Repetitive Bleomycin-Based Electrochemotherapy Improves Antitumor Effectiveness in 3D Tumor Models of Conjunctival Melanoma.

BACKGROUND: Conjunctival melanoma (CM) is associated with a high rate of local recurrence and poor survival rate. Novel therapeutic options are needed to reduce recurrence rate. The objective of the study was to demonstrate the improved effectiveness of electrochemotherapy (ECT) on CM using repetitive application. METHODS: Tumor spheroids of three CM cell lines (CRMM1, CRMM2, CM2005.1) were treated repetitively with ECT using the chemotherapeutic agent bleomycin on days 3, 5, and 7 of culture. Application of bleomycin alone and electroporation alone served as controls. The cytotoxic effect was analyzed on day 10 compared to untreated control using an independent t-test. The spheroid outgrowth rate was measured. RESULT: CM tumor spheroid size (median value: 78%, SD: 32%) and viability (median value: 11%, SD: 11%) were dramatically reduced after repetitive ECT treatment (p-value < 0.001). Decreased proliferation capacity (down to 8%) and an increase of apoptotic cells were observed. In most repetitive ECT-treated spheroids, no viable or proliferating cells were detected. Only 33-40% of repetitive ECT-treated spheroids exhibited single outgrowing cells with a delay of time up to 38 days. CONCLUSION: Repetitive ECT application effectively induces cytotoxic effects in CM spheroids by inducing apoptosis, inhibiting proliferation and decreasing the percentage of surviving tumor cells. Thus, repetitive ECT results in improved antitumor effectiveness in CM and could be an alternative therapy option.

Abandoning node dissection for desmoplasia-negative encapsulated unifocal sporadic medullary thyroid cancer.

BACKGROUND: Predictive criteria to determine the absence of node metastases from thyroid specimens are scarce for sporadic medullary thyroid cancer. METHODS: Histopathologic stratification of patients with unifocal sporadic medullary thyroid cancer ≤25 mm with ≥10 neck nodes at thyroidectomy to evaluate the suitability of desmoplasia (7 increments) and tumor capsule integrity (5 decrements) for intraoperative prediction of node metastasis in unifocal sporadic medullary thyroid cancer. RESULTS: Paraffin-embedded thyroid specimens were available for 139 eligible patients. Significant (P < .001) associations were found between increasing desmoplasia and decreasing tumor capsule integrity and nodal disease (from 0 to 79% and 0 to 62%); the number of node metastases (medians, from 0 to 3 and 0 to 2 nodes); and biochemical cure (from 100 to 36% and 100 to 58%). Desmoplasia (low-moderate to high, with fibrosis >10%) and breach of the tumor capsule (>3 extensions; 1 extension >3 mm in width; or diffuse growth without tumor capsule) yielded excellent sensitivity and negative predictive value (100%), with moderate specificity (57 and 48%) and positive predictive value (50 and 46%). In retrospect, node dissection proved unnecessary in 55 (57%) and 47 (48%) patients who harbored desmoplasia-negative and encapsulated tumors. When available frozen sections were histopathologically compared with matching paraffin-embedded thyroid tumor specimens, concordance was 98% (53 of 54 pairs): 1 of 7 upgrades changed the diagnosis to desmoplasia, whereas 1 of 3 downgrades shifted the diagnosis of tumor capsule breach from "present" to "absent." CONCLUSIONS: Patients with desmoplasia-negative encapsulated sporadic medullary thyroid cancer may forgo node dissection at specialist centers.

Multispectral Fluorescence Imaging Allows for Distinctive Topographic Assessment and Subclassification of Tumor-Infiltrating and Surrounding Immune Cells.

Histomorphology has significantly changed over the last decades due to technological achievements in immunohistochemistry (IHC) for the visualization of specific proteins and in molecular pathology, particularly in the field of in situ hybridization of small oligonucleotides and amplification of DNA and RNA amplicons. With an increased availability of suitable methods, the demands regarding the observer of histomorphological slides were the supply of complex quantitative data as well as more information about protein expression and cell-cell interactions in tissue sections. Advances in fluorescence-based multiplexed IHC techniques, such as multispectral imaging (MSI), allow the quantification of multiple proteins at the same tissue section. In histopathology, it is a well-known technique for over a decade yet harboring serious problems concerning quantitative preciseness and tissue autofluorescence of multicolor staining when using formalin-fixed, paraffin-embedded (FFPE) tissue specimen. In recent years, milestones in tissue preparation, fluorescent dyes, hardware imaging, and software analysis were achieved including automated tissue segmentation (e.g., tumor vs. stroma) as well as in cellular and subcellular multiparameter analysis.This chapter covers the role that MSI plays in anatomic pathology for the analysis of FFPE tissue sections, discusses the technical aspects of MSI, and provides a review of its application in the characterization of immune cell infiltrates and beyond regarding its prognostic and predictive value and its use for guidance of clinical decisions for immunotherapeutic strategies.