Acute exanthemas: a prospective study of 98 adult patients with an emphasis on cytokinic and metagenomic investigation.

BACKGROUND: Acute exanthemas (AEs) are frequently seen; they can be caused by drugs or viruses but often the cause is unknown. OBJECTIVES: To describe the clinical, virological and histological aspects of AEs and explore their cytokinic and metagenomic profiles. METHODS: This prospective study examined 98 patients with AE, from February to July 2014. Clinical data were recorded in a standardized chart. Virological investigation and skin biopsies were performed. In addition, blood and skin samples were analysed for cytokines and then by a shotgun metagenomic approach. We identified five groups of patients: those with maculopapular exanthemas (MPEs) that were virally induced (group 1); those with drug-induced MPEs (group 2), those with MPEs that were both viral and drug induced (group 3), those with idiopathic MPEs (group 4) and those with pityriasis rosea (group 5). RESULTS: A virus was identified in 29 cases (human herpesvirus 6, 72%). Cytokinic analysis of the skin (n = 23 MPEs) showed higher levels of interferon-γ and interleukin-1 receptor-α in viral MPEs, higher interleukin-33 levels in idiopathic MPEs, and higher macrophage inflammatory protein 1α levels in drug-induced MPEs. By metagenomics analysis (n = 10 MPEs), viruses identified with routine practice methods were not found in group 1 (n = 4 MPEs). However, Enterovirus A was detected in two cases, especially in a group 1 patient for whom metagenomic analysis rectified the diagnosis of the culprit agent. CONCLUSIONS: Human herpesvirus 6 was the virus most frequently identified, and histology did not discriminate MPEs. In addition, the level of interleukin-33 seen in idiopathic MPEs suggests that an environmental factor may be the trigger for these. The results bring into question the utility of routine polymerase chain reaction analysis and viral serology for determining cause in AE. What's already known about this topic? Acute exanthemas, especially maculopapular exanthemas, are a frequent reason for patients consulting emergency and dermatology departments. It is difficult to evaluate the aetiology of acute exanthema based on the clinical aspects. Few data are available on the investigations needed in routine practice, and no prospective series have been published. What does this study add? Our study provides a global and prospective description of acute exanthemas. Cytokine analysis could help to investigate the pathophysiology of idiopathic eruptions. Metagenomic analysis provides new insights about the value of routine practice virological investigations. We show for the first time the feasibility of metagenomics analysis in the skin, which results question the interest of routine PCR and viral sérologies for the exploration of such acute exanthemas.

Frequency and prognostic value of cutaneous molecular residual disease in mycosis fungoides: a prospective multicentre trial of the Cutaneous Lymphoma French Study Group.

BACKGROUND: Monoclonal T-cell receptor (TCR) rearrangement is detected in 57-75% of early-stage mycosis fungoides (MF) at diagnosis. A retrospective study showed molecular residual disease (MRD) in 31% of patients in complete clinical remission (CR) after 1 year of treatment. OBJECTIVES: To confirm the frequency of MRD at 1 year and to determine its prognostic value for further relapse. METHODS: Patients with T1-, T2- or T4-stage MF were prospectively included in this multicentre study. At diagnosis, clinical lesions and healthy skin were biopsied. After 1 year of topical treatment, previously involved skin of patients in CR was biopsied for histology and analysis of TCR-γ gene rearrangement. The results were compared with the clinical status each year for 4 years. RESULTS: We included 214 patients, 133 at T1, 78 at T2 and three at T4 stage. At diagnosis, 126 of 204 cases (61·8%) showed TCR clonality in lesional skin. After 1 year, 83 of 178 patients (46·6%) still being followed up were in CR and 13 of 63 (21%) showed MRD. At 4 years, 55 of 109 patients (50·5%) still being followed up were in CR and 44 of 109 (40·4%) were in T1 stage. MRD did not affect clinical status at 4 years (CR vs. T1/T2, P = 1·0; positive predictive value 36·4%; negative predictive value 67·6%). CONCLUSIONS: T-cell clonality at diagnosis and MRD at 1 year are not prognostic factors of clinical status at 4 years.

Histopathology of drug rash with eosinophilia and systemic symptoms syndrome: a morphological and phenotypical study.

BACKGROUND: The histopathological features of drug rash with eosinophilia and systemic symptoms (DRESS) syndrome remain poorly characterized. OBJECTIVES: To better characterize the histopathological features of DRESS syndrome, and define the phenotype of the effector cells in the skin and compare it with maculopapular rash (MPR). METHODS: We conducted a retrospective study on 50 skin biopsies from patients with DRESS syndrome (n = 36). Histopathological and immunophenotypical features were studied and compared with a series of MPRs (n = 20). RESULTS: Foci of interface dermatitis, involving cutaneous adnexae, were frequently seen in cases of DRESS. Eosinophils were seen in only 20% of cases and neutrophils in 42%. Eczematous (40%), interface dermatitis (74%), acute generalized exanthematic pustulosis-like (20%) and erythema multiforme-like (24%) patterns were observed. The association of two or three of these patterns in a single biopsy was significantly more frequent in cases of DRESS than in a series of nondrug-induced dermatoses (P < 0.01), and appeared to be more marked in DRESS syndrome with severe cutaneous lesions (P = 0.01) than in less severe cases of DRESS and MPR. A higher proportion of CD8(+) and granzyme B(+) lymphocytes was observed in cases of DRESS with severe cutaneous eruptions (erythroderma and/or bullae). Atypical lymphocytes were found in 28% of biopsies, and expressed CD8 in most cases; a cutaneous T-cell clone was rarely found (6%). CONCLUSIONS: The histopathology of DRESS syndrome highlights various associated inflammatory patterns in a single biopsy. Cutaneous effector lymphocytes comprise a high proportion of polyclonal CD8(+) granzyme B(+) T lymphocytes.

Folliculotropic T-cell infiltrates associated with B-cell chronic lymphocytic leukaemia or MALT lymphoma may reveal either true mycosis fungoides or pseudolymphomatous reaction: seven cases and review of the literature.

BACKGROUND: Mycosis fungoides (MF) and pseudo-MF (or MF simulant) can be associated with B-cell malignancies, but distinction between a true neoplasm and a reactive process may be difficult. OBJECTIVES: To report seven patients with B-cell malignancy and folliculotropic MF or pseudo-MF and emphasize on criteria allowing distinction between the two conditions. METHODS: We retrospectively and prospectively included seven patients with B-cell malignancy who presented skin lesions histologically consisting in a folliculotropic T-cell infiltrate and reviewed the literature on the topic. RESULTS: Four men and three women had a chronic lymphocytic leukaemia (n = 6) or a MALT-type lymphoma (n = 1). Five patients had localized papules, and two had patches and plaques. Histological examination showed in all cases a diffuse dermal T-cell infiltrate with folliculotropic involvement and follicular mucinosis associated with clusters of the B-cell lymphoma, without significant expression of follicular helper T-cell markers. T-cell rearrangement studies showed a polyclonal pattern in the patients with papules and a monoclonal pattern in the cases of patches and plaques. Papular lesions had an indolent evolution, whereas patches and plaques persisted or worsened into transformed MF. CONCLUSION: Folliculotropic T-cell infiltrates associated with B-cell malignancies can be either a true folliculotropic MF or a pseudo-MF. The distinction between both conditions cannot rely only on the histopathological aspect, but needs both a clinical pathological correlation and the search for a dominant T-cell clone. Whether the neoplastic T and B cells derive from a common ancestor or the T-cell proliferation is promoted by the underlying B-cell lymphoma remains unsolved, but interaction between B and T cell in the skin does not appear to be dependent on a TFH differentiation of the T-cell infiltrate.

EuroClonality/BIOMED-2 guidelines for interpretation and reporting of Ig/TCR clonality testing in suspected lymphoproliferations.

PCR-based immunoglobulin (Ig)/T-cell receptor (TCR) clonality testing in suspected lymphoproliferations has largely been standardized and has consequently become technically feasible in a routine diagnostic setting. Standardization of the pre-analytical and post-analytical phases is now essential to prevent misinterpretation and incorrect conclusions derived from clonality data. As clonality testing is not a quantitative assay, but rather concerns recognition of molecular patterns, guidelines for reliable interpretation and reporting are mandatory. Here, the EuroClonality (BIOMED-2) consortium summarizes important pre- and post-analytical aspects of clonality testing, provides guidelines for interpretation of clonality testing results, and presents a uniform way to report the results of the Ig/TCR assays. Starting from an immunobiological concept, two levels to report Ig/TCR profiles are discerned: the technical description of individual (multiplex) PCR reactions and the overall molecular conclusion for B and T cells. Collectively, the EuroClonality (BIOMED-2) guidelines and consensus reporting system should help to improve the general performance level of clonality assessment and interpretation, which will directly impact on routine clinical management (standardized best-practice) in patients with suspected lymphoproliferations.

[Management of cutaneous T-cell lymphomas: Recommendations of the French Cutaneous Lymphoma Group]. / Prise en charge des lymphomes T cutanés : recommandations du Groupe français d'étude des lymphomes cutanés.

BACKGROUND: Classification of diagnostic methods, of initial staging and of the treatment of primary cutaneous T-cell lymphomas, particularly the most common epidermotropic forms, constitutes an essential step in rationalising therapeutic practice and in evaluating the results of treatment. PATIENTS AND METHODS: We carried out an analysis of the literature and of existing recommendations in order to create recommendations regarding the diagnosis, initial staging and treatment of primary T-cell lymphomas. RESULTS: We present the key elements of diagnosis and initial staging. The selected therapeutic strategy, which necessarily changes over time, must avoid both unnecessarily aggressive early treatment as well as an overly timid therapeutic approach that could allow lesions to rapidly progress towards more advanced stages. Regular reassessment of the benefit/risk ratio is necessary and involves the use of first- and second-line measures, in which it is difficult to establish any hierarchy, with the current tendency favouring in particular combined therapy as second-line treatment in order to limit the toxicity of each individual constituent drug within the combination. The creation of a national SPC marks significant progress in difficult cases. CONCLUSION: As a result of the offer, limited level of proof in existing studies, which are generally unsatisfactory in terms of methodology, the new recommendations described herein are timely and should be updated regularly in accordance with advances in knowledge. The organisation of clinical trials and validation of the scoring systems currently being developed should be encouraged.

[Management of cutaneous B-cell lymphoma: recommendations of the French cutaneous lymphoma study group]. / Prise en charge des lymphomes B cutanés : recommandations du Groupe français d'étude des lymphomes cutanés.

AIMS: To provide recommendations for the treatment of cutaneous B-cell lymphomas (CBCL). METHODS: Literature review and expert opinions from the French Cutaneous Lymphoma Study Group. RESULTS: Diagnosis of marginal zone BCL (MZ BCL), centrofollicular BCL (CF BCL) or cutaneous large B-cell lymphoma, leg type (CLBCL, LT) is based on combination of clinical signs and histopathological features, together with B-cell clonality analyses whenever possible. Staging relies on straightforward laboratory examinations and imaging, completed in selected cases with bone marrow biopsy. Treatment may be topical, including excision, curative radiotherapy (30Gray) or adjunctive/low dose (4Gray) radiotherapy, topical corticosteroids, interferon or intralesional rituximab; or systemic, using chemotherapy and/or intravenous rituximab. For indolent forms of the disease (MZ CBCL and CF CBCL), curative (30Gray) may be given as first-line treatment in patients with localized lesions or few scattered skin lesions. For more numerous slow-growing lesions with a low tumour burden, simple monitoring with adjunctive ad hoc local treatment of individual lesions is acceptable. For multiple growing lesions, systemic rituximab or chlorambucil may be proposed. Polychemotherapy should only be used for progressive forms unresponsive to previous therapies. CLBCL LT forms are more aggressive and occur in older subjects. These lymphomas are best treated with age-adapted combinations of polychemotherapies and rituximab. CONCLUSION: Appropriate clinical trials are still needed to strengthen the levels of evidence of current recommendations.

Peripheral blood natural killer cell count is associated with clinical outcome in patients with aaIPI 2-3 diffuse large B-cell lymphoma.

BACKGROUND: Lymphocytopenia is a prognostic factor in Hodgkin's disease. In diffuse large B-cell lymphoma (DLBCL), data are much less established, in spite of numerous reports on immune system-lymphoma interactions. This study addresses the prognostic value of blood lymphocyte subsets at diagnosis in DLBCL. PATIENTS AND METHODS: Absolute values of blood lymphocyte subsets and monocytes were prospectively determined by flow cytometry in 140 patients with 2 or 3 adverse age-adjusted International Prognostic Index (aaIPI) factors included in a Groupe d'Etude des Lymphomes de l'Adulte protocol (LNH98B3). Absolute cell counts at diagnosis and aaIPI were evaluated with regard to clinical outcome. RESULTS: Low median cell counts of 337, 211, and 104/mul were evidenced for the CD4+, CD8+ T, and natural killer (NK) cells, respectively. In univariate analysis, only NK cell count [odds ratio (OR) = 1.81 (1.27, 2.57), P = 0.001] and aaIPI [OR = 2.29 (0.95, 5.45), P = 0.06] were associated with induction treatment response. Low NK cell count [Hazard ratio (HR) = 1.27 (1.06, 1.52), P = 0.01] and aaIPI 3 [HR = 1.95 (1.20, 3.16), P = 0.01] were also associated with a shorter event free survival (EFS). In multivariate analysis, NK cell count was associated with response [OR = 1.77 (1.24, 2.54), P = 0.002] and EFS [HR = 1.25 (1.04, 1.50) P = 0.02] independently of aaIPI. CONCLUSIONS: This study shows an association between circulating NK cell number and clinical outcome in DLBCL, possibly important in the context of the broadening use of rituximab, a likely NK-dependent therapy.

Powerful strategy for polymerase chain reaction-based clonality assessment in T-cell malignancies Report of the BIOMED-2 Concerted Action BHM4 CT98-3936.

Polymerase chain reaction (PCR) assessment of clonal T-cell receptor (TCR) and immunoglobulin (Ig) gene rearrangements is an important diagnostic tool in mature T-cell neoplasms. However, lack of standardized primers and PCR protocols has hampered comparability of data in previous clonality studies. To obtain reference values for Ig/TCR rearrangement patterns, 19 European laboratories investigated 188 T-cell malignancies belonging to five World Health Organization-defined entities. The TCR/Ig spectrum of each sample was analyzed in duplicate in two different laboratories using the standardized BIOMED-2 PCR multiplex tubes accompanied by international pathology panel review. TCR clonality was detected in 99% (143/145) of all definite cases of T-cell prolymphocytic leukemia, T-cell large granular lymphocytic leukemia, peripheral T-cell lymphoma (unspecified) and angioimmunoblastic T-cell lymphoma (AILT), whereas nine of 43 anaplastic large cell lymphomas did not show clonal TCR rearrangements. Combined use of TCRB and TCRG genes revealed two or more clonal signals in 95% of all TCR clonal cases. Ig clonality was mostly restricted to AILT. Our study indicates that the BIOMED-2 multiplex PCR tubes provide a powerful strategy for clonality assessment in T-cell malignancies assisting the firm diagnosis of T-cell neoplasms. The detected TCR gene rearrangements can also be used as PCR targets for monitoring of minimal residual disease.

Significantly improved PCR-based clonality testing in B-cell malignancies by use of multiple immunoglobulin gene targets. Report of the BIOMED-2 Concerted Action BHM4-CT98-3936.

Polymerase chain reaction (PCR) assessment of clonal immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements is an important diagnostic tool in mature B-cell neoplasms. However, lack of standardized PCR protocols resulting in a high level of false negativity has hampered comparability of data in previous clonality studies. In order to address these problems, 22 European laboratories investigated the Ig/TCR rearrangement patterns as well as t(14;18) and t(11;14) translocations of 369 B-cell malignancies belonging to five WHO-defined entities using the standardized BIOMED-2 multiplex PCR tubes accompanied by international pathology panel review. B-cell clonality was detected by combined use of the IGH and IGK multiplex PCR assays in all 260 definitive cases of B-cell chronic lymphocytic leukemia (n=56), mantle cell lymphoma (n=54), marginal zone lymphoma (n=41) and follicular lymphoma (n=109). Two of 109 cases of diffuse large B-cell lymphoma showed no detectable clonal marker. The use of these techniques to assign cell lineage should be treated with caution as additional clonal TCR gene rearrangements were frequently detected in all disease categories. Our study indicates that the BIOMED-2 multiplex PCR assays provide a powerful strategy for clonality assessment in B-cell malignancies resulting in high Ig clonality detection rates particularly when IGH and IGK strategies are combined.

Identification of circulating CD10 positive T cells in angioimmunoblastic T-cell lymphoma.

In most cases of lymphomas with blood dissemination, the careful cytological analysis of peripheral blood smears provides a rapid orientation to diagnosis, even if the final subtyping is achieved by histology and eventually other techniques. Here, we evaluated if the analysis of blood smears may suggest the blood dissemination of angioimmunoblastic T-cell lymphoma (AITL) and if CD10 expression on neoplastic T cells, as recently reported on AITL, may contribute to the diagnosis. In all, 11 lymph nodes and six peripheral blood samples from 12 patients with AITL were studied using four-colour flow cytometry associated to histological, cytological and molecular data. According to previous results, a fraction of T cells expressed CD10 in 10/11 lymph nodes. Interestingly, all blood smears showed atypical lymphoid cells and a fraction of T cells expressed CD10 with a mean percentage of 18.75% (range 5.00-47.00%), regardless of lymphocytosis level and of rate of CD10 T cells in corresponding lymph node. In contrast, in all control samples (100), none CD10-positive T cell was identified. This is to our knowledge the first description of circulating CD10 neoplastic T cells in AITL. Therefore, they ought to be explored in further studies when aggressive lymphoma, in particular with lymphopenia and circulating atypical cells, is suspected.

Metachronous gastric MALT lymphoma and early gastric cancer: is residual lymphoma a risk factor for the development of gastric carcinoma?

BACKGROUND: Helicobacter pylori plays a major role in the pathogenesis of primary gastric MALT lymphoma (GML) and gastric carcinoma. The occurrence of these two diseases metachronously in a same patient is a rare event. PATIENTS AND METHODS: Gastric biopsies and gastrectomy resection specimens of four patients who developed GML and early gastric cancer (EGC) were analysed by morphology, immunohistochemistry and molecular biology. RESULTS: Four patients (three males and one female; mean age 48 years) were diagnosed with GML. Helicobacter pylori infection was observed in three cases. Two patients had localized disease (stages IE and IIE, respectively) and were treated with H. pylori eradication therapy followed by an alkylating agent for one patient. Two patients had disseminated disease (stage IV), and were treated with an alkylating agent. Three cases were t(11;18) positive. All patients achieved initially complete lymphoma remission. Long-term endoscopic surveillance detected an EGC at the same location as the lymphoma in all patients at a mean time of 9.5 years (range 2.5-17 years) after lymphoma diagnosis. Gastrectomy specimens showed residual GML in all cases. CONCLUSION: Prolonged residual GML could constitute an additional risk factor for the development of gastric carcinoma. Long-term endoscopic surveillance is mandatory in patients treated conservatively for gastric MALT lymphoma.

Specific antibodies modulate the interactions of adenovirus type 5 with dendritic cells.

Adenovirus type 5 (Ad5) is able to induce an efficient CD8+ T lymphocyte (CTL) response against a transgene product, a property thought to be linked to its ability to transduce dendritic cells (DCs). Little, however, is known about the capacity of Ad5 to interact with DCs in the presence of specific antibodies, although most people test positive for antibodies directed against Ad5. In the present study, we found that in the presence of Ad5 antibodies, a large fraction of Ad5 binds very efficiently to DCs, and that this binding is FcgammaRII/FcgammaRIII dependent. Nevertheless, in the presence of high levels of antibodies against the whole virion, Ad5 entry was inhibited. Increased binding led to increased entry in DCs in the presence of fiber-specific antibodies or in the presence of low amounts of a whole antiserum raised against whole virions, showing that the relative concentration of antibodies directed against fiber and penton base plays a major role in entry efficacy. Nevertheless, mice previously immunized with virions or purified fiber developed a lower transgene-specific CD8+ T cell response than naive mice, although their serum appeared to increase virus entry into DCs in vitro.

Efficiency of in vivo purging with rituximab prior to autologous peripheral blood progenitor cell transplantation in B-cell non-Hodgkin's lymphoma: a single institution study.

BACKGROUND: Rituximab induces clinical response in advanced B-cell lymphoma and is efficient in removing circulating B-cell from peripheral blood. We therefore postulated that rituximab might be a useful in vivo purging agent before high-dose therapy in this setting. PATIENTS AND METHODS: Fourteen patients with relapsed follicular, marginal zone and mantle cell lymphomas (11, two and one cases, respectively) and a PCR-detectable molecular marker were treated first with rituximab, then a mobilization chemotherapeutic regimen, followed by high-dose therapy with peripheral blood stem cell transplantation. PCR analyses were performed in peripheral blood before rituximab and during follow-up, and in harvest. RESULTS: Harvests were free of PCR-detectable molecular marker in nine of the 11 studied cases (82%). After high-dose therapy, clinical complete remission was obtained in 13 (93%) patients and molecular remission in 11 (79%). With a median follow-up of 3 years, the 14 transplanted patients were alive, 11 of them remaining in clinical complete remission and eight in molecular remission at last follow-up. CONCLUSION: Rituximab treatment followed by high dose therapy appears to be effective in achieving complete clinical and molecular response. In vivo harvest purging is predictive of prolonged clinical and molecular remission.

Minimal residual disease in mycosis fungoides follow-up can be assessed by polymerase chain reaction.

BACKGROUND: T-cell receptor (TCR) gene rearrangement analysis, i.e. T-cell clonality, using polymerase chain reaction (PCR) is a routine method used to assess the presence of a cutaneous dominant T-cell clone in mycosis fungoides (MF). OBJECTIVES: To compare the outcome of cutaneous lesions of MF after treatment with the fate of the cutaneous T-cell clonality, and to determine whether minimal residual disease can be detected in patients in clinical complete remission. METHODS: Fifty-one patients histologically diagnosed as having MF (17 stage IA, 21 stage IB and 13 stage III) were included in this retrospective study. T-cell clonality was analysed by GC-clamp multiplex PCRgamma-denaturing gradient gel electrophoresis. Every patient had two cutaneous biopsies at least 3 months apart. The second biopsy was performed at the site of a treated lesion. RESULTS: The presence or absence of a dominant T-cell clone in the skin remained identical in 26 of the 31 (84%) patients with persistent disease. Thirteen patients with a detectable dominant T-cell clone at diagnosis went into complete clinical remission. In nine of these 13 (69%) patients, the T-cell clone was no longer detectable after treatment. The remaining four (31%) patients had an unchanged T-cell clonality. CONCLUSIONS: The TCR gene rearrangement imprint is a stable and reliable tumour marker of MF disease. One-third of patients in complete clinical remission had a cutaneous molecular residual disease, the prognostic value of which will be analysed in an ongoing prospective study.

Oligoclonal T-cells in blood and target tissues of patients with anti-Hu syndrome.

T-cell clones of unknown significance (TCUS), assessed by monoclonal or oligoclonal T-cell patterns in PCR-DGGE, were detected in blood of 7/9 patients with anti-Hu syndrome. Clonal patterns were also detected in 2/2 neoplastic lymph nodes, and in 2/2 inflamed dorsal root ganglia from three patients. Only some T-cell clones found in target tissues were also detected in blood or non-target tissues, and likely corresponded to TCUS. In one patient, an identical T-cell clone was found in both neoplastic lymph node tissue and dorsal root ganglia, but not in blood. Dorsal root-infiltrating lymphocytes were cytotoxic CD8(+) TIA-1(+) T-cells. They were often found in close contact to sensory neurons, most of which expressed MHC-1. Taken together, these data support a direct effector role of cytotoxic CD8(+) T-cells, the same clones being likely operative in sensory neuron damage and immune-mediated tumor growth control.