Paving the Path for Immune Enhancing Nutrition in Colon Cancer: Modulation of Tumor Microenvironment and Optimization of Outcomes and Costs.

INTRODUCTION: Published evidence suggests that immunonutrition has the potential to decrease postoperative complications and reduce length of stay in patients undergoing surgery for colorectal cancer. However, only a few studies have analyzed the effects of immunonutrition on tumor microenvironment and evaluated its prognostic impact. MATERIAL AND METHODS: This is a single center retrospective study enrolling 50 patients undergoing elective surgery for colorectal cancer managed with immunonutrition and 50 patients managed with standard nutrition for comparison. Tumor microenvironment was analyzed before (on the biopsy at the time of diagnosis) and after (on the matched surgical specimen) administration of immunonutrition. Immune function related indicators, including cytotoxic T-lymphocytes, helper T-cells, antigen presenting cells, natural killer cells, T-exhausted lymphocytes, T-regulatory cells, M1 and M2 tumor associated macrophages and PD-L1 expression were assessed by immunohistochemistry. For both groups, clinicopathological data were collected and a 5-year follow-up was available. RESULTS: We found that immunonutrition significantly activated the T-cell response against cancer, alter tumor microenvironment phenotype towards M2 polarization and inhibits the PD1/PD-L1 axis. A lower rate of postoperative complications and a shorter length of stay (p = 0.04) were observed in the immune nutrition group. Compared to standard nutrition group, patients managed wit immune nutrition showed a higher 5-year overall survival (p = 0.001). Finally, immune nutrition allowed to reduce the hospital care costs. CONCLUSIONS: Immunonutrition modulates tumor microenvironment by improving immune function and could prolong survival in patients undergoing elective surgery for colorectal cancer. Further studies are needed to optimize IN protocols and confirm their prognostic impact.

Spike mutation resilient scFv76 antibody counteracts SARS-CoV-2 lung damage upon aerosol delivery.

The uneven worldwide vaccination coverage against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and emergence of variants escaping immunity call for broadly effective and easily deployable therapeutic agents. We have previously described the human single-chain scFv76 antibody, which recognizes SARS-CoV-2 Alpha, Beta, Gamma and Delta variants. We now show that scFv76 also neutralizes the infectivity and fusogenic activity of the Omicron BA.1 and BA.2 variants. Cryoelectron microscopy (cryo-EM) analysis reveals that scFv76 binds to a well-conserved SARS-CoV-2 spike epitope, providing the structural basis for its broad-spectrum activity. We demonstrate that nebulized scFv76 has therapeutic efficacy in a severe hACE2 transgenic mouse model of coronavirus disease 2019 (COVID-19) pneumonia, as shown by body weight and pulmonary viral load data. Counteraction of infection correlates with inhibition of lung inflammation, as observed by histopathology and expression of inflammatory cytokines and chemokines. Biomarkers of pulmonary endothelial damage were also significantly reduced in scFv76-treated mice. The results support use of nebulized scFv76 for COVID-19 induced by any SARS-CoV-2 variants that have emerged so far.

Immunohistochemical Markers of the Epithelial-to-Mesenchymal Transition (EMT) Are Related to Extensive Lymph Nodal Spread, Peritoneal Dissemination, and Poor Prognosis in the Microsatellite-Stable Diffuse Histotype of Gastric Cancer.

Background: Although the prognostic value of the epithelial-to-mesenchymal transition (EMT) in gastric cancer has been reported in several studies, the strong association with the diffuse type may represent a confounding factor. Our aim is to investigate potential correlations among EMT status, tumor advancement, and prognosis in diffuse gastric cancer. Methods: Between 1997 and 2012, 84 patients with microsatellite-stable (MSS) diffuse-type tumors underwent surgery. The EMT phenotype was assessed with the E-cadherin, CD44, and zinc finger E-box binding homeobox 1 (ZEB-1) immunohistochemical markers. Results: Forty-five out of 84 cases (54%) were EMT-positive; more advanced nodal status (p = 0.010), pTNM stage (p = 0.032), and vascular invasion (p = 0.037) were observed in this group. The median numbers of positive nodes (13 vs. 5) and involved nodal stations (4 vs. 2) were higher in the EMT-positive group. The cancer-related survival time was 26 months in EMT-positive cases vs. 51 in negative cases, with five-year survival rates of 17% vs. 51%, respectively (p = 0.001). The EMT status had an impact on the prognosis of patients with <70 years, R0 resections, or treatment with adjuvant chemotherapy. Tumor relapses after surgery and peritoneal spread were significantly higher in the EMT-positive tumors. Conclusions: EMT status, when assessed through immunohistochemistry, identified an aggressive phenotype of MSS diffuse-type tumors with extensive lymph nodal spread, peritoneal dissemination, and worse long-term outcomes.

A look towards the clonal origin of metastatic pulmonary carcinosarcoma: Report of a patient with an unexpected long-term survival.

INTRODUCTION: Pulmonary carcinosarcoma is a rare histological subtype of non-small cell lung cancer, defined by the combination of epithelial and mesenchimal elements. Prognosis is usually dismal, with a median survival of about 6 months. The use of immunotherapy by blockade of PD1/PD-L1 immune checkpoint signaling has been shown to improve patients' survival. However, local aggressiveness and distant metastases are frequent. Spread to the gastrointestinal tract is seldom reported. The genetic landscape of the disease has only recently begun to emerge, pointing at TP53, KRAS, EGFR and MET as the most common mutated genes. CASE DESCRIPTION: We describe the case of a metastatic patient with 37 months overall survival, treated by an aggressive multimodal approach combining surgery, chemotherapy, radiotherapy and immunotherapy. To shed new light on the molecular basis for sarcomatoid component in lung carcinoma, we performed next generation sequencing analysis of the squamous and sarcomatoid component by the two sites. We demonstrated a clonal origin and hypermutability of the sarcomatous elements that may account for the good response to immunotherapy. Moreover, we identified some mutations involving TP53 and EGFR genes, targetable by already available drugs. CONCLUSIONS: We depicted a model of how a squamous cell carcinoma can differentiate during its natural history into sub-clonal populations with different features and may ultimately result in a neoplasm (i.e. pulmonary carcinosarcoma) showing clonal heterogeneity. Our data might contribute to a better understanding of the pathogenesis and molecular mechanisms of this rare tumor and open new ways for a more tailored approach.

Preoperative oral immunonutrition in gastrointestinal surgical patients: How the tumour microenvironment can be modified.

BACKGROUND & AIMS: This study is focused on the impact of enteral immunonutrition on the cell-mediated immune response in the microenvironment of gastric and colorectal cancers. METHODS: This is a prospective pilot study approved by the local Ethics Committee. The immunophenotypic structure of the immune cells before (on the biopsy) and after (on the surgical sample) the administration of the immunonutrition in 16 patients is compared with 8 patients receiving regular diet. The samples of non-tumour tissue from sleeve-gastrectomy are used as non-neoplastic control. Antibodies were tested: CD4, CD8, PD-1, FOX-P3, CD68, CD163, CD80, CD21, CD56, PD-L1. We applied already well-known scoring systems for the evaluation of the immunohistochemistry and compared our data in the different groups by statistical analysis. RESULTS: In treated patients, we detected a modulation of the immune response with higher number of cytotoxic and helper T-lymphocytes in the tumour microenvironment of the surgical specimens compared to the pre-operative biopsy, and a lower number of lymphocytes presenting an exhausted (i.e. double positive CD8 and PD-1 lymphocytes) and regulatory (i.e. double positive CD4 and FOX-P3 lymphocytes) phenotype. Moreover we observed the M1 polarization with a lower number of CD163 positive macrophages and the inhibition of the PD-1/PD-L1 pathway in treated patients. CONCLUSIONS: The immunonutrition impacts on the tumoral microenvironment of gastric and colorectal cancer activating the inflammatory pathway, in terms of humoral and cellular response.

Occult malignancy underlying a case of "hyperacute" onset of severe pulmonary hypertension.

Investigations in a patient with new-onset pulmonary hypertension should include screening for undiagnosed malignancy http://bit.ly/2mrLmGM.

Correction: Evaluation of tenascin-C by tenatumomab in T-cell non-Hodgkin lymphomas identifies a new target for radioimmunotherapy.

[This corrects the article DOI: 10.18632/oncotarget.23919.].

Evaluation of tenascin-C by tenatumomab in T-cell non-Hodgkin lymphomas identifies a new target for radioimmunotherapy.

The clinical outcome of T-cell non-Hodgkin lymphoma (NHL) is poor and innovative treatments are needed. Tenascin-C is a large extracellular glycoprotein not expressed under physiological conditions, but overexpressed in cancer. Aim of the study was to evaluate tenascin-C expression within pathologic tissue of T-cell NHL and determine its clinical significance. We used an immunohistochemistry approach using the anti-tenascin-C monoclonal antibody Tenatumomab in 75 systemic T-cell NHL (including 72 mature and 3 precursor T-cell NHL), and 25 primary cutaneous T-cell NHL. Data were analyzed in terms of staining intensity, proportion of involved areas and histologic pattern, and results were correlated with clinical characteristics and outcome. Ninety-three percent of the cases were tenascin-C positive and 59% of systemic diseases were characterized by a predominant involvement (>50%). Stromal expression was detected in all the cases while vascular and vascular plus cytoplasmic expression was present in 49% and 23%. The constant overexpression of the tenascin-C gene was observed in two independent publicly available T-cell NHL gene expression datasets. In conclusions, tenascin-C represents an attractive target that sets the rationale to investigate the therapeutic activity of radiolabeled Tenatumomab in T-cell NHL.

Carnitine palmitoyl transferase-1A (CPT1A): a new tumor specific target in human breast cancer.

Transcriptional mechanisms epigenetically-regulated in tumoral tissues point out new targets for anti-cancer therapies. Carnitine palmitoyl transferase I (CPT1) is the rate-limiting enzyme in the transport of long-chain fatty acids for ß-oxidation. Here we identified the tumor specific nuclear CPT1A as a product of the transcript variant 2, that doesn't retain the classical transferase activity and is strongly involved in the epigenetic regulation of cancer pro-survival, cell death escaping and tumor invasion pathways. The knockdown of CPT1A variant 2 by small interfering RNAs (siRNAs), was sufficient to induce apoptosis in MCF-7, SK-BR3 and MDA-MB-231 breast cancer cells. The cell death triggered by CPT1A silencing correlated with reduction of HDAC activity and histone hyperacetylation. Docking experiments and molecular dynamics simulations confirmed an high binding affinity of the variant 2 for HDAC1. The CPT1A silenced cells showed an up-regulated transcription of pro-apoptotic genes (BAD, CASP9, COL18A1) and down-modulation of invasion and metastasis related-genes (TIMP-1, PDGF-A, SERPINB2). These findings provide evidence of the CPT1 variant 2 involvement in breast cancer survival, cell death escape and invasion. Thus, we propose nuclear CPT1A as a striking tumor specific target for anticancer therapeutics, more selective and effective as compared with the well-known HDAC inhibitors.

Intra-tumor AvidinOX allows efficacy of low dose systemic biotinylated Cetuximab in a model of head and neck cancer.

For locally advanced and metastatic head and neck squamous cell carcinoma (HNSCC), the current clinical use of Cetuximab in chemo/radiotherapy protocols is often associated to severe systemic toxicity. Here we report in vitro data in human FaDu pharynx SCC cells, showing that inactive concentrations of biotinylated Cetuximab (bCet) become active upon anchorage to AvidinOX on the surface of tumor cells. AvidinOX-anchored bCet induces apoptosis and DNA damage as well as specific inhibition of signaling, degradation and abrogation of nuclear translocation of EGFR. In the mouse model of FaDu cancer, we show that intra-tumor injection of AvidinOX allows anti-tumor activity of an otherwise inactive, intraperitoneally delivered, low dose bCet. Consistently with in vitro data, in vivo tumor inhibition is associated to induction of apoptosis, DNA damage and reduced angiogenesis. AvidinOX is under clinical investigation for delivering radioactive biotin to inoperable tumors (ClinicalTrials.gov NCT02053324) and present data support its use for the local treatment of HNSCC in combination with systemic administration of low dose bCet.

[Corrigendum] Trastuzumab and docetaxel in a preclinical organotypic breast cancer model using tissue slices from mammary fat pad: Translational relevance.

Oncol Rep 34: [Related article:] 1146­1152, 2015; DOI: 10.3892/or.2015.4074 After the publication of the article, the authors decided to add an Acknowledgements section: Acknowledgments Research activity leading to the results shown in the paper is the continuation of the IMI Predect program that received support from the Innovative Medicines initiative Joint Undertaking under grant agreement no. 115188, resources of which are composed of financial contribution from the European Union's Seventh Framework Programme (FP7/2007­2013) and EFPIA companies in kind contribution. Present results and conclusions are not endorsed by the Predect Consortium.

Trastuzumab and docetaxel in a preclinical organotypic breast cancer model using tissue slices from mammary fat pad: Translational relevance.

With the ever-increasing number of drugs approved to treat cancers, selection of the optimal treatment regimen for an individual patient is challenging. Breast cancer complexity requires novel predictive methods and tools. In the present study, we set up experimental conditions to obtain an 'ex vivo' organotypic culture from xenotransplanted mice aiming at recapitulating the human clinical condition. The effect of trastuzumab (large biological molecule) and docetaxel (small chemical entity) was subsequently investigated on this organotypic model and compared with in vivo and in vitro activity on tumor cells. Tissue slices of 200 µm were obtained from mammary fat pad of SCID mice xenotransplanted with human MCF-7 breast cancer cells. Viability and proliferation were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) colorimetric assay and Ki-67 immunohistochemistry,and apoptosis by cleaved caspase-3 immunohistochemistry. In vivo antitumor activity of trastuzumab and docetaxel was determined by caliper measurement of tumor volume and Ki-67 expression on explanted masses by immunohistochemistry. A Teflon support and normoxia were necessary experimental conditions to obtain high viability of excised breast cancer infiltrated mammary fat pad slices upon 48 h cultivation, as shown by MTT proliferation assay, and Ki-67 expression. Breast cancer tissue slices treated for 48 h with trastuzumab or docetaxel showed a significant dose­dependent reduction of viability by MTT assay. Consistently, both drugs down-modulated Ki-67 and increased cleaved caspase-3. Tumor masses collected from docetaxel- or trastuzumab­treated mice showed a similar reduction of proliferation markers. By contrast, MCF-7 cell cultures were significantly inhibited by docetaxel but not by trastuzumab. Tumor tissue slices represent a more predictive experimental cancer model compared to cell cultures for both small and large molecule antitumor efficacy. This observation supports the relevance of microenvironment in the overall tumor biology and response to therapeutics.

Radionuclide Therapy of Unresectable Tumors with AvidinOX and (90)Y-biotinDOTA: Tongue Cancer Paradigm.

Local treatment of unresectable tumors is challenging, particularly with radioactivity. Current practice relies on external beam irradiation or on a variety of medical devices for brachytherapy. Both approaches proved useful in controlling tumor growth, but are characterized by poor compliance of the patient, significant side-effects, high costs, and technological complexity, which hamper widespread use. The authors recently described a novel form of radionuclide therapy based on the oxidized form of avidin that, chemically reacting with tissue proteins, can secure radioactive biotin within the injected tissue, either when precomplexed or when taken from the blood stream after intravenous administration. AvidinOX-pretargeted (177)Lu-biotinDOTA ((177)Lu-ST2210) is currently under clinical investigation for the treatment of liver oligometastases from colorectal cancer (clinicaltrials.gov/NCT02053324). In the present work, the authors show that injected AvidinOX can link tissues of various natures such as prostate, kidney, breast, or brain and can react by contact with scraped tissues such as skin or urinary bladder. AvidinOX injected into human OSC19 tongue cancer masses orthotopically transplanted in nude mice takes up intravenously administered (90)Y-ST2210, which exerts significant antitumor activity, while preserving the integrity and functionality of the tongue. Present data confirm that AvidinOX-based radionuclide therapy is an innovative and promising approach for the local treatment of inoperable tumors.

Microcalcifications in breast cancer: an active phenomenon mediated by epithelial cells with mesenchymal characteristics.

BACKGROUND: Mammary microcalcifications have a crucial role in breast cancer detection, but the processes that induce their formation are unknown. Moreover, recent studies have described the occurrence of the epithelial-mesenchymal transition (EMT) in breast cancer, but its role is not defined. In this study, we hypothesized that epithelial cells acquire mesenchymal characteristics and become capable of producing breast microcalcifications. METHODS: Breast sample biopsies with microcalcifications underwent energy dispersive X-ray microanalysis to better define the elemental composition of the microcalcifications. Breast sample biopsies without microcalcifications were used as controls. The ultrastructural phenotype of breast cells near to calcium deposits was also investigated to verify EMT in relation to breast microcalcifications. The mesenchymal phenotype and tissue mineralization were studied by immunostaining for vimentin, BMP-2, ß2-microglobulin, ß-catenin and osteopontin (OPN). RESULTS: The complex formation of calcium hydroxyapatite was strictly associated with malignant lesions whereas calcium-oxalate is mainly reported in benign lesions. Notably, for the first time, we observed the presence of magnesium-substituted hydroxyapatite, which was frequently noted in breast cancer but never found in benign lesions. Morphological studies demonstrated that epithelial cells with mesenchymal characteristics were significantly increased in infiltrating carcinomas with microcalcifications and in cells with ultrastructural features typical of osteoblasts close to microcalcifications. These data were strengthened by the rate of cells expressing molecules typically involved during physiological mineralization (i.e. BMP-2, OPN) that discriminated infiltrating carcinomas with microcalcifications from those without microcalcifications. CONCLUSIONS: We found significant differences in the elemental composition of calcifications between benign and malignant lesions. Observations of cell phenotype led us to hypothesize that under specific stimuli, mammary cells, which despite retaining a minimal epithelial phenotype (confirmed by cytokeratin expression), may acquire some mesenchymal characteristics transforming themselves into cells with an osteoblast-like phenotype, and are able to contribute to the production of breast microcalcifications.

Transglutaminase 2 as a biomarker of osteoarthritis: an update.

Osteoarthritis is a progressive joint disease characterized by cartilage degradation and bone remodelling. Under physiologic conditions, articular cartilage displays a stable chondrocyte phenotype, whereas in osteoarthritis a chondrocyte hypertrophy develops near the sites of cartilage surface damage and associates to the pathologic expression of type X collagen. Transglutaminases (TGs) include a family of Ca(2+)-dependent enzymes that catalyze the formation of γ-glutamyl cross-links. Their substrates include a variety of intracellular and extracellular macromolecular components. TGs are ubiquitously and abundantly expressed and implicated in a variety of physiopathological processes. TGs activity is modulated by inflammatory cytokines. TG2 (also known as tissue transglutaminase) mediates the hypertrophic differentiation of joint chondrocytes and interleukin-1-induced calcification. Histomorphometrical and biomolecular investigations document increased TG2 expression in human and experimental osteoarthritis. Consequently, the level of TG2 expression may represent an adjuvant additional marker to monitor tissue remodelling occurring in osteoarthritic joint tissue. Experimental induction of osteoarthritis in TG2 knockout mice is followed from reduced cartilage destruction and increased osteophyte formation compared to wild-type mice, suggesting a different influence on joint bone and cartilage remodelling. The capacity of transamidation by TG2 to regulate activation of latent TGF-ß seems to have a potential impact on the regulation of inflammatory response in osteoarthritic tissues. Additional studies are needed to define TG2-regulated pathways that are differently modulated in osteoblasts and chondrocytes during osteoarthritis.

Platelet-rich plasma greatly potentiates insulin-induced adipogenic differentiation of human adipose-derived stem cells through a serine/threonine kinase Akt-dependent mechanism and promotes clinical fat graft maintenance.

The potential plasticity and therapeutic utility in tissue regeneration of human adipose-derived stem cells (ASCs) isolated from adult adipose tissue have recently been highlighted. The use of autologous platelet-rich plasma (PRP) represents an alternative strategy in regenerative medicine for the local release of multiple endogenous growth factors. Here we investigated the signaling pathways and effects of PRP and human recombinant insulin on proliferation and adipogenic differentiation of ASCs in vitro. PRP stimulated proliferation (EC(50) = 15.3 ± 1.3% vol/vol), whereas insulin's effect was the opposite (IC(50) = 3.0 ± 0.5 µM). Although PRP alone did not increase adipogenesis, in association with insulin it prevented ASC proliferative arrest, greatly enhanced intracytoplasmic lipid accumulation, strongly increased serine/threonine kinase Akt phosphorylation and mouse monoclonal anti-sterol regulatory element binding protein-1 accumulation, and downregulated Erk-1 activity; adipogenic effects were markedly prevented by the Akt inhibitor wortmannin. PRP with insulin synergistically upregulated fibroblast growth factor receptor (FGFR) and downregulated epidermal growth factor receptor (ErbB) expression; moreover, PRP in association prevented insulin-induced insulin-like growth factor-1 receptor and insulin receptor downregulation. The inhibition of FGFR-1, epidermal growth factor receptor (EGFR), and epidermal growth factor receptor-2 (ErbB2) activity reduced ASC proliferation, but only that of FGFR-1 reduced adipogenesis and Akt phosphorylation, whereas the ErbB2 inhibition effects were the opposite. However, EGFR activity was needed for ErbB2-mediated inhibition of ASC adipogenesis. Clinically, the injection of insulin further ameliorated patients' 1-year PRP-induced fat graft volume maintenance and contour restoring. Our results ascertain that PRP in association with insulin greatly potentiates adipogenesis in human ASCs through a FGFR-1 and ErbB2-regulated Akt mechanism. The ameliorated clinical fat graft maintenance suggests additional useful translational applications of combined PRP-insulin treatment in regenerative medicine.

miR-24 triggers epidermal differentiation by controlling actin adhesion and cell migration.

During keratinocyte differentiation and stratification, cells undergo extensive remodeling of their actin cytoskeleton, which is important to control cell mobility and to coordinate and stabilize adhesive structures necessary for functional epithelia. Limited knowledge exists on how the actin cytoskeleton is remodeled in epithelial stratification and whether cell shape is a key determinant to trigger terminal differentiation. In this paper, using human keratinocytes and mouse epidermis as models, we implicate miR-24 in actin adhesion dynamics and demonstrate that miR-24 directly controls actin cable formation and cell mobility. miR-24 overexpression in proliferating cells was sufficient to trigger keratinocyte differentiation both in vitro and in vivo and directly repressed cytoskeletal modulators (PAK4, Tks5, and ArhGAP19). Silencing of these targets recapitulated the effects of miR-24 overexpression. Our results uncover a new regulatory pathway involving a differentiation-promoting microribonucleic acid that regulates actin adhesion dynamics in human and mouse epidermis.