Peter Arne Gerber

Epidermal EGFR Controls Cutaneous Host Defense and Prevents Inflammation

Epidermal EGFR regulates skin inflammation and contributes to skin barrier function and host defense. Skin-Deep Search for the Effects of EGFR Inhibitors The goal of all medical interventions is to treat disease while minimizing the damage to healthy tissues in the body. This can be difficult to achieve for cancer drugs, however, especially when the effectiveness of a drug directly correlates with its side effects, as is the case for inhibitors of the epidermal growth factor receptor (EGFR). EGFR inhibitors are particularly known for causing a severe rash and skin damage, which sometimes forces patients to prematurely stop their treatments. Now, two papers by Mascia et al. and Lichtenberger et al. help clarify the mechanism of rash formation induced by EGFR inhibitors and uncover some of the skin components that contribute to this phenomenon. Both sets of authors used mouse models that lack EGFR only in the skin to replicate the pattern of injury seen in patients treated with EGFR inhibitors. They characterized the changes in chemokine expression in the skin of treated patients and study animals and examined the effects of EGFR inhibition on skin defenses and bacteria. They also investigated the effects of crossing mice that lack EGFR in the skin with mice deficient in different immune pathways and immune cell types to determine which ones are necessary for the rash phenotype. The findings of these two studies suggest that EGFR signaling is important for normal skin barrier function and antimicrobial defense, and that skin macrophages may contribute to the adverse effects of EGFR inhibitors. Additional work will be necessary to further expand our understanding of EGFR inhibitor toxicity and to continue the search for ways to prevent this disruptive side effect. The current studies provide mechanistic insights that should help guide further investigation in this area. The epidermal growth factor receptor (EGFR) plays an important role in tissue homeostasis and tumor progression. However, cancer patients treated with EGFR inhibitors (EGFRIs) frequently develop acneiform skin toxicities, which are a strong predictor of a patient’s treatment response. We show that the early inflammatory infiltrate of the skin rash induced by EGFRI is dominated by dendritic cells, macrophages, granulocytes, mast cells, and T cells. EGFRIs induce the expression of chemokines (CCL2, CCL5, CCL27, and CXCL14) in epidermal keratinocytes and impair the production of antimicrobial peptides and skin barrier proteins. Correspondingly, EGFRI-treated keratinocytes facilitate lymphocyte recruitment but show a considerably reduced cytotoxic activity against Staphylococcus aureus. Mice lacking epidermal EGFR (EGFRΔep) show a similar phenotype, which is accompanied by chemokine-driven skin inflammation, hair follicle degeneration, decreased host defense, and deficient skin barrier function, as well as early lethality. Skin toxicities were not ameliorated in a Rag2-, MyD88-, and CCL2-deficient background or in mice lacking epidermal Langerhans cells. The skin phenotype was also not rescued in a hairless (hr/hr) background, demonstrating that skin inflammation is not induced by hair follicle degeneration. Treatment with mast cell inhibitors reduced the immigration of T cells, suggesting that mast cells play a role in the EGFRI-mediated skin pathology. Our findings demonstrate that EGFR signaling in keratinocytes regulates key factors involved in skin inflammation, barrier function, and innate host defense, providing insights into the mechanisms underlying EGFRI-induced skin pathologies.

Tumor immune escape by the loss of homeostatic chemokine expression

The novel keratinocyte-specific chemokine CCL27 plays a critical role in the organization of skin-associated immune responses by regulating T cell homing under homeostatic and inflammatory conditions. Here we demonstrate that human keratinocyte-derived skin tumors may evade T cell-mediated antitumor immune responses by down-regulating the expression of CCL27 through the activation of epidermal growth factor receptor (EGFR)–Ras–MAPK-signaling pathways. Compared with healthy skin, CCL27 mRNA and protein expression was progressively lost in transformed keratinocytes of actinic keratoses and basal and squamous cell carcinomas. In vivo, precancerous skin lesions as well as cutaneous carcinomas showed significantly elevated levels of phosphorylated ERK compared with normal skin, suggesting the activation of EGFR–Ras signaling pathways in keratinocyte-derived malignancies. In vitro, exogenous stimulation of the EGFR–Ras signaling pathway through EGF or transfection of the dominant-active form of the Ras oncogene (H-RasV12) suppressed whereas an EGFR tyrosine kinase inhibitor increased CCL27 mRNA and protein production in keratinocytes. In mice, neutralization of CCL27 led to decreased leukocyte recruitment to cutaneous tumor sites and significantly enhanced primary tumor growth. Collectively, our data identify a mechanism of skin tumors to evade host antitumor immune responses.

Chemokine receptors in head and neck cancer: association with metastatic spread and regulation during chemotherapy.

Head and neck carcinomas are histologically and clinically heterogeneous. While squamous cell carcinomas (SCC) are characterized by lymphogenous spread, adenoid cystic carcinomas (ACC) disseminate preferentially hematogenously. To study cellular and molecular mechanisms of organ‐specific metastasis, we used SCC and ACC cell lines and tumor tissues, obtained from patients with primary or metastatic disease. Comprehensive analysis at the mRNA and protein level of human chemokine receptors showed that SCC and ACC cells exhibited distinct and nonrandom expression profiles for these receptors. SCC predominantly expressed receptors for chemokines homeostatically expressed in lymph nodes, including CC chemokine receptor (CCR) 7 and CXC chemokine receptor (CXCR)5. No difference in expression of chemokine receptors was seen in primary SCC and corresponding lymph node metastases. In contrast to SCC, ACC cells primarily expressed CXCR4. In chemotaxis assays, ACC cells were responsive to CXCL12, the ligand for CXCR4. Exposure of ACC cells to cisplatin resulted in upregulation of CXCR4 on the cell surface, which was repressed by the transcriptional inhibitor, α‐amanitin. Treatment of ACC cells with CXCL12 resulted in the activation of Akt and ERK1/2 pathways. Furthermore, CXCL12 suppressed the rate of apoptosis induced by cisplatin in ACC cells, suggesting that signaling via CXCR4 may be part of a tumor cell survival program. Discrimination of the chemokine receptor profile in SCC and ACC in vitro and in tissues provided insights into their distinct biologic and clinical characteristics as well as indications that chemokine receptors might serve as future therapeutic targets in these malignancies.

Development and Management of Severe Cutaneous Side Effects in Head-and-Neck Cancer Patients during Concurrent Radiotherapy and Cetuximab

Background:The concurrent administration of cetuximab to radiotherapy has recently been shown to improve the clinical outcome of head-and-neck cancer (HNC) patients. An aggravation of the radiation-induced skin toxicity was not described. Here, however, two cases with severe skin toxicity during the combined treatment are reported.Clinical Observations:In a small group of five patients with locally advanced HNC treated with irradiation and concurrent cetuximab, two cases of unusually severe radiation dermatitis were observed. Both patients developed confluent moist desquamations confined to the irradiation field at a dose of 40 Gy (CTC [Common Toxicity Criteria] grade 3), which progressed into an ulcerative dermatitis (grade 4) at 58 Gy and 46 Gy, respectively. Histopathology showed a vacuolic degeneration of basal keratinocytes, subepidermal blister formation, and mixed perivascular and interstitial inflammatory infiltrates leading to a complete loss of the epidermis. These cutaneous side effects led to the discontinuation of radiotherapy. Topical corticosteroids and systemic antibiotic treatment resulted in wound healing, which allowed the continuation of radiotherapy.Conclusion:These findings indicate that cetuximab may have the potential to enhance the severity of radiation dermatitis in HNC patients. A systematic monitoring of cutaneous side effects during radiotherapy plus cetuximab is advised in order to reliably estimate the frequency of severe (grade 3/4) radiation dermatitis.Hintergrund:Die simultane Applikation von Cetuximab zur Strahlentherapie verbessert nach einer kürzlich publizierten Studie die Prognose von Patienten mit Kopf-Hals-Tumoren (HNC). Hinweise auf eine Verstärkung der strahlenbedingten Hautreaktion ergaben sich nicht. Im Folgenden wird allerdings über zwei Fälle mit schwersten Hautreaktionen während der kombinierten Behandlung berichtet.Beobachtungen:In einer kleinen Gruppe von fünf Patienten mit HNC, die mit Bestrahlung und simultanem Cetuximab behandelt wurden, war bei zwei Patienten eine ungewöhnlich starke Radiodermatitis zu beobachten. Beide Patienten entwickelten während der Behandlung bei einer Dosis von 40 Gy zunächst konfluierende feuchte Epitheliolysen in den Bestrahlungsfeldern (CTC [Common Toxicity Criteria] Grad 3), welche bei einer Dosis von 58 Gy bzw. 46 Gy in ulzerative Dermatitiden übergingen (CTC Grad 4). Histopathologisch zeigten sich vakuolische Degenerationen der basalen Keratinozyten, subepidermale Blasenbildungen sowie perivaskuläre und interstitielle inflammatorische Infiltrate mit komplettem Verlust der Epidermis. Diese Nebenwirkungen führten in beiden Fällen zu einer Unterbrechung der Radiotherapie. Durch eine intensive Therapie mit topischen Glukokortikosteroiden sowie eine systemische antibiotische Behandlung kam es zur kompletten Abheilung, was die Fortsetzung der Bestrahlung ermöglichte.Schlussfolgerung:Diese Beobachtungen zeigen, dass Cetuximab das Potential haben könnte, den Grad der Radiodermatitis bei Patienten mit HNC wesentlich zu verstärken. Ein systematisches Monitoring der Hautnebenwirkungen während der Radiotherapie in Kombination mit Cetuximab ist erforderlich, um die Häufigkeit der schweren Radiodermatitis (Grad 3/4) verlässlich einschätzen zu können.

Bevacizumab as a Treatment Option for Radiation-Induced Cerebral Necrosis

Radiation necrosis of normal CNS tissue represents one of the main risk factors of brain irradiation, occurring more frequently and earlier at higher total doses and higher doses per fraction. At present, it is believed that the necrosis results due to increasing capillary permeability caused by cytokine release leading to extracellular edema. This process is sustained by endothelial dysfunction, tissue hypoxia, and subsequent necrosis. Consequently, blocking the vascular endothelial growth factor (VEGF) at an early stage could be an option to reduce the development of radiation necrosis by decreasing the vascular permeability. This might help to reverse the pathological mechanisms, improve the symptoms and prevent further progression. A patient with radiationinduced necrosis was treated with an anti-VEGF antibody (bevacizumab), in whom neurologic signs and symptoms improved in accordance with a decrease in T1-weighted fluid-attenuated inversion recovery signals. Our case report together with the current literature suggests bevacizumab as a treatment option for patients with symptoms and radiological signs of cerebral necrosis induced by radiotherapy.Die strahleninduzierte Radionekrose des Gehirns stellt eine schwerwiegende Komplikation der Strahlentherapie dar und tritt bei hohen Gesamtdosen oder hohen fraktionierten Einzeldosen häufiger und früher auf. Es wird vermutet, dass hierfür eine erhöhte Freisetzung von Zytokinen ursächlich ist, die zu einer erhöhten Kapillarpermeabiliät und in der Folge zu einem extrazellulären Ödem führt. Dieser Prozess wird durch die endotheliale Dysfunktion sowie eine Gewebehypoxie weiter verstärkt und kann letztendlich zu einer Nekrose führen. Eine Blockade des vaskulären endothelialen Wachstumsfaktors (vascular endothelial growthfactor; VEGF) könnte diese verstärkte vaskuläre Permeabilität vermindern. Somit könnten pathologische Prozesse umgekehrt, neurologische Ausfallerscheinungen vermindert und ein Fortschreiten der Problematik verhindert werden. Wir präsentieren hier einen Fall einer strahleninduzierten Nekrose, bei dem sich unter Therapie mit einem anti-VEGF Antikörper (Bevacizumab) eine Besserung der neurologischen Zeichen und Symptome in Analogie zu einer MR-morphologischen Abnahme des T2-Signals zeigte. Eine Zusammenschau dieses Falles und der aktuellen verfügbaren Literatur lässt den Schluss zu, dass Bevacizumab eine Behandlungsoption für Patienten mit Symptomen und radiologischen Zeichen einer strahleninduzierten zerebralen Nekrose sein kann.

Rosacea: the Cytokine and Chemokine Network

Rosacea is one of the most common dermatoses of adults. Recent studies have improved our understanding of the pathophysiology of rosacea. Current concepts suggest that known clinical trigger factors of rosacea such as UV radiation, heat, cold, stress, spicy food, and microbes modulate Toll-like receptor signaling, induce reactive oxygen species, as well as enhance antimicrobial peptide and neuropeptide production. Downstream of these events cytokines and chemokines orchestrate an inflammatory response that leads to the recruitment and activation of distinct leukocyte subsets and induces the characteristic histopathological features of rosacea. Here we summarize the current knowledge of the cytokine and chemokine network in rosacea and propose pathways that may be of therapeutic interest.

Chemokines in tumor-associated angiogenesis.

Abstract Tumor growth is dependent on several key factors. Apart from immune escape and an efficient blockade of apoptotic signals, tumors require oxygen and nutrients to grow past a diameter of 2 μm. Therefore, it is of vital importance for the tumor to facilitate tumor-associated angiogenesis, e.g., the de novo formation of new blood vessels. In addition to established and key angiogenic factors, such as vascular endothelial growth factor, chemokines, a superfamily of cytokine-like proteins that bind to seven transmembrane-spanning G-protein-coupled receptors, have been associated with angiogenesis under homeostatic conditions. Chemokines were initially identified as key factors that control the directional migration of leukocytes, stem cells and cancer cells in vitro and which critically regulate their trafficking in vivo. Recently their role in establishing a favorable microenvironment for tumor-associated angiogenesis, a process that requires complex bidirectional interactions of the tumor and associated vessels, has been the focus of research. Chemokine-promoted angiogenesis not only facilitates tumor growth by supplying nutrients and oxygen but it is also a prerequisite to tumor metastasis. Hence, the pharmacologic control of tumor angiogenesis presents a promising strategy for novel anticancer therapeutics. Here, we discuss the current pathogenetic concepts of tumor-associated angiogenesis in the context of chemokines and their receptors and highlight promising therapeutic strategies.

Methylated APC and GSTP1 genes in serum DNA correlate with the presence of circulating blood tumor cells and are associated with a more aggressive and advanced breast cancer disease.

BackgroundTumor-related methylated DNA and circulating tumor cells (CTC) in the peripheral blood might be of prognostic importance in breast cancer. Thus, the aim of our study was to examine free methylated DNA and CTC in the blood from breast cancer patients and to correlate it with clinicopathological features known to influence prognosis.Materials and methodsWe prospectively obtained serum samples from 85 patients with breast cancer and 22 healthy volunteers. Sera were analysed by methylation specific PCR (MethyLight PCR) for five genes: adenomatous polyposis coli (APC), ras association domain family protein 1A (RASSF1A), estrogen receptor 1 (ESR1), CDKN2A (p16) and glutathione s-transferase pi 1 (GSTP1). Beta actin (ACTB) served as control. In parallel matched peripheral blood of 63 patients was used to assay for circulating tumor cells in the peripheral blood by a modified immunomagnetic AdnaTest BreastCancerSelect with PCR detection for EPCAM, MUC1, MGB1 and SPDEF.ResultsWe found a hypermethylation in the APC gene in 29% (25/85), in RASSF1A in 26% (22/85), in GSTP1 in 18% (14/76) and in ESR1 in 38% (32/85) of all breast cancer patients. No hypermethylation of CDKN2A was found (0/25). Blood samples of patients were defined CTC positive by detecting the EPCAM 13% (8/63), MUC1 16% (10/63), MGB 9% (5/55), SPDEF 12% (7/58) and in 27% detecting one or more genes (15/55). A significant difference was seen in methylated APC DNA between cancer patients and healthy volunteers. Moreover, methylated APC, RASSF1 and CTC were significantly different in metastatic versus non-metastatic disease. In addition, the presence of methylated APC, RASSF1A and CTC correlated significantly with AJCC-staging (p = 0.001, p = 0.031 and 0.002, respectively). High incidences of methylations were found for the genes RASSF1 and ESR1 in healthy individuals (both 23% 5/22). Methylated GSTP1 was predominantly found in the serum of patients with large primaries (p = 0.023) and was highly significantly correlated with positive Her2/neu status (p = 0.003). Elevated serum CA15.3 was strongly correlated with methylated APC and CTC detection (both p = 0.000). Methylated ESR1 failed to exhibit significant correlations with any of the above mentioned parameters. The presence of CTC in peripheral blood was significantly associated with methylated APC (p = 0.012) and methylated GSTP1 (p = 0.001).ConclusionThe detection of methylated APC and GSTP1 DNA in serum correlated with the presence of CTC in the blood of breast cancer patients. Both methylated DNA and CTC correlated with a more aggressive tumor biology and advanced disease.

Density of Demodex folliculorum in Patients Receiving Epidermal Growth Factor Receptor Inhibitors

Background: Rosacea-like papulopustular eruptions (rash) are considered the most frequent toxicities associated with the use of inhibitors of the epidermal growth factor receptor (EGFR). Recently, evidence has been accumulating of infectious complications in patients suffering from these adverse effects. Objective: We sought to analyze the density of Demodex folliculorum (DF) in cutaneous lesions of patients presenting with EGFR-inhibitor (EGFRI)-induced rashes. Methods: This is a retrospective study of 19 adult patients presenting with EGFRI rashes. Patients were reviewed for the density of DF (Demodex density, Dd; mites per square centimeter) by standardized skin surface biopsy. Results: In our patient collective the mean Dd of 4.7/cm2 significantly exceeded the mean Dd reported for the healthy adult population (Dd = 0.7/cm2). Limitations: The retrospective nature of the study. Conclusions: EGFRI patients have an increased susceptibility to DF colonization or infection, respectively. Our results support the recent concept that EGFRI may induce an impairment of antimicrobial defense mechanisms.

Glioblastoma Multiforme Metastasis Outside the CNS: Three Case Reports and Possible Mechanisms of Escape

Introduction Primary brain and CNS tumor incidence is approximately 19 per 100,000 individuals per year in the United States compared with seven per 100,000 individuals worldwide. Worldwide this accounts for 2% of all primary tumors and 7% of years of life lost from cancer before the age of 70 years. Glioblastoma multiforme (GBM) is also the most aggressive brain tumor with poor prognosis; patients with GBM have a median survival time of about 14 months. GBM metastases outside the CNS are rare, so therapeutic experience with these types of tumors is limited. Normally the brain is immunologically and anatomically separated from the body by the blood brain barrier. Herein, we present the cases of three patients with GBM with extra-CNS metastasis. The variety of metastasis locations demonstrated in these cases helps to illustrate the various mechanism and corresponding risk factors that allow GBM to escape the CNS.