Background: Cancer immunotherapy has revolutionized melanoma treatment, but the high number of non- responders still emphasizes the need for improvement of therapy. One potential avenue for enhancing antitumor treatment is through the modulation of coagulation and platelet activity. Both have been found to play an important role in the tumor microenvironment, tumor growth and metastasis. Preclinical studies indicate a beneficial effect, clinical data has been inconsistent. Methods: We examined a cohort of advanced, non-resectable melanoma patients (n = 2419) derived from the German prospective multicenter skin cancer registry ADOReg, who were treated with immune checkpoint inhibitors (ICI). The patients were classified based on whether it was documented that they received platelet aggregation inhibition (PAI) (n = 137) (acetylsalicylic acid (ASA) or clopidogrel), anticoagulation (AC) (n = 185) (direct oral anticoagulation (DOAC), phenprocoumon, heparins) at the start of ICI or no antithrombotic medication (n = 2097) at any point during ICI treatment. The study endpoints were best overall response (BOR), progression-free survival (PFS) and overall survival (OS). Results: A significantly improved PFS was observed in patients documented to receive ASA (15.1 vs 6.4 months, HR 0.67, 95 % CI: 0.5 to 0.88, p = 0.0047) as well as in patients to receive AC (15.1 vs. 6.4 months, HR 0.7, 95 % CI: 0.53 to 0.91, p = 0.01) compared to patients for whom no antithrombotic medication was documented. Multivariate analysis of OS showed significant risk reduction in patients who received DOAC (HR 0.68, 95 % CI: 0.49 to 0.92, p = 0.0170) or phenprocoumon (HR: 0.44, 95 % CI: 0.19 to 0.85, p = 0.0301). Conclusion: Our study indicates a positive prognostic effect of anticoagulant and antiplatelet concomitant medication in melanoma patients receiving ICI. Further studies are needed to confrim the cancer-related benefit of adding anticoagulation or platelet inhibition to ICI treatment.

BackgroundAngioedema is a rare but potentially life-threatening adverse drug reaction in patients receiving angiotensin-converting enzyme inhibitors (ACEis). Research suggests that susceptibility to ACEi-induced angioedema (ACEi-AE) involves both genetic and nongenetic risk factors. Genome- and exome-wide studies of ACEi-AE have identified the first genetic risk loci. However, understanding of the underlying pathophysiology remains limited.ObjectiveWe sought to identify further genetic factors of ACEi-AE to eventually gain a deeper understanding of its pathophysiology.MethodsBy combining data from 8 cohorts, a genome-wide association study meta-analysis was performed in more than 1000 European patients with ACEi-AE. Secondary bioinformatic analyses were conducted to fine-map associated loci, identify relevant genes and pathways, and assess the genetic overlap between ACEi-AE and other traits. Finally, an exploratory cross-ancestry analysis was performed to assess shared genetic factors in European and African-American patients with ACEi-AE.ResultsThree genome-wide significant risk loci were identified. One of these, located on chromosome 20q11.22, has not been implicated previously in ACEi-AE. Integrative secondary analyses highlighted previously reported genes (BDKRB2 [bradykinin receptor B2] and F5 [coagulation factor 5]) as well as biologically plausible novel candidate genes (PROCR [protein C receptor] and EDEM2 [endoplasmic reticulum degradation enhancing alpha-mannosidase like protein 2]). Lead variants at the risk loci were found with similar effect sizes and directions in an African-American cohort.ConclusionsThe present results contributed to a deeper understanding of the pathophysiology of ACEi-AE by (1) providing further evidence for the involvement of bradykinin signaling and coagulation pathways and (2) suggesting, for the first time, the involvement of the fibrinolysis pathway in this adverse drug reaction. An exploratory cross-ancestry comparison implicated the relevance of the associated risk loci across diverse ancestries.

Supplementary Methods, Video Legends, Tables 1-2, Figures 1-8 from Cep63 Recruits Cdk1 to the Centrosome: Implications for Regulation of Mitotic Entry, Centrosome Amplification, and Genome Maintenance

Supplementary Figure 6 from Identification of Griseofulvin as an Inhibitor of Centrosomal Clustering in a Phenotype-Based Screen

Cancer of unknown primary has a dismal prognosis, especially following failure of platinum-based chemotherapy. 10-20% of patients have a high tumor mutational burden (TMB), which predicts response to immunotherapy in many cancer types. In this prospective, non-randomized, open-label, multicenter Phase II trial (EudraCT 2018-004562-33; NCT04131621), patients relapsed or refractory after platinum-based chemotherapy received nivolumab and ipilimumab following TMB high vs. TMB low stratification. Progression-free survival (PFS) represented the primary endpoint; overall survival (OS), response rates, duration of clinical benefit and safety were the secondary endpoints. The trial was prematurely terminated in March 2021 before reaching the preplanned sample size ( n = 194). Among 31 evaluable patients, 16% had a high TMB ( > 12 mutations/Mb). Overall response rate was 16% (95% CI 6-34%), with 7.7% (95% CI 1-25%) vs. 60% (95% CI 15-95%) in TMB low and TMB high , respectively. Although the primary endpoint was not met, high TMB was associated with better median PFS (18.3 vs. 2.4 months) and OS (18.3 vs. 3.6 months). Severe immune-related adverse events were reported in 29% of cases. Assessing on-treatment dynamics of circulating tumor DNA using combined targeted hotspot mutation and shallow whole genome sequencing as part of a predefined exploratory analysis identified patients benefiting from immunotherapy irrespective of initial radiologic response.

Background Despite the availability of effective systemic therapies, a significant number of advanced melanoma patients develops brain metastases. This study investigated differences in incidence and time to diagnosis of brain metastasis and survival outcomes dependent on the type of first-line therapy.Methods Patients with metastatic, non-resectable melanoma (AJCCv8 stage IIIC–V) without brain metastasis at start of first-line therapy (1L-therapy) were identified from the prospective multicenter real-world skin cancer registry ADOREG. Study endpoints were incidence of brain metastasis, brain metastasis-free survival (BMFS), progression-free survival (PFS), and overall survival (OS).Results Of 1704 patients, 916 were BRAF wild-type (BRAFwt) and 788 were BRAF V600 mutant (BRAFmut). Median follow-up time after start of 1L-therapy was 40.4 months. BRAFwt patients received 1L-therapy with immune checkpoint inhibitors (ICI) against CTLA-4+PD-1 (n=281) or PD-1 (n=544). In BRAFmut patients, 1L-therapy was ICI in 415 patients (CTLA-4+PD-1, n=108; PD-1, n=264), and BRAF+MEK targeted therapy (TT) in 373 patients. After 24 months, 1L-therapy with BRAF+MEK resulted in a higher incidence of brain metastasis compared with PD-1±CTLA-4 (BRAF+MEK, 30.3%; CTLA-4+PD-1, 22.2%; PD-1, 14.0%). In multivariate analysis, BRAFmut patients developed brain metastases earlier on 1L-therapy with BRAF+MEK than with PD-1±CTLA-4 (CTLA-4+PD-1: HR 0.560, 95% CI 0.332 to 0.945, p=0.030; PD-1: HR 0.575, 95% CI 0.372 to 0.888, p=0.013). Type of 1L-therapy, tumor stage, and age were independent prognostic factors for BMFS in BRAFmut patients. In BRAFwt patients, tumor stage was independently associated with longer BMFS; ECOG Performance status (ECOG-PS), lactate dehydrogenase (LDH), and tumor stage with OS. CTLA-4+PD-1 did not result in better BMFS, PFS, or OS than PD-1 in BRAFwt patients. For BRAFmut patients, multivariate Cox regression revealed ECOG-PS, type of 1L-therapy, tumor stage, and LDH as independent prognostic factors for PFS and OS. 1L-therapy with CTLA-4+PD-1 led to longer OS than PD-1 (HR 1.97, 95% CI 1.122 to 3.455, p=0.018) or BRAF+MEK (HR 2.41, 95% CI 1.432 to 4.054, p=0.001), without PD-1 being superior to BRAF+MEK.Conclusions In BRAFmut patients 1L-therapy with PD-1±CTLA-4 ICI resulted in a delayed and less frequent development of brain metastasis compared with BRAF+MEK TT. 1L-therapy with CTLA-4+PD-1 showed superior OS compared with PD-1 and BRAF+MEK. In BRAFwt patients, no differences in brain metastasis and survival outcomes were detected for CTLA-4+PD-1 compared with PD-1.

Supplementary Figure 6 from Identification of Griseofulvin as an Inhibitor of Centrosomal Clustering in a Phenotype-Based Screen

Supplementary Video 1 from Cep63 Recruits Cdk1 to the Centrosome: Implications for Regulation of Mitotic Entry, Centrosome Amplification, and Genome Maintenance

BACKGROUND:Melanomas frequently harbour somatic mutations in BRAF (40%) or NRAS (20%). Impact of NRAS mutations on the therapeutic outcome of immune checkpoint inhibitors (ICI) remains controversial. Potential correlation of the NRAS mutational status and programmed cell death ligand-1 (PD-L1) expression in melanoma is unknown. PATIENTS AND METHODS:Advanced, non-resectable melanoma patients with known NRAS mutation status treated with first-line ICI between 06/2014 and 05/2020 in the prospective multicenter skin cancer registry ADOREG were included. Overall response rate (ORR), progression-free survival (PFS), and overall survival (OS) according to NRAS status were analysed. A multivariate Cox model was used to analyse factors associated with PFS and OS; survival was analysed using the Kaplan-Meier approach. RESULTS:Among 637 BRAF wild-type patients, 310 (49%) had an NRAS mutation with Q61R (41%) and Q61K (32%). NRAS-mutated (NRASmut) melanomas were significantly more often located on the lower extremities and trunk (p = 0.001); nodular melanoma was the most common subtype (p < 0.0001). No significant differences were found for PFS and OS for anti-PD1 monotherapy (2-year PFS 39%, [95% confidence interval (CI), 33-47] in NRASmut patients and 41% [95% CI, 35-48] in NRAS-wild type (NRASwt) patients; 2-year OS was 54% [95% CI, 48-61] in NRASmut patients and 57% [95% CI, 50-64] in NRASwt patients) and anti-PD1 plus anti-CTLA4 therapy between both cohorts (2-year PFS was 54% [95% CI, 44-66] in NRASmut patients and 53% [95% CI, 41-67] in NRASwt patients; 2-year OS was 58% [95% CI, 49-70] in NRASmut patients and 62% [95% CI, 51-75] in NRASwt patients). The ORR to anti-PD1 was 35% for NRASwt patients and 26% for NRASmut patients and 34% compared to 32% for combinational therapy. Data on PD-L1 expression was available in 82 patients (13%). PD-L1 expression (>5%) was not correlated to NRAS mutational status. In multivariate analysis, elevated lactate dehydrogenase, Eastern Cooperative Oncology Group performance status ≥ 1, and brain metastases were significantly associated with a higher risk of death in all patients. CONCLUSIONS:The PFS and OS were not affected by NRAS mutational status in patients treated with anti-PD1-based ICI. Similar ORR was seen in NRASwt and NRASmut patients. Tumour PD-L1 expression did not correlate with NRAS mutational status.

In daily dermatological practice, the distinction between eczema and psoriasis in dermatoses of the hands can be difficult. However, a clear diagnosis is necessary to initiate optimal therapy and management. In recent years, the so-called molecular classifier has been developed for optimized differentiation of eczema and psoriasis. An occupational dermatological cohort has been established at Heidelberg University Hospital since 2020. It is funded by the German Statutory Accident Insurance. The aim is to follow-up patients over 3 years where this new diagnostic method is used and to compare the results with a retrospective occupational dermatological cohort. Recruitment ended in December 2022. The current analysis reports participants' occupational activity, insurance status, disease progression, and number of sick days. A total of 287 patients were included; mean age was 50.4 years and 63.5% (n = 181) were undergoing treatment at the expense of the liable statutory accident insurance at the start of the study. About 50% of the patients worked in health professions, metal industry, or construction. The average duration of occupational dermatosis was 6.5 years. In 38.9% of the patients, the clinical diagnosis had been classified as unclear by the treating dermatologist. By using the molecular classifier, the diagnosis could be clarified in 98% of the cases (eczema vs. psoriasis). The first analyses demonstrate that the molecular classifier contributes to improving therapy by optimizing the diagnosis.

Supplementary Video 2 from Cep63 Recruits Cdk1 to the Centrosome: Implications for Regulation of Mitotic Entry, Centrosome Amplification, and Genome Maintenance

In der täglichen dermatologischen Praxis kann die Unterscheidung von Ekzem und Psoriasis bei Dermatosen der Hände sehr schwierig sein. Zur Einleitung der optimalen Therapie und zum Management ist jedoch eine klare Diagnosestellung notwendig. In den vergangenen Jahren hat sich der sog. molekulare Klassifikator zur optimierten Differenzierung von Ekzem und Psoriasis entwickelt. Es wurde daher eine berufsdermatologische Kohorte am Universitätsklinikum Heidelberg im Rahmen der von der Deutschen Gesetzlichen Unfallversicherung geförderten FB323-Studie aufgebaut, bei der diese neue diagnostische Methode zum Einsatz kommt. Ziel ist dabei, in einer Langzeitbeobachtung mit einem Nachbeobachtungszeitraum von 3 Jahren die Kohorte zu verfolgen und die gewonnenen Ergebnisse mit einer retrospektiven, berufsdermatologischen Kohorte zu vergleichen. Die Rekrutierung wurde im Dezember 2022 beendet. Die aktuelle Analyse berichtet über die berufliche Tätigkeit der Studienteilnehmer, den versicherungsrechtlichen Status, den Krankheitsverlauf und die Anzahl der Krankheitsfehltage. Es wurden 287 Patienten eingeschlossen, das durchschnittliche Lebensalter betrug 50,4 Jahre; 63,5

Supplementary Video 2 from Cep63 Recruits Cdk1 to the Centrosome: Implications for Regulation of Mitotic Entry, Centrosome Amplification, and Genome Maintenance

2,4,6-Tris(dimethylaminomethyl)phenol (tris-DMP; CAS No. 90-72-2) is a hardener/accelerator frequently used in epoxy resin systems (ERS), especially at low ambient temperature.1 First cases of contact sensitization to tris-DMP have been observed at the Finnish Institute of Occupational Health (FIOH) around 1990.2 Since then, contact sensitization to tris-DMP has been described in several case reports from various European countries.1 In the FIOH, 14 cases of contact sensitization to tris-DMP have been registered from 1991 to 2013, rendering it the second most frequent sensitizer among epoxy resin hardeners.1 Since 2016, tris-DMP is available as patch test preparation at 0.5% in pet. from Chemotechnique, Vellinge, Sweden. The German Contact Dermatitis Research Group (DKG) included tris-DMP in the ‘adhesives and glues’ series and the ‘building trade’ series in October 2016. Tris-DMP 0.5% pet. has been patch tested following DKG guidelines3 in the departments of dermatology joining the Information Network of Departments of Dermatology (IVDK) in 5467 patients in the years 2017–2021. Results of these tests were retrospectively analysed. Patch test results are shown in Table 1. Positive reactions to tris-DMP occurred in 28 patients (0.5%). Seven positive reactions (25%) would have been missed if patch tests would have been read until Day 3 (D3) only. In six patients, positive reactions occurred at D4, and in one patient at D7. In two patients, a weak positive reaction (+) at D3 evolved into a strong positive reaction (++) at D4. Patch tests had been read until D4 or even D7 in 1964 patients (35.9%). Of the 28 tris-DMP-positive patients, 23 were males; the mean age was 47.0 years. Four of the tris-DMP positive patients were floor layers, four construction workers or bricklayers, three tile setters, and three painters. Additional details about those sensitized to tris-DMP, compared to those with a negative tris-DMP patch test, can be found in Table S1. Of the 28 tris-DMP positive patients, 27 were or had previously been patch tested with epoxy resin based on bisphenol A diglycidyl ether (DGEBA). Of these, 23 (85.2%) reacted to the DGEBA resin. In all, concomitant sensitizations to other ERS components occurred in 26 of the 28 tris-DMP positive patients (Table S2). For comparison, in the study period, sensitization to m-xylylenediamine (MXDA) was diagnosed in 63 of 5978 tested patients (1.1%), and to isophorone diamine (IPDA) in 32 out of 6097 patients (0.5%). Consistent with the FIOH data,1 we found sensitizations to tris-DMP less frequently than sensitizations to MXDA, but with a similar frequency as sensitizations to IPDA. Sensitization to tris-DMP without sensitization to other ERS components occurred less, but not significantly, frequently than in the FIOH (IVDK: 2/28 pet.; FIOH: 3/14 pet). However, 9 out of 14 FIOH patients (64%) sensitized to tris-DMP, but only 3 of the 28 tris-DMP positive IVDK patients (11%), were painters. Among our tris-DMP sensitized patients, there were more floor layers, bricklayers, construction workers, and tile setters. One quarter of the 28 positive reactions to tris-DMP in our study did not occur before D4. As only about one third of the patch tests have been read after D3, the true frequency of tris-DMP sensitization might have been higher. In the FIOH, late readings at D4, D5, or D6 had been done routinely,1 suggesting that their (higher) reported frequency of sensitization (1.9%) may be more realistic. However, it should be considered that far more occupational dermatitis patients are patch tested in the FIOH than in the IVDK,1 which may explain the higher proportion of positive reactions. According to information from ERS formulators, tris-DMP is used at higher concentrations in ERS designed for colder ambient temperatures. Thus, it may be that the products used by the Central European patients patch tested in the IVDK contained less tris-DMP than the products used by the Finnish patients tested in the FIOH. Unfortunately, this is not always evident from the safety data sheets, since usually no specific concentration, but rather a concentration range, is declared for tris-DMP. Summing up, we can confirm that tris-DMP is a relevant sensitizer in ERS. In order not to miss cases of sensitization, late readings at D4 and D7 should be done when patch testing with tris-DMP. Johannes Geier: Conceptualization; investigation; writing – original draft. Heinrich Dickel: Validation; writing – review and editing. Claudia Schröder-Kraft: Validation; writing – review and editing. Michal Gina: Validation; visualization; writing – review and editing. Dagmar Simon: Validation; writing – review and editing. Elke Weisshaar: Validation; writing – review and editing. Claudia Lang: Validation; writing – review and editing. Richard Brans: Validation; writing – review and editing. Andrea Bauer: Validation; writing – review and editing. Harald Löffler: Validation; writing – review and editing. Steffen Schubert: Validation; writing – review and editing; supervision; data curation. Open Access funding enabled and organized by Projekt DEAL. The authors declare no conflict of interest. Appendix S1. Supporting Information Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

ObjectivesForty to sixty percent of patients with advanced melanoma show primary resistance to PD-1-based immunotherapy, 30–40% of initial responders also progress. Here, we evaluated the outcome of second-line targeted therapy (TT) after progression on PD-1-based immune checkpoint inhibition (ICI) in BRAFV600-mutated melanoma. In addition, we report data on the activity of re-exposure with PD-1-based regimes.MethodsPatients with advanced (non-resectable stage III or IV, AJCC 2017, 8th edition) melanoma progressing on PD-1-based ICI (nivolumab, pembrolizumab or ipilimumab plus nivolumab) and receiving second-line BRAF plus MEK inhibition were identified from the prospective multicenter skin cancer registry ADOREG.ResultsWe identified 108 patients with unresectable stage III or stage IV melanoma progressing on first-line ICI (nivolumab, pembrolizumab or ipilimumab plus nivolumab) and receiving second-line combined BRAF/MEK inhibition. Seventy-three percent of the cohort presented with primary PD-1 resistant disease. Median progression-free survival (PFS) on ICI was 2.6 (95% CI 2.2–2.9) months. Median PFS on subsequent TT was 6.6 (95% CI 5.4–7.8) months. Median OS from start of second-line TT was 16.0 (95% CI 11.2–20.8) months. The 3-year PFS and OS rates on second-line TT were 16% and 30%. The objective response rate (ORR) and disease control rate (DCR) to TT were 42.6% and 55.6%. In patients with brain metastases, the ORR and DCR were 31.4% and 43.1%. Patients without brain metastases showed an ORR and DCR of 52.6% and 66.7%, respectively. Response to first-line ICI was associated with a numerically higher ORR and DCR to second-line TT and improved OS on TT. Twenty-three patients received third-line ICI of whom two patients showed an objective response.ConclusionsBRAF plus MEK inhibition shows meaningful activity and outcome in patients with advanced melanoma resistant to anti-PD-1-based immunotherapy. Rates of long-term benefit and survival in our study were similar to those reported for treatment-naïve patients receiving first-line MAPKi.

Angioedema is a relatively rare but potentially life-threatening adverse reaction to angiotensin-converting enzyme inhibitors (ACEi) and angiotensin receptor blockers (ARBs). As with hereditary forms of angioedema (HAE), this adverse reaction is mediated by bradykinin. Research suggests that ACEi/ARB-induced angioedema has a multifactorial etiology. In addition, recent case reports suggest that some ACEi/ARB-induced angioedema patients may carry pathogenic HAE variants. The aim of the present study was to investigate the possible association between ACEi/ARB-induced angioedema and HAE genes via systematic molecular genetic screening in a large cohort of ACEi/ARB-induced angioedema cases. Targeted re-sequencing of five HAE-associated genes ( SERPING1 , F12 , PLG , ANGPT1 , and KNG1 ) was performed in 212 ACEi/ARB-induced angioedema patients recruited in Germany/Austria, Sweden, and Denmark, and in 352 controls from a German cohort. Among patients, none of the identified variants represented a known pathogenic variant for HAE. Moreover, no significant association with ACEi/ARB-induced angioedema was found for any of the identified common [minor allele frequency (MAF) >5%] or rare (MAF < 5%) variants. However, several non-significant trends suggestive of possible protective effects were observed. The lowest p -value for an individual variant was found in PLG (rs4252129, p.R523W, p = 0.057, p .adjust > 0.999, Fisher’s exact test). Variant p.R523W was found exclusively in controls and has previously been associated with decreased levels of plasminogen, a precursor of plasmin which is part of a pathway directly involved in bradykinin production. In addition, rare, potentially functional variants (MAF < 5%, Phred-scaled combined annotation dependent depletion score >10) showed a nominally significant enrichment in controls both: 1) across all five genes; and 2) in the F12 gene alone. However, these results did not withstand correction for multiple testing. In conclusion, our results suggest that HAE-associated mutations are, at best, a rare cause of ACEi/ARB-induced angioedema. Furthermore, we were unable to identify a significant association between ACEi/ARB-induced angioedema and other variants in the investigated genes. Further studies with larger sample sizes are warranted to draw more definite conclusions concerning variants with limited effect sizes, including protective variants.