Effect of topical imiquimod as primary treatment for lentigo maligna: the LIMIT-1 study.

BACKGROUND: Topical imiquimod is sometimes used for lentigo maligna (LM) in situ melanoma instead of surgery, but frequency of cure is uncertain. Pathological complete regression (pCR) is a logical surrogate marker for cure after imiquimod, although residual LM and atypical melanocytic hyperplasia may not be reliably distinguished. A trial comparing imiquimod vs. surgery might be justified by a high imiquimod pCR rate. OBJECTIVES: Primary: to estimate the pCR rate for LM following imiquimod. Secondary: to assess the accuracy of prediction of pCR, using clinical complete regression (cCR) plus negative post-treatment biopsies, tolerability, resource use, patients' preferences and induced melanoma immunity. METHODS: This was a single-arm phase II trial of 60 imiquimod applications over 12 weeks for LM then radical resection. A pCR rate ≥ 25 out of 33 would reliably discriminate between pCR rates < 60% and ≥ 85%. Clinical response was assessed and biopsies taken after imiquimod. Patients recorded adverse events in diaries. Patient preference was measured after surgery using a standard gamble tool. RESULTS: The pCR rate was 10 of 27 (37%, 95% confidence interval 19-58%). The rate of cCR plus negative biopsies was 12 of 28, of whom seven of 11 had pCR on subsequent surgery. The median dose intensity was 86·7%. Of the 16 surveyed patients, eight preferred primary imiquimod over surgery if the cure rate for imiquimod was 80%, and four of 16 if it was ≤ 40%. CONCLUSIONS: The pCR rate was insufficient to justify phase III investigation of imiquimod vs. SURGERY: Clinical complete response and negative targeted biopsies left uncertainty regarding pathological clearance. Some patients would trade less aggressive treatment of LM against efficacy.

Phase I/II trial of a dendritic cell vaccine transfected with DNA encoding melan A and gp100 for patients with metastatic melanoma.

This trial tested a dendritic cell (DC) therapeutic cancer vaccine in which antigen is loaded using a novel non-viral transfection method enabling the uptake of plasmid DNA condensed with a cationic peptide. Proof of principle required the demonstration of diverse T lymphocyte responses following vaccination, including multiple reactivities restricted through both major histocompatibility complex (MHC) class I and II. Patients with advanced melanoma were offered four cycles of vaccination with autologous DC expressing melan A and gp100. Disease response was measured using Response Evaluation Criteria in Solid Tumours. Circulating MHC class I- and II-restricted responses were measured against peptide and whole antigen targets using interferon-γ ELIspot and enzyme-linked immunosorbent assay assays, respectively. Responses were analyzed across the trial population and presented descriptively for some individuals. Twenty-five patients received at least one cycle. Vaccination was well tolerated. Three patients had reduction in disease volume. Across the trial population, vaccination resulted in an expansion of effector responses to both antigens, to the human leukocyte antigen A2-restricted modified epitope, melan A ELAGIGILTV, and to a panel of MHC class I- and II-restricted epitopes. Vaccination with mature DC non-virally transfected with DNA encoding antigen had biological effect causing tumour regression and inducing diverse T lymphocyte responses.

Merkel cell carcinoma.

Merkel cell carcinoma (MCC) is a rare neuroendocrine carcinoma. The cellular origin of MCC may include Merkel cell precursors. The incidence of MCC has increased significantly however trends may have been confounded by evolving diagnostic criteria. The two key aetiologies of MCC are ultraviolet radiation and Merkel cell polyoma virus (MCPyV). Both have unique mechanisms of carcinogenesis. MCC presents non-specifically as a rapidly growing, red-to-violet nodule on sun-exposed areas. Diagnostic accuracy has improved through immunohistochemical markers such as CK-20. Lymph nodes should be evaluated in MCC through examination and sentinel biopsy. USS, CT, MRI and CT-PET may be useful in staging. Management depends on tumour location, stage and comorbidities. MCPyV status may guide treatment strategy in the future. Treatment for the primary MCC is commonly wide local excision followed by radiotherapy, guided by anatomical constraints. There is uncertainty about surgical margins. Treatments for nodal disease have not been determined through trials. They include nodal dissection or radiotherapy for clinically or radiologically apparent disease, and adjuvant nodal irradiation for negative nodes, microscopic disease or following nodal dissection for definite disease. Patients with loco-regional advanced inoperable disease should be considered for combination therapy including chemotherapy, radiotherapy, surgery and immunotherapy. Systemic therapy for advanced disease includes immune checkpoint inhibitors targeting the PD-1/PD-L1 pathway. Avelumab can improve survival in metastatic MCC. Immunotherapy may result in longer disease control. Various other immunotherapeutic and molecular agents are undergoing trials. MCC continues to have a high mortality characterized by high recurrence and early metastases.

Epitope focusing in the primary cytotoxic T cell response to Epstein-Barr virus and its relationship to T cell memory.

The relationship between primary and memory cytotoxic T lymphocyte (CTL) responses, and the factors influencing entry into memory, are poorly understood. Here we address this in the context of Epstein-Barr virus (EBV), a persistent human herpesvirus in which memory CTL responses in long-term virus carriers are highly focused on epitopes preferentially drawn from just three of the eight available virus latent proteins, EBNAs 3A, 3B, and 3C. To determine whether this unusual level of focusing is a consequence of long-term virus challenge, we carried out a detailed analysis of EBV antigen/epitope specificities in the primary virus-induced CTL response in 10 infectious mononucleosis (IM) patients of different HLA types. Primary effectors, studied in ex vivo assays and by limiting dilution cloning in vitro, were again highly skewed toward a small number of viral epitopes, almost all derived from the EBNA3 proteins, with CTL to the immunodominant epitope accounting for at least 1% of the circulating CD8+ IM T cell pool. This is the first unequivocal demonstration of an EBV-specific CD8+ CTL response in IM. Prospective studies on individual patients showed that, whereas all of the EBV reactivities found in CTL memory had been detectable earlier during primary infection, the memory population was not simply a scaled down version of the primary response. In particular (a) differences in the relative frequencies of CTL to immunodominant versus subdominant epitopes appeared to be much less marked in memory than in primary populations, and (b) we found at least one clear example in which a significant virus-specific reactivity within the primary response was never detectable in memory.

Immediate early and early lytic cycle proteins are frequent targets of the Epstein-Barr virus-induced cytotoxic T cell response.

Epstein-Barr virus (EBV), a human gamma-herpesvirus, can establish both nonproductive (latent) and productive (lytic) infections. Although the CD8+ cytotoxic T lymphocyte (CTL) response to latently infected cells is well characterized, very little is known about T cell controls over lytic infection; this imbalance in our understanding belies the importance of virus-replicative lesions in several aspects of EBV disease pathogenesis. The present work shows that the primary CD8+ CTL response to EBV in infectious mononucleosis patients contains multiple lytic antigen-specific reactivities at levels at least as high as those seen against latent antigens; similar reactivities are also detectable in CTL memory. Clonal analysis revealed individual responses to the two immediate early proteins BZLF1 and BRLF1, and to three (BMLF1, BMRF1, and BALF2) of the six early proteins tested. In several cases, the peptide epitope and HLA-restricting determinant recognized by these CTLs has been defined, one unusual feature being the number of responses restricted through HLA-C alleles. The work strongly suggests that EBV-replicative lesions are subject to direct CTL control in vivo and that immediate early and early proteins are frequently the immunodominant targets. This contrasts with findings in alpha- and beta-herpesvirus systems (herpes simplex, cytomegalovirus) where viral interference with the antigen-processing pathway during lytic infection renders immediate early and early proteins much less immunogenic. The unique capacity of gamma-herpesvirus to amplify the viral load in vivo through a latent growth-transforming infection may have rendered these agents less dependent upon viral replication as a means of successfully colonizing their hosts.

Cytotoxic T lymphocyte responses to Epstein-Barr virus.

Epstein-Barr virus induces a potent cytotoxic T lymphocyte response in man that is preferentially directed towards a particular subset of the virus latent cycle antigens; the immunodominance of these proteins, apparent in both primary and memory responses, may reflect some differential access to the HLA class I processing pathway. Effector cells recognizing these immunodominant antigens can reverse the growth of virus-induced lymphoproliferative lesions in immunosuppressed patients; however, responses to some of the subdominant latent proteins will be needed to target other virus-positive tumours.

Tumor stroma as a target in cancer.

Solid tumors are composed of the malignant cell itself (most commonly a carcinoma) and supporting cells that comprise the stroma. Significant stromal components include the extracellular matrix, supporting fibroblasts, vessels comprised of endothelium, pericytes and in some cases vascular smooth muscle, lymphatics and usually a major leukocyte infiltration. Indeed, macrophages may constitute up to 50% of the viable cells within the tumor. For many years, researchers have concentrated almost exclusively on the malignant carcinoma and looked for ways to either selectively kill or restrict its growth. In recent years the frustrating lack of advances in cytotoxic cancer therapy provoked a search for more novel strategies and foremost amongst these were anti-angiogenesis and vascular targeting. The purpose of this article is to illustrate how the stroma is now being pursued as an anti-cancer target. The article will briefly touch on anti-angiogenics that are now entering the clinic but concentrate on recent studies looking at vascular disrupting agents, stromal tumor fibroblasts and macrophages. Target identification is illustrated by the search for tumor endothelial markers. Finally, we draw attention to efforts to develop a cancer vaccine. The genetic instability and variation found in carcinoma cells made vaccination in the past a near impossibility. In contrast, genetically stable tumor endothelium with its unique accessibility to blood borne agents, together with recent advances in immunotherapy means that the possibility of a cancer vaccine now takes on a reality not previously recognised.

Dual stimulation of Epstein-Barr Virus (EBV)-specific CD4+- and CD8+-T-cell responses by a chimeric antigen construct: potential therapeutic vaccine for EBV-positive nasopharyngeal carcinoma.

Virus-associated malignancies are potential targets for immunotherapeutic vaccines aiming to stimulate T-cell responses against viral antigens expressed in tumor cells. Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma, a high-incidence tumor in southern China, expresses a limited set of EBV proteins, including the nuclear antigen EBNA1, an abundant source of HLA class II-restricted CD4(+) T-cell epitopes, and the latent membrane protein LMP2, a source of subdominant CD8(+) T-cell epitopes presented by HLA class I alleles common in the Chinese population. We used appropriately modified gene sequences from a Chinese EBV strain to generate a modified vaccinia virus Ankara recombinant, MVA-EL, expressing the CD4 epitope-rich C-terminal domain of EBNA1 fused to full-length LMP2. The endogenously expressed fusion protein EL is efficiently processed via the HLA class I pathway, and MVA-EL-infected dendritic cells selectively reactivate LMP2-specific CD8(+) memory T-cell responses from immune donors in vitro. Surprisingly, endogenously expressed EL also directly accesses the HLA class II presentation pathway and, unlike endogenously expressed EBNA1 itself, efficiently reactivates CD4(+) memory T-cell responses in vitro. This unscheduled access to the HLA class II pathway is coincident with EL-mediated redirection of the EBNA1 domain from its native nuclear location to dense cytoplasmic patches. Given its immunogenicity to both CD4(+) and CD8(+) T cells, MVA-EL has potential as a therapeutic vaccine in the context of nasopharyngeal carcinoma.