Comparison of mutation profiles in primary melanomas and corresponding nodal naevi using next-generation sequencing.

BACKGROUND: Nodal naevi (NN) represent aggregates of melanocytes within peripheral lymph nodes. NN are relatively often found in patients with malignant melanoma (MM), and may mimic metastatic disease. AIM: To study mutation profiles in MM and NN to find out whether NN descend from a primary MM. METHODS: Next-generation sequencing was performed on formalin-fixed paraffin-embedded tissue of 26 pairs of primary MM and corresponding NN detected by sentinel lymph node biopsy, and 29 MM-characteristic genes were investigated. RESULTS: In this study, 90% of mutations were detected exclusively in either MM or NN, but not both, in the same patient; the percentage of identical NN and MM mutations in the same individual was only 10%. The most frequently discovered shared mutations were a C>G substitution in the CDKN2A gene and in-frame deletion in ARID1A. Oncogenic driver mutations were frequently observed in MM but only rarely in NN. About three-quarters of mutations in both MM and NN were characterized by C>T or G>A substitutions. The detected rate of ultraviolet (UV)-related C>T base changes was comparably high in both primary MM (35%) and NN (32%). CONCLUSIONS: Based on our data, it seems that NN descend from previously UV-exposed BRAF wildtype cutaneous melanocytes, rather than from primary MM or arrested progenitor cells.

Aggressive cutaneous squamous cell carcinoma in a hydroxyurea- and ruxolitinib-pretreated patient with polycythaemia vera.

Hydroxyurea and ruxolitinib are frequently used to treat myeloproliferative disorders, including polycythaemia vera, and chronic treatment is associated with many cutaneous adverse effects such as the development of aggressive non-melanoma skin cancer (NMSC). We report an 85-year-old man with a history of hydroxyurea- and ruxolitinib-treated polycythaemia vera who was referred for the management of progressively growing tumours on his scalp. Histopathology of the largest scalp lesion revealed a partly desmoplastic cutaneous squamous carcinoma with perineural invasion. Initial imaging revealed metastatic disease in cervical lymph nodes, bones and lungs. The scalp lesions were successfully treated with bleomycin-based electrochemotherapy. Under initial systemic therapy using four cycles of cetuximab, metastatic disease progressed. Following the approval by the health insurance, compassionate use of pembrolizumab monotherapy was initiated. After three cycles of pembrolizumab, however, metastatic disease further progressed and the patient finally died from global respiratory insufficiency. The present case exemplifies the cutaneous adverse effects of long-term hydroxyurea and ruxolitinib therapy, frequently resulting in highly aggressive NMSCs that are usually not responsive to systemic treatments even such as immune checkpoint inhibitors.

Management of immune-related adverse events in anti-PD-1-treated patients with advanced cutaneous squamous cell carcinoma.

Immune checkpoint inhibitors (ICI) have shown very promising results in the management of patients with inoperable or metastatic cutaneous squamous cell carcinoma (cSCC). However, ICI can cause a range of immune-related adverse events (irAEs) affecting a multitude of organs including skin, gastrointestinal tract, endocrine system, heart, lung, kidneys and the nervous system. In principle, clinical management irAEs does not change significantly with respect to the kind of cancer treated with ICI. However, advanced cSCC typically occurs in a clinically challenging patient population typically presenting with advanced age and/or significant comorbidities such as immunosuppression due to haematological malignancies and their respective treatment. Moreover, many patients with advanced cSCC are organ transplant patients taking immunosuppressants. As a consequence use of ICI per se and management of ICI-induced irAEs generates more complexity and difficulties in patients with cSCC compared to other entities. Here, we provide a brief review on the management of anti-programmed cell death protein 1-induced irAEs in patients with cSCC focusing on the characteristic clinical challenges present in this population.

Development of thoracic sarcoid reactions associated with complete response to anti-PD-1 therapy in a patient with advanced cutaneous squamous cell carcinoma.

In patients with advanced cutaneous squamous cell carcinoma (cSCC), positive efficacy data were reported for anti-PD-1 antibodies. However, anti-PD-1 treatment is associated with a wide range of immune-related adverse events (irAEs). Here, we report on a 78-year-old woman with a huge cSCC on the right cheek spanning from the temporal to the cervical region with evidence for infiltration of the parotid gland, right masseter muscle and right auditory canal. Ultrasound revealed cervical, submandibular and supraclavicular lymph node metastases on patient's right side. On the basis of a medical hardship application, treatment with pembrolizumab was initiated. After two applications, a dramatic regression of the tumour was observed. At this point, the patient was switched to cemiplimab, which, in the meantime, had become available in Germany. After 3 months on cemiplimab, the tumour-related ulcer on the right cheek showed almost complete regression and all previously affected lymph nodes displayed no evidence for malignancy. Thoracic computed tomography (CT) scans revealed enlarged mediastinal and bilateral hilar lymph nodes assessed as primarily reactive. Three months later, however, mediastinal and bilateral hilar lymph nodes further increased in size, accompanied by radiological alterations of the lung parenchyma. Lymph node biopsies revealed sarcoid reactions (SRs) including fibrotic non-caseating epitheloid cell granulomas surrounded by lymphocytes. Since the patient did not display any clinical symptoms, cemiplimab treatment was continued following a 4-week break. Three months later, CT showed significant regression of the described enlarged lymph nodes and parenchymal lung changes. Twenty months after anti-PD-1 treatment, the patient was still in complete remission. In conclusion, we describe, for the first time, the case of a patient with advanced cSCC who developed disseminated thoracic SRs which were associated with dramatic regression of tumour masses. Thus, as with other irAEs, development of SRs might be indicative of an anti-tumour response to anti-PD-1 therapy.

Cutaneous findings following COVID-19 vaccination: review of world literature and own experience.

There is growing evidence that not only the novel coronavirus disease (COVID-19) but also the COVID-19 vaccines can cause a variety of skin reactions. In this review article, we provide a brief overview on cutaneous findings that have been observed since the emerging mass COVID-19 vaccination campaigns all over the world. Unspecific injection-site reactions very early occurring after the vaccination are most frequent. Type I hypersensitivity reactions (e.g. urticaria, angio-oedema and anaphylaxis) likely due to allergy to ingredients may rarely occur but can be severe. Type IV hypersensitivity reactions may be observed, including delayed large local skin lesions ("COVID arm"), inflammatory reactions in dermal filler or previous radiation sites or even old BCG scars, and more commonly morbilliform and erythema multiforme-like rashes. Autoimmune-mediated skin findings after COVID-19 vaccination include leucocytoclastic vasculitis, lupus erythematosus and immune thrombocytopenia. Functional angiopathies (chilblain-like lesions, erythromelalgia) may also be observed. Pityriasis rosea-like rashes and reactivation of herpes zoster have also been reported after COVID-19 vaccination. In conclusion, there are numerous cutaneous reaction patterns that may occur following COVID-19 vaccination, whereby many of these skin findings are of immunological/autoimmunological nature. Importantly, molecular mimicry exists between SARS-CoV-2 (e.g. the spike-protein sequences used to design the vaccines) and human components and may thus explain some COVID-19 pathologies as well as adverse skin reactions to COVID-19 vaccinations.

Decline of programmed death-1-positive circulating T regulatory cells predicts more favourable clinical outcome of patients with melanoma under immune checkpoint blockade.

BACKGROUND: The role of T regulatory lymphocytes (Tregs) and their immunosuppressive mechanisms in the context of programmed death (PD)-1 blockade is not completely understood. OBJECTIVES: To assess the impact of PD-1-blocking antibody treatment on Treg subpopulations in the blood. METHODS: We studied circulating Treg subpopulations in patients with melanoma under nivolumab or pembrolizumab treatment using flow cytometry and correlated these findings with clinical outcomes. RESULTS: These analyses revealed that the frequency of CD4+  CD25++  CD127-  PD-1+ lymphocytes (PD-1+ Tregs) significantly decreased after the first cycle of immunotherapy (23% vs. 8·6%, P = 0·043). Compared with patients who did not show a significant decline of PD-1+ Tregs after the first treatment, those who did had better clinical outcomes with respect to progression-free survival (PFS, P = 0·022) and melanoma-specific death (MSD, P = 0·0038). Multivariate analysis confirmed that a significant decline of PD-1+ Tregs in peripheral blood after the first treatment cycle is a significant predictor of more favourable PFS and MSD (P = 0·04 and 0·017, respectively). Interestingly, the occurrence of immune-related adverse events was also an independent predictor for decreased risk of MSD (P = 0·047; odds ratio 0·064, 95% confidence interval 0·0042-0·97). CONCLUSIONS: We provide preliminary evidence that circulating PD-1+ Tregs rapidly decline after the initiation of treatment with PD-1-blocking antibodies, which is associated with reduced risk of melanoma progression and MSD. Patients showing no decrease of these PD-1+ Tregs in peripheral blood are characterized by an impaired response to immune checkpoint blockade and worse outcome. What's already known about this topic? Programmed death (PD)-1-blocking antibodies are highly effective in melanoma treatment. However, more than half of patients do not benefit from this therapy and to date it is difficult to predict which patients will respond to it. What does this study add? PD-1-blocking antibody therapy rapidly results in a decline of circulating PD-1+ T regulatory cells (Tregs). What is the translational message? Patients showing a decrease of PD-1+ Tregs appear to have better clinical outcome under PD-1 treatment.

[Reality of inpatient vasoactive treatment with prostacyclin derivatives in patients with acral circulation disorders due to systemic sclerosis in Germany]. / Versorgungsrealität der stationären vasoaktiven Therapie mit Prostazyklinderivaten bei Patienten mit akralen Durchblutungsstörungen bei systemischer Sklerose in Deutschland.

BACKGROUND: Raynaud's phenomenon and the frequently ensuing digital ulcerations represent an early and very distressing symptom in patients with systemic sclerosis (scleroderma, SSc) causing significant limitations in the ability to work and quality of life. The use of vasoactive drugs (especially intravenous prostacyclin derivatives) is recommended to reduce the risk of hypoxic tissue damage up to the loss of fingers. METHODS: In order to obtain information about the current state of treatment of patients with prostacyclin derivatives in routine clinical life in Germany, a survey was conducted among the centers affiliated to the German Network for Systemic Scleroderma (DNSS). In addition, a separate patient survey was conducted by the schleroderma self-help group (Sklerodermie Selbsthilfe e. V.), which only covered the symptoms Raynaud's syndrome, digital ulcers and the use of intravenous prostacyclin derivatives. RESULTS: Of the 433 patients surveyed 56% stated that they had already been treated with prostacyclin derivatives (iloprost/alprostadil) because of their illness and symptoms. A total of 61% received the treatment for severe Raynaud's phenomenon and 39% for digital ulcerations. Most respondents not only experienced an improvement in Raynaud's phenomenon and digital ulcers but also a significant improvement of limitations in everyday life. They also needed significantly less outside help and absenteeism from work was much lower. CONCLUSION: Patients consistently reported a positive effect of treatment with prostacyclin derivatives on Raynaud's phenomenon, acral ulcerations, pain and daily restrictions and felt well and safely cared for during inpatient treatment. These positive effects in the patients' perceptions provide crucial information supporting and confirming the current European and international treatment recommendations.

Baseline laboratory parameters predicting clinical outcome in melanoma patients treated with ipilimumab: a single-centre analysis.

BACKGROUND: Overall response rates (ORRs) for ipilimumab in advanced melanoma are only about 10%. Hence, it is important to explore biomarkers predicting ipilimumab responders. OBJECTIVE: We aimed to explore biomarkers to predict therapy outcome in melanoma patients who have undergone standard ipilimumab therapy in a real-world setting. METHODS: Databases of cutaneous melanoma patients (n = 52) who had received ipilimumab were reviewed and data collected on patient characteristics and diverse laboratory parameters. We performed univariate and multivariate statistics including logistic regression analysis and Cox proportional-hazards regression. RESULTS: Baseline leucocytes, lymphocytes, eosinophils, thrombocytes, neutrophil/lymphocyte ratio, thrombocytes/lymphocyte ratio, eosinophil/lymphocyte ratio and serum vitamin D levels were not significantly associated with ORR, progression-free survival (PFS) and melanoma-specific survival (MSS). Multivariate analysis confirmed anti-PD-1 pretreatment as significant predictor for ORR following ipilimumab therapy. Low-LDH levels and more than two ipilimumab cycles turned out to be significant independent predictors for prolonged PFS. Low-S100B levels and anti-PD-1 treatment before or after ipilimumab were significant independent predictors for improved MSS. All aforementioned parameters and faecal calprotectin did not turn out to be predictors for ipilimumab-induced autoimmune-related adverse events and autoimmune colitis, respectively. CONCLUSIONS: Low serum LDH before ipilimumab treatment is an independent predictor for improved PFS. Furthermore, low serum S100B is an independent predictor for MSS. The number of ipilimumab cycles (>2) is significantly associated with prolonged PFS. Pretreatment calprotectin does not predict the occurrence of autoimmune colitis under ipilimumab therapy.

T regulatory cells and other lymphocyte subsets in patients with bullous pemphigoid.

BACKGROUND: Bullous pemphigoid (BP) is the most common autoimmune blistering disease, and is associated with autoantibodies to the hemidesmosomal BP autoantigens BPAG1 and BPAG2. AIM: We aimed to investigate the significance of T regulatory cells and other lymphocyte subsets in patients with BP. METHODS: In total, 31 inpatients with BP were treated with systemic prednisolone in a tapered dose regimen, while 28 healthy individuals matched for age and sex served as the healthy control (HC) group., Blood samples were taken at baseline and after treatment, and levels of inducer/helper and suppressor/cytotoxic T lymphocytes, B lymphocytes, natural killer cells, CD4+CD25++CD127- cells were assessed by flow cytometry, while CD4, CD8, and FOXP3 positivity were assessed by immunohistochemistry, and FOXP3 mRNA was assessed by reverse transcription (RT)-PCR. RESULTS: Flow cytometry showed that numbers of CD8+ and CD4+CD25++CD127- cells were significantly increased, while the number of CD4+ cells and the CD4/CD8 ratio were significantly decreased at baseline and after therapy in patients with BP compared with HCs. Immunohistology revealed that CD4+, CD8+ and FOXP3+ cells were significantly increased at baseline and post-treatment in patients with BP compared with HCs. FOXP3 mRNA levels were significantly increased in the blood of patients with BP compared with HCs. CONCLUSION: These results indicate that increased numbers of CD8+, CD4+CD25++CD127- cells and FOXP3+ cells may play a pathogenetic role during the course of BP.

Glutathione-S-transferase T1 genotyping and phenotyping in psoriasis patients receiving treatment with oral fumaric acid esters.

BACKGROUND: Glutathione S-transferases (GSTs) are involved in detoxification of xenobiotics such as fumaric acid esters (FAE). OBJECTIVES: To perform GSTT1 geno- and phenotyping in psoriasis patients treated with FAE to find out whether the responder status and/or occurrence of side-effects are associated with allelic variants and enzymatic activity of GSTT1. METHODS: We treated 106 psoriasis patients with FAE. GSTT1 genotyping was performed using PCR, phenotyping was carried out by means of a validated high performance liquid chromatography assay at baseline and under treatment. RESULTS: The distribution of GSTT1 genotypes was as follows: 31% *A/*A; 49% *A/*0; 20% *0/*0. GSTT1 phenotypes as expressed in enzyme activity significantly differed between conjugators classes. (P < 0.001). GSTT1 activity under treatment was significantly (P = 0.0001) increased when compared with baseline. There were no significant associations between the aforementioned GSTT1 pheno- and genotypes and clinical parameters such as psoriasis area and severity index (PASI)50, adverse effects and FAE dosage (P > 0.05), except for the frequent occurrence of reduction (>50%) of circulating lymphocytes in patients with *0/*0 GSTT1 status (P = 0.036; odds ratio: 6, 95% CI: 1.1-32). CONCLUSION: GSTT1 geno- and phenotypes significantly correlate in psoriasis patients and do not substantially differ from healthy controls. Response to FAE does likely not depend on GSTT1. However, *0/*0 GSTT1 status is a predictor for the occurrence of marked reduction of lymphocyte counts under FAE therapy. Notably, FAE seem to enhance GSTT1 enzyme activity in high and low conjugators.

[Clinical presentation and treatment of herpes zoster and postherpetic neuralgia]. / Klinik und Therapie des Herpes zoster und der postherpetischen Neuralgie.

Herpes zoster (HZ) is a common disease caused by reactivation of varicella zoster virus. Diagnosis is usually based on the typical clinical presentation. Standard treatment includes antiviral, topical and analgesic therapies. As a complication, postherpetic neuralgia (PHN) can result from acute HZ infection, particularly in older and/or immunocompromised people. This can seriously impair the quality of life of those affected and requires adequate analgesia. In addition to the genesis, clinical presentation and treatment recommendations for HZ and PHN, this article also deals in particular with the vaccination prophylaxis recommended by the standing vaccination commission of the Robert Koch Institute (STIKO).

Expression of DNA mismatch repair proteins in melanoma patients treated with immune checkpoint inhibitors.

PURPOSE: To investigate the protein expression of DNA mismatch repair (MMR) proteins in patients with cutaneous melanoma (CM) under immune checkpoint inhibitor (ICI) therapy. METHODS: Immunohistochemistry was performed on tumor tissue for MMR proteins MLH1, MSH2, MSH6, and PMS2 in 50 metastatic CM patients treated with ICI (ipilimumab, nivolumab, pembrolizumab). RESULTS: Best overall response (BOR) rate was 48% (24/50). Reduced MMR protein expression (nuclear expression in < 80% of tumor cells) was observed in 8 patients (16%). Compared to other clinical parameters, baseline neutrophil/lymphocyte ratio and reduced intratumoral MMR protein expression (P = 0.0033) were determined as the only parameters significantly associated with favorable BOR. However, in this small study population, reduced MMR protein expression did not reach statistical significance in multivariate analysis. CONCLUSION: Reduced MMR protein expression is observed in CM and might predict favorable BOR in patients treated with ICI, as was observed for other entities. However, these findings need to be substantiated in larger patient cohorts.

The pan-immune-inflammation value and systemic immune-inflammation index in advanced melanoma patients under immunotherapy.

PURPOSE: To evaluate the pan-immune-inflammation value (PIV) and systemic immune-inflammation index (SII) in patients with cutaneous melanoma (CM) under immune checkpoint inhibitor (ICI) therapy. METHODS: PIV and SII were calculated before the start of ICI therapy and at time of progression/death in patients with metastatic CM (stage III/IV). Sex-age-matched CM patients in stage I/II and healthy subjects (HC) served as controls. RESULTS: The median PIV of stage III/IV patients was significantly (P = 0.0011) higher than in stage I/II patients and HC. SII was significantly (P = 0.00044) lower in HC than in CM patients. At baseline, PIV and SII did significantly correlate with lactate dehydrogenase (P = 0.045/0.017). However, ROC curve statistics revealed that SII and PIV were not significantly associated with clinical parameters, including best response to ICI treatment (P = 0.87/0.64), progression-free survival (P = 0.73/0.91), and melanoma-specific survival (P = 0.13/0.17). Moreover, there were no significant changes of PIV and SII from baseline to progression/death (P = 0.38/0.52). CONCLUSIONS: Even though both immune-inflammation biomarkers showed some power to differentiate between CM stages and HC, respectively, PIV and SII seem not to be significant predictors for clinical outcome measures of CM patients under ICI therapy.

Pan-immune-inflammation value independently predicts disease recurrence in patients with Merkel cell carcinoma.

PURPOSE: We aimed to determine whether the pan-immune-inflammation value (PIV) of patients with Merkel cell carcinoma (MCC) at primary diagnosis differs from controls and whether it is associated with disease stage and outcome. METHODS: In this retrospective study, we recruited MCC patients with stage I-III. PIV was calculated from absolute complete blood cell counts obtained within one week at MCC diagnosis as follows: [neutrophils (103/mm3) × platelets (103/mm3) × monocytes (103/mm3)]/lymphocytes (103/mm3). As controls, we studied age-gender-matched cutaneous melanoma (CM, stage I-III) patients and healthy controls (HC). Univariate and multivariate statistics were used. RESULTS: The median PIV in MCC patients was significantly increased compared to both CM patients as well as healthy controls. PIV of MCC patients in stage II and III was significantly higher compared to stage I patients. ROC analysis revealed that MCC recurrence was significantly associated with a PIV greater than 372 [p < 0.0001, Youden index 0.58; hazard ratio: 4 (95% confidence interval: 1.7 to 9.2)]. In multivariate analysis, only a PIV greater than 372 and higher MCC stage were determined as independent predictors for disease recurrence. CONCLUSION: We determined, for the first time, the prognostic ability of the promising blood-based biomarker PIV in MCC patients and observed that PIV is increased in MCC patients in dependence on disease stage and independently predicts MCC recurrence.

Protein expression of prognostic genes in primary melanoma and benign nevi.

PURPOSE: To evaluate the protein expression characteristics of genes employed in a recently introduced prognostic gene expression assay for patients with cutaneous melanoma (CM). METHODS: We studied 37 patients with CM and 10 with benign (melanocytic) nevi (BN). Immunohistochemistry of primary tumor tissue was performed for eight proteins: COL6A6, DCD, GBP4, KLHL41, KRT9, PIP, SCGB1D2, SCGB2A2. RESULTS: The protein expression of most markers investigated was relatively low (e.g., DCD, KRT9, SCGB1D2) and predominantly cytoplasmatic in melanocytes and keratinocytes. COL6A6, GBP4, and KLHL41 expression was significantly enhanced in CM when compared to BN. DCD protein expression was significantly correlated with COL6A6, GBP4, and KLHL41. GBP4 was positively correlated with KLHL41 and inversely correlated with SCGB2B2. The latter was also inversely correlated with serum S100B levels at time of initial diagnosis. The presence of SCGB1D2 expression was significantly associated with ulceration of the primary tumor. KRT9 protein expression was significantly more likely found in acral lentiginous melanoma. The presence of DCD expression was less likely associated with superficial spreading melanoma subtype but significantly associated with non-progressive disease. The absence of SCGB2A2 expression was significantly more often observed in patients who did not progress to stage III or IV. CONCLUSIONS: The expression levels observed were relatively low but differed in part with those found in BN. Even though we detected some significant correlations between the protein expression levels and clinical parameters (e.g., CM subtype, course of disease), there was no major concordance with the protective or risk-associated functions of the corresponding genes included in a recently introduced prognostic gene expression assay.