Clear cell renal cell carcinoma (ccRCC) shows variable chromosomal abnormalities. The aim of this study was to assess the prognostic role of ccRCC chromosomal abnormalities in a single-center cohort ...with an extended follow-up.
A systematic cytogenetic analysis was performed in 283 consecutive surgically-treated patients for renal masses between 1997 and 2002. Kaplan-Meier and multivariable Cox regression (MCR) models were used to calculate cancer specific survival (CSS).
Among 174 ccRCC patients, the most common abnormality was deletion in chromosome 3 (54.6%). At a median follow-up of 119 months, 38 patients (21.8%) died from RCC. At MCR models, worse CSS was independently predicted by deletions in chromosomes 2, 19, 20 or 22 and insertions in chromosome 18.
Specific ccRCC chromosomal abnormalities are independently associated with worse CSS. Cytogenetic evaluation may direct further genetic analysis for personalized prognostic stratification.
Background. We recently reported that Circulating Endothelial Cell (CEC) count changes represent a promising marker to monitor endothelial damage in patients undergoing allogeneic hematopoietic stem ...cell transplant (allo-HSCT), potentially becoming a valuable tool in the diagnostic definition of GVHD. Besides confirming an increase of CEC counts at GVHD onset, we repeatedly documented at time of engraftment statistically significant higher numbers of CEC in patients who will not manifest GVHD in comparison to patients in which GVHD will be diagnosed (Transplantation 2014,98:706-12; Bone Marrow Transplantation 2017,52:1637-42; Scientific Reports 2019,9:1-12). Recent knowledges in organ transplant pointed out that endothelial cells from the grafted organ, besides being a continuous source of alloantigens, can downregulate alloreactivity exerting tolerogenic responses. By inference to the allo-HSCT field, it could be envisaged that presence of donor CEC could induce protective effects on alloreactivity.
Methods. We planned a study to test the hypothesis that at time of engraftment, CEC present in peripheral blood (PB), besides coming from cells shedding from patient vasculature, could partly belong to donor, originating from the cellular graft. Therefore, in an exploratory set, we performed FISH analysis on flowcytometry-sorted CEC (CD45neg/CD34bright/CD146pos, Lyotube #623920, BD Biosciences) (n=3) and on whole PB derived culture-expanded CEC (n=3) (EGM-2 BulletKit, Lonza), obtained at engraftment in sex-mismatched allo-HSCT. In the confirmatory set (n=15), single CEC were recovered from PB, at engraftment (T1) and at 90 days (T2) after allo-HSCT, through the DEPArrayTM technology (Menarini Silicon Biosystems), after preliminary bulk separation step carried out with the CellSearch® System. Single recovered CEC was whole genome amplified (Ampli1™ WGA Kit) and short tandem repeat (STR) profile determined (Ampli 1TM STR kit) on each single CEC. To confirm host/donor origin, single CEC STR profile was compared to that determined on patient and donor cells before allo-HSCT. Moreover, donor CEC presence was evaluated by CISH analysis on formaline fixed and paraffin-embedded biopsy sections obtained at least three months after sex mismatched allo-HSCT.
Results. By positive findings of the exploratory set, we proved, at the single cell level in the confirmatory set, the presence of donor CEC at engraftment (T1) in 4 out of 15 patients (Table 1). Of them, 2 did not manifested GVHD, despite a GVHD risk score of 2, and the other 2 presented GVHD grade I. On the contrary, among the 10 patients in whom no donor CEC were detected, 6 experienced GVHD grade II-III, while 4 did not manifested GVHD, despite a 1-3 GVHD risk score.
Conclusions. Our data represent the proof of principle that donor CEC may flow in host PB early on from hematopoietic recovery and seldom persist thereafter at steady-state conditions, being potentially embedded in host vascular wall. These puzzling findings suggest that neovascularization takes place in parallel with hematopoietic engraftment and could provide further clues on shedding light on tissue tolerance in the context of GVHD, opening up paradoxical scenarios on the protective role potentially played by donor CEC.
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Fontana:Menarini Silicon Biosystem: Employment. Rotta:BD Biosciences Italia: Employment. Manaresi:Menarini SIlicon Biosystem: Employment, Membership on an entity's Board of Directors or advisory committees.
Pancreatic mucinous cystic tumors (MCT) are proliferations of mucin-producing epithelia supported by an ovarian-like stroma. They are classified into adenomas (MCA), borderline (MCB) and noninvasive ...or invasive carcinomas (MCC). The molecular mechanisms underlying their clinical behavior are poorly understood, partly due to the lack of cellular models. We report the establishment of MCC1, the first cell line from a pancreatic MCT, deriving from the highly dysplastic cell component of a noninvasive MCC. MCC1 has mutations in codon 12 of K-RAS (GGT>GAT), codon 58 of P16 (CGA>TGA) and codon 132 of P53 (AAG>AGG). The FHIT and DPC4 genes are unaltered. Immunohistochemistry shows abnormal expression of MUC1 and p53, loss of p16 and retention of Fhit and Dpc4 in both the cell line and the highly dysplastic cells of the primary lesion. The morphological and molecular features of MCC1 and its corresponding primary tumor are consistent with a model for non-invasive MCC, where K-RAS, P16, P53 and MUC1 alterations are pre-invasive changes associated with progression of malignancy of MCT from adenoma to carcinoma. MCC1 is sensitive to 5-fluorouracil, representing the first assessment of drug sensitivity for MCC. Finally, MCC1 is a suitable model for preclinical studies, as it grows in immunodeficient mice.
The aim of this study was to investigate the biological and clinical significance of epidermal growth factor receptor (EGFR) signaling pathway in follicular dendritic cell sarcoma (FDC-S).
Expression ...of EGFR and cognate ligands as well as activation of EGFR signaling components was assessed in clinical samples and in a primary FDC-S short-term culture (referred as FDC-AM09). Biological effects of the EGFR antagonists cetuximab and panitumumab and the MEK inhibitor UO126 on FDC-S cells were determined in vitro on FDC-AM09. Direct sequencing of KRAS, BRAF, and PI3KCA was conducted on tumor DNA.
We found a strong EGFR expression on dysplastic and neoplastic FDCs. On FDC-AM09, we could show that engagement of surface EGFR by cognate ligands drives the survival and proliferation of FDC-S cells, by signaling to the nucleus mainly via MAPK and STAT pathways. Among EGFR ligands, heparin-binding EGF-like growth factor, TGF-α and Betacellulin (BTC) are produced in the tumor microenvironment of FDC-S at RNA level. By extending this finding at protein level we found that BTC is abundantly produced by FDC-S cells and surrounding stromal cells. Finally, direct sequencing of tumor-derived genomic DNA showed that mutations in KRAS, NRAS, BRAF, and PI3KCA, which predicts resistance to anti-EGFR MoAb in other cancer models, are not observed in FDC-S.
Activation of EGFR by cognate ligands produced in the tumor microenvironment sustain viability and proliferation of FDC-S indicating that the receptor blockade might be clinically relevant in this neoplasm.
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Introduction: Patients with aggressive B cell lymphomas carrying concomitant c-Myc and Bcl-2 and/or Bcl-6 gene rearrangement (DHL/THL) have a poor outcome when treated with standard R-CHOP regimen. ...Better results were reported in patients treated with more intensive programs, currently used in highly proliferative diseases such as Burkitt lymphoma. In particular, R-DA-EPOCH obtained the best result in a retrospective study comparing different intensive regimens (Oki et al. BJH 2014). We report the experience with this program of two hematologic centres in Northern Italy in patients with dysregulation of c-Myc gene associated with Bcl-2 and/or Bcl-6 abnormalities.
Methods: From January 2014, all consecutive fit patients aged less than 80 years, with diffuse large B cell lymphoma (DLBCL) or lymphoma with intermediate features between DLBCL and Burkitt (BCLU), including 10 patients with histological features suggesting transformation from follicular lymphoma, and showing DHL or THL by fluorescent in situ hybridization (FISH), were treated with R-DA-EPOCH and central nervous system prophylaxis with intrathecal methotrexate +/- cytarabine . Patients with c-Myc amplification (more than 4 signals) plus Bcl-2 and/or Bcl-6 gene rearrangement or amplification were also included (AMPL). Pre-treatment with one cycle of R-CHOP was allowed in patients in need of urgent treatment, pending the results of FISH analysis. Autologous stem cell transplantation (ASCT) was planned in one of the two centres for patients who reached at least a partial remission after six R-DA-EPOCH courses. Cell of origin (COO) was defined by immunohistochemistry (IHC) according to Hans's algorithm.
Results: Thirty-five patients (30 DLBCL and 5 BCLU) were treated. They received a median of 6 courses of R-DA-EPOCH (range 1-6) administered either to inpatients or to outpatients using a single portable infusion pump. Six patients were pre-treated with R-CHOP and 10 patients received consolidation with ASCT. According to FISH analysis, 18 cases were DHL, 6 THL and 11 AMPL. The median age of the whole group was 63 years (range 34-79). Twenty-nine patients were male (83%). Thirty-two patients (91%) had Ann-Arbor stage III/IV and 25 (71%) had high-intermediate/high risk score according to International Prognostic Index (IPI), with extranodal presentation in 74%, mainly in bone and gastrointestinal tract. According to Hans's algorithm, 85% of patients were of germinal center COO. By IHC double expression of myc (>40%) and of bcl-2 protein (>50%) was reported in 75% of patients. Median lymphoma cells proliferation fraction assessed by Ki-67 IHC was 80% (range 35-100%). Three patients died of infectious complications during chemotherapy. The overall response rate for the entire cohort was 71%. Of seven refractory patients, five have died of lymphoma, one is disease free after allogeneic transplantation and one is still on treatment. Among 25 patients who achieved a response to front-line therapy, one died of suicide and 24 are still in remission after a median of 15 months. The 1-year PFS and OS rates were 69% and 73%, respectively. There was no significant association of PFS or OS with age, COO, type of histology, transformation from FL, extranodal involvement, Ki-67. THL had significant worse survival comparing to DHL and AMPL (20% vs 88% vs 68% p 0.03) (Fig.1).
Conclusions: These results confirm R-DA-EPOCH as a feasible program in unselected fit patients up to the age of 80. Short-term efficacy was high, considering the poor prognosis of double or triple-hit aggressive B-cell lymphoma. Few data are available to define the role of consolidative ASCT and its utility is still unclear. Treatment response seems to favourably impact on patient survival. A longer follow-up on larger number of patients is ongoing to confirm these promising results.
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Corradini:Takeda: Honoraria; Sanofi: Honoraria; Novartis: Honoraria; Roche: Honoraria; Janssen: Honoraria; Celgene: Honoraria; Amgen: Honoraria; Gilead: Honoraria. Rossi:Teva: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees.
Background: MYC rearrangements occur approximately in 5-15% of diffuse large B-cell lymphoma (DLBCL), and are known to unfavorably impact on patients (pts) survival, especially if associated to BCL2 ...or/and BCL6 rearrangements (double/triple-hit). Previous studies have shown that also a MYC gene ICN may confer a worse prognosis to these pts (Yoon, Histopathology 2008), but only few groups have transferred this observation into the clinical practice. However, this finding may account for those cases that, despite the absence of MYC rearrangements, show a similar aggressive clinical course. For this reason, a careful study of all aberrations involving MYC geneshould be widely applied. We report a single center experience on the clinical outcome of pts with MYC ICN in the setting of aggressive B-cell lymphoma.
Methods: In the study period (Aug 2011-Aug 2015) we performed FISH analysis in all consecutive pts with de novo or transformed DLBCL or B-cell lymphoma, unclassifiable (BCLU) that displayed Ki67 >80% or intense bcl2 protein expression. Interphase FISH has been performed on 23 μm thick sections of formalin fixed paraffin embedded tissues, using splitsignal DNA probes (Dako) specific for the following loci: 8q24 (MYC), 18q21 (BCL2) and 3q27 (BCL6). For each sample, 60 evaluable nuclei with complete FISH signals were scored. ICN was considered when 3 or more copies of the gene studied were identified.
Almost all pts with a MYC translocation have been treated according to Burkitt lymphoma (BL)-like regimens as first line therapy (FLT); pts with MYC ICN have been treated with BL-like regimens from 2013, whereas standard ICT (R-CHOP or R-CHOPlike) has been used for these pts diagnosed before 2013. Overall survival (OS) was evaluated by Kaplan Meier method.
Results: In the 4-year study period, 317 consecutive pts were diagnosed with de novo or transformed DLBCL or BCLU. Twenty-one (7%) showed a translocation of MYC, in single (14%), double (67%) or triple hit (19%), with respect to BCL2 and BCL6 rearrangements. Interestingly, 8 (2.5%) pts with no MYC translocation showed MYC ICN, ranging from 3 up to 10 gene copies per cell. This aberrant gene copy number was observed in > 80% of the analyzed nuclei in 7 cases. The characteristics of the pts with MYC gene rearrangement or ICN are summarized in Table 1.
Of the 8 pts with MYC ICN, 5 had de novo DLBCL, 2 had transformed DLBCL and 1 had BCLU. Seven pts also showed BCL2 and/or BCL6 rearrangement/ICN. Three pts were treated with standard ICT and 5 with a BLlike regimen. Two pts (25%) responded to the FLT, showing one complete and one partial remission (CR and PR, respectively); both received BLlike regimen. The patient with PR progressed after 7 months, and received a second line treatment likewise the 6 nonresponders to FLT. None of them had a response and eventually died, with a median OS of 10.5 months. Cause of death was lymphoma in all cases. The patient who obtained a CR was still in remission at 29 months from the end of the FLT. The survival of these pts, compared to pts with MYC rearrangement, is shown in Figure 1 (respectively 12% vs 57% at 2 years, p 0.04).
Conclusion: These preliminary data show a dismal prognosis of pts with aggressive B cell lymphoma and MYC ICN, significantly worse than pts harboring a MYC translocation, independently of the treatment received. To better understand the genetic defect underlying this FISH pattern, further analysis using a specific centromere probe for chromosome 8 is ongoing on these and new cases. A previous study distinguished gain from amplification of MYC based on the number of gene copies, identifying 2 groups with different prognosis (Valera, Hematologica 2013). In our hands, there was a wider range of gene copy number in each patient, and the outcome was homogeneously poor. Overall, the biological consequence of this alteration is still to be fully explained, and only a few studies have addressed this point so far (Stasik, Hematologica 2010). Further investigation on larger cohort groups is necessary to confirm the unfavorable prognostic role of MYC ICN in aggressive B cell lymphoma. Such insights would help practitioners to determine more accurate therapeutic approaches, in order to improve the outcome of this specific subset of pts.
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No relevant conflicts of interest to declare.
Inflammatory myofibroblastic tumor (IMT) is a locally aggressive neoplasm, most frequently occurring in the abdominal cavity as multiple recurrent nodules. We report a case of IMT in a 24-year-old ...male presenting as multiple nodules involving the omentum, the liver, and the colon. Spindle tumor cells expressed ALK with a cytoplasmic granular distribution, the CLTC-ALK fusion gene was demonstrated by reverse-transcriptase polymerase chain reaction analysis, and break-apart fluorescence in situ hybridization (FISH) probes for the ALK gene showed a pathological pattern (single red signal associated with 1/2 normal fused signals) highly suggestive for combined gene fusion and deletion. To reduce the surgically unresectable liver mass, the patient was treated with crizotinib, and after 4 months of treatment the disease was defined stable according to RECIST criteria. Interestingly, ALK and FISH/FICTION analysis revealed that tumor cells were widely dispersed as multiple microscopic foci or as single cells beneath the omental mesothelium. These findings indicate that IMT multifocality might result either from dissemination from the main tumor mass or development of multiple independent neoplastic foci; furthermore, they underline the need of omentectomy in abdominal IMT to obtain surgical radicality.
Introduction: Diffuse Large B-cell lymphoma (DLBCL) is an heterogeneous disease in terms of biology, clinical presentation and prognosis. According to the putative Cell-Of-Origin (COO), DLBCL of the ...general HIV negative population can be classified into three molecular subtypes by gene expression profiling (GEP): the germinal center B-cell type (GCB), the activated B-cell type (ABC) and the unclassifiable type (UC). Immuno-histochemistry (IHC) models routinely surrogate GEP in the COO assignment, identifying GCB and non-GCB type. Hans algorithm is one of the most widely used IHC models and its concordance with GEP varies from 70 to 90% (Meyer PN et al, JCO 2011). NanoString (NS) 20-genes assay has emerged as a feasible technique for DLBCL molecular subgroups identification and shows a high concordance with GEP (Scott DW et al, Blood 2014). DLBCL classification according to COO categories has shown prognostic impact in the HIV negative population, since clinical outcome of non-GCB type is inferior if compared to GCB type in patients (pts) receiving CHOP or Rituximab-CHOP. DLBCL in HIV positive population have shown different biological features. COO subtypes distribution and its prognostic impact has not been extensively studied in HIV setting and still needs to be defined.
Aim of the study: To evaluate the proportion of COO subtypes in HIV-associated DLBCL according to IHC and NS assay, the concordance of these two different diagnostic methods, and the prognostic impact of COO assignment; to analyze BCL-2 and MYC protein expression and their correlation with outcome; to determine the prognostic value of baseline clinical characteristics, such as stage, LDH value and CD4+ lymphocyte count.
Methods: We retrospectively evaluated 66 cases of HIV positive pts with newly diagnosed DLBCL from 2000 to 2016, in 5 Italian Institutions. Histological samples were centrally reviewed by a panel of expert Pathologists for DLBCL diagnosis confirmation, BCL-2 and MYC protein expression and COO assignment according to Hans algorithm (Hans CP et al, Blood 2004). The cut off considered for BCL-2 and MYC overexpression was 70% and 40% respectively. NS Lymph2Cx assay for COO was performed in all pts. Clinical data were gathered from pts medical records.
Results: Pts were mostly male (73%) and median age at DLBCL onset was 45 years (range: 28-83). 80% of pts had advanced stage lymphoma and 34% showed a high-intermediate or high IPI score. HIV first detection was concomitant to DLBCL diagnosis in 30% of pts. At DLBCL diagnosis median CD4+ cell count was 188/microL (range: 8-1172) and 34/66 pts (52%) had detectable HIV viral load. COO assigned by IHC was GCB in 31/66 pts (47%) and non-GCB in 35/66 (53%); COO allocation by NS assay was 57% GCB (n=34/60 pts) and 43% non-GCB (ABC n=11, 18%; UC n=15, 25%). IHC algorithm and NS assay concordantly assigned COO subtypes in 80% of pts with a Cohen's kappa=0,604 (p<0.0005). Twenty pts (30%) had MYC and 23 pts (33%) had BCL-2 overexpression, respectively. An anthracycline-containing first-line chemotherapy was delivered to 97% of pts, with the addition of Rituximab in 86% of cases. Two pts (3%) could receive only palliative treatment. Two-years progression-free survival (PFS) and overall survival (OS) of the entire series were 61% and 56%, respectively, after a median follow-up of 50 months (range 4-184). No difference was found in OS and PFS according to COO subtypes, either determined by IHC or NS. Notably, bcl-2 overexpression had a negative impact on OS (2-years OS 44% vs 70%, p=0.018, HR: 2,34) and PFS (2-years PFS 45% vs 62%, p= 0,12, HR: 1,71), while MYC expression had no significant correlation with survival. A baseline CD4+ count > 200/microL has a protective role both for progression and death (2-years PFS: 62% vs 42%, p=0.02, HR: 0.41; 2-years OS: 72% vs 42%, p=0.002, HR: 0.30). Stage IV disease and elevated baseline LDH were associated with inferior PFS (p=0.0158 and p=0.02) and OS (p=0.0076 and p=0.034).
Conclusions: This analysis of 66 HIV-associated DLBCL confirmed that GCB is the more frequent subtype identified by NS, as previously reported (Baptista MJ et al, Hemat Oncol 2017). We found an high concordance between Hans algorithm and NS in the COO subtypes allocation. However, COO classification showed no impact on pts outcome. FISH assays for detection of MYC, BCL-2 and BCL-6 rearrangements are ongoing in the entire study population.
Rusconi:Celgene: Research Funding.
Plasmacytoid dendritic cells (PDC) belong to a subtype of dendritic cells that are normally absent in healthy skin. In some inflammatory diseases of the skin, especially lupus erythematosus (LE), ...these cells are occasionally recruited in great amounts, which can be used as a helpful clue for diagnosis. Rarely, PDC may also accumulate in the skin of patients with myeloid leukemia, a yet poorly known condition currently called ‘tumor‐forming PDC associated with myeloid neoplasms’. In this study, we describe a patient with unsuspected chronic myelomonocytic leukemia who developed cutaneous lesions characterized by a dermal infiltrate rich in PDC. Similarly to LE, such neoplastic PDC were accompanied by interface dermatitis‐like changes, but displayed an aberrant phenotype and shared the same chromosomal abnormality with the leukemic cells identified in the bone marrow, thus revealing the neoplastic nature of the process. This observation illustrates that tumor‐forming PDC associated with myeloid neoplasms may microscopically mimic LE in some patients. Accordingly, a hematologic workup is recommended in any skin lesion featuring excessive numbers of PDC, even if morphological alterations suggestive of interface dermatitis are found.
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