Heme catabolism by tumor-associated macrophages controls metastasis formation.

Although the pathological significance of tumor-associated macrophage (TAM) heterogeneity is still poorly understood, TAM reprogramming is viewed as a promising anticancer therapy. Here we show that a distinct subset of TAMs (F4/80hiCD115hiC3aRhiCD88hi), endowed with high rates of heme catabolism by the stress-responsive enzyme heme oxygenase-1 (HO-1), plays a critical role in shaping a prometastatic tumor microenvironment favoring immunosuppression, angiogenesis and epithelial-to-mesenchymal transition. This population originates from F4/80+HO-1+ bone marrow (BM) precursors, accumulates in the blood of tumor bearers and preferentially localizes at the invasive margin through a mechanism dependent on the activation of Nrf2 and coordinated by the NF-κB1-CSF1R-C3aR axis. Inhibition of F4/80+HO-1+ TAM recruitment or myeloid-specific deletion of HO-1 blocks metastasis formation and improves anticancer immunotherapy. Relative expression of HO-1 in peripheral monocyte subsets, as well as in tumor lesions, discriminates survival among metastatic melanoma patients. Overall, these results identify a distinct cancer-induced HO-1+ myeloid subgroup as a new antimetastatic target and prognostic blood marker.

The prognostic role of mitotic rate in cutaneous malignant melanoma: Evidence from a multicenter study on behalf of the Italian Melanoma Intergroup.

BACKGROUND: This study aimed to improve the understanding of the prognostic value of tumor mitotic rate (TMR) in cutaneous melanoma and assessed its significance as a predictor for overall, melanoma-specific, and recurrence-free survival. PATIENTS AND METHODS: This is a retrospective multicenter Italian cohort study of 13,016 patients diagnosed with and treated for invasive primary melanoma between 2005 and 2020 with median follow-up of 5.5 years. The survival probability was assessed by Kaplan-Meier method, hazard ratios (HRs), and corresponding 95% confidence interval (CI) of all-cause mortality and recurrence/death by multivariable Cox proportional hazards models. RESULTS: Higher dermal mitoses number was associated with decreased overall survival. Among patients with TMR 0/mm2 , 1/mm2 , 2/mm2 -3/mm2 , 4/mm2 -10/mm2 , and >10/mm2 , 5-year overall survival (OS) was 97.3%, 93.6%, 88.3%, 73.0%, and 60.9%, respectively. In multivariate analyses, compared to TMR of 0/mm2 , HRs for all-cause mortality were 1.35 (95% CI, 1.08-1.68), 1.70 (95% CI, 1.40-2.07), 2.04 (95% CI, 1.67-2.49), and 2.39 (95% CI, 1.90-3.00) for 1 mitoses/mm2 , 2 mitoses/mm2 -3 mitoses/mm2 , 4 mitoses/mm2 -10 mitoses/mm2 , and >10 mitoses/mm2 , respectively. A similar increase in risks was observed in melanoma-specific survival (MSS) and recurrence-free survival (RFS). The HRs for MSS and RFS for the highest compared to the lowest TMR category were 3.01 (95% CI, 2.20-4.11) and 2.26 (95% CI, 1.88-2.73), respectively. Sentinel lymph-node biopsy positivity was significantly associated with TMR increase even with adjustment for several potential confounders. CONCLUSIONS: A clear association was demonstrated between an increasing TMR and decreased OS, MSS, and RFS, suggesting a reconsideration of TMR prognostic role for future inclusion in the melanoma staging system. PLAIN LANGUAGE SUMMARY: The 8th American Joint Committee on Cancer for melanoma staging removed tumor mitotic rate (TMR) from the staging criteria for T1 melanomas, giving way to ulceration and tumor thickness as stronger prognostic predictors. However, it is still recommended that TMR should be assessed and recorded in all primary invasive melanomas. In a large retrospective multicenter study on primary invasive melanomas, we investigated the prognostic value of TMR to assess its significance as survival predictor. Our results showed a clear association between increasing TMR and decreased patients' survival, suggesting that TMR should be considered for inclusion in the melanoma staging system.

Clinical management of melanocytic tumours of uncertain malignant potential (MelTUMPs), including melanocytomas: A systematic review and meta-analysis.

Little guidance is currently available for managing patients with melanocytic tumours of uncertain or low malignant potential (MelTUMPs, including melanocytomas), in particular the optimal excision margins and whether to offer sentinel node biopsy (SNB). The objective of this review was to evaluate excision margins and the prognostic utility of SNB by systematic review of the literature and meta-analysis. PRISMA guidelines were followed. Medline, EMBASE and Cochrane databases were searched to October 2021 for studies of patients with MelTUMPs reporting excision margins and/or SNB-positivity. Meta-analysis was performed on the SNB-positivity rate using a random effects model, followed by sensitivity analyses on subgroups. 111 primary studies reported excision margins and/or SNB data for 1962 patients. Follow-up was available for 1649 patients: 1561 (94.7%) were alive without disease at last review, 53 (3.2%) had developed further disease, 29 (1.8%) had died of metastatic disease (melanoma) and six (0.4%) died of unrelated causes. SNB was performed in 837 patients. The pooled positivity rate on meta-analysis was 32% (95% CI: 23-44%). Clinical outcome could be correlated with excision margin in only 171 patients (60% of those with known follow up) and was therefore not analysed further. Evidence indicating the ideal excision margins for MelTUMPs was lacking. SNB had a high positivity rate despite very low rates of recurrence or melanoma-related death. Consequently, SNB should not be offered routinely for MelTUMPs (including melanocytomas), due to its lack of prognostic utility for this tumour type (high certainty of evidence).

Similar local recurrence and survival in patients with T1 radial growth phase melanoma on head and neck treated with 5 or 10 mm margins: A retrospective study.

BACKGROUND: Melanoma guidelines recommend surgical excision with 10 mm margins for T1 melanomas (invasive melanomas with Breslow thickness ≤1 mm), including those in radial growth phase, which are without metastatic potential; however, such margins may be problematic on head-and-neck. OBJECTIVE: We compared outcomes of wide (10 mm margins) versus narrow (5 mm margins) excisions in patients with radial growth phase T1 melanoma on head-and-neck including face. METHODS: We retrospectively examined 610 consecutive patients excised with wide versus narrow margins, from 2001 to 2018, at six European centres. In all cases, radial growth phase, and clear margins with 5 or 10 mm of clearance, were ascertained histologically. Multivariable models investigated associations of margins and other factors with overall survival and local recurrence. RESULTS: Three hundred and sixteen (51.8%) patients received wide excision, 219 (69.3%) with primary wound closure, 97 (30.7%) with reconstruction; 294 (48.2%) patients received narrow excision, 264 (89.8%) with primary wound closure, 30 (10.2%) with reconstruction (p < 0.001). Median follow-ups were 88 months (wide) and 187 months (narrow) (inter-quartile ranges 43-133 and 79-206, respectively). Ten-year overall survival (95% confidence interval) was 96.7% (94.2%-99.3%) in wide and 98.2% (96.4%-100%) in narrow patients. Ten-year local recurrence incidence was 6.4% (4.1%-10.1%) in wide and 7.8% (5.3%-11.6%) in narrow groups. Lentigo maligna melanoma subtype appeared associated with increased risk of local recurrence in narrow versus wide patients (15.0% vs. 7.5%; p = 0.190). CONCLUSIONS: Narrower excision margins for T1 radial growth phase melanoma are not associated with worse overall survival (hazard ratio 0.97, p = 0.996) or increased local recurrence (subdistribution hazard ratio: 0.87; p = 0.751) compared to wider margins, and may be safely applied to such lesions, although caution may be required in the presence of lentigo maligna melanoma.

The role of sentinel node tumor burden in modeling the prognosis of melanoma patients with positive sentinel node biopsy: an Italian melanoma intergroup study (N = 2,086).

BACKGROUND: The management of melanoma patients with metastatic melanoma in the sentinel nodes (SN) is evolving based on the results of trials questioning the impact of completion lymph node dissection (CLND) and demonstrating the efficacy of new adjuvant treatments. In this landscape, new prognostic tools for fine risk stratification are eagerly sought to optimize the therapeutic path of these patients. METHODS: A retrospective cohort of 2,086 patients treated with CLND after a positive SN biopsy in thirteen Italian Melanoma Centers was reviewed. Overall survival (OS) was the outcome of interest; included independent variables were the following: age, gender, primary melanoma site, Breslow thickness, ulceration, sentinel node tumor burden (SNTB), number of positive SN, non-sentinel lymph nodes (NSN) status. Univariate and multivariate survival analyses were performed using the Cox proportional hazard regression model. RESULTS: The 3-year, 5-year and 10-year OS rates were 79%, 70% and 54%, respectively. At univariate analysis, all variables, except for primary melanoma body site, were found to be statistically significant prognostic factors. Multivariate Cox regression analysis indicated that older age (P < 0.0001), male gender (P = 0.04), increasing Breslow thickness (P < 0.0001), presence of ulceration (P = 0.004), SNTB size (P < 0.0001) and metastatic NSN (P < 0.0001) were independent negative predictors of OS. CONCLUSION: The above results were utilized to build a nomogram in order to ease the practical implementation of our prognostic model, which might improve treatment personalization.

Melanoma recurrence patterns and management after adjuvant targeted therapy: a multicentre analysis.

BACKGROUND: Adjuvant targeted therapy (TT) improves relapse free survival in patients with resected BRAF mutant stage III melanoma. The outcomes and optimal management of patients who relapse after adjuvant TT is unknown. METHODS: Patients from twenty-one centres with recurrent melanoma after adjuvant TT were included. Disease characteristics, adjuvant therapy, recurrence, treatment at relapse and outcomes were examined. RESULTS: Eighty-five patients developed recurrent melanoma; nineteen (22%) during adjuvant TT. Median time to first recurrence was 18 months and median follow-up from first recurrence was 31 months. Fifty-eight (68%) patients received immunotherapy (IT) or TT as 1st line systemic therapy at either first or subsequent recurrence and had disease that was assessable for response. Response to anti-PD-1 (±trial agent), combination ipilimumab-nivolumab, TT rechallenge and ipilimumab monotherapy was 63%, 62% 25% and 10% respectively. Twenty-eight (33%) patients had died at census, all from melanoma. Two-year OS was 84% for anti-PD-1 therapy (±trial agent), 92% for combination ipilimumab and nivolumab, 49% for TT and 45% for ipilimumab monotherapy (p = 0.028). CONCLUSIONS: Patients who relapse after adjuvant TT respond well to subsequent anti-PD-1 based therapy and have outcomes similar to those seen when first line anti-PD-1 therapy is used in stage IV melanoma.

Liquid Biopsy and Radiological Response Predict Outcomes Following Discontinuation of Targeted Therapy in Patients with BRAF Mutated Melanoma.

BACKGROUND: Outcomes of patients with metastatic melanoma discontinuing BRAF-targeted therapy for cumulative toxicity after sustained response are unknown. MATERIALS AND METHODS: This retrospective case series analysis conducted at a single Cancer Center in Italy included patients with BRAF mutated metastatic melanoma treated with a BRAF inhibitor as a single agent or in combination with a MEK inhibitor between June 1, 2011 and January 1, 2020 and interrupted treatment due to cumulative toxicity after achieving complete response (CR) or long-lasting partial response (PR; i.e. >12 months). RESULTS: We included 24 patients with a median treatment duration of 59.4 months (95% confidence interval [CI], 55.4-63.4; range, 12-88). CR and PR were achieved in 71% and 29% of patients, respectively. At a median follow-up after treatment discontinuation of 37.8 months (95% CI, 33.7-41.9), the 12-month progression-free survival after discontinuation (dPFS) rate was 70.8% (95% CI 54.8-91.6) and 24-month dPFS rate was 58.3% (95% CI, 41.6-81.8). Baseline patient and tumor characteristics as well as treatment duration and best response did not significantly impact on dPFS. Patients with CR and negative circulating tumor DNA (ctDNA) at time of discontinuation had a significantly improved dPFS compared with patients with either radiological residual disease or ctDNA positivity (p = .007). No patient in CR with undetectable ctDNA experienced progression. CONCLUSION: The risk of progression is high even in patients with sustained sensitivity to BRAF/MEK inhibitors. Integration of liquid biopsy in clinical trials investigating the optimal management of patients with sustained sensitivity to BRAF/MEK inhibitors is warranted. IMPLICATIONS FOR PRACTICE: Outcomes of patients with metastatic melanoma discontinuing BRAF-targeted therapy for cumulative toxicity are unknown. This study analyzed patients with sustained responses (median treatment duration 59.4 months). Twelve- and 24-month progression-free survival following discontinuation were 70.8% and 58.3%, respectively. Complete response and negative circulating tumor DNA at time of discontinuation are promising prognostic biomarkers in this setting.

Survival in Patients With Sentinel Node-Positive Melanoma With Extranodal Extension.

BACKGROUND: Prognostic parameters in sentinel node (SN)-positive melanoma are important indicators to identify patients at high risk of recurrence who should be candidates for adjuvant therapy. We aimed to evaluate the presence of melanoma cells beyond the SN capsule-extranodal extension (ENE)-as a prognostic factor in patients with positive SNs. METHODS: Data from 1,047 patients with melanoma and positive SNs treated from 2001 to 2020 at the Istituto Nazionale dei Tumori in Milano, Italy, were retrospectively investigated. Kaplan-Meier survival and crude cumulative incidence of recurrence curves were estimated. A multivariable logistic model was used to investigate the association between ENE and selected predictive factors. Cox models estimated the effect of the selected predictors on survival endpoints. RESULTS: Median follow-up was 69 months. The 5-year overall survival rate was 62.5% and 71.7% for patients with positive SNs with and without ENE, respectively. The 5-year disease-free survival rate was 54.0% and 64.0% for patients with positive SNs with and without ENE, respectively. The multivariable logistic model showed that age, size of the main metastatic focus in the SN, and numbers of metastatic non-SNs were associated with ENE (all P<.0001). The multivariable Cox regression models showed the estimated prognostic effects of ENE associated with age, ulceration, size of the main metastatic focus in the SN, and number of metastatic non-SNs (all P<.0001) on disease-free survival and overall survival. CONCLUSIONS: ENE was a significant prognostic factor in patients with positive-SN melanoma. This parameter may be useful in clinical practice as a selection criterion for adjuvant treatment in patients with stage IIIA disease with a tumor burden <1 mm in the SN. We recommend its inclusion as an independent prognostic determinant in future updates of melanoma guidelines.

Analysis of Sentinel Node Biopsy and Clinicopathologic Features as Prognostic Factors in Patients With Atypical Melanocytic Tumors.

BACKGROUND: Atypical melanocytic tumors (AMTs) include a wide spectrum of melanocytic neoplasms that represent a challenge for clinicians due to the lack of a definitive diagnosis and the related uncertainty about their management. This study analyzed clinicopathologic features and sentinel node status as potential prognostic factors in patients with AMTs. PATIENTS AND METHODS: Clinicopathologic and follow-up data of 238 children, adolescents, and adults with histologically proved AMTs consecutively treated at 12 European centers from 2000 through 2010 were retrieved from prospectively maintained databases. The binary association between all investigated covariates was studied by evaluating the Spearman correlation coefficients, and the association between progression-free survival and all investigated covariates was evaluated using univariable Cox models. The overall survival and progression-free survival curves were established using the Kaplan-Meier method. RESULTS: Median follow-up was 126 months (interquartile range, 104-157 months). All patients received an initial diagnostic biopsy followed by wide (1 cm) excision. Sentinel node biopsy was performed in 139 patients (58.4%), 37 (26.6%) of whom had sentinel node positivity. There were 4 local recurrences, 43 regional relapses, and 8 distant metastases as first events. Six patients (2.5%) died of disease progression. Five patients who were sentinel node-negative and 3 patients who were sentinel node-positive developed distant metastases. Ten-year overall and progression-free survival rates were 97% (95% CI, 94.9%-99.2%) and 82.2% (95% CI, 77.3%-87.3%), respectively. Age, mitotic rate/mm2, mitoses at the base of the lesion, lymphovascular invasion, and 9p21 loss were factors affecting prognosis in the whole series and the sentinel node biopsy subgroup. CONCLUSIONS: Age >20 years, mitotic rate >4/mm2, mitoses at the base of the lesion, lymphovascular invasion, and 9p21 loss proved to be worse prognostic factors in patients with ATMs. Sentinel node status was not a clear prognostic predictor.

Pediatric patients with cutaneous melanoma: A European study.

INTRODUCTION: Cutaneous melanoma is rare in childhood and published studies have mainly been retrospective single-institution series or small case series. Given the absence of clinical protocols dedicated to pediatric melanoma, the treatment approach is generally extrapolated from the ones applied to adults. METHODS: Coordinated by the European Cooperative Study Group for Pediatric Rare Tumors (EXPeRT), this study collected patients prospectively registered between 2002 and 2012 under national cooperative projects dedicated to rare pediatric tumors in Italy, Poland, Germany, and France. Additional cases were collected from dermatology registries in Germany and Israel. RESULTS: A total of 219 patients aged 0-18 years (median 14.4) were included in the analysis. Sentinel lymph node biopsy was performed in 112 patients (76% of those with Breslow thickness > 0.75 mm) and was positive in 37.5%. Systemic therapy was used in 33 cases. In stage III cases, survival rates were similar for patients who received (23 cases) or not (21 cases) adjuvant therapy. For the whole series, 3-year overall and disease-free survival rates were 91.4% and 84.0%, respectively (median follow-up 41.8 months). Tumor site, tumor stage, and ulceration influenced survival rates. Patients treated by pediatric oncologists (n = 140) were more likely to have advanced disease than those treated by dermatologists (n = 79). DISCUSSION: This study would suggest that the clinical history of melanoma in children and adolescents might resemble that of adult counterpart. Cooperative efforts are needed to make new drugs more readily available to pediatric patients to increase the outcome of patient with advanced disease.

Multiple primary melanomas (MPMs) and criteria for genetic assessment: MultiMEL, a multicenter study of the Italian Melanoma Intergroup.

BACKGROUND: Multiple primary melanoma (MPM), in concert with a positive family history, is a predictor of cyclin-dependent kinase (CDK) inhibitor 2A (CDKN2A) germline mutations. A rule regarding the presence of either 2 or 3 or more cancer events (melanoma and pancreatic cancer) in low or high melanoma incidence populations, respectively, has been established to select patients for genetic referral. OBJECTIVE: We sought to determine the CDKN2A/CDK4/microphthalmia-associated transcription factor mutation rate among Italian patients with MPM to appropriately direct genetic counseling regardless of family history. METHODS: In all, 587 patients with MPM and an equal number with single primary melanomas and control subjects were consecutively enrolled at the participating centers and tested for CDKN2A, CDK4, and microphthalmia-associated transcription factor. RESULTS: CDKN2A germline mutations were found in 19% of patients with MPM versus 4.4% of patients with single primary melanoma. In familial MPM cases the mutation rate varied from 36.6% to 58.8%, whereas in sporadic MPM cases it varied from 8.2% to 17.6% in patients with 2 and 3 or more melanomas, respectively. The microphthalmia-associated transcription factor E318K mutation accounted for 3% of MPM cases altogether. LIMITATIONS: The study was hospital based, not population based. Rare novel susceptibility genes were not tested. CONCLUSION: Italian patients who developed 2 melanomas, even in situ, should be referred for genetic counseling even in the absence of family history.

Armed antibodies for cancer treatment: a promising tool in a changing era.

Advances in the understanding of tumor immunology and molecular biology of melanoma cells have favored a larger application of immunotherapy and targeted therapies in the clinic. Several selective mutant gene inhibitors and immunomodulating antibodies have been reported to improve overall survival or progression-free survival in metastatic melanoma patients. However, despite impressive initial responses, patients treated with selective inhibitors relapse quickly, and toxicities associated to the use of immunomodulating antibodies are not easily manageable. In this sense, the concept of using antibodies as delivery vehicles for the preferential in vivo localization of the drug at the site of disease with reduction of side effects has raised particular interest. Antibody-cytokine fusion proteins (termed immunocytokines) represent a new simple and effective way to deliver the immunomodulatory payload at the tumor site, with the aim of inducing both local and systemic antitumoral immune responses and limiting systemic toxicities. Several clinical trials have been conducted and are actually ongoing with different immunocytokines, in several tumor histotypes. In metastatic melanoma patients, different drug delivery modalities such as systemic, loco-regional and intratumoral are under investigation. In this review, the rationale for the use of L19-IL2 and L19-TNF, two clinical stage immunocytokines produced by the Philogen group, as well as opportunities for their future development will be discussed.

Intralesional administration of L19-IL2/L19-TNF in stage III or stage IVM1a melanoma patients: results of a phase II study.

The intratumoral injection of cytokines, in particular IL2, has shown promise for cutaneous melanoma patients with unresectable disease or continuous recurrence despite surgery. We recently reported that the intralesional injection of L19-IL2, an immunocytokine combining IL2 and the human monoclonal antibody fragment L19, resulted in efficient regional control of disease progression, increased time to distant metastasis and evidence of effect on circulating immune cell populations. We have also shown in preclinical models of cancer a remarkable synergistic effect of the combination of L19-IL2 with L19-TNF, a second clinical-stage immunocytokine, based on the same L19 antibody fused to TNF. Here, we describe the results of a phase II clinical trial based on the intralesional administration of L19-IL2 and L19-TNF in patients with stage IIIC and IVM1a metastatic melanoma, who were not candidate to surgery. In 20 efficacy-evaluable patients, 32 melanoma lesions exhibited complete responses upon intralesional administration of the two products, with mild side effects mainly limited to injection site reactions. Importantly, we observed complete responses in 7/13 (53.8 %) non-injected lesions (4 cutaneous, 3 lymph nodes), indicating a systemic activity of the intralesional immunostimulatory treatment. The intralesional administration of L19-IL2 and L19-TNF represents a simple and effective method for the local control of inoperable melanoma lesions, with a potential to eradicate them or make them suitable for a facile surgical removal of the residual mass.

A large de novo 9p21.3 deletion in a girl affected by astrocytoma and multiple melanoma.

BACKGROUND: Association of melanoma, neural system tumors and germ line mutations at the 9p21 region in the CDKN2A, CDKN2B and CDKN2BAS genes has been reported in a small number of families worldwide and described as a discrete syndrome in melanoma families registered as a rare disease, the melanoma-astrocytoma syndrome. CASE PRESENTATION: We here studied two young patients developing melanoma after radiotherapy for astrocytoma, both reporting lack of family history for melanoma or neural system tumors at genetic counselling. Patient A is a girl treated for anaplastic astrocytoma at 10 years and for multiple melanomas on the scalp associated to dysplastic nevi two years later. Her monozygotic twin sister carried dysplastic nevi and a slow growing, untreated cerebral lesion. Direct sequencing analysis showed no alterations in melanoma susceptibility genes including CDKN2A, CDK4, MC1R and MITF or in TP53. By microsatellite analysis, multiplex ligation-dependent probe amplification, and array comparative genomic hybridization a deletion including the CDKN2A, CDKN2B and CDKN2BAS gene cluster was detected in both twin sisters, encompassing a large region at 9p21.3 and occurring de novo after the loss of one paternal allele.Patient B is a boy of 7 years when treated for astrocytoma then developing melanoma associated to congenital nevi on the head 10 years later: sequencing and multiplex ligation-dependent probe amplification revealed a normal profile of the CDKN2A/CDKN2B/CDKN2BAS region. Array comparative genomic hybridization confirmed the absence of deletions at 9p21.3 and failed to reveal known pathogenic copy number variations. CONCLUSIONS: By comparison with the other germ line deletions at the CDKN2A, CDKN2B and CDKN2BAS gene cluster reported in melanoma susceptible families, the deletion detected in the two sisters is peculiar for its de novo origin and for its extension, as it represents the largest constitutive deletion at 9p21.3 region identified so far.In addition, the two studied cases add to other evidence indicating association of melanoma with exposure to ionizing radiation and with second neoplasm after childhood cancer. Melanoma should be considered in the monitoring of pigmented lesions in young cancer patients.

Cutaneous melanoma in children and adolescents: the Italian rare tumors in pediatric age project experience.

OBJECTIVE: To describe a series of cutaneous melanoma in children collected by the Italian Rare Tumors in Pediatric Age project. STUDY DESIGN: From 2000 to 2012, 54 patients younger than 18 years of age were prospectively registered and treated at 12 Italian pediatric centers on the basis of the same diagnostic/therapeutic recommendations and with the same forms to record clinical data. RESULTS: Considering the estimated annual incidence in Italy, the registered cases accounted for 30% of those expected in children and 10% of adolescents. Clinically, 47% of the tumors were amelanotic and 81% were raised, 39% of cases had tumor thickness >2 mm, and 36% had lymph node involvement. For the whole series, 5-year event-free survival and overall survival rates were 75.2% and 84.6%, respectively. Patient survival correlated with tumor stage and ulceration. No relapses were recorded for T1-2 (thickness <2 mm), N0, and stage 0-I-II cases. CONCLUSION: We suggest that the variables influencing survival in children with melanoma are the same as for adults, the clinical approach used in adults is feasible in children, and pediatric cases are more likely to have advanced disease at diagnosis but similar survival. New effective drugs are needed for advanced disease, and biological studies and international cooperative schemes are warranted.

Association of promoter polymorphism -765G>C in the PTGS2 gene with malignant melanoma in Italian patients and its correlation to gene expression in dermal fibroblasts.

Prostaglandins, especially prostaglandin E synthetase (PGE2), influence carcinogenesis by promoting cell proliferation, inhibiting apoptosis, stimulating angiogenesis and mediating immune suppression. Cyclooxygenase-2, coded by the PTGS2 gene, is the key enzyme in the production of prostaglandins. In melanoma, Cox-2 is over expressed in primary malignant melanoma (MM) and in their corresponding metastases. Polymorphisms in the promoter region of PTGS2 gene can modulate gene expression and could modify individual susceptibility to MM. Two hundred and forty melanoma patients and 342 controls were genotyped for polymorphisms -765G>C (rs20417) and -1195A>G (rs689466). Allele -765C frequency was significantly higher in melanoma patients. No allele frequency differences for -1195A>G polymorphism were observed. Haplotype analysis revealed that the haplotypes carrying the minor alleles were associated to a higher risk of melanoma (P = 0.02). Expression analysis showed that allele -765C is associated to a higher gene expression and could represent a risk allele by affecting the functionality of the promoter.

Accuracy and prognostic value of sentinel lymph node biopsy in head and neck melanomas.

BACKGROUND: Debate remains around the accuracy and prognostic implications of sentinel lymph node biopsy (SLNB) for melanoma arising in the head and neck (HN) areas because several analyses have shown discordances between clinically predicted lymphatic drainage pathways and those identified by lymphoscintigraphy. This study assesses the accuracy and prognostic value of SLNB in this critical anatomic region. METHODS: Retrospective review of a prospectively collected melanoma database identified 331 patients with HN melanomas from January 2000 to December 2012. Primary end points included SLNB result, time to recurrence, site of recurrence, and survival. Multivariate models were constructed for analyses. RESULTS: A sentinel lymph node (SLN) was identified in all 331 patients. There were 59 patients with a positive SLN (17.8%) with a recurrence rate of 88.1% compared with 22.4% in SLN-negative patients (P < 0.0001). The 5-y overall survival was 91.2% for SLN-negative patients and 48.7% for SLN-positive patients (P < 0.0001). Patients with scalp melanoma had thicker lesions and an elevated risk of SLN positivity, recurrence, and death compared with those with other sites. Among the 272 SLN-negative patients, four patients developed regional nodal disease in the same basin and had undergone a previous SLNB procedure for a false-omission rate of 1.45%. Risks for false-negative SLN occurrences included thick and scalp melanomas. Multivariate analysis on prognostic factors affecting relapse-free survival showed positive SLNB status to be the most prognostic clinicopathologic predictor of recurrence (hazard ratio, 20.56; P < 0.0001). CONCLUSIONS: SLNB for patients with HN melanomas is an accurate procedure and has prognostic value.

Effects of cyclophosphamide and IL-2 on regulatory CD4+ T cell frequency and function in melanoma patients vaccinated with HLA-class I peptides: impact on the antigen-specific T cell response.

The frequency and function of regulatory T cells (Tregs) were studied in stage II-III melanoma patients who were enrolled in a phase II randomized trial of vaccination with HLA-A*0201-modified tumor peptides versus observation. The vaccinated patients received low-dose cyclophosphamide (CTX) and low-dose interleukin-2 (IL-2). Tregs were analyzed in the lymph nodes (LNs) of stage III patients who were undergoing complete lymph node dissection and in peripheral blood mononuclear cells (PBMCs) collected before vaccination and at different time points during the vaccination period. The LNs of the vaccinated patients, which were surgically removed after two rounds of vaccination and one dose of CTX, displayed a low frequency of Tregs and a less immunosuppressive environment compared with those of the untreated patients. The accurate time-course analysis of the PBMCs of patients enrolled in the vaccination arm indicated a limited and transient modulation in the frequencies of Tregs in PBMCs collected after low-dose CTX administration and a strong Treg boost in those PBMCs collected after low-dose IL-2 administration. However, a fraction of the IL-2-boosted Tregs was functionally modulated to a Th-1-like phenotype in the vaccinated patients. Moreover, low-dose IL-2 promoted the concomitant expansion of conventional activated CD4(+) T cells. Despite the amplification of Tregs, IL-2 administration maintained or further increased the number of antigen-specific CD8(+) T cells that were induced by vaccination as demonstrated by the ex vivo human leukocyte antigen-multimer staining and IFN-γ ELISpot assays. Our study suggests that the use of CTX as a Treg modulator should be revised in terms of the administration schedule and of patients who may benefit from this drug treatment. Despite the Treg expansion that was observed in this study, low-dose IL-2 is not detrimental to the functional activities of vaccine-primed CD8(+) T cell effectors when used in the inflammatory environment of vaccination.

CDKN2A and MC1R variants influence dermoscopic and confocal features of benign melanocytic lesions in multiple melanoma patients.

Non-invasive diagnostic tools are effective in the histomorphological study of melanocytic lesions. The role of melanoma susceptibility genes on melanocytic nevi histopathological features is not clear. The current study aimed to correlate genetic alterations and histomorphological features of melanocytic nevi. Clinical, dermoscopic and confocal features of 34 multiple melanoma patients and 34 controls were compared. Among patients with melanoma, carriers of CDKN2A mutations and/or MC1R variants, and wild-type genes were also compared. In patients with melanoma, a lighter phototype (P = 0.051), a higher number of nevi (P < 0.01) and clinically atypical nevi (P < 0.01) were observed. At dermoscopy, these nevi showed a complex pattern (P = 0.011), atypical network (P = 0.018) and irregular pigmentation (P = 0.037); at confocal, an irregular meshwork pattern (P = 0.026) with atypical nests (P = 0.016) and an inflammatory infiltrate (P = 0.048) were observed. Among patients with melanoma genetically tested, CDKN2A G101W mutation carriers were more frequently younger (P = 0.023), with clinically atypical nevi (P = 0.050), with cytological atypia (P = 0.033) at confocal. G101W mutation and MC1R variants carriers showed hypopigmented nevi (P = 0.002) and, at confocal, roundish cells infiltrating the junction (P = 0.019). These data suggest an influence of CDKN2A mutation and MC1R variants in the development of dysplastic melanocytic lesions. Non-invasive histomorphological evaluation, together with genetic studies, improves melanoma risk identification and early diagnosis, for a patient-tailored management.

Isolated limb perfusion with the tumor-targeting human monoclonal antibody-cytokine fusion protein L19-TNF plus melphalan and mild hyperthermia in patients with locally advanced extremity melanoma.

BACKGROUND: L19-TNF is a tumor-targeting immunocytokine composed of the human L19 antibody binding to extra domain B (ED-B) of fibronectin of newly formed blood vessels, and of human TNF. This exploratory trial evaluates safety and clinical activity of L19-TNF plus melphalan-containing isolated limb perfusion (ILP) in extremity melanoma patients. METHODS: Seven and 10 patients received 325 µg and 650 µg of L19-TNF, respectively, during the ILP. Patients were studied for safety, tolerability, and clinical activity of this experimental L19-TNF ILP procedure. RESULTS: Non-hematologic toxicity of L19-TNF ILP was very low, but severe myelosuppression was seen in four patients. Although L19-TNF was administered at a TNF-equivalent dose of only 3.13 and 6.25% of the approved TNF (Beromun®) dose of 4 mg, L19-TNF ILP induced objective responses in 86 and 89% of patients, respectively, including a complete response (CR) in 5/10 patients treated with L19-TNF ILP at 650 µg that was durable at 12 months in four patients. No CR was seen at 325 µg of L19-TNF. CONCLUSIONS: ILP with L19-TNF had a favorable safety and a promising activity profile at a dose of 650 µg of L19-TNF, supporting the exploration of higher L19-TNF doses and a Phase II trial comparing L19-TNF ILP with standard melphalan-containing ILP.