Background:Up to 10% of patients with chronic lymphocytic leukemia (CLL) undergo a high-grade transformation called Richter Syndrome (RS), most commonly of the diffuse large B cell lymphoma type. In ...~80% of cases, the lymphoma originates from the CLL clone and is associated with a median survival inferior to 1 year after diagnosis. Prognosis following standard immunochemotherapy remains very poor and new therapeutic targets for RS are urgently needed.
Methods:Only samples from patients with newly diagnosed RS recruited into the UK CHOP-OR (CHOP in combination with Ofatumumab, EudraCT number 2009-016459-23) were included (Eyre et al, Br J Haematol. 2016). DNA was extracted from 35 peripheral blood mononuclear cells (leukemic phase) and 21 FFPE-lymph node slides (Richter phase). For 15 patients, paired CLL and RS phase were available. A TruSeq Custom Amplicon panel (TSCA, Illumina) was designed targeting 28 recurrently mutated genes in CLL based on recent publications. Average sequencing depth was 2311x. Single nucleotide variant (SNV) were filtered with a variant allele frequency (VAF) >10%, with a depth >20, with an altered variant read >6 and by excluding synonymous mutation and mutation with a frequency >5% in the general population. Mutation frequencies were compared to those of Landau et al based on a CLL population of 538 patients.
Results: Pathogenic mutations were identified in 28/30 CLL phase (93%) and 18/18 (100%) of RS phase. In the majority of paired cases (80%), mutations identified in the CLL phase were confirmed in the RS phase. Additional mutations were acquired in the RS phase compared to the CLL phase (277 versus 82) with a median of 5.5 mutations/RS phase versus 3 mutations/CLL phase (P=0.0003). When compared to the frequency in a general CLL population, we observed a significant increase of mutations in ATM (15%, 57%, 78% for general CLL population, CLL phase of RS, RS phase, respectively), TP53 (7%, 23%, 56%), NOTCH1 (6%, 23%, 50%), CDKN2A (<1%, 20%, 50%), SAMHD1 (2%, 13%, 33%), XPO1 (4%, 7%, 28%) and IRF4 (2%, 3%, 28%). IgHV analysis showed that all CLL/Richter samples were clonally related. The most frequently acquired mutations in the RS phase were TP53 and CDKN2A followed by SAMHD1, XPO1 and MED12. Central review of biopsy histology by two independent pathologists showed that 5 patients did not fulfill WHO diagnostic criteria of RS (Soilleux et al, Histopathology 2016) and were excluded from further analyses. However, mutation analysis of these biopsies revealed mutations associated with chemo-refractory CLL (3/5 TP53, 1/5 SAMHD1, 1/5 POT1) and clinical outcome of these patients was similar to patients with classical RS. Finally, we correlated the presence of mutations with progression-free (PFS) and overall (OS) survival (n=28). The median PFS was 5.8 months while OS after randomization was 11.5 months. None of the mutations alone was able to significantly stratify patient in terms of PFS and OS. Therefore, we combined the presence of high-risk mutations in any of the following 6 genes in the leukemic phase (NOTCH1, SF3B1, SAMHD1, DDX3X, FBXW7, KLHL6): patient with no mutation in these genes present a median PFS of 14.9 months compared to patients with at least one mutated gene who had a median PFS of 5.3 month (P=0.0022). Similar results were observed for OS (median OS >23.1 months vs 6.6 months, P=0.0242).
Conclusion: We show that CLL patients developing RS carry a high mutation burden and recurrent driver mutations in the leukemic clone and that additional mutations are acquired in the RS phase. We confirm the association of mutations in TP53, NOTCH1 and CDKN2A with RS and show in addition that SAMHD1 mutations are also frequently seen in RS. Finally, we propose that patients with biopsy findings of classical RS but no mutation in any of the 6 selected genes have an improved PFS and OS whereas patients without the classical RS biopsy features but a high-risk mutation profile fare poorly.
Eyre:GSK: Honoraria; Celgene: Other: Travel, Accomodation; Gilead: Honoraria, Other: Travel, Accomodation, Speakers Bureau; Takeda: Honoraria, Other: Travel, Speakers Bureau. Schuh:Gilead: Consultancy, Honoraria, Research Funding; Roche, Janssen, Novartis, Celgene, Abbvie: Consultancy, Honoraria.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
We studied the involvement of NOV/CCN3, whose function is poorly understood, in chondrocyte differentiation. NOV was found to upregulate TGF‐β2 and type X collagen and to act as a downstream effector ...of TGF‐β1 in ATDC5 and primary chondrocytes. Thus, NOV is a positive modulator of chondrogenesis.
Introduction: NOV/CCN3 is a matricellular protein that belongs to the CCN family. A growing body of evidence indicates that NOV could play a role in cell differentiation, particularly in chondrogenesis. During chick embryo development, NOV expression is tightly regulated in cartilage, and a high expression of NOV has been associated with cartilage differentiation in Wilms' tumors. However, a precise role for NOV and potential target genes of NOV in chondrogenesis are unknown.
Materials and Methods: ATDC5 cells and primary chondrocytes were either treated with NOV recombinant protein or transfected with a NOV‐specific siRNA to determine, using quantitative RT‐PCR, the effect of NOV on the expression of several molecules involved in chondrocyte differentiation. Stable ATDC5 clones expressing NOV were also established to show that NOV was a downstream effector of TGF‐β1.
Results: We established that NOV/CCN3 expression increases in ATDC5 cells at early stages of chondrogenic differentiation and precedes the appearance of TGF‐β2 and of several chondrocytic markers such as SOX9 or type X collagen. When exogenously administered, NOV recombinant protein up‐regulates TGF‐β2 and type X collagen mRNA levels both in ATDC5 cells and in primary mouse chondrocytes but does not influence SOX9 expression. This regulation also occurs at the endogenous level because downregulation of NOV expression is correlated with an inhibition of TGF‐β2 and type X collagen in primary chondrocytes. Furthermore, we found that NOV expression is downregulated when chondrocytes are exposed to TGF‐β1‐dedifferentiating treatment in chondrocytes, further providing evidence that NOV may counteract TGF‐β1 effects on chondrocytes.
Conclusions: This study provides the first characterization of two new targets of NOV involved in chondrocyte differentiation, shows that NOV acts with TGF‐β1 in a cascade of gene regulation, and indicates that NOV is a positive modulator of chondrogenesis.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Background & Aims Guidelines recommend 6 months of alcohol abstinence before treating hepatitis C (HCV). Abstinence is difficult for alcohol-dependent patients to achieve. This study evaluated HCV ...treatment in alcoholic patients with ongoing consumption or less than 6 months of abstinence. Methods A multidisciplinary management model was built by a liver unit and two centers involved in the care of addict patients. Patients were included in a prospective observational study of treatment with pegylated interferon and ribavirin if they presented alcohol dependence with ongoing intoxication or abstinence of less than 6 months. Pre-therapeutic evaluation and follow-up were multidisciplinary, and addiction care was personalized to patient condition and willingness. Alcohol abstinence or reduction was encouraged but not mandatory. The primary end point was sustained virological response (SVR). Results were compared to a control group of patients matched for genotype, viral load, fibrosis stage, sex, and age. Results A total of 73 patients treated between 2002 and 2008 were included in the study. Intent to treat analysis showed an SVR in 48% (35/73) of patients versus 49% (36/73) of controls. Low viral load and length of abstinence during treatment were independently associated with SVR. During treatment, 20 (27%) patients were abstinent, 23 (32%) had controlled consumption, and 24 (33%) had excessive consumption. At the end of the follow-up, 22 (30%) patients were durably abstinent. Conclusions A multidisciplinary approach allowed HCV treatment in alcohol-dependent patients with a satisfactory SVR rate and positive effects on addiction behavior.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
We studied the involvement of NOV/CCN3, whose function is poorly understood, in chondrocyte differentiation. NOV was found to upregulate TGF-beta2 and type X collagen and to act as a downstream ...effector of TGF-beta1 in ATDC5 and primary chondrocytes. Thus, NOV is a positive modulator of chondrogenesis.
NOV/CCN3 is a matricellular protein that belongs to the CCN family. A growing body of evidence indicates that NOV could play a role in cell differentiation, particularly in chondrogenesis. During chick embryo development, NOV expression is tightly regulated in cartilage, and a high expression of NOV has been associated with cartilage differentiation in Wilms' tumors. However, a precise role for NOV and potential target genes of NOV in chondrogenesis are unknown.
ATDC5 cells and primary chondrocytes were either treated with NOV recombinant protein or transfected with a NOV-specific siRNA to determine, using quantitative RT-PCR, the effect of NOV on the expression of several molecules involved in chondrocyte differentiation. Stable ATDC5 clones expressing NOV were also established to show that NOV was a downstream effector of TGF-beta1.
We established that NOV/CCN3 expression increases in ATDC5 cells at early stages of chondrogenic differentiation and precedes the appearance of TGF-beta2 and of several chondrocytic markers such as SOX9 or type X collagen. When exogenously administered, NOV recombinant protein up-regulates TGF-beta2 and type X collagen mRNA levels both in ATDC5 cells and in primary mouse chondrocytes but does not influence SOX9 expression. This regulation also occurs at the endogenous level because downregulation of NOV expression is correlated with an inhibition of TGF-beta2 and type X collagen in primary chondrocytes. Furthermore, we found that NOV expression is downregulated when chondrocytes are exposed to TGF-beta1-dedifferentiating treatment in chondrocytes, further providing evidence that NOV may counteract TGF-beta1 effects on chondrocytes.
This study provides the first characterization of two new targets of NOV involved in chondrocyte differentiation, shows that NOV acts with TGF-beta1 in a cascade of gene regulation, and indicates that NOV is a positive modulator of chondrogenesis.
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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