Melanoma 2.0. Skin cancer as a paradigm for emerging diagnostic technologies, computational modelling and artificial intelligence.

We live in an unprecedented time in oncology. We have accumulated samples and cases in cohorts larger and more complex than ever before. New technologies are available for quantifying solid or liquid samples at the molecular level. At the same time, we are now equipped with the computational power necessary to handle this enormous amount of quantitative data. Computational models are widely used helping us to substantiate and interpret data. Under the label of systems and precision medicine, we are putting all these developments together to improve and personalize the therapy of cancer. In this review, we use melanoma as a paradigm to present the successful application of these technologies but also to discuss possible future developments in patient care linked to them. Melanoma is a paradigmatic case for disruptive improvements in therapies, with a considerable number of metastatic melanoma patients benefiting from novel therapies. Nevertheless, a large proportion of patients does not respond to therapy or suffers from adverse events. Melanoma is an ideal case study to deploy advanced technologies not only due to the medical need but also to some intrinsic features of melanoma as a disease and the skin as an organ. From the perspective of data acquisition, the skin is the ideal organ due to its accessibility and suitability for many kinds of advanced imaging techniques. We put special emphasis on the necessity of computational strategies to integrate multiple sources of quantitative data describing the tumour at different scales and levels.

Standardized Computer-Assisted Analysis of PRAME Immunoreactivity in Dysplastic Nevi and Superficial Spreading Melanomas.

PRAME (PReferentially expressed Antigen in MElanoma) is a cancer testis antigen that is frequently expressed in melanoma compared to benign melanocytic proliferations and nevi. However, the interpretation of the intensity and distribution of PRAME immunostaining is not standardized a lot, which makes interpretation difficult. PRAME-stained histological slides of superficial spreading melanomas (SSM) and dysplastic nevi (DN) were digitized and analyzed using the digital pathology and image platform QuPath. t-tests and ROC AUCs were performed with SPSS. A p-value of <0.05 was used for statistical significance, and a ROC AUC score of >0.8 was considered a good result. A cut-off score was defined in an evaluation cohort and subsequently analyzed in an independent validation cohort. In total, 81 PRAME-stained specimens were included. The evaluation cohort included 32 (50%) SSM and 32 (50%) DN, and the mean of PRAME-positive cells/mm2 for the entire lesion was 455.3 (SD 428.2) in SSM and 60.5 (SD 130.1; p < 0.001) in DN. The ROC AUC of PRAME-positive cells of the entire lesion was 0.866, and in the epidermis it was 0.901. The defined cut-off score to distinguish between DN and SSM was 97.67 cells/mm2. In the validation cohort, 16 out of 17 cases (94.1%) were correctly classified by the cut-off score. The computer-aided assessment of PRAME immunostaining is a useful tool in dermatopathology to distinguish between DN and SSM. Lesions with a moderate expression and indifferent morphologic features will remain a challenge for dermatopathologists.

Standardized Computer-Assisted Analysis of 5-hmC Immunoreactivity in Dysplastic Nevi and Superficial Spreading Melanomas.

5-Hydroxymethylcytosine (5-hmC) is an important intermediate of DNA demethylation. Hypomethylation of DNA is frequent in cancer, resulting in deregulation of 5-hmC levels in melanoma. However, the interpretation of the intensity and distribution of 5-hmC immunoreactivity is not very standardized, which makes its interpretation difficult. In this study, 5-hmC-stained histological slides of superficial spreading melanomas (SSM) and dysplastic compound nevi (DN) were digitized and analyzed using the digital pathology and image platform QuPath. Receiver operating characteristic/area under the curve (ROCAUC) and t-tests were performed. A p-value of <0.05 was used for statistical significance, and a ROCAUC score of >0.8 was considered a "good" result. In total, 92 5-hmC-stained specimens were analyzed, including 42 SSM (45.7%) and 50 DN (54.3%). The mean of 5-hmC-positive cells/mm2 for the epidermis and dermo-epidermal junction and the entire lesion differed significantly between DN and SSM (p = 0.002 and p = 0.006, respectively) and showed a trend towards higher immunoreactivity in the dermal component (p = 0.069). The ROCAUC of 5-hmC-positive cells of the epidermis and dermo-epidermal junction was 0.79, for the dermis 0.74, and for the entire lesion 0.76. These results show that the assessment of the epidermal with junctional expression of 5-hmC is slightly superior to dermal immunoreactivity in distinguishing between DN and SSM.

One Website to Gather them All: Usability Testing of the New German SKin Cancer INFOrmation (SKINFO) Website-A Mixed-methods Approach.

Skin cancer patients increasingly search the internet to acquire disease-related information. However, information on the internet may be misleading. Recently, SKINFO has been launched, a website exclusively created for German-speaking skin cancer patients providing information as well as additional resources of verified quality. Here, we describe the results of the first usability test of SKINFO using a mixed-methods approach. Ten adult patients with skin cancer were recruited for usability testing in the skin cancer units of the University Hospitals of Erlangen and Dresden, Germany. Testing consisted of three different scenarios where patients were asked to find specific information on the SKINFO website guided by the think-aloud method. Descriptive analysis and content analyses were performed. All patients would recommend SKINFO and appreciated its content, design, and structure. Think-aloud analysis identified the topics layout, navigation, and content and structure which would benefit from refinement. Major criticism included the navigation through the website, and the desire for more specific information addressing patients' relatives and the latest, up-to-date information. Overall, usability testing showed that the unique web-based information platform has the potential to support patients coping with skin cancer and thus strengthen informed decision-making.

S1-guideline cutaneous and subcutaneous leiomyosarcoma.

Superficial leiomyosarcomas (LMS) are rare skin cancers (2-3% of cutaneous sarcomas) that originate from dermally located hair follicle muscles, dartos or areolar muscles (cutaneous/dermal LMS), or from vascular muscle cells of the subcutaneous adipose tissue (subcutaneous LMS). These superficial LMS are distinct from LMS of the deep soft tissues. Leiomyosarcomas are typically localized at the lower extremities, trunk or capillitium, and present as painful, erythematous to brownish nodules. Diagnosis is made by histopathology. The treatment of choice for primary LMS is complete (R0) microscopically controlled excision, with safety margins of 1 cm in dermal LMS, and 2 cm in subcutaneous LMS, if possible. Non-resectable or metastatic LMS require individual treatment decisions. After R0 resection with 1 cm safety margins, the local recurrence rate of dermal LMS is very low, and metastasis is very rare. Subcutaneous LMS, very large, or incompletely excised LMS recur and metastasize more frequently. For this reason, clinical follow-up examinations are recommended every six months for cutaneous LMS, and every three months for subcutaneous LMS within the first two years (in subcutaneous LMS including locoregional lymph node sonography). Imaging such as CT/MRI is indicated only in primary tumors with special features, recurrences, or already metastasized tumors.

Checkpoint-inhibitor induced Polyserositis with Edema.

BACKGROUND: As immune checkpoint inhibitors (ICI) are increasingly being used due to effectiveness in various tumor entities, rare side effects occur more frequently. Pericardial effusion has been reported in patients with advanced non-small cell lung cancer (NSCLC) after or under treatment with immune checkpoint inhibitors. However, knowledge about serositis and edemas induced by checkpoint inhibitors in other tumor entities is scarce. METHODS AND RESULTS: Four cases with sudden onset of checkpoint inhibitor induced serositis (irSerositis) are presented including one patient with metastatic cervical cancer, two with metastatic melanoma and one with non-small cell lung cancer (NSCLC). In all cases treatment with steroids was successful in the beginning, but did not lead to complete recovery of the patients. All patients required multiple punctures. Three of the patients presented with additional peripheral edema; in one patient only the lower extremities were affected, whereas the entire body, even face and eyelids were involved in the other patients. In all patients serositis was accompanied by other immune-related adverse events (irAEs). CONCLUSION: ICI-induced serositis and effusions are complex to diagnose and treat and might be underdiagnosed. For differentiation from malignant serositis pathology of the punctured fluid can be helpful (lymphocytes vs. malignant cells). Identifying irSerositis as early as possible is essential since steroids can improve symptoms.

Clinical determinants of long-term survival in metastatic uveal melanoma.

This study aimed to identify prognostic factors in patients with metastatic uveal melanoma (UM) that were associated with long-term survival in a real-world setting. A total of 94 patients with metastatic UM were included from German skin cancer centers and the German national skin cancer registry (ADOReg). Data were analyzed for the response to treatment, progression-free survival, and overall survival (OS). Prognostic factors were explored with univariate Cox regression, log-rank, and χ2-tests. Identified factors were subsequently validated after the population was divided into two cohorts of short-term survival (< 2 years OS, cohort A, n = 50) and long-term survival (> 2 years OS, cohort B, n = 44). A poor ECOG performance status (hazard ratio [HR] 2.0, 95% confidence interval [CI] 1.0-3.9) and elevated serum LDH (HR 2.0, 95% CI 1.0-3.8) were associated with a poor OS, whereas a good response to immune checkpoint blockade (ICB, p < 0.001), radiation therapy (p < 0.001), or liver-directed treatments (p = 0.01) were associated with a prolonged OS. Long-term survivors (cohort B) showed a higher median number of organs affected by metastasis (p < 0.001), while patients with liver metastases only were more common in cohort A (40% vs. 9%; p = 0.002). A partial response to ICB was observed in 16% (12/73), being 21% (8/38) for combined ICB, 17% (1/6) for single CTLA4 inhibition, and 10% (3/29) for single PD1 inhibition. One complete response occurred in cohort B with combined ICB. We conclude that the response to ICB and the presence of extrahepatic disease were favorable prognostic factors for long-term survival.

Epidermotropie von Immunzellen unterscheidet Pyoderma gangraenosum vom Ulcus cruris venosum.

HINTERGRUND UND ZIELE: Pyoderma gangraenosum ist eine ulzerierende, autoinflammatorische Erkrankung. Es gibt keine eindeutigen histopathologischen Merkmale zur Differenzierung von anderen Ursachen chronischer Wunden wie dem Ulcus cruris venosum. Ziel dieser Studie war es, histopathologische Merkmale von Pyoderma gangraenosum und Unterschiede zu venösen Ulzerationen zu detektieren. PATIENTEN UND METHODIK: Acht Gewebeproben von Pyoderma gangraenosum, zwölf Proben von Ulcus cruris venosum und sechs Proben von gesunder Haut wurden einer immunhistologischen Multi-Antigen-Analyse unterzogen. Das Immuninfiltrat und seine räumliche Verteilung wurden anhand von Fluoreszenzbildern mit einer Gewebezytometriesoftware analysiert. ERGEBNISSE: Die dichte epidermale Präsenz von CD45RO+ -T-Gedächtnis-Zellen und die Rarefizierung von CD1a+ -Langerhans-Zellen in der Epidermis waren Marker für Pyoderma gangraenosum, welche auch auf eine epidermale Immunreaktion schließen lassen. Darüber hinaus konnte dermal eine hohe Anzahl CD11c+ CD68+ pro-inflammatorischer M1-Makrophagen nachgewiesen werden. Diese überstieg die Anzahl der in venösen Ulzerationen beobachteten Makrophagen deutlich. SCHLUSSFOLGERUNGEN: Die histopathologischen Unterschiede zwischen Pyoderma gangraenosum und Ulcus cruris venosum können zur Unterscheidung der beiden Erkrankungen herangezogen werden und somit eine wichtige Hilfe zur schnellen Einleitung einer adäquaten Therapie sein. Darüber hinaus deuten unsere Daten auf einen antigengesteuerten Prozess in der Epidermis hin, möglicherweise unter Beteiligung von CD1a+ Langerhans-Zellen.

Epidermotropism of inflammatory cells differentiates pyoderma gangrenosum from venous leg ulcers.

BACKGROUND AND OBJECTIVES: Pyoderma gangrenosum is an ulcerative autoinflammatory disease, lacking distinct histopathological characteristics to differentiate from other ulcerating conditions, like venous leg ulcers. The objective of this study was therefore to find histopathological characteristics of pyoderma gangrenosum in a head-to-head comparison to venous leg ulcers. PATIENTS AND METHODS: Eight tissue samples of pyoderma gangrenosum, twelve samples of venous leg ulcers and six samples of healthy skin were stained using an immunohistological multi antigen staining technology. The immune infiltrate and its spatial distribution were analyzed with contextual tissue cytometry software using fluorescence images. RESULTS: The dense epidermal presence of CD45RO+ memory T cells and the rarefication of CD1a+ Langerhans cells in the epidermis were defining markers for pyoderma gangrenosum, implicating an epidermal immune reaction. In addition, high numbers of CD11c+ CD68+ pro-inflammatory M1 macrophages were detected in the dermis, significantly extending the numbers seen in venous leg ulcers. CONCLUSIONS: The histopathological differences found between pyoderma gangrenosum and venous leg ulcer can be used to distinguish between the two diseases and thus provide an important aid for the rapid initiation of adequate therapy. In addition, our data hint at an antigen-driven process in the epidermis, possibly involving CD1a+ Langerhans cells.

S1-guideline atypical fibroxanthoma (AFX) and pleomorphic dermal sarcoma (PDS).

Atypical fibroxanthoma (AFX) and pleomorphic dermal sarcoma (PDS) are rare cutaneous neoplasms representing histomorphological, genetic as well as epigenetic variants of a disease spectrum. Both tumors typically manifest as nonspecific, often ulcerated, skin- to flesh-colored nodules in chronically sun-damaged skin of elderly male patients. AFX is a rather well demarcated, often rapidly growing tumor. PDS tumors are poorly circumscribed and are characterized by aggressive infiltrative growth. Fast as well as slow growth behavior has been reported for both tumors. Histologically, both are composed of spindle-shaped and epithelioid tumor cells with pleomorphic nuclei as well as atypical multinucleated giant cells. Atypical mitoses are common. In contrast to AFX, PDS involves relevant parts of the subcutis and shows areas of tumor necrosis and/or perineural infiltration. Due to the poorly differentiated nature of AFX/PDS (Grade 3), histopathologically similar cutaneous sarcomas, undifferentiated carcinomas, melanomas and other diseases have to be excluded by immunohistochemical analysis. The treatment of choice is micrographically controlled surgery. In cases of AFX, a cure can be assumed after complete excision. Local recurrence rates are low as long as PDS tumors are surgically removed with a safety margin of 2 cm. Metastasis is rare and mostly associated with very thick or incompletely excised tumors; it mainly affects the skin and lymph nodes. Distant metastasis is even more rare. No approved and effective systemic therapy has been established.

"I Feel I'm in Best Hands with You!": A Survey of Patient Satisfac-tion in a German University Skin Cancer Centre.

An important measure of hospital quality is the satisfaction of patients. The aim of this cross-sectional study, performed in the dermato-oncology unit of the university hospital in Erlangen, Germany, was to assess skin cancer patients' degree of satisfaction with healthcare services. Self-administered questionnaires on patient satisfaction regarding contact with staff, need for information, and recommendation of the skin cancer centre were distributed in the day-care unit and the outpatient department to patients between April and June 2017. Results were reported descriptively and subgroup differences were explored using the Mann-Whitney U test, binary logistic regression, or χ2 test. Overall, 496 of 571 questionnaires were returned (86.9%). The median of all satisfaction items ranged between 1 (very good) and 2 (good). The majority of patients wanted more detailed information about skin cancer (46.7%, 142/304). Long waiting times were often criticized (22.8%; 80/351). Particular attention in addressing specific needs and fears may further increase patient satisfaction.

The Role of Immune Checkpoint Blockade in Uveal Melanoma.

Uveal melanoma (UM) represents the most common intraocular malignancy in adults and accounts for about 5% of all melanomas. Primary disease can be effectively controlled by several local therapy options, but UM has a high potential for metastatic spread, especially to the liver. Despite its clinical and genetic heterogeneity, therapy of metastatic UM has largely been adopted from cutaneous melanoma (CM) with discouraging results until now. The introduction of antibodies targeting CTLA-4 and PD-1 for immune checkpoint blockade (ICB) has revolutionized the field of cancer therapy and has achieved pioneering results in metastatic CM. Thus, expectations were high that patients with metastatic UM would also benefit from these new therapy options. This review provides a comprehensive and up-to-date overview on the role of ICB in UM. We give a summary of UM biology, its clinical features, and how it differs from CM. The results of several studies that have been investigating ICB in metastatic UM are presented. We discuss possible reasons for the lack of efficacy of ICB in UM compared to CM, highlight the pitfalls of ICB in this cancer entity, and explain why other immune-modulating therapies could still be an option for future UM therapies.

Senescence markers: Predictive for response to checkpoint inhibitors.

Recent studies suggest that the age-related remodeling of the immune system, known as immunosenescence, could impact the efficacy of immune checkpoint inhibitors in leukemia or nonsmall cell lung cancer. We investigated whether senescence markers can predict response to checkpoint inhibitor therapy in melanoma patients. The peripheral blood of patients with newly diagnosed, untreated metastatic melanoma was analyzed by flow cytometry to correlate the frequency of senescence markers with clinical response as measured by RECIST after 12 weeks of treatment with immune checkpoint inhibitors. The loss of surface markers CD27 and CD28 or the expression of Tim-3 and CD57 on T cells was associated with resistance to checkpoint inhibitor blockade, presenting these phenotypes as possible predictive biomarkers for checkpoint inhibitor therapy. Immunosenescence seems to impact on the response to checkpoint inhibitor therapy in melanoma patients. Thus, lymphocyte phenotyping for senescence markers, with the introduction of immunosenescence panels, could be predictive for checkpoint inhibitor response.

Automated Good Manufacturing Practice-compliant generation of human monocyte-derived dendritic cells from a complete apheresis product using a hollow-fiber bioreactor system overcomes a major hurdle in the manufacture of dendritic cells for cancer vaccines.

BACKGROUND: Although dendritic cell (DC)-based cancer vaccines represent a promising treatment strategy, its exploration in the clinic is hampered due to the need for Good Manufacturing Practice (GMP) facilities and associated trained staff for the generation of large numbers of DCs. The Quantum bioreactor system offered by Terumo BCT represents a hollow-fiber platform integrating GMP-compliant manufacturing steps in a closed system for automated cultivation of cellular products. In the respective established protocols, the hollow fibers are coated with fibronectin and trypsin is used to harvest the final cell product, which in the case of DCs allows processing of only one tenth of an apheresis product. MATERIALS AND RESULTS: We successfully developed a new protocol that circumvents the need for fibronectin coating and trypsin digestion, and makes the Quantum bioreactor system now suitable for generating large numbers of mature human monocyte-derived DCs (Mo-DCs) by processing a complete apheresis product at once. To achieve that, it needed a step-by-step optimization of DC-differentiation, e.g., the varying of media exchange rates and cytokine concentration until the total yield (% of input CD14+ monocytes), as well as the phenotype and functionality of mature Mo-DCs, became equivalent to those generated by our established standard production of Mo-DCs in cell culture bags. CONCLUSIONS: By using this new protocol for the Food and Drug Administration-approved Quantum system, it is now possible for the first time to process one complete apheresis to automatically generate large numbers of human Mo-DCs, making it much more feasible to exploit the potential of individualized DC-based immunotherapy.

Denileukin diftitox (ONTAK) induces a tolerogenic phenotype in dendritic cells and stimulates survival of resting Treg.

Denileukin diftitox (DD), a diphtheria toxin fragment IL-2 fusion protein, is thought to target and kill CD25(+) cells. It is approved for the treatment of cutaneous T-cell lymphoma and is used experimentally for the depletion of regulatory T cells (Treg) in cancer trials. Curiously enough, clinical effects of DD did not strictly correlate with CD25 expression, and Treg depletion was not confirmed unambiguously. Here, we report that patients with melanoma receiving DD immediately before a dendritic cell (DC) vaccine failed to develop a tumor-antigen-specific CD4 and CD8 T-cell immune response even after repeated vaccinations. Analyzing the underlying mechanism, so far we found unknown effects of DD. First, DD modulated DCs toward tolerance by downregulating costimulatory receptors such as CD83 and CD25 while upregulating tolerance-associated proteins/pathways including Stat-3, ß-catenin, and class II transactivator-dependent antigen presentation. Second, DD blocked Stat3 phosphorylation in maturing DCs. Third, only activated, but not resting, Treg internalized DD and were killed. Conversely, resting Treg showed increased survival because of DD-mediated antiapoptotic IL-2 signaling. We conclude that DD exerts functions beyond CD25(+) cell killing that may affect their clinical use and could be tested for novel indications.