Investigating the Mechanobiology of Cancer Cell-ECM Interaction: The Impact of Substrate Stiffness in Breast Cancer Progression

The loss of tissue homeostasis and mechanoreciprocity are considered one of the hallmarks of cancer. Here we have applied a porous type I collagen-based three-dimensional (3D) scaffold to study how breast cancer cells interact with and alter extracellular collagen. We assessed the modifications induced by tumor cells on the micro- and macro- characteristic of extracellular collagen and on its compressive stiffness. Mechanical testing was conducted both with an in-house built low-force compressive device and by Dynamic Mechanical Analysis. The stiffness of single collagen fibers was assessed by Atomic Force Microscopy. When cultured on collagen scaffolds, the two cell lines generated coherent tissue-like structures. MCF7 displayed an epithelial morphology with a tightly cohesive cobblestone appearance, while MDA-MB-231 showed a mesenchymal phenotype with lower cell-to-cell contact. MDA-MB-231, which belongs to the aggressive basal-like subtype, increased scaffold stiffness from 46.9 kPa, to 57.9 kPa, and overexpressed the matrix-modifying enzyme, lysyl oxidase (LOX), whereas luminal A MCF-7 cells did not alter the mechanical characteristics of extracellular collagen. When the activity of LOX was blocked, MDA-MB-231 were unable to alter the scaffold stiffness: the compressive modulus increased by 8.9%, in contrast to the increase observed without LOX inhibition (23%). No significant changes were found between the Young’s modulus of fibers taken from control scaffolds compared to fibers taken from scaffolds after culture with MDA-MB-231. Overall this work provides evidence that invasive, mesenchymal-like breast cancer cells produce high levels of the crosslinking enzyme LOX, and are able to increase the stiffness of extracellular collagen and to alter its structural characteristics. A causal relationship between this behavior of cancer cells and expression of the enzyme LOX was also provided. Our model offers a relevant in vitro tool to reproduce and investigate the biomechanical interplay subsisting between cancer cells and the surrounding ECM

The emerging role of cancer nanotechnology in the panorama of sarcoma

In the field of nanomedicine a multitude of nanovectors have been developed for cancer application. In this regard, a less exploited target is represented by connective tissue. Sarcoma lesions encompass a wide range of rare entities of mesenchymal origin affecting connective tissues. The extraordinary diversity and rarity of these mesenchymal tumors is reflected in their classification, grading and management which are still challenging. Although they include more than 70 histologic subtypes, the first line-treatment for advanced and metastatic sarcoma has remained unchanged in the last fifty years, excluding specific histotypes in which targeted therapy has emerged. The role of chemotherapy has not been completely elucidated and the outcomes are still very limited. At the beginning of the century, nano-sized particles clinically approved for other solid lesions were tested in these neoplasms but the results were anecdotal and the clinical benefit was not substantial. Recently, a new nanosystem formulation NBTXR3 for the treatment of sarcoma has landed in a phase 2-3 trial. The preliminary results are encouraging and could open new avenues for research in nanotechnology. This review provides an update on the recent advancements in the field of nanomedicine for sarcoma. In this regard, preclinical evidence especially focusing on the development of smart materials and drug delivery systems will be summarized. Moreover, the sarcoma patient management exploiting nanotechnology products will be summed up. Finally, an overlook on future perspectives will be provided

When Electrospun Fiber Support Matters: In Vitro Ovine Long-Term Folliculogenesis on Poly (Epsilon Caprolactone) (PCL)-Patterned Fibers

Current assisted reproduction technologies (ART) are insufficient to cover the slice of the population needing to restore fertility, as well as to amplify the reproductive performance of domestic animals or endangered species. The design of dedicated reproductive scaffolds has opened the possibility to better recapitulate the reproductive 3D ovarian environment, thus potentially innovating in vitro folliculogenesis (ivF) techniques. To this aim, the present research has been designed to compare ovine preantral follicles in vitro culture on poly(epsilon-caprolactone) (PCL)-based electrospun scaffolds designed with different topology (Random vs. Patterned fibers) with a previously validated system. The ivF performances were assessed after 14 days under 3D-oil, Two-Step (7 days in 3D-oil and on scaffold), or One-Step PCL protocols (14 days on PCL-scaffold) by assessing morphological and functional outcomes. The results show that Two- and One-Step PCL ivF protocols, when performed on patterned scaffolds, were both able to support follicle growth, antrum formation, and the upregulation of follicle marker genes leading to a greater oocyte meiotic competence than in the 3D-oil system. In conclusion, the One-Step approach could be proposed as a practical and valid strategy to support a synergic follicle-oocyte in vitro development, providing an innovative tool to enhance the availability of matured gametes on an individual basis for ART purposes

Virtual Surgical Planning, 3D-Printing and Customized Bone Allograft for Acute Correction of Severe Genu Varum in Children

Complex deformities of lower limbs are frequent in children with genetic or metabolic skeletal disorders. Early correction is frequently required, but it is technically difficult and burdened by complications and recurrence. Herein, we described the case of a 7-year-old girl affected by severe bilateral genu varum due to spondyloepiphyseal dysplasia. The patient was treated by patient-specific osteotomies and customized structural wedge allograft using Virtual Surgical Planning (VSP) and 3D-printed patient-specific instrumentation (PSI). The entire process was performed through an in-hospital 3D-printing Point-of-Care (POC). VSP and 3D-printing applied to pediatric orthopedic surgery may allow personalization of corrective osteotomies and customization of structural allografts by using low-cost in-hospital POC. However, optimal and definitive alignment is rarely achieved in such severe deformities in growing skeleton through a single operation

Prognostic and Predictive Role of Body Composition in Metastatic Neuroendocrine Tumor Patients Treated with Everolimus: A Real-World Data Analysis

Neuroendocrine tumors (NETs) are rare neoplasms frequently characterized by an up- regulation of the mammalian rapamycin targeting (mTOR) pathway resulting in uncontrolled cell proliferation. The mTOR pathway is also involved in skeletal muscle protein synthesis and in adipose tissue metabolism. Everolimus inhibits the mTOR pathway, resulting in blockade of cell growth and tumor progression. The aim of this study is to investigate the role of body composition in- dexes in patients with metastatic NETs treated with everolimus. The study population included 30 patients with well-differentiated (G1-G2), metastatic NETs treated with everolimus at the IRCCS Romagnolo Institute for the Study of Tumors (IRST) “Dino Amadori”, Meldola (FC), Italy. The body composition indexes (skeletal muscle index [SMI] and adipose tissue indexes) were assessed by measuring on a computed tomography (CT) scan the cross-sectional area at L3 at baseline and at the first radiological assessment after the start of treatment. The body mass index (BMI) was assessed at baseline. The median progression-free survival (PFS) was 8.9 months (95% confidence interval [CI]: 3.4–13.7 months). The PFS stratified by tertiles was 3.2 months (95% CI: 0.9–10.1 months) in patients with low SMI (tertile 1), 14.2 months (95% CI: 2.3 months-not estimable [NE]) in patients with intermediate SMI (tertile 2), and 9.1 months (95% CI: 2.7 months-NE) in patients with high SMI (tertile 3) (p = 0.039). Similarly, the other body composition indexes also showed a statistically significant difference in the three groups on the basis of tertiles. The median PFS was 3.2 months (95% CI: 0.9–6.7 months) in underweight patients (BMI 18.49 kg/m2) and 10.1 months (95% CI: 3.7–28.4 months) in normal-weight patients (p = 0.011). There were no significant differences in terms of overall survival. The study showed a correlation between PFS and the body composition indexes in patients with NETs treated with everolimus, underlining the role of adipose and muscle tissue in these patients

Metastatic neuroendocrine neoplasia treatments in patients over 70 years of age

The incidence of neuroendocrine neoplasia (NEN) is higher in individuals ≥70 years of age (elderly) who are underrepresented in clinical trials because of comorbidities and low performance status. We retrospectively analyzed the outcome of elderly patients with metastatic NEN (mNEN). Comorbidities were summarized by Charlson Comorbidity Index (CCI), Kaplan–Meier method was applied to estimate overall survival (OS) and Cox’s proportional hazard model was used to assess the impact of known prognostic factors. We retrieved data on 145 mNEN patients aged ≥70 years seen at our center from June 2007 to March 2016. Fifty-six (38.6%) were aged ≥75 years. ECOG PS was 0 in 45.7% of cases and CCI was 0 in 41.0% and 1 in 37.4%. A total of 75.4% of patients had grade (G)1/G2 NEN and 24.6%, G3. Octreoscan/Gallium PET/CT and FDG-PET/CT were positive in 94.2% and 70.3% of cases, respectively. Median follow-up was 72.3 (53.2–85.1) months. Seventy-nine patients received first-line somatostatin analogs (SSA), 23 peptide receptor radionuclide therapy (PRRT) and 36 chemotherapy (CHT). Seven did not undergo first-line therapy and 102 received more than one line. Median overall survival (mOS) was 5.1 years (95% CI: 3.4–6.6). No differences in mOS were seen according to CCI. First-line PRRT patients had a mOS of 6.5 years (95% CI: 3.3–not reached (NR)), SSA 5.7 years (95% CI: 4.2–7) and CHT 5.9 years (95% CI: 0.4–NR). mOS in CHT-treated G3 patients was 1.5 years (1.0–2.5). ECOG PS and FDG PET/CT were identified as independent prognostic factors. Results suggest that the above treatments positively impacted OS in elderly mNEN patients, including those aged ≥75 years

HRAS overexpression predicts response to Lenvatinib treatment in gastroenteropancreatic neuroendocrine tumors

IntroductionNeuroendocrine neoplasms (NENs) are a rare group of tumors exceptionally heterogeneous, with clinical presentation ranging from well differentiated more indolent tumors to poorly differentiated very aggressive forms. Both are often diagnosed after the metastatic spread and require appropriate medical treatment. A high priority need in the management of this disease is the identification of effective therapeutic strategies for advanced and metastatic patients. The recent TALENT trial demonstrated the efficacy of lenvatinib, a multi-tyrosine kinase inhibitor, in patients with gastroenteropancreatic neuroendocrine tumors (GEP-NETs) with no other treatment indication. Further development of this drug in advanced NETs is warranted.MethodsWe investigated potential clinical and molecular determinants of lenvatinib response in human primary cultures derived from patients with GEP-NET of different grades and sites of origin. We correlated response to treatment with patient clinical characteristics, with the mutational status of 161-cancer associated genes and with the expression levels of MKI-related genes.ResultsLenvatinib exerted a significant antitumor activity in primary GEP-NET cells, with median survival inhibitions similar or higher than those of standard frontline treatments. Of the 11 primary cultures analyzed in our case series, 6 were classified as responder showing a significant survival inhibition, and 5 as non-responder. We observed that the overexpression of HRAS in the original tumor tissue compared to the matched healthy tissue significantly correlated with responsiveness of primary cells to lenvatinib (p=.048). All 5 non-responder cultures showed normal HRAS expression, while of the 6 responder cultures, 4 had HRAS overexpression. Overexpression of HRAS was not associated with gene mutation. None of the other evaluated clinical variables (grade, Ki67, site of origin and syndromic disease) or molecular markers correlated with response.DiscussionLenvatinib appears to be a highly effective drug for the treatment of NETs. The evaluation of HRAS expression in the tumor tissue might improve patient selection and optimize therapeutic outcome

310 A new proposal for the clinical classification of vulvar lichen sclerosus: an observational prospective study

Introduction/Background Vulvar Lichen Sclerosus (VLS) is a chronic inflammatory disorder which commonly affects the female anogenital epithelium, leading to scarring, anatomical distortions, impaired sexual function, decreased quality of life and increased vulvar cancer risk. An agreement to measure VLS severity in a standard way is yet to be defined and, to our knowledge, no standardized clinical classification of anatomical modifications in VLS has been validated. The purpose of this study was to prepare a clinical classification for VLS aimed at defining the morphological patterns of this condition, while stratifying them into grades. The classification is intended to provide a homogeneous and reproducible description of the different features of this disease. It also serves as an important tool for the evaluation of the course of the disease over time, response to treatment, and for comparison of clinical studies. Methodology A board of seven specialists with expertise in vulvar pathology were asked to outline the anatomical criteria for the definition of VLS severity (phimosis of the clitoris, resorption of the labia minora, involvement of the inter-labial sulcus, and narrowing of the vulvar introitus), identifying five grades to be used to build-up of a score model. The classification was validated by 13 physicians upon pictures of 137 consecutive patients. Each physician individually assigned a grade to each case, according to the abovementioned criteria. Inter-rater agreement among evaluators was analysed by means of ICC (Intraclass Correlation Coefficient). Intra-observer reproducibility and inter-observer concordance in vivo were analysed by means of Kappa index. Results This study provides a new classification of VLS, based on defined anatomical criteria and graded into mutually exclusive progressive classes (table 1). The ICC analysis showed a substantial agreement in the attribution of the grade of VLS among the 137 cases, ICC=0,89 (0.87–0.91), both in the expert and in the non-expert group (ICC=0.92 and 0.87 respectively). An 'almost perfect' agreement was achieved for intra-observer reproducibility and among physicians in vivo (Kappa 0.93). Conclusion Our classification showed a high accuracy in defining morphological modifications in VLS. It is easy to use, reproducible, and can be applied by different health care providers in daily clinical practice and in all clinical settings. Disclosure Nothing to disclose