Clinical and histological study of follicular helper T-cell lymphomas with indolent evolution.

INTRODUCTION: Follicular helper T-cell lymphomas (TFHL) have an aggressive course with a poor outcome. European and US guidelines recommend anthracycline-based chemotherapy as a first-line treatment, but the 5-year overall survival rate is still approximately 30%. We describe here the features of a cohort of TFHL patients who experienced prolonged survival despite the absence of specific treatment or the initiation of steroid-based therapy. PATIENTS AND METHODS: In our study, we describe 15 adult patients who suffered from TFHL and had not received intensive chemotherapy at diagnosis for any reason. Biopsies of these cases were centrally reviewed, and the mutational pattern was determined using next-generation sequencing. RESULTS: These 15 patients had the classic clinical, biological and pathological features of TFHL, angioimmunoblastic-type. TET2 mutations were found in 83% of patients; RHOA G17V, IDH2 R172 and DNMT3A mutations were found in 67%, 42% and 33% of the patients, respectively. Among the 15 patients, 8 did not receive any treatment, and 7 received steroid-based treatment. Ten patients had progression (5 in each group). Four patients died (3 of them from the progression of their lymphoma). The median follow-up in our cohort was 53 months. The 5-year OS was 66%, 100% for untreated patients and 29% for the others. In those 2 groups, the median time to treatment initiation was 22 months from diagnosis. CONCLUSION: We described a series of 15 well-characterized TFHL patients with an indolent outcome, suggesting that a watch-and-wait approach can be proposed in selected patients. Identifying factors predicting such evolution is warranted.

An EMT-primary cilium-GLIS2 signaling axis regulates mammogenesis and claudin-low breast tumorigenesis.

The epithelial-mesenchymal transition (EMT) and primary ciliogenesis induce stem cell properties in basal mammary stem cells (MaSCs) to promote mammogenesis, but the underlying mechanisms remain incompletely understood. Here, we show that EMT transcription factors promote ciliogenesis upon entry into intermediate EMT states by activating ciliogenesis inducers, including FGFR1. The resulting primary cilia promote ubiquitination and inactivation of a transcriptional repressor, GLIS2, which localizes to the ciliary base. We show that GLIS2 inactivation promotes MaSC stemness, and GLIS2 is required for normal mammary gland development. Moreover, GLIS2 inactivation is required to induce the proliferative and tumorigenic capacities of the mammary tumor­initiating cells (MaTICs) of claudin-low breast cancers. Claudin-low breast tumors can be segregated from other breast tumor subtypes based on a GLIS2-dependent gene expression signature. Collectively, our findings establish molecular mechanisms by which EMT programs induce ciliogenesis to control MaSC and MaTIC stemness, mammary gland development, and claudin-low breast cancer formation.

Peritoneal or mesenteric tumours revealing histiocytosis.

OBJECTIVE: Peritoneal or mesenteric tumours may correspond to several tumour types or tumour-like conditions, some of them being represented by histiocytosis. This rare condition often poses diagnostic difficulties that can lead to important time delay in targeted therapies. Our aim was to describe main features of histiocytoses with mesenteric localisation that can improve the diagnostic process. DESIGN: We performed a retrospective study on 22 patients, whose peritoneal/mesenteric biopsies were infiltrated by histiocytes. RESULTS: Abdominal pain was the revealing symptom in 10 cases, and 19 patients underwent surgical biopsies. The diagnosis of histiocytosis was proposed by initial pathologists in 41% of patients. The other initial diagnoses were inflammation (n=7), sclerosing mesenteritis (n=4) and liposarcoma (n=1). The CD163/CD68+CD1a- histiocytes infiltrated subserosa and/or deeper adipose tissues in 16 and 14 cases, respectively. A BRAFV600E mutation was detected within the biopsies in 11 cases, and two others were MAP2K1 mutated. The final diagnosis was histiocytosis in 18 patients, 15 of whom had Erdheim-Chester disease. The median diagnostic delay of histiocytosis was 9 months. Patients treated with BRAF or MEK inhibitors showed a partial response or a stable disease. One patient died soon after surgery, and five died by the progression of the disease. CONCLUSION: Diagnosis of masses arising in the mesentery should be carefully explored as one of the possibilities in histiocytosis. This diagnosis is frequently missed on mesenteric biopsies. Molecular biology for detecting the mutations in BRAF or in genes of the MAP kinase pathway is a critical diagnostic tool.

Phenotypic discordance between primary and metastatic breast cancer in the large-scale real-life multicenter French ESME cohort.

Expression of hormone receptor (HR) for estrogens (ER) and progesterone (PR) and HER2 remains the cornerstone to define the therapeutic strategy for breast cancer patients. We aimed to compare phenotypic profiles between matched primary and metastatic breast cancer (MBC) in the ESME database, a National real-life multicenter cohort of MBC patients. Patients with results available on both primary tumour and metastatic disease within 6 months of MBC diagnosis and before any tumour progression were eligible for the main analysis. Among the 16,703 patients included in the database, 1677 (10.0%) had available biopsy results at MBC diagnosis and on matched primary tumour. The change rate of either HR or HER2 was 27.0%. Global HR status changed (from positive = either ER or PR positive, to negative = both negative; and reverse) in 14.2% of the cases (expression loss in 72.5% and gain in 27.5%). HER2 status changed in 7.8% (amplification loss in 45.2%). The discordance rate appeared similar across different biopsy sites. Metastasis to bone, HER2+ and RH+/HER2- subtypes and previous adjuvant endocrine therapy, but not relapse interval were associated with an HR discordance in multivariable analysis. Loss of HR status was significantly associated with a risk of death (HR adjusted = 1.51, p = 0.002) while gain of HR and HER2 discordance was not. In conclusion, discordance of HR and HER2 expression between primary and metastatic breast cancer cannot be neglected. In addition, HR loss is associated with worse survival. Sampling metastatic sites is essential for treatment adjustment.

Composite and sequential lymphoma between classical Hodgkin lymphoma and primary mediastinal lymphoma/diffuse large B-cell lymphoma, a clinico-pathological series of 25 cases.

Composite and sequential lymphomas involving both classical Hodgkin lymphoma (CHL) and primary mediastinal B-cell lymphoma (PMBCL) are rare phenomena. Beyond the relevant biological interest raised by these cases, treatments and outcome data are poorly covered in the recent literature. This retrospective analysis describes the pathological and clinical characteristics of 10 composite and 15 sequential cases included after a central pathological review. At diagnosis, 70% of the composite lymphomas presented a disseminated and extranodal disease. Among the 15 sequential lymphomas, 12 were CHL at first occurrence and three were PMBCL. Based on their clinical evolution, these sequential lymphomas could be divided into early (i.e., diagnosis of second lymphoma within a year) and late [(i.e., a second lymphoma occurrence occurring after a long period of complete remission]). All composite cases were alive in complete remission after a median follow-up of 34 months. If the early sequential lymphoma presented a particularly poor outcome with a median overall survival shorter than one year, the late cases were efficiently salvaged. Further molecular studies are needed to describe the underlying biology of these rare diseases, possibly representing the extreme of tumour cell plasticity found in grey-zone lymphoma.

Predictive factors associated with involved margins in breast cancer treated with neoadjuvant chemotherapy followed by breast-conserving therapy.

INTRODUCTION: This study sought to identify predictive factors of involved surgical margins in breast-conserving surgery (BCS) after neoadjuvant chemotherapy (NAC) to help guide the surgical procedure. MATERIALS AND METHODS: Retrospective study of patients who had BCS after NAC between January 2008 and December 2013. OUTCOME MEASURE: tumor-involved margin, defined by tumor cells on ink for invasive cancer and tumor-free margin < 2 mm for DCIS. RESULTS: Ninety-seven patients were included. The median age of patients was 46 years old [28-71]. The initial average tumor size was 47.8 mm [+/- 18.6]. Twelve patients (12.4%) had involved tumor margins on final histology after BCS and NAC. According to the multivariate model including only preoperative variables of positive margins, initial ultrasound tumor size ≤ 27 mm (p = 0.045) and low SBR grade (p = 0.009) were independently associated with tumor-involved margins. According to the multivariate model including pre- and postoperative variables of positive margins, ductal carcinomain situ was also independently associated with tumor-involved margins (p = 0.021). CONCLUSION: Initial ultrasound tumor size ≤ 27 mm and low SBR grade were independently associated with tumor-involved margins. These preoperative data were very helpful to guide the surgical procedure in breast cancer.

Tumor analysis: freeze-thawing cycle of triple-negative breast cancer cells alters tumor CD24/CD44 profiles and the percentage of tumor-infiltrating immune cells.

OBJECTIVE: The use of novel methods to characterize living tumor cells relies on well-conceived biobanks. Herein, we raised the question of whether the composition of fresh and freeze/thawed dissociated tumor samples is comparable in terms of quantitative and qualitative profiling. RESULTS: Breast cancer is a heterogeneous disease, encompassing luminal A and B, basal/triple-negative breast cancer (TNBC), and ERBB2-like tumors. We examined living cells dissociated from TNBC and found that a classical freeze/thaw protocol leads to a marked reduction in the number of CD45-CD44LowCD24Low tumor cells. This, in turn, changed the percentage of tumor cells with certain CD44/CD24 expression patterns and changed the percentage of tumor-infiltrating immune cells. These cryopreservation-driven alterations in cellular phenotype make it impossible to compare fresh and frozen samples from the same patient directly. Moreover, the freeze/thaw process changed the transcriptomic signatures of triple-negative cancer stem cells in such a manner that hierarchical clustering no longer ranked them according to expected inter-individual differences. Overall, this study suggests that all analyses of living tumor cells should be conducted only using freshly dissociated tumors if we are to generate a robust scoring system for prognostic/predictive markers.

Impact of Expert Pathologic Review of Lymphoma Diagnosis: Study of Patients From the French Lymphopath Network.

Purpose To prospectively assess the clinical impact of expert review of lymphoma diagnosis in France. Materials and Methods From January 2010 to December 2013, 42,145 samples from patients with newly diagnosed or suspected lymphomas were reviewed, according to the 2008 WHO classification, in real time by experts through the Lymphopath Network. Changes in diagnosis between referral and expert review were classified as major or minor according to their potential impact on patient care. Results The 42,145 reviewed samples comprised 36,920 newly diagnosed mature lymphomas, 321 precursor lymphoid neoplasms, 314 myeloid disorders, and 200 nonhematopoietic neoplasms, with 4,390 benign lesions. There were 4,352 cutaneous and 32,568 noncutaneous lymphomas. The most common mature noncutaneous lymphomas were diffuse large B-cell lymphomas (32.4%), follicular lymphomas (15.3%), classic Hodgkin lymphomas (13%), peripheral T-cell lymphomas (6.3%) of which angioimmunoblastic T-cell lymphomas (2.3%) were the most frequent, and mucosa-associated lymphoid tissue lymphomas (5.8%). A diagnostic change between referral and expert review occurred in 19.7% of patients, with an estimated impact on patient care for 17.4% of patients. This rate was significantly higher for patients sent with a provisional diagnosis seeking expert second opinion (37.8%) than for patients sent with a formal diagnosis (3.7%). The most frequent discrepancies were misclassifications in lymphoma subtype (41.3%), with 12.3% being misclassifications among small B-cell lymphoma entities. Fewer than 2% of changes were between benign and malignant lymphoid conditions. Minor changes (2.3%) mostly consisted of follicular lymphoma misgrading and diffuse large B-cell lymphoma subtype misclassification. Conclusion To our knowledge, this study provides the largest ever description of the distribution of lymphoma entities in a western country and highlights how expert review significantly contributes to a precise lymphoma diagnosis and optimal clinical management in a proportion of patients.

Lectin-like transcript 1 is a marker of germinal center-derived B-cell non-Hodgkin's lymphomas dampening natural killer cell functions.

Non-Hodgkin's lymphomas (NHLs) are malignant neoplasms which are clinically and biologically diverse. Their incidence is constantly increasing and despite treatment advances, there is a need for novel targeted therapies. Here, we identified Lectin-like transcript 1 (LLT1) as a biomarker of germinal center (GC)-derived B-cell NHLs. LLT1 identifies GC B cells in reactive tonsils and lymph nodes and its expression is maintained in B-cell NHLs which derive from GC, including Burkitt lymphoma (BL), follicular lymphoma (FL), and GC-derived diffuse large B-cell lymphoma (DLBCL). We further show that LLT1 expression by tumors dampens natural killer (NK) cell functions following interaction with its receptor CD161, uncovering a potential immune escape mechanism. Our results pinpoint LLT1 as a novel biomarker of GC-derived B-cell NHLs and as a candidate target for innovative immunotherapies.

The locoregional recurrence post-mastectomy for ductal carcinoma in situ: Incidence and risk factors.

BACKGROUND: The objective of this retrospective study was to determine the incidence of recurrence of breast cancer after mastectomy for ductal carcinoma in situ (DCIS) in our institution, and to evaluate the associated risk factors while comparing them to those proposed in the literature. METHODS: The files of 218 patients who had undergone mastectomy for pure DCIS or DCIS with micro-invasion at Centre Eugène Marquis between January 2003 and November 2013 were compared for: age at diagnosis, type of mastectomy and immediate reconstruction, tumor characteristics, and the evaluation of the sentinel axillary lymph node. The mean follow-up period was 30.5 months. RESULTS: In a mean period of 39.13 months, 8 patients (3.67%) developed a recurrence post-mastectomy, 2 of whom with distant metastasis. Two others developed distant metastases subsequently during treatment. All 4 died due to progression of metastases, while the other 4 are alive and disease-free after treatment. The only risk factor was young age at initial diagnosis (<40 years). None of the other factors described in the literature, such as high grade or diffuse disease, comedo-necrosis, positive margins or micro-invasion were statistically significant. CONCLUSION: The recurrence of breast cancer after mastectomy for DCIS is rare, however, it carries a high mortality rate for those who do relapse. Patients who have high risk factors such as young age at diagnosis and high risk tumor factors should be followed closely for signs of recurrence and/or metastasis.

COUP-TFI modifies CXCL12 and CXCR4 expression by activating EGF signaling and stimulates breast cancer cell migration.

BACKGROUND: The orphan receptors COUP-TF (chicken ovalbumin upstream promoter transcription factor) I and II are members of the nuclear receptor superfamily that play distinct and critical roles in vertebrate organogenesis. The involvement of COUP-TFs in cancer development has recently been suggested by several studies but remains poorly understood. METHODS: MCF-7 breast cancer cells overexpressing COUP-TFI and human breast tumors were used to investigate the role of COUP-TFI in the regulation of CXCL12/CXCR4 signaling axis in relation to cell growth and migration. We used Immunofluorescence, western-blot, RT-PCR, Formaldehyde-assisted Isolation of Regulatory Elements (FAIRE) assays, as well as cell proliferation and migration assays. RESULTS: Previously, we showed that COUP-TFI expression is enhanced in breast cancer compared to normal tissue. Here, we report that the CXCL12/CXCR4 signaling pathway, a crucial pathway in cell growth and migration, is an endogenous target of COUP-TFI in breast cancer cells. The overexpression of COUP-TFI in MCF-7 cells inhibits the expression of the chemokine CXCL12 and markedly enhances the expression of its receptor, CXCR4. Our results demonstrate that the modification of CXCL12/CXCR4 expression by COUP-TFI is mediated by the activation of epithelial growth factor (EGF) and the EGF receptor. Furthermore, we provide evidence that these effects of COUP-TFI increase the growth and motility of MCF-7 cells in response to CXCL12. Cell migration toward a CXCL12 gradient was inhibited by AMD3100, a specific antagonist of CXCR4, or in the presence of excess CXCL12 in the cell culture medium. The expression profiles of CXCR4, CXCR7, CXCL12, and COUP-TFI mRNA in 82 breast tumors and control non-tumor samples were measured using real-time PCR. CXCR4 expression was found to be significantly increased in the tumors and correlated with the tumor grade, whereas the expression of CXCL12 was significantly decreased in the tumors compared with the healthy samples. Significantly higher COUP-TFI mRNA expression was also detected in grade 1 tumors. CONCLUSIONS: Together, our mechanistic in vitro assays and in vivo results suggest that a reduction in chemokine CXCL12 expression, with an enhancement of CXCR4 expression, provoked by COUP-TFI, could be associated with an increase in the invasive potential of breast cancer cells.

Atypical hyperplasia of the breast: the black hole of routine breast cancer screening.

AIM: Determination of the prevalence, of the radiological and clinical characteristics, and outcome of atypical hyperplasia (AH) of the breast within a population subjected to routine screening (double-view mammography with double reading, performed every two years between 50 and 75 years of age). PATIENTS AND METHODS: The clinical and radiological records and histological findings of percutaneous and surgical biopsy specimens of sixty-eight patients presenting with AH were reviewed together with patient follow-up data after percutaneous and surgical biopsy. RESULTS: AH incidence in the population was 0.19‰ with the following distribution of lesions: atypical epithelial hyperplasia (AEH, 53%), columnar cell metaplasia with atypia (CCMA, 32%), and lobular intraepithelial neoplasia (LIN, 8%). The mean patient age was 58 years and 24% of patients were receiving hormone replacement therapy. The main radiological finding was the presence of microcalcifications for AEH and CCMA lesions in particular, and the mammograms were valid (correlation between American College of Radiology score and risk of lesion, only 3% of lesions were recognized on the second reading). A total of 13.7% of AH cases were underestimated and a real risk of AH progression was observed, regardless of whether or not surgical biopsy had been performed. CONCLUSION: The clinical and radiological characteristics of AH observed in a population subjected to routine breast cancer screening are identical to those for patients with the same lesions referred to specialist centers. Surgical biopsy remains the recommended option due to the risk of underestimation of lesions by percutaneous biopsy and the risk of progression justifies the need for continued close monitoring.

Sentinel lymph node analysis in breast cancer: contribution of one-step nucleic acid amplification (OSNA).

One-step nucleic acid amplification (OSNA, Sysmex, Kobe, Japan) offers an excellent opportunity for accurate exhaustive sentinel lymph node (SLN) examination in breast cancer patients. Calibrated with conventional postoperative histology, this molecular technique yields comparable results intraoperatively, expressed as micrometastasis, macrometastasis or no metastasis depending on the CK19 mRNA copy number amplified in SLN lysates. We applied OSNA to detect metastasis in 810 SLNs from 367 patients with early stage breast cancer. We compared the rate of OSNA-positive SLNs in patients with invasive breast cancer (< 2 cm) versus the rate observed in a historical cohort using conventional histological examination of SLNs. No significant difference was observed, the OSNA assay was positive in 24.4% of patients, compared with positive histology in 24.8% in the historical cohort if including patients with isolated tumour cell (ITC) and in 23.4% excluding them. Opportunities for optimised patient management using OSNA are discussed: intraoperative detection of OSNA-positive SLNs enables axillary lymph node dissection (ALND) during the same procedure; standard OSNA techniques enable the establishment of homogeneous groups based on examination of whole SLNs for valid comparisons between different centres.

Pure flat epithelial atypia (DIN 1a) on core needle biopsy: study of 60 biopsies with follow-up surgical excision.

Flat epithelial atypia (FEA) is recognized as a precursor of breast cancer and its management (surgical excision or intensive follow-up) remains unclear after diagnosis on core needle biopsy (CNB). The aim of this study was to determine the underestimation rate of pure FEA on CNB and clinical, radiological, and pathological factors of underestimation. 4,062 CNBs from 5 breast cancer centers, performed over a 5-year period, were evaluated. A CNB diagnosis of pure FEA was made in 60 cases (1.5%) (the presence of atypical ductal hyperplasia, lobular neoplasia, radial scars, phyllodes tumor, papillary lesions, ductal carcinoma in situ or invasive carcinoma at CNB were exclusion criteria), and subsequent surgical excision was systematically performed. The histological diagnosis was retrospectively reviewed using standardized criteria and the precise terminology of the World Health Organization by two pathologist physicians. At surgical excision, 6 (10%) ductal carcinoma in situ and 2 (3%) invasive carcinoma were diagnosed. The total underestimation rate was 13%. FEA was associated with atypical ductal hyperplasia in 10 (17%) cases and with lobular neoplasia in 2 (3%) at final pathology. Residual FEA was found in 14 (23%) cases. No clinical, radiological or pathological factors were significantly associated with underestimation. Our data highlight the importance of recognizing and diagnosing FEA in core needle biopsies. Thus, the presence of FEA on CNB, even in isolation, warrants follow-up excision.

Intraoperative touch imprint cytology of axillary sentinel nodes for breast cancer: a series of 355 procedures.

AIMS: Our objective was to evaluate intraoperative sentinel node touch imprint cytology (IOSNTI) for breast cancer. Three hundred and fifty-five patients with invasive breast cancer (pT1N0, lobular or ductal subtype) were included in our study. IOSNTI consists of touching glass slides to the surfaces of interest after gently pressing the spatially localized specimen, taken according to predetermined conditions, in order to perform a final histological examination consisting of H&E and immunohistochemical staining. RESULTS: The total sensitivity (Se) of IOSNTI was 36% and 15% of patients with nodal metastasis went undetected during the intraoperative examination. Sensitivity was significantly lower for the oldest patients (aged over 57 years: 25%), small tumors (smaller than 12 mm: 23.3%), lobular subtypes (8.3%), in the absence of vascular emboli (33%) and for detection of micrometastases (10%). DISCUSSION: This simple, fast and relatively inexpensive method could be combined with intraoperative molecular biology methods in populations in which cytology is less efficient and produces negative results.

[Update of the GEFPICS' recommendations for HER2 status determination in breast cancers in France]. / Mise à jour des recommandations du GEFPICS pour l'évaluation du statut HER2 dans les cancers du sein en France.

In Europe, patients who may benefit from an HER2 targeted drug are currently selected by immunohistochemistry (IHC). In situ hybridization (ISH) techniques should be used for complementary assessment of ambiguous 2+ IHC cases and for the calibration of the IHC technique. Eligibility to an HER2 target treatment is defined by an HER2 positive status being IHC test 3+ or 2+ amplified. Reliable detection of HER2 status is essential to the appropriate usage of HER2 targeted drugs because its specificity is limited to tumors overexpressing HER2. It is essential that the IHC evaluation of the HER2 status of a mammary carcinoma is optimized and reliable. This GEFPICS' guidelines look over the different steps of the IHC technique, the controls and, the rules for interpretation. Once acquired, this knowledge must be perpetuated by the observation of rules of good technical practice (internal and external controls, quality assurance programs).