Long-term follow-up of patients with complex coronary artery disease treated with minimally invasive direct coronary artery bypass.

BACKGROUND: Patients with complex coronary artery disease (CAD) may benefit from surgical myocardial revascularization but weighing the risk of peri-operative complications against the expected merit is difficult. Minimally invasive direct artery bypass (MIDCAB) procedures are less invasive, provide the prognostic advantage of operative revascularization of the left anterior descending artery and may be integrated in hybrid strategies. Herein, the outcomes between patients with coronary 1-vessel disease (1-VD) and patients with 2-VD and 3-VD after MIDCAB procedures were compared in this single-center study. METHODS: Between 1998 and 2018, 1363 patients underwent MIDCAB at the documented institution. 628 (46.1%) patients had 1-VD, 434 (31.9%) patients 2-VD and 300 (22.0%) patients suffered from 3-VD. Data of patients with 2-VD, and 3-VD were pooled as multi-VD (MVD). RESULTS: Patients with MVD were older (66.2 ± 10.9 vs. 62.9 ± 11.2 years; p < 0.001) and presented with a higher EuroScore II (2.10 [0.4; 34.2] vs. 1.2 [0.4; 12.1]; p < 0.001). Procedure time was longer in MVD patients (131.1 ± 50.3 min vs. 122.2 ± 34.5 min; p < 0.001). Post-operatively, MVD patients had a higher stroke rate (17 [2.3%] vs. 4 [0.6%]; p = 0.014). No difference in 30-day mortality was observed (12 [1.6%] vs. 4 [0.6%]; p = 0.128). Survival after 15 years was significantly lower in MVD patients (p < 0.01). Hybrid procedures were planned in 295 (40.2%) patients with MVD and realized in 183 (61.2%) cases. MVD patients with incomplete hybrid procedures had a significantly decreased long-term survival compared to cases with complete revascularization (p < 0.01). CONCLUSIONS: Minimally invasive direct coronary artery bypass procedures are low-risk surgical procedures. If hybrid procedures have been planned, completion of revascularization should be a major goal.

Topical 188Re Ionizing Radiation Therapy Exerts High Efficacy in Curing Nonmelanoma Skin Cancer.

PURPOSE OF THE REPORT: Nonmelanoma skin cancer (NMSC) is the most frequent malignancy. Surgical intervention is the common treatment but may lead to disappointing results; alternative treatment options are needed. METHODS: In this monocentric pilot study, topical 188Re resin was investigated as a treatment for invasive NMSC up to 3-mm thickness. Twenty-two patients with 40 histologically confirmed NMSCs with a median size of 1.25 cm2 (range, 0.04-16.8 cm2) and a median tumor thickness of 0.35 mm (range, 0.1-2.1 mm) were included. Patients were treated once with 188Re resin with a targeted dose of 50 Gy. The median applied activity was 111.4 MBq (range, 21.0-168.0 MBq), and the median treatment time was 89 minutes (range, 38-175 minutes). The response rate, adverse events, and cosmetic outcome were assessed at 14 days, 4 months, and 12 months. RESULTS: Response rate at 12 months was 97.5%, with 95% complete responses (clinically or histologically proven in case of clinical doubt). Most adverse events were reported at 14 days, with 20% itching and 12.5% mostly minor pain. Forty-nine percent of the lesions showed hypopigmentation only at 12 months. Forty-one percent of the lesions were graded as cosmetically superior to the expected result after surgery and 51.3% as comparable to successful surgery. The cosmetic outcome on the head and face was superior compared with the trunk and leg (P = 0.003). CONCLUSION: 188Re resin is a highly effective treatment for NMSC up to 3-mm thickness and a valid alternative to surgery, specifically for tumors located on sensitive areas such as nose or ear.

A Prospective Randomized Controlled Pilot Study to Assess the Response and Tolerability of Cold Atmospheric Plasma for Rosacea.

INTRODUCTION: Rosacea is a common, facial, chronic inflammatory skin disease. Due to its complex pathogenesis, adequate therapy of rosacea can be challenging. An innovative recent therapeutic tool is cold atmospheric plasma (CAP), which is already established in the treatment of chronic wounds and promising in different other skin diseases. METHODS: In a split-face pilot study we investigated dielectric-barrier-discharged CAP in erythemato-telangiectatic (ETR) and/or papulopustular rosacea (PPR). CAP treatment was applied on lesional skin of a randomized side once daily (90 s/area) for 6 weeks. The other untreated side served as control. Co-primary endpoints were ≥1 improvement of the Investigator Global Assessment (IGA) score on the treated side compared to control and a decline of the Dermatology Life Quality Index (DLQI) after 6 weeks. Secondary endpoints included inflammatory lesion count (papules and pustules), skin redness intensity and erythema size. Adverse events (AEs) were recorded constantly. Additionally, participants were weekly assessed for symptoms, skin condition, trigger factors, skin care, treatment success, and local tolerance parameters. All p values were calculated using the Wilcoxon signed-rank test. RESULTS: Twelve subjects (ETR, n = 3; ETR and PPR, n = 9) completed the study. DLQI was significantly improved after 6 weeks (p = 0.007). On the CAP-treated side, lesions (p = 0.007) and erythema size (p = 0.041) were significantly reduced compared to the control. IGA (p = 0.2) and skin redness intensity (p = 0.5) did not differ significantly between control and CAP-treated side. No serious AEs occurred and treatment was well tolerated. CONCLUSION: CAP is a promising new treatment of rosacea, especially for PPR.

Tumor-derived GDF-15 blocks LFA-1 dependent T cell recruitment and suppresses responses to anti-PD-1 treatment.

Immune checkpoint blockade therapy is beneficial and even curative for some cancer patients. However, the majority don't respond to immune therapy. Across different tumor types, pre-existing T cell infiltrates predict response to checkpoint-based immunotherapy. Based on in vitro pharmacological studies, mouse models and analyses of human melanoma patients, we show that the cytokine GDF-15 impairs LFA-1/ß2-integrin-mediated adhesion of T cells to activated endothelial cells, which is a pre-requisite of T cell extravasation. In melanoma patients, GDF-15 serum levels strongly correlate with failure of PD-1-based immune checkpoint blockade therapy. Neutralization of GDF-15 improves both T cell trafficking and therapy efficiency in murine tumor models. Thus GDF-15, beside its known role in cancer-related anorexia and cachexia, emerges as a regulator of T cell extravasation into the tumor microenvironment, which provides an even stronger rationale for therapeutic anti-GDF-15 antibody development.

Real-world systemic treatment of patients with psoriasis: A retrospective study based on German claims data.

BACKGROUND AND OBJECTIVES: Psoriasis is a common skin disorder with a high physical and psychological burden for patients. Up to 30% of the patients are candidates for a systemic treatment. The aim of this study was to describe the characteristics and the real-world systemic treatment of psoriasis patients. PATIENTS AND METHODS: This study was based on German medical claims data. A cross-sectional analysis observed all psoriasis patients in 2020. A longitudinal analysis was conducted, addressing psoriasis patients who newly started a systemic treatment. RESULTS: In total, 116,507 prevalent psoriasis patients and 13,449 newly treated patients were followed. Of all prevalent patients, 15.2% received systemic treatment in 2020 (8.7% systemic corticosteroids). Of the newly treated patients, 95.2% started with conventional treatment (79.2% systemic corticosteroids), 4.0% with biologics and 0.9% with apremilast. The rate of treatment discontinuation/switch after one year was highest for corticosteroids (91.3%) and lowest for biologics (23.1%). CONCLUSIONS: Around 15% of psoriasis patients in Germany received a systemic treatment, with > 50% of these prescribed systemic corticosteroids. Therefore, we conclude that systemic treatment is not in line with guideline recommendations in a substantial number of observed patients. The lowest discontinuation/switch rates for biologics support their wider use.

Immune checkpoint inhibition and targeted therapy for melanoma: A patient-oriented cross-sectional comparative multicentre study.

BACKGROUND: Choosing the adequate systemic treatment for melanoma is driven by clinical parameters and personal preferences. OBJECTIVE: Evaluation of the impact of disease and treatment on the daily life of patients receiving systemic therapy for melanoma. METHODS: A German-wide, cross-sectional comparative study was conducted at 13 specialized skin cancer centres from 08/2020 to 03/2021. A questionnaire was distributed to assess patients' perception of disease and symptoms, the impact of their current treatment on quality of life (QOL) and activities, adverse events (AEs), therapeutic visits, as well as believe in and satisfaction with their current systemic melanoma treatment. Patient-reported outcomes (PROs) were rated on a continuous numerical rating scale or selected from a given list. RESULTS: Four hundred and fourteen patients with systemic melanoma therapy were included. 359 (87%) received immune checkpoint inhibition (ICI) and 55 (13%) targeted therapy (TT). About 1/3 of patients were adjuvantly treated, the remaining because of unresectable/metastatic melanoma. In subgroup analyses, only in the adjuvant setting, TT patients reported a significant decrease in their treatment associated QOL compared to patients with ICI (p = 0.02). Patients with TT were 1.9 times more likely to report AEs than patients with ICI, a difference being significant just for the adjuvant setting (p = 0.01). ICI treatment intervals differed significantly between adjuvant and unresectable/metastatic setting (p = 0.04), though all patients, regardless of their specific ICI drug, evaluated their treatment frequency as adequate. TT patients with dabrafenib/trametinib (n = 37) or encorafenib/binimetinib (n = 15) did not differ regarding the strain of daily pill intake. Patients older than 63 years rated various PROs better than younger patients. CONCLUSIONS: Patients evaluated their treatment mainly positively. ICI might be preferred over TT regarding QOL and patient-reported AEs in the adjuvant setting. Older melanoma patients appeared to be less impacted by their disease and more satisfied with their treatment.

Homozygous CRISPR/Cas9 Knockout Generated a Novel Functionally Active Exon 1 Skipping XPA Variant in Melanoma Cells.

Defects in DNA repair pathways have been associated with an improved response to immune checkpoint inhibition (ICI). In particular, patients with the nucleotide excision repair (NER) defect disease Xeroderma pigmentosum (XP) responded impressively well to ICI treatment. Recently, in melanoma patients, pretherapeutic XP gene expression was predictive for anti-programmed cell death-1 (PD-1) ICI response. The underlying mechanisms of this finding are still to be revealed. Therefore, we used CRISPR/Cas9 to disrupt XPA in A375 melanoma cells. The resulting subclonal cell lines were investigated by Sanger sequencing. Based on their genetic sequence, candidates from XPA exon 1 and 2 were selected and further analyzed by immunoblotting, immunofluorescence, HCR and MTT assays. In XPA exon 1, we established a homozygous (c.19delG; p.A7Lfs*8) and a compound heterozygous (c.19delG/c.19_20insG; p.A7Lfs*8/p.A7Gfs*55) cell line. In XPA exon 2, we generated a compound heterozygous mutated cell line (c.206_208delTTG/c.208_209delGA; p.I69_D70delinsN/p.D70Hfs*31). The better performance of the homozygous than the heterozygous mutated exon 1 cells in DNA damage repair (HCR) and post-UV-C cell survival (MTT), was associated with the expression of a novel XPA protein variant. The results of our study serve as the fundamental basis for the investigation of the immunological consequences of XPA disruption in melanoma.

The Prognostic and Predictive Role of Xeroderma Pigmentosum Gene Expression in Melanoma.

BACKGROUND: Assessment of immune-specific markers is a well-established approach for predicting the response to immune checkpoint inhibitors (ICIs). Promising candidates as ICI predictive biomarkers are the DNA damage response pathway genes. One of those pathways, which are mainly responsible for the repair of DNA damage caused by ultraviolet radiation, is the nucleotide excision repair (NER) pathway. Xeroderma pigmentosum (XP) is a hereditary disease caused by mutations of eight different genes of the NER pathway, or POLH, here together named the nine XP genes. Anecdotal evidence indicated that XP patients with melanoma or other skin tumors responded impressively well to anti-PD-1 ICIs. Hence, we analyzed the expression of the nine XP genes as prognostic and anti-PD-1 ICI predictive biomarkers in melanoma. METHODS: We assessed mRNA gene expression in the TCGA-SKCM dataset (n = 445) and two pooled clinical melanoma cohorts of anti-PD-1 ICI (n = 75). In TCGA-SKCM, we applied hierarchical clustering on XP genes to reveal clusters, further utilized as XP cluster scores. In addition, out of 18 predefined genes representative of a T cell inflamed tumor microenvironment, the TIS score was calculated. Besides these scores, the XP genes, immune-specific single genes (CD8A, CXCL9, CD274, and CXCL13) and tumor mutational burden (TMB) were cross-correlated. Survival analysis in TCGA-SKCM was conducted for the selected parameters. Lastly, the XP response prediction value was calculated for the two pooled anti-PD-1 cohorts by classification models. RESULTS: In TCGA-SKCM, expression of the XP genes was divided into two clusters, inversely correlated with immune-specific markers. A higher ERCC3 expression was associated with improved survival, particularly in younger patients. The constructed models utilizing XP genes, and the XP cluster scores outperformed the immune-specific gene-based models in predicting response to anti-PD-1 ICI in the pooled clinical cohorts. However, the best prediction was achieved by combining the immune-specific gene CD274 with three XP genes from both clusters. CONCLUSION: Our results suggest pre-therapeutic XP gene expression as a potential marker to improve the prediction of anti-PD-1 response in melanoma.

Prevalence of pathological skin-picking in dermatological patients.

OBJECTIVE: The prevalence of pathological skin-picking (PSP) has predominantly been studied in students and the community, but not yet in dermatological patients. However, those may be at increased risk of PSP because it is often triggered by the feel or look of the skin. Thus, its prevalence among patients with a physician-diagnosed dermatological disease remains to be determined. METHODS: A consecutive series of 460 adult patients attending a dermatological university outpatient clinic was administered the 8-item Skin Picking Scale-Revised (SPS-R). They also reported demographic data and rated the severity of their skin disease. The dermatologist evaluating the patient provided his/her diagnosis. RESULTS: PSP as defined by SPS-R scores ≥7 was reported by 121 participants (26.3%). It was significantly more frequent in patients with atopic dermatitis (AD, OR = 3.23; 95% CI: 1.95-5.68) and psoriasis (OR = 1.64; 95% CI: 1.00-2.67), but less frequent in those with malignant epithelial skin tumors (OR = 0.10; 95% CI: 0.02-0.43). PSP was not associated with female gender or younger age. CONCLUSIONS: Our findings indicate that PSP affects about one in four patients with skin disease. In particular, individuals suffering from atopic dermatitis may represent a high-risk population for PSP deserving early recognition and adequate treatment.

[Wound therapy with cold atmospheric plasma in severe recessive dystrophic epidermolysis bullosa : A pilot study]. / Wundtherapie mit kaltem Plasma bei Epidermolysis bullosa dystrophica : Eine Pilotuntersuchung.

BACKGROUND: Cold atmospheric pressure plasma (CAP) has antimicrobial and wound-healing properties. Patients affected by severe autosomal recessive dystrophic epidermolysis bullosa (RDEB) suffer from widespread, difficult-to-treat wounds, which require complex wound management. OBJECTIVE: In a pilot project, we investigated over a period of 5 months the response and tolerability of a CAP wound therapy in a 21-year-old and a 28-year-old female patient with severe generalized RDEB and following cutaneous squamous cell cancer (cSSC) in the older patient. MATERIALS AND METHODS: In both patients, diagnosis of RDEB was confirmed by molecular genetics. Individual- and patient-specific wound therapy was continued during the study period, and additionally CAP therapy with a dielectric barrier discharge (DBE) device was initiated. CAP treatment was performed for 90 s per wound and could be applied every day or every other day. Clinical evaluation included photographic documentation and regular interviews of patients and parents. RESULTS: CAP-treated wounds largely demonstrated improved wound healing and signs of a reduced bacterial contamination. Furthermore, CAP proved to prevent wound chronification. When applied on a polyester mesh, it was well-tolerated on most body sites. CONCLUSION: The introduction of CAP could improve the wound management of EB patients and should be evaluated in clinical studies. The effect of CAP on cSSC development should be particularly studied.

The Response and Tolerability of a Novel Cold Atmospheric Plasma Wound Dressing for the Healing of Split Skin Graft Donor Sites: A Controlled Pilot Study.

INTRODUCTION: Cold atmospheric plasma (CAP) has positive effects on wound healing and antimicrobial properties. However, an ongoing challenge is the development of specific modes of application for different clinical indications. OBJECTIVES: We investigated in a prospective pilot study the response and tolerability of a newly developed CAP wound dressing for the acute healing of split skin graft donor sites compared to conventional therapy. METHODS: We applied both treatments to each patient (n = 10) for 7 days and measured 4 parameters of wound healing every other day (i.e., 1,440 measurements) using a hyperspectral imaging camera. Additionally, we evaluated the clinical appearance and pain levels reported by the patients. RESULTS: The CAP wound dressing was superior to the control (p < 0.001) in the improvement of 3 wound parameters, that is, deep tissue oxygen saturation, hemoglobin distribution, and tissue water distribution. CAP was well tolerated, and pain levels were lower in CAP-treated wound areas. CONCLUSION: CAP wound dressing is a promising new tool for acute wound healing.

Risk Factors for Mortality in Acute Aortic Dissection Type A: A Centre Experience Over 15 Years.

INTRODUCTION: Acute aortic dissection Type A (AADA) is still associated with a high mortality rate and frequent postoperative complications. This study was designed to evaluate the risk factors for mortality in AADA patients. PATIENTS AND METHODS: This retrospective analysis included 344 consecutive patients who underwent surgery for AADA in moderate hypothermic circulatory arrest (20-24°C nasopharyngeal) between 2001 and 2016. RESULTS: The 30-day mortality rate was 18%. Nonsurvivors were significantly older (65.7 ± 12.0 years vs. 62.0 ± 12.5 years; p = 0.034) with significantly higher Euro-score II [15.4% (6.6; 23.0) vs. 4.63% (2.78; 9.88); p < 0.001)]. Intraoperatively, survivors had statistically shorter cardiopulmonary bypass times [163 (134; 206) vs. 198 min (150; 245); p = 0.001]. However, the hypothermic circulatory arrest time was similar between both groups. Postoperatively, the incidence of acute kidney injury (AKI) (55.9 vs. 15.2%; p < 0.001), stroke (27.9 vs. 12.1%; p = 0.002) and sepsis (18.0 vs. 2.1%; p < 0.001) were significantly higher among nonsurvivors. The multi-variable logistic regression confirmed that older age, previous cardiac surgery, preoperative cardiopulmonary resuscitation (CPR), blood transfusion and postoperative acute kidney injury (AKI) were independent risk factors for mortality. CONCLUSION: Our analysis suggested that the reason for mortality was multifactorial, especially age, previous cardiac surgery, CPR, transfusion, as well as postoperative AKI were considered risk factors for mortality.

Influence of moderate hypothermic circulatory arrest on outcome in patients undergoing elective replacement of thoracic aorta.

BACKGROUND: The ideal technique of cerebral protection in the surgical operation of the ascending aorta.is currently controversial. The current analysis evaluates the influence of moderate hypothermic circulatory arrest (MHCA) on elective replacement of the ascending aorta. METHODS: The study included 905 consecutive patients between 2001 and 2015, who underwent replacement of ascending aorta in MHCA. Patients were divided according to the postoperative 30-day mortality into survivor und non-survivor group. RESULTS: The average age was 66.5±11.1 in survivors vs. 70.0±10.5 years in non-survivors (P=0.057). The survivor group had a significantly lower Euro-SCORE II than non-survivors [4.0% (2.3, 6.6) vs. 9.5% (4.8, 20.9); P<0.001)]. The incidence of coronary heart disease (38.0% vs. 58.3%; P=0.022) and chronic renal failure (10.0% vs. 33.3%, P<0.001 was significantly higher in non-survivors. Intraoperatively, the cardiopulmonary bypass time [140 min (112, 185) vs. 194 min (164, 271); P<0.001] and cross-clamping time [91 min (64, 124) vs.119 min (94, 157); P<0.001] were significantly longer in non-survivors. However, the MHCA time was similar in both groups with statistical significance (P=0.023). Postoperatively, re-exploration due to bleeding was highly significant in non-survivors (5.4% vs. 33.3%; P<0.001) with a higher incidence of stroke (4.6% vs. 33.3%; P<0.001). The duration of mechanical ventilation was significantly shorter in survivors than in non-survivors [17 h (12, 26) vs. 147 h (49, 337); P<0.001] with a lower incidence of pulmonary infection (6.0% vs.16.7%; P=0.023). The multivariable logistic regression analysis showed age, female gender, aortic aneurysm, additional CABG, total arch replacement and cardiopulmonary bypass time were independent risk factors for 30-day mortality. CONCLUSIONS: The acceptable morbidity and mortality rates show that MHCA can be considered as a safe technique for cerebral protection in surgical replacement of thoracic aorta.

Treatment Motivations and Expectations in Patients with Actinic Keratosis: A German-Wide Multicenter, Cross-Sectional Trial.

Patient-centered motives and expectations of the treatment of actinic keratoses (AK) have received little attention until now. Hence, we aimed to profile and cluster treatment motivations and expectations among patients with AK in a nationwide multicenter, cross-sectional study including patients from 14 German skin cancer centers. Patients were asked to complete a self-administered questionnaire. Treatment motives and expectations towards AK management were measured on a visual analogue scale from 1-10. Specific patient profiles were investigated with subgroup and correlation analysis. Overall, 403 patients were included. The highest motivation values were obtained for the items "avoid transition to invasive squamous cell carcinoma" (mean ± standard deviation; 8.98 ± 1.46), "AK are considered precancerous lesions" (8.72 ± 1.34) and "treating physician recommends treatment" (8.10 ± 2.37; p < 0.0001). The highest expectation values were observed for the items "effective lesion clearance" (8.36 ± 1.99), "safety" (8.20 ± 2.03) and "treatment-related costs are covered by health insurance" (8.00 ± 2.41; p < 0.0001). Patients aged ≥77 years and those with ≥7 lesions were identified at high risk of not undergoing any treatment due to intrinsic and extrinsic motivation deficits. Heat mapping of correlation analysis revealed four clusters with distinct motivation and expectation profiles. This study provides a patient-based heuristic tool for a personalized treatment decision in patients with AK.

Effect of moderate hypothermic circulatory arrest on neurological outcomes in elderly patients undergoing replacement of the thoracic aorta.

BACKGROUND: Various studies evaluated the relationship between hypothermic circulatory arrest and neurological outcome in patients undergoing replacement of ascending aorta. The current analysis focuses on the effect of moderate hypothermic circulatory arrest (MHCA) on elderly patients. The aim of our study was to evaluate the impact of MHCA on neurological outcomes in elderly patients undergoing replacement of the ascending aorta. RESULTS: We retrospectively analyzed 905 consecutive patients, who underwent elective replacement of ascending aorta in MHCA (24 ± 2 °C, nasopharyngeal) between 2001 and 2015. Patients with acute aortic dissection were excluded from this study. Patients were divided into two groups: those aged 75 years and older (elderly group 22.4%, n = 203) and those younger than 75 years (younger group 77.6%, n = 702). The average age was 63.2 ± 10.2 in the young group vs. 78.7 ± 3.0 years in elderly group (p < 0.001). The elderly group had a significantly higher EuroSCORE II [26.7% (18.1, 36.3) vs. 11.6% (7.4, 19.9); p < 0.001)]. The incidence of coronary heart disease (49.8% vs. 35.6%, p < 0.001) and chronic renal failure (17.2% vs. 9.1%, p = 0.001) was significantly higher in the elderly group. Intraoperatively, the time of MHCA [14 min (12, 17) vs. 15 min (12, 18); p = 0.42], cardiopulmonary bypass [139 min (110, 183) vs. 144 min (113, 189); p = 0.225], and cross-clamping [91 min (63, 116) vs. 92 min (65, 127); p = 0.348] was similar in both groups. Postoperatively, a higher incidence of delirium was significantly reported in the elderly group (24.1% vs. 9.0%, p < 0.001). However, there was no significant difference regarding neurological complications between both groups. A 30-day mortality was acceptable for the elderly group, but significantly higher compared with the younger group (7.1% vs. 3.5%, p = 0.031). CONCLUSIONS: Our study suggests that surgical replacement of the ascending aorta in MHCA can also be applied safely in elderly patients without increasing the risk of severe neurological complications.

IFN-gamma-induced PD-L1 expression in melanoma depends on p53 expression.

BACKGROUND: Immune checkpoint inhibition and in particular anti-PD-1 immunotherapy have revolutionized the treatment of advanced melanoma. In this regard, higher tumoral PD-L1 protein (gene name: CD274) expression is associated with better clinical response and increased survival to anti-PD-1 therapy. Moreover, there is increasing evidence that tumor suppressor proteins are involved in immune regulation and are capable of modulating the expression of immune checkpoint proteins. Here, we determined the role of p53 protein (gene name: TP53) in the regulation of PD-L1 expression in melanoma. METHODS: We analyzed publicly available mRNA and protein expression data from the cancer genome/proteome atlas and performed immunohistochemistry on tumors with known TP53 status. Constitutive and IFN-É£-induced PD-L1 expression upon p53 knockdown in wildtype, TP53-mutated or JAK2-overexpressing melanoma cells or in cells, in which p53 was rendered transcriptionally inactive by CRISPR/Cas9, was determined by immunoblot or flow cytometry. Similarly, PD-L1 expression was investigated after overexpression of a transcriptionally-impaired p53 (L22Q, W23S) in TP53-wt or a TP53-knockout melanoma cell line. Immunoblot was applied to analyze the IFN-É£ signaling pathway. RESULTS: For TP53-mutated tumors, an increased CD274 mRNA expression and a higher frequency of PD-L1 positivity was observed. Interestingly, positive correlations of IFNG mRNA and PD-L1 protein in both TP53-wt and -mutated samples and of p53 and PD-L1 protein suggest a non-transcriptional mode of action of p53. Indeed, cell line experiments revealed a diminished IFN-É£-induced PD-L1 expression upon p53 knockdown in both wildtype and TP53-mutated melanoma cells, which was not the case when p53 wildtype protein was rendered transcriptionally inactive or by ectopic expression of p53L22Q,W23S, a transcriptionally-impaired variant, in TP53-wt cells. Accordingly, expression of p53L22Q,W23S in a TP53-knockout melanoma cell line boosted IFN-É£-induced PD-L1 expression. The impaired PD-L1-inducibility after p53 knockdown was associated with a reduced JAK2 expression in the cells and was almost abrogated by JAK2 overexpression. CONCLUSIONS: While having only a small impact on basal PD-L1 expression, both wildtype and mutated p53 play an important positive role for IFN-É£-induced PD-L1 expression in melanoma cells by supporting JAK2 expression. Future studies should address, whether p53 expression levels might influence response to anti-PD-1 immunotherapy.

The identification of patient-specific mutations reveals dual pathway activation in most patients with melanoma and activated receptor tyrosine kinases in BRAF/NRAS wild-type melanomas.

BACKGROUND: Increasing knowledge of cancer genomes has triggered the development of specific targeted inhibitors, thus providing a valuable therapeutic pool. METHODS: In this report, the authors analyze the presence of targetable alterations in 136 tumor samples from 92 patients with melanoma using a comprehensive approach based on targeted DNA sequencing and supported by RNA and protein analysis. Three topics of high clinical relevance are addressed: the identification of rare, activating alterations; the detection of patient-specific, co-occurring single nucleotide variants (SNVs) and copy number variations (CNVs) in parallel pathways; and the presence of cancer-relevant germline mutations. RESULTS: The analysis of patient-matched blood and tumor samples was done with a custom-designed gene panel that was enriched for genes from clinically targetable pathways. To detect alterations with high therapeutic relevance for patients with unknown driver mutations, genes that are untypical for melanoma also were included. Among all patients, CNVs were identified in one-third of samples and contained amplifications of druggable kinases, such as CDK4, ERBB2, and KIT. Considering SNVs and CNVs, 60% of patients with metastases exhibited co-occurring activations of at least 2 pathways, thus providing a rationale for individualized combination therapies. Unexpectedly, 9% of patients carry potentially protumorigenic germline mutations frequently affecting receptor tyrosine kinases. Remarkably two-thirds of BRAF/NRAS wild-type melanomas harbor activating mutations or CNVs in receptor tyrosine kinases. CONCLUSIONS: The results indicate that the integrated analysis of SNVs, CNVs, and germline mutations reveals new druggable targets for combination tumor therapy.