Epidemiology, management and survival outcomes of primary cutaneous melanoma: a ten-year overview.

BACKGROUND: Malignant melanoma (MM) is the most aggressive type of skin cancer, accounting for 90% of all the skin cancer mortality. The objective of this study was providing an overview of current patient- and tumour characteristics, treatment strategies, complications and survival in patients with MM over the past ten years. Hereby, an up-to-date view of every day clinical practice is obtained. METHODS: Files of patients treated for primary cutaneous melanoma (n = 686) in the VieCuri Medical Centre in the Netherlands between January 2002 and December 2013 were retrospectively reviewed. Relevant patient features, tumour characteristics, and (surgical) outcomes were evaluated. RESULTS: The majority of all the patients presented thin tumours (59.1% stage 1A/in situ melanoma). Men showed more ulceration (17.7% vs. 8.4%, p < .01) and a significantly higher Breslow thickness than women (1.2 mm vs. 0.9 mm, p < .01). 14.6% (40/273) underwent sentinel lymph node biopsy (SLNB); 10/40 (25%) showed nodal metastasis, 50 patients (7.3%) developed distant metastases (M: 10.6%, F: 5%, p < .01). One-, 5- and 10- year disease specific survival rates were 96%, 86% and 84%, respectively. Median survival for stage 4 MM was 3 months. Extensive surgery was uncommon (n = 3). CONCLUSIONS: Patients generally presented with thin melanomas. Lymph node disease and distant metastases remained infrequently observed during following years, and general 1- and 5-year overall disease-specific survival rates exceeded 85%. Small numbers of rescue surgery and palliative medical treatment warrant further centralisation and investigation.

Characterization of recombinant Mal d 4 and its application for component-resolved diagnosis of apple allergy.

BACKGROUND: Profilins are ubiquitous panallergens that have been extensively characterized; yet, their clinical relevance is still unclear. OBJECTIVE: The aim of the present study was to produce recombinant apple profilin (rMal d 4) and to evaluate its allergenic activity and its potency for component-resolved allergy diagnosis. METHODS: Complementary DNA-derived Mal d 4 was cloned, expressed in Escherichia coli and subsequently purified via poly (l-proline) sepharose. A total of 28 sera from apple-allergic patients were used for IgE-ELISA, immunoblot, RAST and basophil histamine release (BHR) test. In addition, skin prick tests (SPTs) were performed in five patients. RESULTS: Four different complementary DNA coding for apple profilin, Mal d 4, each with an open reading frame of 393 nucleotides, were identified. One isoform Mal d 4.0101 was expressed in Escherichia coli and subsequently purified. Mass spectroscopy revealed the expected mass of 13.826 for rMal d 4.0101, and circular dichroism analysis data were typical for a folded protein and small-angle X-ray scattering measurement identified the protein as a monomer. All the serum samples displayed IgE binding to rMal d 4.0101 in IgE ELISA, immunoblot and RAST. In immunoblotting, IgE binding to natural Mal d 4 was partially/completely inhibited by preincubation with rMal d 4.0101, and RAST values to apple extract were significantly reduced upon serum pretreatment with rMal d 4.0101. SPTs and BHR assays using purified rMal d 4.0101 were positive. Purified rMal d 4.0101 was destroyed within seconds when subjected to pepsin digestion. CONCLUSIONS: Apple profilin complementary DNAs were identified. The physicochemical and allergenic properties of purified recombinant Mal d 4.0101 were evaluated showing that the recombinant protein was equal to the natural protein as shown by inhibition assays. Thus, Mal d 4 represents another example suitable for component-resolved diagnosis of food allergy.

A mutant of the major apple allergen, Mal d 1, demonstrating hypo-allergenicity in the target organ by double-blind placebo-controlled food challenge.

BACKGROUND: Allergen-specific immunotherapy for food allergy has been hindered by severe side-effects in the past. Well-characterized hypo-allergenic recombinant food allergens potentially offer a safe solution. OBJECTIVE: To demonstrate hypo-allergenicity of a mutated major food allergen from apple, Mal d 1, in vitro and in vivo. METHODS: A mutant of the major apple allergen, Mal d 1, was obtained by site-directed mutagenesis exchanging five amino acid residues. Fourteen patients with combined birch pollen-related apple allergy were included in the study. Hypo-allergenicity of the mutant rMal d 1 (rMal d 1mut) compared with rMal d 1 was assessed by in vitro methods, i.e. RAST (inhibition), immunoblotting and basophil histamine release (BHR) and in vivo by skin prick test and double-blind placebo-controlled food challenge (DBPCFC). RESULTS: RAST analysis (n = 14) revealed that IgE reactivity to rMal d 1mut was twofold lower than that of the wild-type molecule (95% confidence interval (CI): 1.7-2.4). RAST inhibition (n = 6) showed a 7.8-fold decrease in IgE-binding potency (95% CI: 3.0-12.6). In contrast to this moderate decrease in IgE-binding potency, the biological activity of rMal d 1mut assessed by SPT and BHR decreased 10-200-fold. Hypo-allergenicity was confirmed by DBPCFC (n = 2) with both recombinant molecules. CONCLUSION: A moderate decrease in IgE-binding potency translates into a potent inhibition of biological activity. This is the first study that confirms by DBPCFC that a mutated recombinant major food allergen is clinically hypo-allergenic. This paves the way towards safer immunotherapy for the treatment of food-allergic patients.

Severe allergy to sharon fruit caused by birch pollen.

BACKGROUND: Allergy to sharon fruit (persimmon) has been only rarely reported. Cross-reactivity with pollen (profilin and Bet v 6) appeared to be involved, but Bet v 1 has not been implicated previously. OBJECTIVE: It is our aim to identify whether Bet v 1 sensitization is linked to sharon fruit allergy. METHODS: Two patients with a reaction upon first exposure to sharon fruit were included in the study, as well as 7 patients with birch-pollen-related apple allergy. Sensitivity was assessed by skin prick testing (SPT), a radio-allergosorbent test (RAST) and immunoblotting. RAST analysis was performed for Bet v 1, Bet v 2 and Bet v 6. Cross-reactivity was evaluated by RAST and immunoblot inhibitions. Biological activity of IgE was measured by basophil histamine release. Sharon fruit allergy was evaluated by double-blind placebo-controlled food challenge (DBPCFC) or open challenge (OC). RESULTS: Both sharon-fruit-allergic patients demonstrated positive reactions in the RAST (8.6 and 6.2 IU/ml, respectively) and SPT (wheal area 37 and 36 mm2). Sharon fruit allergy was confirmed by DBPCFC in 1 patient. The second patient refused a challenge because of the severe initial reaction. Sera from both patients were reactive to Bet v 1 and Bet v 6, which was cross-reactive with sharon fruit by inhibition assays. The patient with the severest reactions was reactive to profilin on immunoblotting. However, profilin did not induce significant histamine release, nor did Bet v 6. Bet v 1 induce approximately 60% histamine release. An OC with sharon fruit in 7 patients allergic to birch pollen and apple, who had not eaten sharon fruit previously, was positive in 6/7 cases. CONCLUSIONS: Birch-pollen-related allergy to sharon fruit is mediated by the known cross-reactive pollen allergens including Bet v 1 and may become more of a problem should sharon fruit consumption increase.

Allergy to jackfruit: a novel example of Bet v 1-related food allergy.

BACKGROUND: Jackfruit allergy has been reported just once. It is unknown whether this food allergy is caused by direct sensitization or cross-sensitization to pollen allergens. OBJECTIVE: Establish whether jackfruit allergy is linked to birchpollen allergy. METHODS: Two jackfruit allergic patients and five patients with birchpollen-related apple allergy were recruited. Sensitization to pollen and plant foods was assessed by skin prick test (SPT), radio-allergosorbent test (RAST) and immunoblot. RAST analysis was performed for Bet v 1 and Mal d 1. Cross-reactivity was evaluated by RAST and immunoblot-inhibition. Biological activity of immunoglobulin E (IgE) was measured by basophil histamine release. Allergy to jackfruit was evaluated by double-blind placebo-controlled food challenge (DBPCFC) or open challenge (OC). RESULTS: In both patients DBPCFC confirmed the reported jackfruit allergy. SPT was 41 and 27 mm2 and specific IgE to jackfruit was 5.9 and 0.8 IU/ml, respectively. Immunoblot analysis revealed IgE reactivity at Mr of approximately 17 kDa. The Bet v 1-related nature of this allergen in jackfruit was demonstrated by RAST and immunoblot inhibition. To assess whether jackfruit allergy might be common in patients with combined birchpollen-fruit allergy, five such patients underwent an OC with jackfruit. All five had OA-like symptoms. CONCLUSIONS: Jackfruit allergy can be added to the list of birchpollen-related food allergies. Increased consumption of this fruit will result in a rise in allergic reactions.

Efficacy of birch-pollen immunotherapy on cross-reactive food allergy confirmed by skin tests and double-blind food challenges.

BACKGROUND: The effect of birch-pollen immunotherapy (IT) on cross-reactive food allergies is controversial. OBJECTIVE: The aim of this study was to investigate the effect of birch-pollen IT on apple allergy and to evaluate recombinant allergens and double-blind placebo-controlled food challenges (DBPCFCs) as monitoring tools. METHODS: Twenty-five adult birch-pollen- and apple-allergic patients were randomly divided into two groups, either receiving birch-pollen IT or symptomatic drugs only. IgE and IgG4 antibodies against birch pollen, apple, natural Bet v 1 and Mal d 1 were measured. In addition, skin prick tests (SPT) were performed using recombinant Bet v 1 (rBet v 1) and Mal d 1 (rMal d 1). Clinical outcome was evaluated by DBPCFC. CD4(+)CD25(+) regulatory T cells (Tregs) were isolated from peripheral blood and tested in functional assays. RESULTS: Birch-pollen IT resulted in a significant decrease of SPT reactivity for rBet v 1 (30-fold) and rMal d 1 (10-fold) already after 3 months. IgG4 antibodies were potently induced against Bet v 1, displaying cross-reactivity to Mal d 1. Visual analogue scale scores decreased >10-fold in 9/13 patients of the IT group, with three patients converting to negative. In the control group, no decrease was observed. Birch-pollen IT did not lead to detectable changes in the number or function of the CD4(+)CD25(+) Tregs. CONCLUSIONS: This trial supports the claims that birch-pollen IT also decreases allergy to foods containing Bet v 1-homologous allergens. Recombinant allergens and DBPCFCs have proven to be useful tools for monitoring the effect of birch-pollen IT on linked food allergies.