Clinical and molecular practice of European thoracic pathology laboratories during the COVID-19 pandemic. The past and the near future.

BACKGROUND: This study evaluated the consequences in Europe of the COVID-19 outbreak on pathology laboratories orientated toward the diagnosis of thoracic diseases. MATERIALS AND METHODS: A survey was sent to 71 pathology laboratories from 21 European countries. The questionnaire requested information concerning the organization of biosafety, the clinical and molecular pathology, the biobanking, the workload, the associated research into COVID-19, and the organization of education and training during the COVID-19 crisis, from 15 March to 31 May 2020, compared with the same period in 2019. RESULTS: Questionnaires were returned from 53/71 (75%) laboratories from 18 European countries. The biosafety procedures were heterogeneous. The workload in clinical and molecular pathology decreased dramatically by 31% (range, 3%-55%) and 26% (range, 7%-62%), respectively. According to the professional category, between 28% and 41% of the staff members were not present in the laboratories but did teleworking. A total of 70% of the laboratories developed virtual meetings for the training of residents and junior pathologists. During the period of study, none of the staff members with confirmed COVID-19 became infected as a result of handling samples. CONCLUSIONS: The COVID-19 pandemic has had a strong impact on most of the European pathology laboratories included in this study. Urgent implementation of several changes to the organization of most of these laboratories, notably to better harmonize biosafety procedures, was noted at the onset of the pandemic and maintained in the event of a new wave of infection occurring in Europe.

Whole-exome sequencing in osteosarcoma reveals important heterogeneity of genetic alterations.

BACKGROUND: Whole-genome sequencing studies have recently shown that osteosarcomas (OSs) display high rates of structural variation, i.e. they contain many somatic mutations and copy number alterations. TP53 and RB1 show recurrent somatic alterations in concordant studies, suggesting that they could be key players in bone oncogenesis. PATIENTS AND METHODS: we carried out whole-genome sequencing of DNA from seven high-grade OS samples matched with normal tissue from the same patients. RESULTS: We confirmed the presence of genetic alterations of the TP53 (including novel unreported mutations) and RB1 genes. Most interestingly, we identified a total of 84 point mutations and 4 deletions related to 82 different genes in OS samples, of which only 15 have been previously reported. Interestingly, the number of mutated genes (ranging from 4 to 8) was lower in TP53mut cases compared with TP53wt cases (ranging from 14 to 45). This was also true for the mutated RB1 case. We also observed that a dedifferentiated OS harboring MDM2 amplification did not carry any other mutations. CONCLUSION: This study suggests that bone oncogenesis driven by TP53 or RB1 mutations occurs on a background of relative genetic stability and that the dedifferentiated OS subtype represents a clinico-pathological entity with distinct oncogenic mechanisms and thus requires different therapeutic management.

Breast implant-associated anaplastic large cell lymphoma: two distinct clinicopathological variants with different outcomes.

BACKGROUND: ALK-negative anaplastic large cell lymphoma associated with breast implant (i-ALCL) has been recently recognized as a distinct entity. Among 43 830 lymphomas registered in the French Lymphopath network since 2010, 300 breast lymphomas comprising 25 peripheral T-cell lymphomas (PTCL) were reviewed. Among PTCL, ALK-negative ALCL was the most frequent and all of them were associated with breast implants. PATIENTS AND METHODS: Since 2010, all i-ALCL cases were collected from different institutions through Lymphopath. Immuno-morphologic features, molecular data and clinical outcome of 19 i-ALCLs have been retrospectively analyzed. RESULTS: The median age of the patients was 61 years and the median length between breast implant and i-ALCL was 9 years. Most implants were silicone-filled and textured. Implant removal was performed in 17 out of 19 patients with additional treatment based on mostly CHOP or CHOP-like chemotherapy regimens (n = 10/19) or irradiation (n = 1/19). CHOP alone or ABVD following radiation without implant removal have been given in two patients. The two clinical presentations, i.e. effusion and less frequently tumor mass correlated with distinct histopathologic features: in situ i-ALCL (anaplastic cell proliferation confined to the fibrous capsule) and infiltrative i-ALCL (pleomorphic cells massively infiltrating adjacent tissue with eosinophils and sometimes Reed-Sternberg-like cells mimicking Hodgkin lymphoma). Malignant cells were CD30-positive, showed a variable staining for EMA and were ALK negative. Most cases had a cytotoxic T-cell immunophenotype with variable T-cell antigen loss and pSTAT3 nuclear expression. T-cell receptor genes were clonally rearranged in 13 out of 13 tested cases. After 18 months of median follow-up, the 2-year overall survival for in situ and infiltrative i-ALCL was 100% and 52.5%, respectively. CONCLUSIONS: In situ i-ALCLs have an indolent clinical course and generally remain free of disease after implant removal. However, infiltrative i-ALCLs could have a more aggressive clinical course that might require additional therapy to implant removal.

Specific infiltration pattern of FOXP3+ regulatory T cells in chronic histiocytic intervillositis of unknown etiology.

INTRODUCTION: Chronic histiocytic intervillositis of unknown etiology (CIUE) is a rare placental lesion characterized by an intervillous mononuclear inflammatory infiltrate of maternal origin. Although the mechanism and origin of these lesions are currently not understood, they appear to be related to an immune conflict between mother and fetus cells. AIM: To clarify the inflammatory cell profile and evaluate the T regulatory lymphocyte (Treg) status in CIUE. MATERIALS AND METHODS: All cases of CIUE that occurred over an 8-year period were analyzed using immunohistochemistry. RESULTS: The inflammatory profile of CIUE was characterized by a clearly predominant component of histiocytic cells (80% ± 6.9) associated with some T cells (24% ± 5.7). The ratio of CD4+ versus CD8+ T cells was close to 1. This profile differs from infectious disease and chronic histiocytic villitis, the main differential diagnoses of CIUE. As for normal pregnancies most regulatory T cells were localized in the decidua basalis. Nevertheless, their appearance was also noted in the intervillous space. In both the intervillous space and the deciduas the number of Tregs gradually increased from grade 1 to 3. CONCLUSION: We found that CIUE is associated with an increase in Treg lymphocytes in the decidua basalis and the intervillous space. Contrary to previously published data on human miscarriage, this result appears to be specific to CIUE and would support the hypothesis of an immunopathological disorder for CIUE.

B-cell regulator of immunoglobulin heavy-chain transcription (Bright)/ARID3a is a direct target of the oncomir microRNA-125b in progenitor B-cells.

B-cell acute lymphoblastic leukemia (B-ALL) is often associated with chromosomal translocations leading to the deregulation of proto-oncogenes. MicroRNAs can also be affected by chromosomal alterations and thus contribute to carcinogenesis. The microRNA, miR-125b-1, is overexpressed in B-ALL cases with the t(11;14)(q24;q32) translocation; therefore, we sought to determine the role of this microRNA in B-cell fate. We used murine pre-BI cells alongside murine and human leukemic B-cell lines to show that miR-125b expression enhances proliferation by targeting B-cell regulator of immunoglobulin heavy-chain transcription (Bright)/ARID3a, an activator of immunoglobulin heavy-chain transcription. Accordingly, this target gene was downregulated in B-ALL patients with the t(11;14)(q24;q32) translocation. Repression of Bright/ARID3a blocked differentiation and conferred a survival advantage to Ba/F3 cells under interleukin-3 starvation. In addition, overexpression of miR-125b protected pre-BI and leukemic B-cell lines from apoptosis by blockade of caspase activation by a mechanism that was independent of p53 and BAK1. In summary, miR-125b can act as an oncogene in B-ALL by targeting ARID3a and mediating its repression, thus leading to a blockage in differentiation, increased proliferation and inhibition of apoptosis.

Specific small nucleolar RNA expression profiles in acute leukemia.

Apart from microRNAs, little is known about the regulation of expression of non-coding RNAs in cancer. We investigated whether small nucleolar RNAs (snoRNAs) accumulation displayed specific signatures in acute myeloblastic and acute lymphoblastic leukemias. Using microarrays and high-throughput quantitative PCR (qPCR), we demonstrate here that snoRNA expression patterns are negatively altered in leukemic cells compared with controls. Interestingly, a specific signature was found in acute promyelocytic leukemia (APL) with ectopic expression of SNORD112-114 snoRNAs located at the DLK1-DIO3 locus. In vitro experiments carried out on APL blasts demonstrate that transcription of these snoRNAs was lost under all-trans retinoic acid-mediated differentiation and induced by enforced expression of the PML-RARalpha fusion protein in negative leukemic cell lines. Further experiments revealed that the SNORD114-1 (14q(II-1)) variant promoted cell growth through cell cycle modulation; its expression was implicated in the G0/G1 to S phase transition mediated by the Rb/p16 pathways. This study thus reports three important observations: (1) snoRNA regulation is different in normal cells compared with cancer cells; (2) a relationship exists between a chromosomal translocation and expression of snoRNA loci; and (3) snoRNA expression can affect Rb/p16 cell cycle regulation. Taken together, these data strongly suggest that snoRNAs have a role in cancer development.

[Clear-cell sarcoma of the kidney: about a paediatric case]. / Sarcome à cellules claires du rein : à propos d'un cas pédiatrique.

Clear-cell sarcoma of the kidney (CCSK) is a rare malignant tumor of childhood, known for its aggressiveness, its tendency to recurrence and to metastasis to bone. We report an observation of a child of 48 months carrying a large abdominal mass. The diagnosis of the SCCR was made on biopsy, since imaging remained uncertain as to the renal origin of the mass. Indeed, our observation underlines the difficulty of its diagnosis. Excepting the morphological aspect, there is no criterion for its recognition. Its prognosis has been improved by the new treatments.

PAX5 mutations occur frequently in adult B-cell progenitor acute lymphoblastic leukemia and PAX5 haploinsufficiency is associated with BCR-ABL1 and TCF3-PBX1 fusion genes: a GRAALL study.

Adult and child B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) differ in terms of incidence and prognosis. These disparities are mainly due to the molecular abnormalities associated with these two clinical entities. A genome-wide analysis using oligo SNP arrays recently demonstrated that PAX5 (paired-box domain 5) is the main target of somatic mutations in childhood BCP-ALL being altered in 38.9% of the cases. We report here the most extensive analysis of alterations of PAX5 coding sequence in 117 adult BCP-ALL patients in the unique clinical protocol GRAALL-2003/GRAAPH-2003. Our study demonstrates that PAX5 is mutated in 34% of adult BCP-ALL, mutations being partial or complete deletion, partial or complete amplification, point mutation or fusion gene. PAX5 alterations are heterogeneous consisting in complete loss in 17%, focal deletions in 10%, point mutations in 7% and translocations in 1% of the cases. PAX5 complete loss and PAX5 point mutations differ. PAX5 complete loss seems to be a secondary event and is significantly associated with BCR-ABL1 or TCF3-PBX1 fusion genes and a lower white blood cell count.

Detection of different clonal EBV strains in Hodgkin lymphoma and nasopharyngeal carcinoma tissues from the same patient.

The ubiquitous herpesvirus Epstein-Barr virus (EBV) is linked to the development of several malignancies, including nasopharyngeal carcinoma (UCNT) and Hodgkin lymphoma (HL). Despite the well-known oncogenic properties of the EBV latent membrane protein 1 (LMP-1), the different oncogenic pathways involved in the pathogenesis of each disease remain unclear. This study reported, for the first time, the case of a patient with sequential development of UCNT and HL. Polymerase chain reaction was used to determine the LMP-1 gene sequence and demonstrate that the two tumours contained different clonal viral genomes, suggesting a central and specific role of EBV infection.

Diagnostic value of combining immunostaining for CD3 and nuclear morphometry in mycosis fungoides.

BACKGROUND: Mycosis fungoides (MF) is the most common skin lymphoid neoplasm. In initial stages, differential diagnosis of MF from other benign dermal lymphoid infiltrates (BDLI) may be impossible on morphological basis alone. In previous studies, only deletion of CD7 in MF proved to be of diagnostic help, but not the ratio between immunoexpression of CD4 and CD8. METHODS: 30 cases of MF and 11 cases of BDLI were analysed, in order to compare morphometric parameters, which could be of diagnostic aid. As CD7 is frequently deleted in MF, immunohistochemical detection of T-cells was made using an antibody to CD3. Images of 100 CD3-positive cells per case in both groups were captured and analysed using a simple computer program for nuclear perimeter, area, diameter and nuclear contour index. RESULTS: All parameters showed statistically significant higher values for MF. Area was the variable with the strongest discriminating power between the two groups of patients. Thus even if morphological evaluation is not accurate to distinguish benign versus malignant dermal lymphoid infiltrates, due to the variability of size and shape of these cells, a more sensitive method promptly shows this difference. CONCLUSION: Results suggest that morphometry of CD3-positive lymphoid cells may add valuable information in the differential diagnosis of MF and benign dermatoses.

[Psycho-pathological risk factor of arterial disease (prevalence, superposition or vinculation with other factors)]. / Estudio oncativo: factores de riesgo psicopatológicos de enfermedad arterial (prevalencia, superposición o vínculo con otros factores).

Alexitimia and depression may or not coexist with others risk factors (comportment o physical). Frecuently they have relation with socio-echonomic status and with ethnia. Sometimes are determinants of the atherosclerotic process by increasing the vascular reactivity by the alteration of the evolution. There is no information in our country about this problem in general population. The present study result of the investigation of these aspects and the comportamental and physical factors of arterial disease, in a population of Cordoba province (Argentina Republic).

Estudio oncativo: factores de riesgo psicopatológicos de enfermedad arterial (prevalencia, superposición o vínculo con otros factores) / Psycho-pathological risk factor of arterial disease (prevalence, superposition or vinculation with other factors)

Entidades psicopatológicas como alexitimia y depresión, pueden coexistir o no con factores de riesgo de enfermedad arterial o ser entidades independientes. Tienen frecuentemente relación con aspectos socio-económico-culturales y con la etnia; a veces son determinantes del proceso ateroesclerótico por alterar la reactividad vascular (Alexitimai) o agravadores del proceso evolutivo (depresión). Materiales y métodos: Existe escasa información sobre esta tematica en nuestro país: la existente se refiere a los mismos en muestras seleccionadas, no de poblaciones generales (urbanas o rurales). Resultados: se exponen los resultados obtenidos de lainvestigación de esos factores en una población de la Provincia de Córdoba (RA), donde simultáneamente se estudiaron factores de orden fisico y conductual. Conclusiones: La prevalencia de la depresión es más elevada que en otras poblaciones

Alexitimia and depression may or not coexist with others risk factors (comportment o physical). Frecuently they have relation with socio-echonomic status and with ethnia. Sometimes are determinants of the atherosclerotic process by increasing the vascular reactivity by the alteration of the evolution. There is no information in our country about this problem in general population. The present study result of the investigation of these aspects and the comportamental and physical factors of arterial disease, in a population of Cordoba province (Argentina Republic).

"In situ-like" mantle cell lymphoma: a report of two cases.

Mantle cell lymphoma (MCL) is a B cell neoplasm that most often shows a diffuse growth pattern. Two cases of MCL are reported here, both with a previous diagnosis of lymphoid hyperplasia. Morphologically, germinal centres are hyperplasic with a normal or discretely enlarged mantle zone, where foci of irregularly shaped small lymphocytes are seen. These are positive for CD20, CD5 and cyclin D1, confirming a diagnosis of in situ-like MCL. This type differs from the mantle zone pattern in that the neoplastic mantle zone is very thin and there is very little or no spread of tumour cells into interfollicular areas. To the best of our knowledge, this is the first report on such a pattern of MCL, which is important to recognise, as it can be confused with lymphoid hyperplasia.

Gene expression profiling identifies molecular subgroups among nodal peripheral T-cell lymphomas.

The classification of peripheral T-cell lymphomas (PTCL) is still a matter of debate. To establish a molecular classification of PTCL, we analysed 59 primary nodal T-cell lymphomas using cDNA microarrays, including 56 PTCL and three T-lymphoblastic lymphoma (T-LBL). The expression profiles could discriminate angioimmunoblastic lymphoma, anaplastic large-cell lymphoma and T-LBL. In contrast, cases belonging to the broad category of 'PTCL, unspecified' (PTCL-U) did not share a single molecular profile. Using a multiclass predictor, we could separate PTCL-U into three molecular subgroups called U1, U2 and U3. The U1 gene expression signature included genes known to be associated with poor outcome in other tumors, such as CCND2. The U2 subgroup was associated with overexpression of genes involved in T-cell activation and apoptosis, including NFKB1 and BCL-2. The U3 subgroup was mainly defined by overexpression of genes involved in the IFN/JAK/STAT pathway. It comprised a majority of histiocyte-rich PTCL samples. Gene Ontology annotations revealed different functional profile for each subgroup. These results suggest the existence of distinct subtypes of PTCL-U with specific molecular profiles, and thus provide a basis to improve their classification and to develop new therapeutic targets.