Treatment management for BRAF-mutant melanoma patients with tumor recurrence on adjuvant therapy: a multicenter study from the prospective skin cancer registry ADOREG.

BACKGROUND: Adjuvant therapy with immune-checkpoint inhibitors (CPI) or BRAF/MEK-directed targeted therapy (TT) improves recurrence-free survival (RFS) for patients with advanced, BRAFV600-mutant (BRAFmut) resected melanoma. However, 40% of these patients will develop distant metastases (DM) within 5 years, which require systemic therapy. Little data exist to guide the choice of upfront adjuvant therapy or treatment management upon DM. This study evaluated the efficacy of subsequent treatments following tumor recurrence upon upfront adjuvant therapy. METHODS: For this multicenter cohort study, we identified 515 BRAFmut patients with resected stage III melanoma who were treated with PD-1 inhibitors (anti-PD1) or TT in the adjuvant setting. Disease characteristics, treatment regimens, details on tumor recurrence, subsequent treatment management, and survival outcomes were collected within the prospective, real-world skin cancer registry ADOReg. Primary endpoints included progression-free survival (PFS) following DM and best tumor response to first-line (1L) treatments. RESULTS: Among 515 eligible patients, 273 patients received adjuvant anti-PD1 and 242 adjuvant TT. At a median follow-up of 21 months, 54.6% of anti-PD1 patients and 36.4% of TT patients recurred, while 39.6% (anti-PD1) and 29.3% (TT) developed DM. Risk of recurrence was significantly reduced in patients treated with TT compared with anti-PD1 (adjusted HR 0.52; 95% CI 0.40 to 0.68, p<0.001). Likewise, median RFS was significantly longer in TT-treated patients (31 vs 17 months, p<0.001). Patients who received TT as second adjuvant treatment upon locoregional recurrence had a longer RFS2 as compared with adjuvant CPI (41 vs 6 months, p=0.009). Patients who recurred at distant sites following adjuvant TT showed favorable response rates (42.9%) after switching to 1L ipilimumab+nivolumab (ipi+nivo). Patients with DM during adjuvant anti-PD1 achieved response rates of 58.7% after switching to 1L TT and 35.3% for 1L ipi+nivo. Overall, median PFS was significantly longer in patients who switched treatments for stage IV disease (median PFS 9 vs 5 months, p=0.004). CONCLUSIONS: BRAFmut melanoma patients who developed DM upon upfront adjuvant therapy achieve favorable tumor control and prolonged PFS after switching treatment modalities in the first-line setting of stage IV disease. Patients with locoregional recurrence benefit from complete resection of recurrence followed by a second adjuvant treatment with TT.

Response to primary chemoradiotherapy of locally advanced oropharyngeal carcinoma is determined by the degree of cytotoxic T cell infiltration within tumor cell aggregates.

Background: Effective anti-tumor immune responses are mediated by T cells and require organized, spatially coordinated interactions within the tumor microenvironment (TME). Understanding coordinated T-cell-behavior and deciphering mechanisms of radiotherapy resistance mediated by tumor stem cells will advance risk stratification of oropharyngeal cancer (OPSCC) patients treated with primary chemoradiotherapy (RCTx). Methods: To determine the role of CD8 T cells (CTL) and tumor stem cells for response to RCTx, we employed multiplex immunofluorescence stains on pre-treatment biopsy specimens from 86 advanced OPSCC patients and correlated these quantitative data with clinical parameters. Multiplex stains were analyzed at the single-cell level using QuPath and spatial coordination of immune cells within the TME was explored using the R-package Spatstat. Results: Our observations demonstrate that a strong CTL-infiltration into the epithelial tumor compartment (HR for overall survival, OS: 0.35; p<0.001) and the expression of PD-L1 on CTL (HR: 0.36; p<0.001) were both associated with a significantly better response and survival upon RCTx. As expected, p16 expression was a strong predictor of improved OS (HR: 0.38; p=0.002) and correlated with overall CTL infiltration (r: 0.358, p<0.001). By contrast, tumor cell proliferative activity, expression of the tumor stem cell marker CD271 and overall CTL infiltration, regardless of the affected compartment, were not associated with response or survival. Conclusion: In this study, we could demonstrate the clinical relevance of the spatial organization and the phenotype of CD8 T cells within the TME. In particular, we found that the infiltration of CD8 T cells specifically into the tumor cell compartment was an independent predictive marker for response to chemoradiotherapy, which was strongly associated with p16 expression. Meanwhile, tumor cell proliferation and the expression of stem cell markers showed no independent prognostic effect for patients with primary RCTx and thus requires further study.

Combination of immune-checkpoint inhibitors and targeted therapies for melanoma therapy: The more, the better?

The approval of immune-checkpoint inhibitors (CPI) and mitogen activated protein kinase inhibitors (MAPKi) in recent years significantly improved the treatment management and survival of patients with advanced malignant melanoma. CPI aim to counter-act receptor-mediated inhibitory effects of tumor cells and immunomodulatory cell types on effector T cells, whereas MAPKi are intended to inhibit tumor cell survival. In agreement with these complementary modes of action preclinical data indicated that the combined application of CPI and MAPKi or their optimal sequencing might provide additional clinical benefit. In this review the rationale and preclinical evidence that support the combined application of MAPKi and CPI either in concurrent or consecutive regimens are presented. Further, we will discuss the results from clinical trials investigating the sequential or combined application of MAPKi and CPI for advanced melanoma patients and their implications for clinical practice. Finally, we outline mechanisms of MAPKi and CPI cross-resistance which limit the efficacy of currently available treatments, as well as combination regimens.

Impaired Treg-DC interactions contribute to autoimmunity in leukocyte adhesion deficiency type 1.

Leukocyte adhesion deficiency type 1 (LAD-1) is a rare disease resulting from mutations in the gene encoding for the common ß-chain of the ß2-integrin family (CD18). The most prominent clinical symptoms are profound leukocytosis and high susceptibility to infections. Patients with LAD-1 are prone to develop autoimmune diseases, but the molecular and cellular mechanisms that result in coexisting immunodeficiency and autoimmunity are still unresolved. CD4+FOXP3+ Treg are known for their essential role in preventing autoimmunity. To understand the role of Treg in LAD-1 development and manifestation of autoimmunity, we generated mice specifically lacking CD18 on Treg (CD18Foxp3), resulting in defective LFA-1 expression. Here, we demonstrate a crucial role of LFA-1 on Treg to maintain immune homeostasis by modifying T cell-DC interactions and CD4+ T cell activation. Treg-specific CD18 deletion did not impair Treg migration into extralymphatic organs, but it resulted in shorter interactions of Treg with DC. In vivo, CD18Foxp3 mice developed spontaneous hyperplasia in lymphatic organs and diffuse inflammation of the skin and in multiple internal organs. Thus, LFA-1 on Treg is required for the maintenance of immune homeostasis.

Response to First-Line Treatment with Immune-Checkpoint Inhibitors in Patients with Advanced Cutaneous Squamous Cell Carcinoma: A Multicenter, Retrospective Analysis from the German ADOReg Registry.

Cutaneous squamous cell carcinoma (cSCC) is a common malignancy of the skin and has an overall favorable outcome, except for patients with an advanced stage of the disease. The efficacy of checkpoint inhibitors (CPI) for advanced cSCC has been demonstrated in recent clinical studies, but data from real-world cohorts and trial-ineligible cSCC patients are limited. We retrospectively investigated patients with advanced cSCC who have been treated with CPI in a first-line setting at eight German skin cancer centers registered within the multicenter registry ADOReg. Clinical outcome parameters including response, progression-free (PFS) and overall survival (OS), time-to-next-treatment (TTNT), and toxicity were analyzed and have been stratified by the individual immune status. Among 39 evaluable patients, the tumor response rate (rwTRR) was 48.6%, the median PFS was 29.0 months, and the median OS was not reached. In addition, 9 patients showed an impaired immune status due to immunosuppressive medication or hematological diseases. Our data demonstrated that CPI also evoked tumor responses among immunocompromised patients (rwTRR: 48.1 vs. 50.0%), although these responses less often resulted in durable remissions. In line with this, the median PFS (11 vs. 40 months, p = 0.059), TTNT (12 months vs. NR, p = 0.016), and OS (29 months vs. NR, p < 0.001) were significantly shorter for this patient cohort. CPI therapy was well tolerated in both subcohorts with 15% discontinuing therapy due to toxicity. Our real-world data show that first-line CPI therapy produced strong and durable responses among patients with advanced cSCC. Immunocompromised patients were less likely to achieve long-term benefit from anti-PD1 treatment, despite similar tumor response rates.

The Role of the Immune Phenotype in Tumor Progression and Prognosis of Patients with Mycosis Fungoides: A Quantitative Immunohistology Whole Slide Approach.

BACKGROUND AND OBJECTIVES: Mycosis fungoides (MF) is the most common type of cutaneous T-cell lymphomas, characterized by mature, skin-tropic CD4+ T-helper cells. In order to study the immune tumor microenvironment in MF patients, we performed immunohistochemical stains on MF biopsies, digitized whole-slide tissue sections, and performed quantitative analysis of the different immune cell subsets to correlate tissue parameters with the clinical data of patients, such as progression-free survival or overall survival. PATIENTS AND METHODS: Overall, 35 patients who were treated between 2009 and 2019 and for whom one or more paraffin tissue blocks were available have been included in the present study (58 tissue specimens in total). Conventional immunohistochemistry stains for CD3, CD4, CD8, CD20 and CD30 were used for the analysis of the immune phenotype, and quantitative analysis was performed using QuPath as a quantitative digital pathology tool for bioimage analysis of whole slides. RESULTS: Analysis of tissue parameters for prognostic significance revealed that patients with a stronger infiltration by CD8+ lymphocytes within the tumor cell compartment had a higher risk of disease progression (p = 0.031) and showed a shorter progress-free survival (p = 0.038). Furthermore, a significant association of the percentage of CD30+ cells (median: 7.8%) with the risk of disease progression (p = 0.023) and progression-free survival (p = 0.023) was found. In relation to the clinical features of our patient cohort, a higher risk of disease progression (p = 0.015) and a shorter progression-free survival (p = 0.032) for older patients (>61 years) were observed. CONCLUSIONS: Our results demonstrated the prognostic relevance of large-cell transformation in mycosis fungoides and its strong association with the presence of CD30+ lymphocytes. Unlike previous reports, our study suggests an adverse prognostic role for CD8+ T cells in patients with mycosis fungoides. Moreover, our data indicate that the immune phenotype within the tumor microenvironment shows strong temporal heterogeneity and is altered in the course of tumor progression.

Protease- and cell type-specific activation of protease-activated receptor 2 in cutaneous inflammation.

BACKGROUND: Protease-activated receptor 2 (PAR2) signaling controls skin barrier function and inflammation, but the roles of immune cells and PAR2-activating proteases in cutaneous diseases are poorly understood. OBJECTIVE: To dissect PAR2 signaling contributions to skin inflammation with new genetic and pharmacological tools. METHODS/RESULTS: We found markedly increased numbers of PAR2+ infiltrating myeloid cells in skin lesions of allergic contact dermatitis (ACD) patients and in the skin of contact hypersensitivity (CHS) in mice, a murine ACD model for T cell-mediated allergic skin inflammation. Cell type-specific deletion of PAR2 in myeloid immune cells as well as mutation-induced complete PAR2 cleavage insensitivity significantly reduced skin inflammation and hapten-specific Tc1/Th1 cell response. Pharmacological approaches identified individual proteases involved in PAR2 cleavage and demonstrated a pivotal role of tissue factor (TF) and coagulation factor Xa (FXa) as upstream activators of PAR2 in both the induction and effector phase of CHS. PAR2 mutant mouse strains with differential cleavage sensitivity for FXa versus skin epithelial cell-expressed proteases furthermore uncovered a time-dependent regulation of CHS development with an important function of FXa-induced PAR2 activation during the late phase of skin inflammation. CONCLUSIONS: Myeloid cells and the TF-FXa-PAR2 axis are key mediators and potential therapeutic targets in inflammatory skin diseases.

ß2 Integrins on Dendritic Cells Modulate Cytokine Signaling and Inflammation-Associated Gene Expression, and Are Required for Induction of Autoimmune Encephalomyelitis.

Heterodimeric ß2 integrin surface receptors (CD11a-d/CD18) are specifically expressed by leukocytes that contribute to pathogen uptake, cell migration, immunological synapse formation and cell signaling. In humans, the loss of CD18 expression results in leukocyte adhesion deficiency syndrome (LAD-)1, largely characterized by recurrent severe infections. All available mouse models display the constitutive and ubiquitous knockout of either α or the common ß2 (CD18) subunit, which hampers the analysis of the cell type-specific role of ß2 integrins in vivo. To overcome this limitation, we generated a CD18 gene floxed mouse strain. Offspring generated from crossing with CD11c-Cre mice displayed the efficient knockdown of ß2 integrins, specifically in dendritic cells (DCs). Stimulated ß2-integrin-deficient splenic DCs showed enhanced cytokine production and the concomitantly elevated activity of signal transducers and activators of transcription (STAT) 1, 3 and 5, as well as the impaired expression of suppressor of cytokine signaling (SOCS) 2-6 as assessed in bone marrow-derived (BM) DCs. Paradoxically, these BMDCs also showed the attenuated expression of genes involved in inflammatory signaling. In line, in experimental autoimmune encephalomyelitis mice with a conditional DC-specific ß2 integrin knockdown presented with a delayed onset and milder course of disease, associated with lower frequencies of T helper cell populations (Th)1/Th17 in the inflamed spinal cord. Altogether, our mouse model may prove to be a valuable tool to study the leukocyte-specific functions of ß2 integrins in vivo.

eHealth Literacy in German Skin Cancer Patients.

The global incidence of skin cancer has steadily increased in recent years, and malignant melanoma still has one of the fastest-growing incidence rates among all malignant tumors in the western world. Thus, newly diagnosed patients have an increased need for health information concerning their disease. Using a standardized questionnaire, our study aims to investigate our patients' primary sources of health-related information as well as their self-proclaimed eHealth literacy. We received 714 questionnaires. Regardless of age, the primary source of information was the treating dermato-oncologist, followed by the treating general practitioner and the Internet. However, with increasing age, the usage of the Internet decreased. Hence, younger participants were better equipped to find health-related information while using the Internet. Additionally, comprehending health-related information and gaining medical knowledge was significantly increased in better-educated participants. Overall, our study shows that with increased use of eHealth services, accessing web-based information increased, correlating with a better eHealth literacy of our patients. eHealth technologies are increasingly becoming more prevalent as a primary source of information in our modern health care system. Thus, it is crucial to educate cancer patients in eHealth literacy to make autonomous, informed decisions and gain more confidence in dealing with their disease.

Neutrophil-Specific Knockdown of ß2 Integrins Impairs Antifungal Effector Functions and Aggravates the Course of Invasive Pulmonal Aspergillosis.

ß2-integrins are heterodimeric surface receptors that are expressed specifically by leukocytes and consist of a variable α (CD11a-d) and a common ß-subunit (CD18). Functional impairment of CD18, which causes leukocyte adhesion deficiency type-1 results in an immunocompromised state characterized by severe infections, such as invasive pulmonary aspergillosis (IPA). The underlying immune defects have largely been attributed to an impaired migratory and phagocytic activity of polymorphonuclear granulocytes (PMN). However, the exact contribution of ß2-integrins for PMN functions in-vivo has not been elucidated yet, since the mouse models available so far display a constitutive CD18 knockout (CD18-/- or CD18hypo). To determine the PMN-specific role of ß2-integrins for innate effector functions and pathogen control, we generated a mouse line with a Ly6G-specific knockdown of the common ß-subunit (CD18Ly6G cKO). We characterized CD18Ly6G cKO mice in-vitro to confirm the PMN-specific knockdown of ß2-integrins. Next, we investigated the clinical course of IPA in A. fumigatus infected CD18Ly6G cKO mice with regard to the fungal burden, pulmonary inflammation and PMN response towards A. fumigatus. Our results revealed that the ß2-integrin knockdown was restricted to PMN and that CD18Ly6G cKO mice showed an aggravated course of IPA. In accordance, we observed a higher fungal burden and lower levels of proinflammatory innate cytokines, such as TNF-α, in lungs of IPA-infected CD18Ly6G cKO mice. Bronchoalveolar lavage revealed higher levels of CXCL1, a stronger PMN-infiltration, but concomitantly elevated apoptosis of PMN in lungs of CD18Ly6G cKO mice. Ex-vivo analysis further unveiled a strong impairment of PMN effector function, as reflected by an attenuated phagocytic activity, and a diminished generation of reactive oxygen species (ROS) and neutrophil-extracellular traps (NET) in CD18-deficient PMN. Overall, our study demonstrates that ß2-integrins are required specifically for PMN effector functions and contribute to the clearance of A. fumigatus by infiltrating PMN, and the establishment of an inflammatory microenvironment in infected lungs.

Nanodrugs Targeting T Cells in Tumor Therapy.

In contrast to conventional anti-tumor agents, nano-carriers allow co-delivery of distinct drugs in a cell type-specific manner. So far, many nanodrug-based immunotherapeutic approaches aim to target and kill tumor cells directly or to address antigen presenting cells (APC) like dendritic cells (DC) in order to elicit tumor antigen-specific T cell responses. Regulatory T cells (Treg) constitute a major obstacle in tumor therapy by inducing a pro-tolerogenic state in APC and inhibiting T cell activation and T effector cell activity. This review aims to summarize nanodrug-based strategies that aim to address and reprogram Treg to overcome their immunomodulatory activity and to revert the exhaustive state of T effector cells. Further, we will also discuss nano-carrier-based approaches to introduce tumor antigen-specific chimeric antigen receptors (CAR) into T cells for CAR-T cell therapy which constitutes a complementary approach to DC-focused vaccination.

The Role of Treatment Sequencing with Immune-Checkpoint Inhibitors and BRAF/MEK Inhibitors for Response and Survival of Patients with BRAFV600-Mutant Metastatic Melanoma-A Retrospective, Real-World Cohort Study.

The advent of immune-checkpoint inhibitors (CPI) and BRAF/MEK-directed targeted therapy (TT) has improved the treatment landscape of patients with BRAFV600-mutant metastatic melanoma. While TT allows for rapid disease control, the development of secondary TT resistance limits the duration of responses. Responses to CPI have a slower onset but can be durable in a subset of patients. To date, little prospective data is available for the optimal sequencing of these agents in melanoma patients. In this retrospective, single-center, real-world analysis, we identified 135 patients with BRAF-mutated, metastatic melanoma who received consecutive treatment with TT followed by CPI, or vice versa, as first and second-line therapy, respectively. We collected data on clinical-pathological factors, treatment duration, best overall response, progression-free survival and overall survival (OS). Our data revealed that front-line treatment with CPI, followed by TT, showed a non-significant trend towards better OS compared to front-line TT (median OS: 35.0 vs. 18.0 months, p = 0.070). This association was confirmed in a subgroup of patients without systemic pre-treatments (median OS: 41.0 vs. 14.0 months, p = 0.02). Further, we observed significantly better objective response rates to second-line treatments for patients receiving front-line CPI (18.4 vs. 37.8%, p = 0.024). Last, our results indicated that rapid disease progression was less common in patients treated with front-line CPI (27.6% vs. 16.2%) and that subsequent treatment with TT resulted in favorable survival outcomes. Our real-world data indicate that sequential treatment with front-line CPI is associated with favorable tumor control and overall survival in a subgroup of previously untreated BRAF-mutant metastatic melanoma patients.

Immunomodulatory Properties of Immune Checkpoint Inhibitors-More than Boosting T-Cell Responses?

The approval of immune checkpoint inhibitors (ICI) that serve to enhance effector T-cell anti-tumor responses has strongly improved success rates in the treatment of metastatic melanoma and other tumor types. The currently approved ICI constitute monoclonal antibodies blocking cytotoxic T-lymphocyte-associated protein (CTLA)-4 and anti-programmed cell death (PD)-1. By this, the T-cell-inhibitory CTLA-4/CD80/86 and PD-1/PD-1L/2L signaling axes are inhibited. This leads to sustained effector T-cell activity and circumvents the immune evasion of tumor cells, which frequently upregulate PD-L1 expression and modulate immune checkpoint molecule expression on leukocytes. As a result, profound clinical responses are observed in 40-60% of metastatic melanoma patients. Despite the pivotal role of T effector cells for triggering anti-tumor immunity, mounting evidence indicates that ICI efficacy may also be attributable to other cell types than T effector cells. In particular, emerging research has shown that ICI also impacts innate immune cells, such as myeloid cells, natural killer cells and innate lymphoid cells, which may amplify tumoricidal functions beyond triggering T effector cells, and thus improves clinical efficacy. Effects of ICI on non-T cells may additionally explain, in part, the character and extent of adverse effects associated with treatment. Deeper knowledge of these effects is required to further develop ICI treatment in terms of responsiveness of patients to treatment, to overcome resistance to ICI and to alleviate adverse effects. In this review we give an overview into the currently known immunomodulatory effects of ICI treatment in immune cell types other than the T cell compartment.

Systemically Administered TLR7/8 Agonist and Antigen-Conjugated Nanogels Govern Immune Responses against Tumors.

The generation of specific humoral and cellular immune responses plays a pivotal role in the development of effective vaccines against tumors. Especially the presence of antigen-specific, cytotoxic T cells influences the outcome of therapeutic cancer vaccinations. Different strategies, ranging from delivering antigen-encoding mRNAs to peptides or full antigens, are accessible but often suffer from insufficient immunogenicity and require immune-boosting adjuvants as well as carrier platforms to ensure stability and adequate retention. Here, we introduce a pH-responsive nanogel platform as a two-component antitumor vaccine that is safe for intravenous application and elicits robust immune responses in vitro and in vivo. The underlying chemical design allows for straightforward covalent attachment of a model antigen (ovalbumin) and an immune adjuvant (imidazoquinoline-type TLR7/8 agonist) onto the same nanocarrier system. In addition to eliciting antigen-specific T and B cell responses that outperform mixtures of individual components, our two-component nanovaccine leads in prophylactic and therapeutic studies to an antigen-specific growth reduction of different tumors expressing ovalbumin intracellularly or on their surface. Regarding the versatile opportunities for functionalization, our nanogels are promising for the development of highly customized and potent nanovaccines.

Anticoagulation with Factor Xa Inhibitors Is Associated with Improved Overall Response and Progression-Free Survival in Patients with Metastatic Malignant Melanoma Receiving Immune Checkpoint Inhibitors-A Retrospective, Real-World Cohort Study.

Immune checkpoint inhibitors (ICI) significantly improved the prognosis of advanced melanoma patients. However, many patients do not derive long-term benefit from ICI therapy due to primary and acquired resistance. In this regard, it has been shown that coagulation factors contribute to cancer immune evasion and might therefore promote resistance to ICI. In particular, recent observations in murine systems demonstrated that myeloid-derived factor Xa (FXa) impedes anti-tumor immunity in the tumor microenvironment and that the oral FXa inhibitor (FXa-i) rivaroxaban synergizes with ICI. The synergistic effect of FXa inhibitors with clinical ICI therapy is unknown. We performed a retrospective study of 280 metastatic melanoma patients who were treated with ICI and stratified them for concomitant anticoagulation (AC) by medical chart review. Data on baseline patient characteristics, specific AC treatment, ICI therapy, other tumor-targeting therapies, and clinical outcomes were analyzed. Of 280 patients who received ICI, 76 received concomitant AC during initial ICI therapy. Patients on AC were treated either with heparins (n = 29), vitamin K antagonists (VKA) (n = 20), or FXa-i (n = 27). Patients requiring AC during ICI therapy showed no significantly reduced objective response rate (ORR) (p = 0.27), or progression-free (PFS; median PFS 4 vs. 4 months; p = 0.71) or overall survival (OS; median OS: 39 vs. 51 months; p = 0.31). Furthermore, patients who underwent AC did not show significantly more bleeding complications (p = 0.605) than those who were not anticoagulated. Remarkably, stratification of patients by the class of AC revealed that patients receiving FXa-i were more likely to obtain CR (26.9 vs. 12.6%, p = 0.037), and showed better ORR (69.2 vs. 36.4%, p = 0.005), PFS (median PFS: 12 months vs. 3 months; p = 0.006), and OS (median OS not reached vs. 42 months; p = 0.09) compared to patients not receiving FXa-i. Patient demographics and tumor characteristics in this patient subcohort did not significantly differ from patients not on FXa-i. In summary, our study provides first clinical evidence that the clinical application of FXa-i may enhance the efficacy of ICI therapy via the restoration of anti-tumor immunity, while patients who received FXa-i were not more likely to encounter bleeding complications.

Immunomodulatory Properties of BRAF and MEK Inhibitors Used for Melanoma Therapy-Paradoxical ERK Activation and Beyond.

The advent of mitogen-activated protein kinase (MAPK) inhibitors that directly inhibit tumor growth and of immune checkpoint inhibitors (ICI) that boost effector T cell responses have strongly improved the treatment of metastatic melanoma. In about half of all melanoma patients, tumor growth is driven by gain-of-function mutations of BRAF (v-rat fibrosarcoma (Raf) murine sarcoma viral oncogene homolog B), which results in constitutive ERK activation. Patients with a BRAF mutation are regularly treated with a combination of BRAF and MEK (MAPK/ERK kinase) inhibitors. Next to the antiproliferative effects of BRAF/MEKi, accumulating preclinical evidence suggests that BRAF/MEKi exert immunomodulatory functions such as paradoxical ERK activation as well as additional effects in non-tumor cells. In this review, we present the current knowledge on the immunomodulatory functions of BRAF/MEKi as well as the non-intended effects of ICI and discuss the potential synergistic effects of ICI and MAPK inhibitors in melanoma treatment.

The Status of Adjuvant and Neoadjuvant Melanoma Therapy, New Developments and Upcoming Challenges.

The global incidence of malignant melanoma, the leading cause of skin cancer death, has steadily increased in recent years. Surgical excision is the treatment of choice for early-stage melanoma. However, 40-60% of patients with high-risk melanoma or with nodal involvement eventually experience loco-regional relapse or tumor progression. Adjuvant therapy aims to reduce the rate of recurrence in radically operated high-risk patients with melanoma and thus improves survival. Interferon-α has long been the only approved drug for adjuvant melanoma therapy, despite an unclear survival benefit. The landmark success of immune-checkpoint inhibitors and BRAF/MEK-directed targeted therapies in the treatment of patients with stage IV melanoma led to the initiation of clinical trials in the adjuvant setting. These trials demonstrated the efficacy of immune-checkpoint inhibitors and targeted therapies for the adjuvant treatment of high-risk patients with melanoma, as shown both by an increase in recurrence-free survival and the emergence of long-term survivors, finally resulting in the approval of the cytotoxic T-lymphocyte antigen 4 inhibitor ipilimumab, PD1 inhibitors (nivolumab, pembrolizumab), and BRAF/MEK inhibitors for adjuvant melanoma therapy. This review aims to delineate the advances in adjuvant melanoma therapy, issuing particularly recent results from clinical trials. Moreover, we also discuss pending issues and future challenges, which comprise the adequate selection of adjuvant regimens for patient subgroups and the identification of markers likely to predict the individual response to adjuvant treatments. Last, we outline the role of emerging neoadjuvant approaches, which may complement adjuvant strategies and are currently investigated in clinical trials.

Discontinuation of BRAF/MEK-Directed Targeted Therapy after Complete Remission of Metastatic Melanoma-A Retrospective Multicenter ADOReg Study.

The advent of BRAF/MEK inhibitors (BRAFi/MEKi) has significantly improved progression-free (PFS) and overall survival (OS) for patients with advanced BRAF-V600-mutant melanoma. Long-term survivors have been identified particularly among patients with a complete response (CR) to BRAF/MEK-directed targeted therapy (TT). However, it remains unclear which patients who achieved a CR maintain a durable response and whether treatment cessation might be a safe option in these patients. Therefore, this study investigated the impact of treatment cessation on the clinical course of patients with a CR upon BRAF/MEK-directed-TT. We retrospectively selected patients with BRAF-V600-mutant advanced non-resectable melanoma who had been treated with BRAFi ± MEKi therapy and achieved a CR upon treatment out of the multicentric skin cancer registry ADOReg. Data on baseline patient characteristics, duration of TT, treatment cessation, tumor progression (TP) and response to second-line treatments were collected and analyzed. Of 461 patients who received BRAF/MEK-directed TT 37 achieved a CR. TP after initial CR was observed in 22 patients (60%) mainly affecting patients who discontinued TT (n = 22/26), whereas all patients with ongoing TT (n = 11) maintained their CR. Accordingly, patients who discontinued TT had a higher risk of TP compared to patients with ongoing treatment (p < 0.001). However, our data also show that patients who received TT for more than 16 months and who discontinued TT for other reasons than TP or toxicity did not have a shorter PFS compared to patients with ongoing treatment. Response rates to second-line treatment being initiated in 21 patients, varied between 27% for immune-checkpoint inhibitors (ICI) and 60% for BRAFi/MEKi rechallenge. In summary, we identified a considerable number of patients who achieved a CR upon BRAF/MEK-directed TT in this contemporary real-world cohort of patients with BRAF-V600-mutant melanoma. Sustained PFS was not restricted to ongoing TT but was also found in patients who discontinued TT.

The Functional Crosstalk between Myeloid-Derived Suppressor Cells and Regulatory T Cells within the Immunosuppressive Tumor Microenvironment.

Immune checkpoint inhibitors (ICI) have led to profound and durable tumor regression in some patients with metastatic cancer diseases. However, many patients still do not derive benefit from immunotherapy. Here, the accumulation of immunosuppressive cell populations within the tumor microenvironment (TME), such as myeloid-derived suppressor cells (MDSC), tumor-associated macrophages (TAM), and regulatory T cells (Treg), contributes to the development of immune resistance. MDSC and Treg expand systematically in tumor patients and inhibit T cell activation and T effector cell function. Numerous studies have shown that the immunosuppressive mechanisms exerted by those inhibitory cell populations comprise soluble immunomodulatory mediators and receptor interactions. The latter are also required for the crosstalk of MDSC and Treg, raising questions about the relevance of cell-cell contacts for the establishment of their inhibitory properties. This review aims to outline the current knowledge on the crosstalk between these two cell populations, issuing particularly the potential role of cell adhesion molecules. In this regard, we further discuss the relevance of ß2 integrins, which are essential for the differentiation and function of leukocytes as well as for MDSC-Treg interaction. Lastly, we aim to describe the impact of such bidirectional crosstalk for basic and applied cancer research and discuss how the targeting of these pathways might pave the way for future approaches in immunotherapy.

Role of Hypoxia and the Adenosine System in Immune Evasion and Prognosis of Patients with Brain Metastases of Melanoma: A Multiplex Whole Slide Immunofluorescence Study.

Following the introduction of immune checkpoint inhibitors, a substantial prolongation of the overall survival has been achieved for many patients with multiple brain metastases from melanoma. However, heterogeneity between individual tumor responses is incompletely understood. In order to determine the impact of the individual tumor phenotype on the prognosis of melanoma patients, we examined surgical sections from 33 patients who were treated with radiotherapy (whole-brain radiotherapy, WBRT, stereotactic radiotherapy, STX, or both) and Ipilimumab. We analyzed multiplex staining of the hypoxia marker GLUT-1, the adenosine (ADO)-associated enzymes CD73 and CD39, and CD8, a marker of cytotoxic T lymphocytes (CTL) on a single-cell basis using QuPath. Additionally, the MOSAIC interaction analysis algorithm was used to explore the hypothesis that CTL systematically avoid GLUT-1high tumor areas. Our results revealed, that a strong GLUT-1 expression, low numbers of CTL, or exclusion of CTL from the tumor were correlated with significant prognostic detriment. Hypoxic tumors overall have smaller amounts of CTL, and spatial analysis revealed a repellent effect of hypoxia on CTL. In contrast to in vitro studies, specific upregulation of ADO-related enzymes CD73 and CD39 in GLUT-1high tumor regions was never observed. In this study, we could show direct in vivo evidence for hypoxia-mediated immunosuppression in melanoma. Moreover, this study suggests a significant prognostic relevance of the tumor immune phenotype, the strength of CD8 infiltration in the tumor, and the expression of hypoxia marker GLUT-1 on melanoma cells. Last, our results suggest a temporal stability of the microenvironment-mediated immunosuppressive phenotype in melanoma.