Liver-targeted polymeric prodrugs delivered subcutaneously improve tafenoquine therapeutic window for malaria radical cure.

Approximately 3.3 billion people live with the threat of Plasmodium vivax malaria. Infection can result in liver-localized hypnozoites, which when reactivated cause relapsing malaria. This work demonstrates that an enzyme-cleavable polymeric prodrug of tafenoquine addresses key requirements for a mass administration, eradication campaign: excellent subcutaneous bioavailability, complete parasite control after a single dose, improved therapeutic window compared to the parent oral drug, and low cost of goods sold (COGS) at less than $1.50 per dose. Liver targeting and subcutaneous dosing resulted in improved liver:plasma exposure profiles, with increased efficacy and reduced glucose 6-phosphate dehydrogenase-dependent hemotoxicity in validated preclinical models. A COGS and manufacturability analysis demonstrated global scalability, affordability, and the ability to redesign this fully synthetic polymeric prodrug specifically to increase global equity and access. Together, this polymer prodrug platform is a candidate for evaluation in human patients and shows potential for P. vivax eradication campaigns.

The effect of percentage of ideal body weight on outcomes in ileo-anal pull through for ulcerative colitis.

PURPOSE: Ileo-anal pull through (IAPT) is a commonly performed operation for the surgical management of ulcerative colitis. The effect of body weight on outcomes for patients undergoing this operation has not been extensively studied. METHODS: This was a prospective cohort study at a single tertiary care inflammatory bowel disease (IBD) center. A total of 457 patients who were operated on at the Mount Sinai Medical Center between 1983 and 2015 were included. Demographic characteristics, the patients' body weight at the time of IAPT, and postoperative outcome data were collected. RESULTS: For each patient, body weight was calculated as a percentage of the ideal body weight (IBW) for that patient's height. The mean percentage of ideal body weight was 93.9% with a standard deviation of 20%. The range for the population was 53.1 to 175%. Four hundred forty (96%) of the patients had a weight within two standard deviations of the mean, indicating a normal distribution. Seventy-nine patients developed a Clavien-Dindo class III complication necessitating a procedural treatment. The most common of these was a stricture at the anastomotic site (n = 54). Our study identified an association between a percentage of ideal body weight in the lowest quartile of our population and development of an anastomotic stricture. This association was statistically significant on multivariate analysis. CONCLUSION: Low body weight at the time of ileo-anal pull through for treatment of UC may be a risk factor for development of anastomotic stricture requiring dilation.

Slow and Steady Wins the Race: A Solid Case for a 3-Stage Approach in Ulcerative Colitis.

BACKGROUND: Approximately 10% to 20% of patients with ulcerative colitis require surgery during their disease course, of which the most common is the staged restorative proctocolectomy with IPAA. OBJECTIVE: The aim was to compare the rates of anastomotic leaks among all staged restorative proctocolectomy with IPAA procedures. DESIGN: This was a retrospective cohort study. SETTINGS: This study was conducted at a single tertiary care IBD center. PATIENTS: All patients with ulcerative colitis or IBD-unspecified who underwent a primary total proctocolectomy with IPAA for medically refractory disease or dysplasia between 2008 and 2017 were identified. MAIN OUTCOME MEASURES: The primary outcome was anastomotic leak within a 6-month postoperative period. Univariate and multivariate logistic regression were used to compare patients with and without anastomotic leaks. RESULTS: The sample was composed of 584 nonemergent patients, of whom 50 (8.6%) underwent 1-stage, 162 (27.7%) underwent 2-stage, 58 (9.9%) underwent modified 2-stage, and 314 (53.7%) underwent a 3-stage total proctocolectomy with IPAA. The primary indication was medically refractory disease in 488 patients and dysplasia/cancer in 101 patients. Anastomotic leak occurred in 10 patients (3.2%) after 3-stage, 14 patients (8.6%) after 2-stage, 6 patients (10.3%) after modified 2-stage, and 10 patients (20.0%) after a 1-stage procedure. A 3-stage procedure had fewer leaks and additional procedures for leaks compared with 1- and modified 2-stage procedures (p < 0.03). The 3-stage procedure had fewer combined anastomotic leaks and pelvic abscesses than all of the other staged procedures (p < 0.05). LIMITATIONS: This study was limited by its retrospective design and evolving electronic medical charts system. CONCLUSIONS: The 3-stage total proctocolectomy with IPAA is the optimal staged method in ulcerative colitis to reduce leaks and related complications. See Video Abstract at http://links.lww.com/DCR/B693. LENTO Y CONSTANTE GANA LA CARRERA UN CASO SLIDO PARA UN ENFOQUE DE TRES ETAPAS EN LA COLITIS ULCEROSA: ANTECEDENTES:Aproximadamente el 10-20% de los pacientes con colitis ulcerosa requieren cirugía durante el curso de su enfermedad, de los cuales la más común es la proctocolectomía restauradora escalonada con anastomosis con bolsa ileo-anal.OBJETIVO:El objetivo fue comparar las tasas de fugas anastomóticas entre todos los procedimientos de proctocolectomía restauradora por etapas con procedimiento de anastomosis con bolsa ileo-anal.DISEÑO:Este fue un estudio de cohorte retrospectivo.ENTORNO CLÍNICO:Este estudio se llevó a cabo en un único centro de atención terciaria de tercer nivel para enfermedades inflamatorias del intestino.PACIENTES:Se identificaron todos los pacientes con colitis ulcerosa o enfermedad inflamatoria intestinal inespecífica que se sometieron a una proctocolectomía total primaria mas anastomosis con bolsa ileo-anal por enfermedad médicamente refractaria o displasia entre 2008 y 2017.PRINCIPALES MEDIDAS DE RESULTADO:El resultado primario fue la fuga anastomótica dentro de un período posoperatorio de seis meses. Se utilizó regresión logística univariante y multivariante para comparar pacientes con y sin fugas anastomóticas.RESULTADOS:La muestra estuvo compuesta por 584 pacientes no emergentes, de los cuales 50 (8,6%) se sometieron a una etapa, 162 (27,7%) se sometieron a dos etapas, 58 (9,9%) se sometieron a modificación en dos etapas y 314 (53,7%) se sometieron a una proctocolectomía total en tres tiempos mas anastomosis con bolsa ileo-anal. La indicación principal fue enfermedad médicamente refractaria en 488 pacientes y displasia / cáncer en 101 pacientes. Se produjo una fuga anastomótica en 10 (3,2%) pacientes después de tres etapas, 14 (8,6%) pacientes después de dos etapas, 6 (10,3%) pacientes después de dos etapas modificadas y 10 (20,0%) pacientes después de una etapa procedimiento. Un procedimiento de tres etapas tuvo menos fugas y procedimientos adicionales para las fugas en comparación con los procedimientos de una y dos etapas modificadas (p <0.03). El procedimiento de tres etapas tuvo menos fugas anastomóticas y abscesos pélvicos combinados que todos los demás procedimientos por etapas (p <0,05).LIMITACIONES:Este estudio estuvo limitado por su diseño retrospectivo y su sistema de registros médicos electrónicos en evolución.CONCLUSIONES:La proctocolectomía total en tres etapas mas anastomosis con bolsa ileo-anal es el método óptimo por etapas en la colitis ulcerosa para reducir las fugas y las complicaciones relacionadas. Consulte Video Resumen en http://links.lww.com/DCR/B693.

A novel histopathological scoring system to distinguish urticarial vasculitis from chronic spontaneous urticaria.

BACKGROUND: Urticarial vasculitis (UV) is defined by long-lasting urticarial lesions combined with the histopathologic findings of leukocytoclastic vasculitis. As one of the major unmet needs in UV, diagnostic criteria are rather vague and not standardized. Moreover, there seems to be considerable overlap with chronic spontaneous urticaria (CSU), particularly for the normocomplementemic variant of UV. Therefore, this study aimed to develop a diagnostic scoring system that improves the histopathologic discrimination between UV and CSU. METHODS: Lesional skin sections of patients with clinical and histopathologic diagnosis of UV (n = 46) and CSU (n = 51) were analyzed (blinded to the diagnosis) for the following pre-defined criteria: presence of leukocytoclasia, erythrocyte extravasation, fibrin deposits, endothelial cell swelling, ectatic vessels, blurred vessel borders, dermal edema, intravascular neutrophil, and eosinophil numbers and numbers of dermal neutrophils, macrophages and mast cells. RESULTS: The greatest differences between UV and CSU samples were observed for leukocytoclasia (present in 76% of UV vs. 3.9% of CSU samples; p < 0.0001), erythrocyte extravasation (present in 41.3% of UV vs. 2.0% of CSU samples; p < 0.0001), and fibrin deposits (present in 27.9% of UV vessels vs. 9.7% of CSU vessels; p < 0.0001). Based on these findings, we developed a diagnostic score, the urticarial vasculitis score (UVS), which correctly assigned 37 of 46 cases of UV and 49 of 51 cases of CSU to the previously established diagnosis. CONCLUSION: Our results suggest that the UVS, a combined quantitative assessment of the three criteria leukocytoclasia, fibrin deposits and extravasated erythrocytes, distinguishes UV from CSU in skin histopathology. The UVS, if validated in larger patient samples, may help to improve the diagnostic approach to UV.

Liver-targeted polymeric prodrugs of 8-aminoquinolines for malaria radical cure.

Primaquine and tafenoquine are the two 8-aminoquinoline (8-AQ) antimalarial drugs approved for malarial radical cure - the elimination of liver stage hypnozoites after infection with Plasmodium vivax. A single oral dose of tafenoquine leads to high efficacy against intra-hepatocyte hypnozoites after efficient first pass liver uptake and metabolism. Unfortunately, both drugs cause hemolytic anemia in G6PD-deficient humans. This toxicity prevents their mass administration without G6PD testing given the approximately 400 million G6PD deficient people across malarial endemic regions of the world. We hypothesized that liver-targeted delivery of 8-AQ prodrugs could maximize liver exposure and minimize erythrocyte exposure to increase their therapeutic window. Primaquine and tafenoquine were first synthesized as prodrug vinyl monomers with self-immolative hydrolytic linkers or cathepsin-cleavable valine-citrulline peptide linkers. RAFT polymerization was exploited to copolymerize these prodrug monomers with hepatocyte-targeting GalNAc monomers. Pharmacokinetic studies of released drugs after intravenous administration showed that the liver-to-plasma AUC ratios could be significantly improved, compared to parent drug administered orally. Single doses of the liver-targeted, enzyme-cleavable tafenoquine polymer were found to be as efficacious as an equivalent dose of the oral parent drug in the P. berghei causal prophylaxis model. They also elicited significantly milder hemotoxicity in the humanized NOD/SCID mouse model engrafted with red blood cells from G6PD deficient donors. The clinical application is envisioned as a single subcutaneous administration, and the lead tafenoquine polymer also showed excellent bioavailability and liver-to-blood ratios exceeding the IV administered polymer. The liver-targeted tafenoquine polymers warrant further development as a single-dose therapeutic via the subcutaneous route with the potential for broader patient administration without a requirement for G6PD diagnosis.

Diode-pumped cw Alexandrite laser with temporally stable 6.5 W in TEM00 operation with prospect of power scaling.

We present the design of a longitudinally diode-pumped Alexandrite laser in continuous-wave operation and resulting performance data. A laser power of 6.5 W in fundamental mode operation was measured, which is, to the best of our knowledge, the highest laser power in fundamental mode operation yet reported. The laser crystal was pumped by two diode modules emitting at 637 nm. The pump radiation was polarization-combined and spatially symmetrized. The laser operates at an output power of 6.5 W with an optical-to-optical efficiency of 26%, temporally stable output with stability of 8% on ms timescale, a beam quality of M2 = 1.1 in both spatial directions and emission of an output wavelength of 752 nm. Measurements of the thermal dioptric power at pumping intensities up to 9.5 kW/cm2 support the appropriate approach of the design. Based on our results, we estimate the potential and show our concept for future scaling of the output power.

MTL-CEBPA, a Small Activating RNA Therapeutic Upregulating C/EBP-α, in Patients with Advanced Liver Cancer: A First-in-Human, Multicenter, Open-Label, Phase I Trial.

PURPOSE: Transcription factor C/EBP-α (CCAAT/enhancer-binding protein alpha) acts as a master regulator of hepatic and myeloid functions and multiple oncogenic processes. MTL-CEBPA is a first-in-class small activating RNA oligonucleotide drug that upregulates C/EBP-α. PATIENTS AND METHODS: We conducted a phase I, open-label, dose-escalation trial of MTL-CEBPA in adults with advanced hepatocellular carcinoma (HCC) with cirrhosis, or resulting from nonalcoholic steatohepatitis or with liver metastases. Patients received intravenous MTL-CEBPA once a week for 3 weeks followed by a rest period of 1 week per treatment cycle in the dose-escalation phase (3+3 design). RESULTS: Thirty-eight participants have been treated across six dose levels (28-160 mg/m2) and three dosing schedules. Thirty-four patients were evaluable for safety endpoints at 28 days. MTL-CEBPA treatment-related adverse events were not associated with dose, and no maximum dose was reached across the three schedules evaluated. Grade 3 treatment-related adverse events occurred in nine (24%) patients. In 24 patients with HCC evaluable for efficacy, an objective tumor response was achieved in one patient [4%; partial response (PR) for over 2 years] and stable disease (SD) in 12 (50%). After discontinuation of MTL-CEBPA, seven patients were treated with tyrosine kinase inhibitors (TKIs); three patients had a complete response with one further PR and two with SD. CONCLUSIONS: MTL-CEBPA is the first saRNA in clinical trials and demonstrates an acceptable safety profile and potential synergistic efficacy with TKIs in HCC. These encouraging phase I data validate targeting of C/EBP-α and have prompted MTL-CEBPA + sorafenib combination studies in HCC.

Gene activation of CEBPA using saRNA: preclinical studies of the first in human saRNA drug candidate for liver cancer.

Liver diseases are a growing epidemic worldwide. If unresolved, liver fibrosis develops and can lead to cirrhosis and clinical decompensation. Around 5% of cirrhotic liver diseased patients develop hepatocellular carcinoma (HCC), which in its advanced stages has limited therapeutic options and negative survival outcomes. CEPBA is a master regulator of hepatic function where its expression is known to be suppressed in many forms of liver disease including HCC. Injection of MTL-CEBPA, a small activating RNA oligonucleotide therapy (CEBPA-51) formulated in liposomal nanoparticles (NOV340- SMARTICLES) upregulates hepatic CEBPA expression. Here we show how MTL-CEBPA therapy promotes disease reversal in rodent models of cirrhosis, fibrosis, hepatosteatosis, and significantly reduces tumor burden in cirrhotic HCC. Restoration of liver function markers were observed in a carbon-tetrachloride-induced rat model of fibrosis following 2 weeks of MTL-CEBPA therapy. At 14 weeks, animals showed reduction in ascites and enhanced survival rates. MTL-CEBPA reversed changes associated with hepatosteatosis in non-alcoholic methionine and cholic-deficient diet-induced steaotic liver disease. In diethylnitrosamine induced cirrhotic HCC rats, MTL-CEBPA treatment led to a significant reduction in tumor burden. The data included here and the rapid adoption of MTL-CEBPA into a Phase 1 study may lead to new therapeutic oligonucleotides for undruggable diseases.

Efficacy and safety of 4 months of sublingual immunotherapy with recombinant Mal d 1 and Bet v 1 in patients with birch pollen-related apple allergy.

BACKGROUND: Birch pollen-related apple allergy is among the most prevalent food allergies in adolescent/adult subjects and mainly results from sensitization to the major birch pollen allergen Bet v 1 and subsequent cross-reaction with the apple protein Mal d 1. However, specific immunotherapy with birch pollen has inconsistent effects on apple allergy. OBJECTIVE: We sought to compare the safety and efficacy of sublingual immunotherapy (SLIT) with 2 formulations containing either rMal d 1 or rBet v 1 on birch pollen-related apple allergy. METHODS: Sixty participants with birch pollen-related apple allergy were randomized to daily sublingual application of placebo (n = 20) or 25 µg of rMal d 1 (n = 20) or rBet v 1 (n = 20) for 16 weeks. Adverse events were regularly recorded. Sublingual challenges with standardized doses of rMal d 1, skin prick tests with recombinant allergens, and measurements of allergen-specific IgE and IgG4 antibodies were performed before and after treatment. RESULTS: Both formulations caused comparable, mainly local adverse events. No systemic reactions occurred. Compared with the placebo and rBet v 1-treated groups, SLIT with rMal d 1 reduced rMal d 1-induced oral symptoms (P = .001 and P = .038) accompanied by longitudinally reduced rMal d 1-specific cutaneous reactions (P = .022) and enhanced IgG4/IgE ratios (P = .012). SLIT with rBet v 1 neither improved the clinical reactivity to rMal d 1 nor enhanced rMal d 1-specific IgG4/IgE ratios. Participants receiving placebo showed no allergen-specific changes. CONCLUSION: Sublingual treatment with a recombinant food allergen was safe and clinically effective, as determined by using standardized challenges. We present a promising approach for the effective treatment of birch pollen-related apple allergy.

Development and Mechanism of Small Activating RNA Targeting CEBPA, a Novel Therapeutic in Clinical Trials for Liver Cancer.

Small activating RNAs (saRNAs) are short double-stranded oligonucleotides that selectively increase gene transcription. Here, we describe the development of an saRNA that upregulates the transcription factor CCATT/enhancer binding protein alpha (CEBPA), investigate its mode of action, and describe its development into a clinical candidate. A bioinformatically directed nucleotide walk around the CEBPA gene identified an saRNA sequence that upregulates CEBPA mRNA 2.5-fold in human hepatocellular carcinoma cells. A nuclear run-on assay confirmed that this upregulation is a transcriptionally driven process. Mechanistic experiments demonstrate that Argonaute-2 (Ago2) is required for saRNA activity, with the guide strand of the saRNA shown to be associated with Ago2 and localized at the CEBPA genomic locus using RNA chromatin immunoprecipitation (ChIP) assays. The data support a sequence-specific on-target saRNA activity that leads to enhanced CEBPA mRNA transcription. Chemical modifications were introduced in the saRNA duplex to prevent activation of the innate immunity. This modified saRNA retains activation of CEBPA mRNA and downstream targets and inhibits growth of liver cancer cell lines in vitro. This novel drug has been encapsulated in a liposomal formulation for liver delivery, is currently in a phase I clinical trial for patients with liver cancer, and represents the first human study of an saRNA therapeutic.

Epicutaneous allergen application preferentially boosts specific T cell responses in sensitized patients.

The effects of epicutaneous allergen administration on systemic immune responses in allergic and non-allergic individuals has not been investigated with defined allergen molecules. We studied the effects of epicutaneous administration of rBet v 1 and rBet v 1 fragments on systemic immune responses in allergic and non-allergic subjects. We conducted a clinical trial in which rBet v 1 and two hypoallergenic rBet v 1 fragments were applied epicutaneously by atopy patch testing (APT) to 15 birch pollen (bp) allergic patients suffering from atopic dermatitis, 5 bp-allergic patients suffering from rhinoconjunctivitis only, 5 patients with respiratory allergy without bp allergy and 5 non-allergic individuals. Epicutaneous administration of rBet v 1 and rBet v 1 fragments led to strong and significant increases of allergen-specific T cell proliferation (CLA+ and CCR4+T cell responses) only in bp-allergic patients with a positive APT reaction. There were no relevant changes of Bet v 1-specific IgE and IgG responses. No changes were noted in allergic subjects without bp allergy and in non-allergic subjects. Epicutaneous allergen application boosts specific T cell but not antibody responses mainly in allergic, APT-positive patients suggesting IgE-facilitated allergen presentation as mechanism for its effects on systemic allergen-specific immune responses.

Extracorporeal IgE Immunoadsorption in Allergic Asthma: Safety and Efficacy.

BACKGROUND: Prevention of IgE-binding to cellular IgE-receptors by anti-IgE (Omalizumab) is clinically effective in allergic asthma, but limited by IgE threshold-levels. To overcome this limitation, we developed a single-use IgE immunoadsorber column (IgEnio). IgEnio is based on a recombinant, IgE-specific antibody fragment and can be used for the specific extracorporeal desorption of IgE. OBJECTIVE: To study safety and efficacy of IgEnio regarding the selective depletion of IgE in a randomized, open-label, controlled pilot trial in patients with allergic asthma and to investigate if IgEnio can bind IgE-Omalizumab immune complexes. METHODS: Fifteen subjects were enrolled and randomly assigned to the treatment group (n=10) or to the control group (n=5). Immunoadsorption was done by veno-venous approach, processing the twofold calculated plasma volume during each treatment. A minimum average IgE-depletion of 50% after the last cycle in the intention-to-treat population was defined as primary endpoint. Safety of the treatment was studied as secondary endpoint. In addition, possible changes in allergen-specific sensitivity were investigated, as well as clinical effects by peak flow measurement and symptom-recording. The depletion of IgE-Omalizumab immune complexes was studied in vitro. The study was registered at clinicaltrials.gov (NCT02096237) and conducted from December 2013 to July 2014. RESULTS: IgE immunoadsorption with IgEnio selectively depleted 86.2% (±5.1% SD) of IgE until the end of the last cycle (p<0.0001). Removal of pollen allergen-specific IgE was associated with a reduction of allergen-specific basophil-sensitivity and prevented increases of allergen-specific skin-sensitivity and clinical symptoms during pollen seasons. IgEnio also depleted IgE-Omalizumab immune complexes in vitro. The therapy under investigation was safe and well-tolerated. During a total of 81 aphereses, 2 severe adverse events (SAE) were recorded, one of which, an episode of acute dyspnea, possibly was related to the treatment and resolved after administration of antihistamines and corticosteroids. CONCLUSIONS: This pilot study indicates that IgE immunoadsorption with IgEnio may be used to treat patients with pollen-induced allergic asthma. Furthermore, the treatment could render allergic patients with highly elevated IgE-levels eligible for the administration of Omalizumab and facilitate the desorption of IgE-Omalizumab complexes. This study was funded by Fresenius Medical Care Deutschland GmbH, Bad Homburg, Germany.

MMP8 Is Increased in Lesions and Blood of Acne Inversa Patients: A Potential Link to Skin Destruction and Metabolic Alterations.

Acne inversa (AI; also designated as hidradenitis suppurativa) is a chronic inflammatory disease with still unknown pathogenesis that affects the intertriginous skin of perianal, inguinal, and axillary sites. It leads to painful nodules, abscesses, and fistulas with malodorous secretion and is frequently associated with metabolic alterations. Here, we demonstrate that one of the most highly upregulated molecules in AI lesions is matrix metalloproteinase 8 (MMP8), an enzyme specialized in the degradation of extracellular matrix components and the HDL component apolipoprotein A-I. Granulocytes, which were present in AI lesions, secreted high amounts of MMP8 especially after TNF-α stimulation. Furthermore, activated fibroblasts but not keratinocytes were found to express MMP8. The high lesional MMP8 levels were accompanied by elevated blood levels that positively correlated with TNF-α blood levels and disease severity assessed by Sartorius score, especially with the number of regions with inflammatory nodules/abscesses and fistulas. Additionally, we found a negative correlation between blood MMP8 and HDL-cholesterol levels, suggesting a contributory role of MMP8 in metabolic alterations in AI. In summary, we demonstrate elevated MMP8 levels in AI lesions, suggest their role in skin destruction and metabolic alterations, and recommend the use of MMP8 as blood biomarker for AI disease activity assessment.

Biological Effects of Ingenol Mebutate Gel in Moderate to Severe Actinic Fields Assessed by Reflectance Confocal Microscopy: A Phase I Study.

Ingenol mebutate represents a topical treatment for fields with actinic keratosis (AK). The biological effects of ingenol mebutate in AK, subclinical (SC)-AK, and reference-skin were assessed and graded by in vivo reflectance confocal microscopy (RCM) and histology. Patients with AK and SC-AK lesions in one 25 cm2 field on hands or forearms, and with an area of reference skin on the inner upper arm, were included. The two fields were each treated with ingenol mebutate 0.05% gel (n=16), or vehicle (n=8), on 2 consecutive days; clinical and RCM assessments were performed on days 1, 2, 3, 8, and 57, and biopsies on day 3. Local skin responses were more pronounced in AK fields (6.1 (mean) ± 2.6 (SD)) compared with reference skin (3.5 ± 1.5). The clinical AK lesion reduction was 43.8% and 6.3% with ingenol mebutate and vehicle, respectively. RCM and histology evaluations showed that ingenol mebutate induced a significant pronounced cell death and immune response in AK and SC-AK lesions, compared with reference skin. Ingenol mebutate induced RCM-measured reduction in (investigator-1/investigator-2): AK lesions (34/28%), SC-AK lesions (72/56%), and solar elastosis in AK fields (mean, -0.22/-0.25). In conclusion, ingenol mebutate showed selective pronounced biological responses in AK and SC-AK as compared with reference skin.

J Drugs Dermatol. 2016;15(10):1181-1189.

Mechanisms, safety and efficacy of a B cell epitope-based vaccine for immunotherapy of grass pollen allergy.

BACKGROUND: We have developed a recombinant B cell epitope-based vaccine (BM32) for allergen-specific immunotherapy (AIT) of grass pollen allergy. The vaccine contains recombinant fusion proteins consisting of allergen-derived peptides and the hepatitis B surface protein domain preS as immunological carrier. METHODS: We conducted a randomized, double-blind, placebo-controlled AIT study to determine safety, clinical efficacy and immunological mechanism of three subcutaneous injections of three BM32 doses adsorbed to aluminum hydroxide versus aluminum hydroxide (placebo) applied monthly to grass pollen allergic patients (n=70). Primary efficacy endpoint was the difference in total nasal symptom score (TNSS) through grass pollen chamber exposure before treatment and 4weeks after the last injection. Secondary clinical endpoints were total ocular symptom score (TOSS) and allergen-specific skin response evaluated by titrated skin prick testing (SPT) at the same time points. Treatment-related side effects were evaluated as safety endpoints. Changes in allergen-specific antibody, cellular and cytokine responses were measured in patients before and after treatment. RESULTS: Sixty-eight patients completed the trial. TNSS significantly decreased with mean changes of -1.41 (BM32/20µg) (P=0.03) and -1.34 (BM32/40µg) (P=0.003) whereas mean changes in the BM32/10µg and placebo group were not significant. TOSS and SPT reactions showed a dose-dependent decrease. No systemic immediate type side effects were observed. Only few grade 1 systemic late phase reactions occurred in BM32 treated patients. The number of local injection site reactions was similar in actively and placebo-treated patients. BM32 induced highly significant allergen-specific IgG responses (P<0.0001) but no allergen-specific IgE. Allergen-induced basophil activation was reduced in BM32 treated patients and addition of therapy-induced IgG significantly suppressed T cell activation (P=0.0063). CONCLUSION: The B cell epitope-based recombinant grass pollen allergy vaccine BM32 is well tolerated and few doses are sufficient to suppress immediate allergic reactions as well as allergen-specific T cell responses via a selective induction of allergen-specific IgG antibodies. (ClinicalTrials.gov number, NCT01445002.).

Relationship between Histological and Clinical Course of Psoriasis: A Pilot Investigation by Reflectance Confocal Microscopy during Goeckerman Treatment.

Alterations of the skin microvasculature are known to play an important role in the development and maintenance of psoriatic skin lesions. In this study, we investigated lesional skin in 11 psoriatic patients during a modified Goeckerman treatment using reflectance confocal microscopy (RCM) to study the relationship between clinical clearance and histological normalization of psoriatic skin and the significance of histological abnormalities on the course of disease. The treatment regimen resulted in a significant reduction of the Psoriasis Area and Severity Index (PASI) as well as capillary and papillary diameters (p < 0.0001). The capillary and papillary diameters were still enlarged when compared to those in normal skin (p < 0.001). Capillary and papillary diameters correlated with each other prior to and after treatment (correlation coefficient = 0.63 and 0.64, p = 0.01 and 0.002, respectively) but not with the PASI. Capillary and papillary diameters after treatment and percentage reduction of the PASI during treatment seemed to be better predictors for the clinical course of relapse than the PASI after treatment. These findings make the subclinical changes of psoriatic skin vessels and dermal papillae a legitimate target for treatment. Further investigations of a large group of patients are needed to evaluate the potential of RCM findings as successor of the PASI in the monitoring of psoriasis.

Large molecular systems landscape uncovers T cell trapping in human skin cancer.

Immune surveillance of tumour cells is an important function of CD8 T lymphocytes, which has failed in cancer for reasons still unknown in many respect but mainly related to cellular processes in the tumour microenvironment. Applying imaging cycler microscopy to analyse the immune contexture in a human skin cancer we could identify and map 7,000 distinct cell surface-associated multi-protein assemblies. The resulting combinatorial geometry-based high-functional resolution led to discovery of a mechanism of T cell trapping in the epidermis, which involves SPIKE, a network of suprabasal keratinocyte projections piercing and interconnecting CD8 T cells. It appears initiated by clusters of infrabasal T and dendritic cells connected via cell projections across a fractured basal lamina to suprabasal keratinocytes and T lymphocytes.

Frequent occurrence of T cell-mediated late reactions revealed by atopy patch testing with hypoallergenic rBet v 1 fragments.

BACKGROUND: Late allergic reactions are common in the course of allergen-specific immunotherapy and even occur with allergy vaccines with reduced IgE reactivity. OBJECTIVE: We sought to study atopy patch test (APT) reactions and T-cell responses to the recombinant birch pollen allergen Bet v 1 and recombinant hypoallergenic T-cell epitope-containing Bet v 1 fragments in patients with birch pollen allergy with and without atopic dermatitis (AD). METHODS: A clinical study was conducted in 15 patients with birch pollen allergy with AD (group 1), 5 patients with birch pollen allergy without AD (group 2), 5 allergic patients without birch pollen allergy (group 3), and 5 nonallergic subjects (group 4) by performing skin prick tests and APTs with rBet v 1 and hypoallergenic rBet v 1 fragments. T-cell, cutaneous lymphocyte antigen (CLA)(+) and CCR4(+) T-cell and cytokine responses were studied by thymidine uptake, carboxyfluorescein diacetate succinimidyl ester staining, and Luminex technology, respectively. RESULTS: rBet v 1 and hypoallergenic rBet v 1 fragments induced APT reactions in not only most of the patients with birch pollen allergy with AD (11/15) but also in most of those without AD (4/5). Patients with birch pollen allergy with AD had higher Bet v 1-specific proliferation of CLA(+) and CCR4(+) T cells compared with patients with birch pollen allergy without AD. There were no differences in Bet v 1-specific CLA(+) and CCR4(+) proliferation and cytokine secretion in patients with and without APT reactions. CONCLUSION: Hypoallergenic rBet v 1 fragments induce T cell-dependent late reactions not only in patients with birch pollen allergy with AD but also in those without AD, which can be determined based on APT results but not based on in vitro parameters.

Comparison of morphologic criteria for actinic keratosis and squamous cell carcinoma using in vivo multiphoton tomography.

The routine diagnostic procedure of actinic keratosis (AK) and invasive squamous cell carcinoma (SCC) is a histological examination after taking a biopsy. In the past decades, non-invasive optical methods for skin examination have been developed. Patients with clinical diagnosis of AK or SCC were examined. The morphological criteria were determined for healthy, AK and SCC skin and compared for statistically significant differences. In this study, the applicability of multiphoton tomography (MPT) as an in vivo diagnostic tool for AK and SCC was evaluated. Changes in the morphology of the keratinocytes such as broadened epidermis, large intercellular spaces, enlarged nucleus and a large variance in cell shape could easily be recognized. The cell nuclei of AK and SCC were significantly larger compared to healthy skin cells in all cell layers. The nucleus-cytoplasm ratio was also significantly higher for AK and SCC than for the healthy skin cells. It was even higher in SCC compared to spinous and basal cell layer of AK. The cell density in AK and SCC was significantly lower than in the basal and spinous cell layers of healthy skin. In SCC, the cell density was significantly lower than in AK. Concerning the intercellular spaces, significant differences were found for AK and healthy skin in spinous and basal cell layer and for SCC compared to AK and healthy skin. In this study, MPT proved to be a valuable non-invasive imaging method for in vivo detection and discrimination of AK and SCC from healthy skin.