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Immune cells infiltrating the microenvironment of melanoma metastases may either limit or promote tumor progression, but the characteristics that distinguish these effects are obscure. In this study, we systematically evaluated the composition and organization of immune cells that infiltrated melanoma metastases in human patients. Three histologic patterns of immune cell infiltration were identified, designated immunotypes A, B, and C. Immunotype A was characterized by no immune cell infiltrate. Immunotype B was characterized by infiltration of immune cells limited only to regions proximal to intratumoral blood vessels. Immunotype C was characterized by a diffuse immune cell infiltrate throughout a metastatic tumor. These immunotypes represented 29%, 63%, and 8% of metastases with estimated median survival periods of 15, 23, and 130 months, respectively. Notably, from immunotypes A to C, there were increasing proportions of B cells and decreasing proportions of macrophages. Overall, the predominant immune cells were T cells (53%), B cell lineage cells (33%), and macrophages (13%), with natural killer and mature dendritic cells only rarely present. Whereas higher densities of CD8+ T cells correlated best with survival, a higher density of CD45+ leukocytes, T cells, and B cells also correlated with increased survival. Together, our findings reveal striking differences in the immune infiltrate in melanoma metastases in patients, suggesting microenvironmental differences in immune homing receptors and ligands that affect immune cell recruitment. These findings are important, not only by revealing how the immune microenvironment can affect outcomes but also because they reveal characteristics that may help improve individualized therapy for patients with metastatic melanoma. [ABSTRACT FROM AUTHOR]

The strength and durability of systemic anti-tumor immune responses induced by cancer vaccines depends on adjuvants to support an immunogenic vaccine site microenvironment (VSME). Adjuvants include water-in-oil emulsions with incomplete Freund's adjuvant (IFA) and combinations of toll-like receptor (TLR) agonists, including a preparation containing TLR4 and TLR9 agonists with QS-21 (AS15). IFA-containing vaccines can promote immune cell accumulation at the VSME, whereas effects of AS15 are largely unexplored. Therefore, we assessed innate and adaptive immune cell accumulation and gene expression at the VSME after vaccination with AS15 and compared to effects with IFA. We hypothesized that AS15 would promote less accumulation of innate and adaptive immune cells at the VSME than IFA vaccines. In two clinical trials, patients with resected high-risk melanoma received either a multipeptide vaccine with IFA or a recombinant MAGE-A3 protein vaccine with AS15. Vaccine site biopsies were obtained after one or multiple vaccines. T cells accumulated early after vaccines with AS15, but this was not durable or of the same magnitude as vaccination in IFA. Vaccines with AS15 increased durable expression of DC- and T cell-related genes, as well as PD-L1 and IDO1, suggesting complex activation and regulation of innate and adaptive immune function with AS15. These changes were generally greater with vaccines containing IFA, but IFA induced reduction in myeloid suppressor cells markers. Evidence of tertiary lymphoid structure (TLS) formation was observed with both adjuvants. Our findings highlight adjuvant-dependent changes in immune features at the VSME that may impact systemic immune responses. [ABSTRACT FROM AUTHOR]

Background: Cancer vaccines require adjuvants to induce effective immune responses; however, there is no consensus on optimal adjuvants. We hypothesized that toll-like receptor (TLR)3 agonist polyICLC or TLR4 agonist lipopolysaccharide (LPS), combined with CD4 T cell activation, would support strong and durable CD8+ T cell responses, whereas addition of an incomplete Freund's adjuvant (IFA) would reduce magnitude and persistence of immune responses. Patients and methods: Participants with resected stage IIB-IV melanoma received a vaccine comprised of 12 melanoma peptides restricted by Class I MHC (12MP), plus a tetanus helper peptide (Tet). Participants were randomly assigned 2:1 to cohort 1 (LPS dose-escalation) or cohort 2 (polyICLC). Each cohort included 3 subgroups (a-c), receiving 12MP + Tet + TLR agonist without IFA (0), or with IFA in vaccine one (V1), or all six vaccines (V6). Toxicities were recorded (CTCAE v4). T cell responses were measured with IFNγ ELIspot assay ex vivo or after one in vitro stimulation (IVS). Results: Fifty-three eligible patients were enrolled, of which fifty-one were treated. Treatment-related dose-limiting toxicities (DLTs) were observed in 0/33 patients in cohort 1 and in 2/18 patients in cohort 2 (11%). CD8 T cell responses to 12MP were detected ex vivo in cohort 1 (42%) and in cohort 2 (56%) and in 18, 50, and 72% for subgroups V0, V1, and V6, respectively. T cell responses to melanoma peptides were more durable and of highest magnitude for IFA V6. Conclusions: LPS and polyICLC are safe and effective vaccine adjuvants when combined with IFA. Contrary to the central hypothesis, IFA enhanced T cell responses to peptide vaccines when added to TLR agonists. Future studies will aim to understand mechanisms underlying the favorable effects with IFA. [ABSTRACT FROM AUTHOR]

Objective To characterize epidemiologic and clinical features of red primary amelanotic melanomas, an atypical presentation of melanoma that is underemphasized in patient and physician education. Patients and methods Review of a prospectively collected melanoma database identified 46 patients with red amelanotic melanomas, whose clinical features were compared with 329 patients with pigmented melanomas from the same database and same timeframe from January 1964 to September 2005. Results Red amelanotic melanomas represented 3.9% of all melanomas recorded in our database and accounted for nearly 70% of amelanotic melanomas. Melanoma was included in the clinical differential diagnosis in 32% of red amelanotic melanomas vs. 94% of pigmented melanomas ( P < 0.001). Red amelanotic melanomas more commonly underwent shave biopsy (55 vs. 12%, P < 0.001) and more likely had positive deep margins (35 vs. 9%, P < 0.001) but had comparable risks of metastasis and mortality. Conclusion Red amelanotic melanomas are often misdiagnosed clinically but carry a mortality risk comparable to pigmented melanomas. Clinicians screening for melanoma should be more vigilant in considering melanoma in the differential diagnosis of red skin lesions. [ABSTRACT FROM AUTHOR]

Background: Cancer vaccines require adjuvants to induce effective immune responses; however, there is no consensus on optimal adjuvants. We hypothesized that toll-like receptor (TLR)3 agonist polyICLC or TLR4 agonist lipopolysaccharide (LPS), combined with CD4 T cell activation, would support strong and durable CD8+ T cell responses, whereas addition of an incomplete Freund's adjuvant (IFA) would reduce magnitude and persistence of immune responses. Patients and methods: Participants with resected stage IIB-IV melanoma received a vaccine comprised of 12 melanoma peptides restricted by Class I MHC (12MP), plus a tetanus helper peptide (Tet). Participants were randomly assigned 2:1 to cohort 1 (LPS dose-escalation) or cohort 2 (polyICLC). Each cohort included 3 subgroups (a-c), receiving 12MP + Tet + TLR agonist without IFA (0), or with IFA in vaccine one (V1), or all six vaccines (V6). Toxicities were recorded (CTCAE v4). T cell responses were measured with IFNγ ELIspot assay ex vivo or after one in vitro stimulation (IVS). Results: Fifty-three eligible patients were enrolled, of which fifty-one were treated. Treatment-related dose-limiting toxicities (DLTs) were observed in 0/33 patients in cohort 1 and in 2/18 patients in cohort 2 (11%). CD8 T cell responses to 12MP were detected ex vivo in cohort 1 (42%) and in cohort 2 (56%) and in 18, 50, and 72% for subgroups V0, V1, and V6, respectively. T cell responses to melanoma peptides were more durable and of highest magnitude for IFA V6. Conclusions: LPS and polyICLC are safe and effective vaccine adjuvants when combined with IFA. Contrary to the central hypothesis, IFA enhanced T cell responses to peptide vaccines when added to TLR agonists. Future studies will aim to understand mechanisms underlying the favorable effects with IFA. Trial registration: The clinical trial Mel58 was performed with IRB (#15781) and FDA approval and is registered with Clinicaltrials.gov on April 25, 2012 (NCT01585350). Patients provided written informed consent to participate. Enrollment started on June 24, 2012. [ABSTRACT FROM AUTHOR]

Abstract: Background: Differentiating melanoma metastasis from benign cutaneous lesions currently requires biopsy or costly imaging, such as positron emission tomography scans. Melanoma metastases have been observed to be subjectively warmer than similarly appearing benign lesions. We hypothesized that infrared (IR) thermography would be sensitive and specific in differentiating palpable melanoma metastases from benign lesions. Materials and methods: Seventy-four patients (36 females and 38 males) had 251 palpable lesions imaged for this pilot study. Diagnosis was determined using pathologic confirmation or clinical diagnosis. Lesions were divided into size strata for analysis: 0–5, >5–15, >15–30, and >30 mm. Images were scored on a scale from −1 (colder than the surrounding tissue) to +3 (significantly hotter than the surrounding tissue). Sensitivity and specificity were calculated for each stratum. Logistical challenges were scored. Results: IR imaging was able to determine the malignancy of small (0–5 mm) lesions with a sensitivity of 39% and specificity of 100%. For lesions >5–15 mm, sensitivity was 58% and specificity 98%. For lesions >15–30 mm, sensitivity was 95% and specificity 100%, and for lesions >30 mm, sensitivity was 78% and specificity 89%. The positive predictive value was 88%–100% across all strata, and the negative predictive value was 95% for >15–30 mm lesions and 80% for >30 mm lesions. Conclusions: Malignant lesions >15 mm were differentiated from benign lesions with excellent sensitivity and specificity. IR imaging was well tolerated and feasible in a clinic setting. This pilot study shows promise in the use of thermography for the diagnosis of malignant melanoma with further potential as a noninvasive tool to follow tumor responses to systemic therapies. [Copyright &y& Elsevier]