: autosomal dominant polycystic kidney disease) who underwent kidney biopsy for worsening renal function and new onset hypertension. Histologic findings showed severe microvascular inflammation with intense glomerulitis and presence of intracapillary multinucleated cells, positive on immunostaining for endothelial marker ETS-related gene (ERG). Focal intense peritubular capillaritis and early glomerular basement membrane reduplication, C4d negative, were observed, consistent with early chronic active ABMR. HLA-DSA were absent, but high level of anti-angiotensin II type-1 receptor (AT1R) antibodies (Ab) were detected (78 U/L, normal levels < 10 U/L). Two subsequent biopsies showed intense microvascular inflammation with diffuse peritubular capillaritis, and multinucleated, ERG-positive, endothelial cells were still seen in glomerular capillary loops. The patient was started on angiotensin receptor blockers (ARBs) and plasma exchange (PEX) sessions obtaining normalization of blood pressure and AT1R Ab and proteinuria reduction, but, after subsequent liver transplant, rituximab therapy failed to maintain remission and the patient remained PEX-dependent., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

We propose a pilot, prospective, translational study with the aim of identifying possible molecular markers underlying metastatic prostate cancer (PC) evolution with the use of liquid biopsy. Twenty-eight castrate sensitive, oligometastatic PC patients undergoing bone and/or nodal stereotactic body radiotherapy (SBRT) were recruited. Peripheral blood samples were collected before the commencement of SBRT, then they were processed for circulating cell free DNA (cfDNA) extraction. Deep targeted sequencing was performed using a custom gene panel. The primary endpoint was to identify differences in the molecular contribution between the oligometastatic and polymetastatic evolution of PC to same-first oligo-recurrent disease presentation. Seventy-seven mutations were detected in 25/28 cfDNA samples: ATM in 14 (50%) cases, BRCA2 11 (39%), BRCA1 6 (21%), AR 13 (46%), ETV4, and ETV6 2 (7%). SBRT failure was associated with an increased risk of harboring the BRCA1 mutation (OR 10.5) ( p = 0.043). The median cfDNA concentration was 24.02 ng/mL for ATM mutation carriers vs. 40.04 ng/mL for non-carriers ( p = 0.039). Real-time molecular characterization of oligometastatic PC may allow for the identification of a true oligometastatic phenotype, with a stable disease over a long time being more likely to benefit from local, curative treatments or the achievement of long-term disease control. A prospective validation of our promising findings is desirable for a better understanding of the real impact of liquid biopsy in detecting tumor aggressiveness and clonal evolution.

We consider simple methods to improve the growth nowcasts and forecasts obtained by mixed-frequency MIDAS and UMIDAS models with a variety of indicators during the Covid-19 crisis and recovery period, such as combining forecasts across various specifications for the same model and/or across different models, extending the model specification by adding MA terms, enhancing the estimation method by taking a similarity approach, and adjusting the forecasts to put them back on track using a specific form of intercept correction. Among these methods, adjusting the original nowcasts and forecasts by an amount similar to the nowcast and forecast errors made during the financial crisis and subsequent recovery seems to produce the best results for the US, notwithstanding the different source and characteristics of the financial crisis. In particular, the adjusted growth nowcasts for 2020Q1 get closer to the actual value, and the adjusted forecasts based on alternative indicators become much more similar, all unfortunately indicating a much slower recovery than without adjustment, and very persistent negative effects on trend growth. Similar findings also emerge for forecasts by institutions, for survey forecasts, and for the other G7 countries., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2020 International Institute of Forecasters. Published by Elsevier B.V. All rights reserved.)

Detection of BRAF V600E within cell free tumor DNA (ctDNA) is emerging as a promising means to improve patients' stratification or enable BRAF inhibitor (BRAFi) therapeutic monitoring in a minimally invasive manner. Here, we investigated whether extracellular vesicle-(EV)-associated-DNA (EV-DNA) has value as an alternative source of circulating BRAF V600E . To do so, we identified a clinical practice-compatible protocol for the isolation of EV-DNA and assessed BRAF gene status on plasma samples from metastatic melanoma patients at the beginning and during BRAFi therapy. This protocol uses a peptide with high affinity for EVs and it has been found to recover more mutant DNA from plasma than standard ultracentrifugation. Molecular analyses revealed that mutant DNA is largely unprotected from nuclease digestion, interacting with the outer side of the EV membrane or directly with the peptide. When used on clinical samples, we found that the protocol improves the detection of BRAF V600E gene copies in comparison to the reference protocol for ctDNA isolation. Taken together, these findings indicate that EVs are a promising source of mutant DNA and should be considered for the development of next-generation liquid biopsy approaches.

We evaluated the advantages and the reliability of novel protocols for the enrichment of tumor extracellular vesicles (EVs), enabling a blood-based test for the noninvasive parallel profiling of multiple androgen receptor ( AR) gene alterations. Three clinically relevant AR variants related to response/resistance to standard-of-care treatments ( AR -V7 transcript, AR T878A point mutation and AR gene amplification) were evaluated by digital PCR in 15 samples from patients affected by Castration-Resistant Prostate Cancer (CRPC). Plasma was processed to obtain circulating RNA and DNA using protocols based on tumor EVs enrichment through immuno-affinity and peptide-affinity compared to generic extraction kits. Our results showed that immuno-affinity enrichment prior to RNA extraction clearly outperforms the generic isolation method in the detection of AR -V7, also allowing for a distinction between responder (R) and non-responder (NR) patients. The T878A mutation was detected, overall, in nine out of 15 samples and no approach alone was able to reveal mutations in all harboring samples, showing that the employed methods complement each other. AR amplification was detected in the majority of CRPC samples analysed using either cell-free DNA (cfDNA) or exosome isolation kits (80%). We demonstrated that selective isolation of a subset of circulating exosomes enriched for tumor origin, rather than analysis of total plasma exosomes, or total plasma nucleic acids, increases sensitivity and specificity for the detection of specific alterations.

Due to the discovery of their role in intra‑cellular communications, exosomes, which carry information specific to the cell of origin, have garnered considerable attention in cancer research. Moreover, there is evidence to suggest the possibility of isolating different exosome sub‑populations based on target antigens at the cell surface. Philadelphia chromosome‑positive (Ph+) chronic myeloid leukemia (CML) is a clonal myeloproliferative neoplasia characterized by the breakpoint cluster region‑proto‑oncogene 1 tyrosine‑protein kinase (BCR‑ABL1) fusion‑gene, derived from the t (9;22) translocation. Tyrosine kinase inhibitors (TKIs) target BCR‑ABL1 protein and induce major or deep molecular responses in the majority of patients. Despite the fact that several studies have demonstrated the persistence of leukemic cells in the bone marrow niche, even following treatment, TKIs prolong patient survival time and facilitate treatment‑free remission. These characteristics render CML a plausible model for investigating the feasibility of tumor‑derived exosome fraction enrichment. In the present study, patients in the chronic phase (CP) of CML were treated with TKIs, and the quantification of the BCR‑ABL1 exosomal transcript was performed using digital PCR (dPCR). The possibility of tumor‑derived exosomes enrichment was confirmed, and for the first time, to the best of our knowledge, the detection of the BCR‑ABL1 transcript highlighted the presence of active leukemic cells in patients with CP‑CML. According to these findings, tumor‑derived exosomes may be considered a novel tool for the identification of active leukemic cells, and for the assessment of innovative monitoring focused on the biological functions of exosomes in CML.

Background: Circulating tumor cells have been described in prostate cancer patients at diagnosis and in the metastatic phase but little is known on their role at biochemical PSA recurrence., Patients and Methods: Patients radically cured with either prostatectomy or radiotherapy were sequentially included at PSA recurrence. The presence of CTCs was evaluated by the CellSearch system., Results: Twenty-nine patients were accrued at PSA recurrence. Median PSA at recurrence was 7.2 ng/ml (range=3.86-51.0 ng/ml). The median time to PSA progression was 4.66 years (range=0.1-16 years). CTCs were detected in one patient (3%) with low numbers (1 CTC/7.5 ml)., Conclusion: In patients radically cured for prostate cancer at biochemical recurrence, CTCs are detected at very low levels in a minority of patients. Further studies are required to investigate alternative methods of CTC detection and the possible role of the bone marrow pre-metastatic niche at biochemical recurrence., (Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Purpose: To assess whether the combination of letrozole, metronomic cyclophosphamide and sorafenib (LCS) is well tolerated and shows activity in primary breast cancer (BC)., Methods: Thirteen oestrogen receptor-positive, postmenopausal, T2-4, N0-1 BC patients received the LCS combination for 6 months. In these patients we examined the pharmacokinetics of sorafenib and cyclophosphamide, toxicity of the regimen, the clinical response to therapy and changes in the levels of biologically relevant biomarkers., Results: Adequate plasma concentrations of sorafenib were achieved in patients when it was dosed in combination with L+C. The mean plasma concentrations of C were consistently lower following administration of LCS, compared with administration of L+C only. The most common drug-related grade 3/4 adverse events were skin rash (69.3%), hand-foot skin reaction (69.3%) and diarrhoea (46.1%). According to RECIST Criteria, a clinical complete response was observed in 6 of 13 patients. A significant reduction in tumour size, evaluated with MRI, was also observed between baseline and 14 days of treatment in all 13 patients (P=0.005). A significant reduction in SUV uptake, measured by (18)FDG-PET/CT, was observed in all patients between baseline and 30 days of treatment (P=0.015) and between baseline and definitive surgery (P=0.0002). Using modified CT Criteria, a response was demonstrated in 8 out of 10 evaluable patients at 30 days and in 11 out of 13 evaluable patients at the definitive surgery. A significant reduction in Ki67 expression was observed in all patients at day 14 compared with baseline (P<0.00001) and in 9 out of 13 patients at the definitive surgery compared with baseline (P<0.03). There was also a significant suppression of CD31 and VEGF-A expression in response to treatment (P=0.01 and P=0.007, respectively)., Conclusions: The LCS combination is feasible and tolerable. The tumour response and target biomarker modulation indicate that the combination is clinically and biologically active.

The study investigated the anti-tumour effect of zoledronic acid (ZA) administered alone in a biological window therapy in naïve bone-only metastatic and locally advanced breast cancer (LABC) patients. 33 patients with LABC (Group 1) and 20 patients with a first diagnosis of bone metastasis only (Group 2) received 4 mg single dose of ZA, 14 days (biological window) before starting any treatment. In Group 1, Ki67, CD34, p53/bcl-2 and caspase 3 expression along with the adenosine triphosphate (ATP) levels and RNA disruption index were evaluated as markers of tumor growth in tumour specimens obtained before and after ZA administration (basal, day 14). In Group 2, the total enumeration of circulating tumour cells (CTCs), and of M30+ve CTCs along with the soluble marker of cell death (M30/M65) were carried-out as markers of tumor dissemination at baseline, at 48 h and day 14th. In Group 1, there was a significant reduction in Ki67, CD34, bcl-2 expression after 14 days ZA based-treatment (p = 0.0032; p = 0.0001, p < 0.00001 respectively). ZA showed a significant increase of RNA disruption (p < 0.0076). In Group 2, we observed a significant reduction of CTCs number after 48 h (p = 0.0012), followed by a significant rebound at 14 days (p = 0.012). The apoptotic CTCs/M30+ve and M65 levels significantly increased under treatment (p = 0.018 and p = 0.039 respectively) after drug administration when compared to the baseline. These results are the first prospective in vivo data showing the direct pure anti-tumour effect (either on the tumour cell or on CTCs) of ZA.

DNA repair is a key determinant in the cellular response to therapy and tumor repair status could play an important role in tailoring patient therapy. Our goal was to evaluate the mRNA of 13 genes involved in different DNA repair pathways (base excision, nucleotide excision, homologous recombination, and Fanconi anemia) in paraffin embedded samples of triple negative breast cancer (TNBC) compared to luminal A breast cancer (LABC). Most of the genes involved in nucleotide excision repair and Fanconi Anemia pathways, and CHK1 gene were significantly less expressed in TNBC than in LABC. PARP1 levels were higher in TNBC than in LABC. In univariate analysis high level of FANCA correlated with an increased overall survival and event free survival in TNBC; however multivariate analyses using Cox regression did not confirm FANCA as independent prognostic factor. These data support the evidence that TNBCs compared to LABCs harbour DNA repair defects.

Epithelial-mesenchymal transition (EMT) is defined by the loss of epithelial characteristics and the acquisition of a mesenchymal phenotype. In this process, cells acquire molecular alterations that facilitate dysfunctional cell-cell adhesive interactions and junctions. These processes may promote cancer cell progression and invasion into the surrounding microenvironment. Such transformation has implications in progression of breast carcinoma to metastasis, and increasing evidences support most tumors contain a subpopulation of cells with stem-like and mesenchymal features that is resistant to chemotherapy. This review focuses on the physiological and pathological role of EMT process, its molecular related network, its putative role in the metastatic process and its implications in response/resistance to the current and/or new approaching drugs in the clinical management of breast cancer., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Breast cancer is a heterogeneous disease. Predictive molecular markers are crucial in patient management, but the only recommended predictive biomarkers are estrogen and progesterone receptors and HER2. There are many new targeted therapies, and although the target pathway expression is readily analyzed on conventional pathology, the dynamic response cannot be assessed and pathway expression is no guarantee it has a major driver role, even if mutated. Selecting therapies requires considering the patient, the molecular characteristics of the tumor, and the microenvironment of the tumor. Thus, the integration of molecular pathology, imaging, and early tumor biological response to therapy may provide evidence of drug activity and allow more rapid changes of therapy. The adaptive response of the tumor is a key resistance mechanism that can be assessed readily in the neoadjuvant setting. Although there are no markers that meet all surrogacy criteria, their use could provide crucial information on mechanisms of drug sensitivity/resistance. Validation of such markers requires a major emphasis on neoadjuvant trials to relate early-biomarker response to outcome.

Neural stem cells (NSCs) in the murine subventricular zone (SVZ) niche allow life-long neurogenesis. During the first postnatal month and throughout aging, the decrease of neuroblasts and the rise of astrocytes results in diminished neurogenesis and increased astrocyte:neuron ratio. Also, a different neurogenic activity characterizes the SVZ periventricular region (LV, lateral ventricle) as compared to its rostral extension (RE). In order to investigate whether and to what extent these physiological modifications may be ascribed to intrinsic changes of the endogenous NSC/progenitor features, we performed a functional analysis on NSCs isolated and cultured from LV and RE tissues at distinct postnatal stages that are marked by striking modifications to the SVZ niche in vivo. We evaluated the effect of age and brain region on long-term proliferation and multipotency, and characterized the cell type composition of NSC-derived progeny, comparing this make-up to that of region- and age-matched primary neural cultures. Furthermore, we analyzed the effect of prolonged in vitro expansion on NSC functional properties. We documented age- and region-dependent differences on the clonogenic efficiency and on the long-term proliferative capacity of NSCs. Also, we found age- and region-dependent quantitative changes in the cell composition of NSC progeny (decreased quantity of neurons and oligodendrocytes; increased amount of astroglial cells) and these differences were maintained in long-term cultured NSC populations. Overall, these data strengthen the hypothesis that age- and region-dependent differences in neurogenesis (observed in vivo) may be ascribed to the changes in the intrinsic developmental program of the NSC populations.

Recent observations have suggested that extensive culturing of adult neural stem cells (ANSCs) by exploiting the NeuroSphere assay might select for aggressive cell clones, endowed with neoplastic potential, that overgrow the rest of the native stem cells. However, a detailed study of the propensity of ANSCs to transform has never been thoroughly undertaken. Here, we report the first demonstration that ANSCs can be propagated in vitro for over a year, maintaining a strikingly stable profile with regard to self-renewal, differentiation, growth factor dependence, karyotype, and molecular profiling. Most importantly, the long-term culturing of ANSCs did not result in the formation of tumors in vivo, even when ANSCs were transduced with Myc and Ras oncogenes. The cancer resistance could depend on specific mechanisms aimed at protecting ANSCs and preserved by optimal nonstressful culture conditions. In conclusion, besides a plentiful and safe source of cells for therapeutic applications, ANSCs provide an ideal model to study aging and cancer in the context of stemness.

Reliable procedures to induce neural commitment of totipotent undifferentiated embryonic stem (ES) cells have provided new tools for investigating the molecular mechanisms underlying cell fate choices. We extensively characterized the developmental potential of ES-induced neural cells obtained using an adaptation of the multistep induction protocol. We provided evidence that ES-derived neural proliferating cells are endowed with stem cell properties such as extensive self-renewal capacity and single-cell multipotency. In differentiating conditions, cells matured exclusively into neurons, astrocytes, and oligodendrocytes. All these features have been previously described in only somatic neural stem cells (NSCs). Therefore, we consider it more appropriate to rename our cells ES-derived NSCs. These similarities between the two NSC populations induced us to carefully compare their proliferation ability and differentiation potential. Although they were very similar in overall behavior, we scored specific differences. For instance, ES-derived NSCs proliferated at higher rate and consistently generated a higher number of neurons compared with somatic NSCs. To further investigate their relationships, we carried out a molecular analysis comparing their transcriptional profiles during proliferation. We observed a large fraction of shared expressed transcripts, including genes previously described to be critical in defining somatic NSC traits. Among the genes differently expressed, candidate genes possibly responsible for divergences between the two cell types were selected and further investigated. In particular, we showed that an enhanced MAPK (mitogen-activated protein kinase) signaling is acting in ES-induced NSCs, probably triggered by insulin-like growth factor-II. This may contribute to the high proliferation rate exhibited by these cells in culture.

To understand global features of gene expression changes during in vitro neural differentiation, we carried out the microarray analysis of embryonic stem cells (ESCs), embryonal carcinoma cells, and adult neural stem/progenitor (NS) cells. Expression profiling of ESCs during differentiation in monolayer culture revealed three distinct phases: undifferentiated ESCs, primitive ectoderm-like cells, and neural progenitor cells. Principal component (PC) analysis revealed that these cells were aligned on PC1 over the course of 6 days. This PC1 represents approximately 4,000 genes, the expression of which increased with neural commitment/differentiation. Furthermore, NS cells derived from adult brain and their differentiated cells were positioned along this PC axis further away from undifferentiated ESCs than embryonic stem-derived neural progenitors. We suggest that this PC1 defines a path to neural fate, providing a scale for the degree of commitment/differentiation.

Transformed stem cells have been isolated from some human cancers. We report that, unlike other brain cancers, the lethal glioblastoma multiforme contains neural precursors endowed with all of the critical features expected from neural stem cells. Similar, yet not identical, to their normal neural stem cell counterpart, these precursors emerge as unipotent (astroglial) in vivo and multipotent (neuronal-astroglial-oligodendroglial) in culture. More importantly, these cells can act as tumor-founding cells down to the clonal level and can establish tumors that closely resemble the main histologic, cytologic, and architectural features of the human disease, even when challenged through serial transplantation. Thus, cells possessing all of the characteristics expected from tumor neural stem cells seem to be involved in the growth and recurrence of adult human glioblastomas multiforme.