Tamara Bittolo

Despite the development of robust prognostic biomarkers for risk stratification in newly diagnosed diffuse large B-cell lymphoma (DLBCL), namely international prognostic index (IPI), cell-of-origin (COO), circulating tumor DNA (ctDNA) and molecular clusters, none of these capture potential host-related features. In this study, untargeted plasma metabolomics was performed in a discovery cohort of 96 newly diagnosed R-CHOP-treated DLBCL patients. The key finding was validated in an independent cohort of 42 DLBCL with comparable clinical characteristics and outcomes. None of the 280 identified plasma metabolites was associated with COO classification subtypes. Conversely, the metabolomic profile appeared to show variations across ctDNA-defined molecular clusters. After an elastic net regression-based survival analysis, 2-hydroxybutyric acid (2HB) was selected as the only endogenous metabolite to be associated with worse progression-free survival (PFS) in the discovery cohort. The impact of 2HB levels on PFS was confirmed in the validation cohort. Also, 2HB levels (HR 2.09, 95% CI 1.04-4.19, p=0.04) maintained an independent association with shorter PFS when adjusted for higher ctDNA levels (HR 2.08, 95% CI 1.06-4.08, p=0.03). Therefore, 2HB and ctDNA levels were included in a two-factor prognostic score based entirely on minimally invasive plasma-derived biomarkers. This hypothesis-generating score improved outcome prediction compared with ctDNA assessment alone and enabled the identification of high-risk patients who might benefit from early restaging and potential treatment intensification (24-month PFS 28.8%). In conclusion, plasma metabolomics revealed a prognostically relevant role and 2HB sorted out as a potential host-related biomarker able to refine risk stratification in newly diagnosed DLBCL.

B-cell receptor (BCR) signaling is a key determinant of chronic lymphocytic leukemia (CLL) pathophysiology. CD49d, the α4 subunit of the very late antigen 4 integrin, can be activated by BCR signals; however, its role in modulating BCR functionality remains unknown. We used knockout mouse models and primary human CLL stratified by CD49d expression to address this aspect. CD49d was required for bone marrow (BM) infiltration and shaped BM infiltration patterns and patient outcomes in human CLL. In TCL1 transplantation models, the loss of CD49d abrogated BM homing and leukemic cell positioning within splenic niches. At the cellular level, CD49d-deficient murine TCL1 transgenic cells and human CD49d-low CLL cells failed to form efficient immune synapses with antigen-presenting membranes. Transcriptome analyses identified CD49d-dependent regulation of actin-associated pathways and distinct signatures of BCR responsiveness in both human and mouse cells. Consistently, CD49d-low human CLL cells displayed aberrant actin remodeling following BCR stimulation, and a second aggressive murine CLL model reproduced the actin and engraftment defects. Kinome profiling linked impaired antigen-induced BCR responses in CD49d-deficient murine cells to altered kinase activity, and pharmacologic actin perturbation phenocopied CD49d loss. In human CD49d-low CLL cells, a desynchronization of BCR-related downstream Syk and PLCɣ2 activation was found. Mechanistically, the CD49d-BCR interplay involved their colocalization, and CD49d converged with BCR signaling on a focal adhesion kinase–actin axis. In summary, our findings establish CD49d as a key regulator of BCR functionality in CLL, linking integrins to cytoskeletal dynamics and antigen responsiveness.

BTK inhibitors deeply affect the proliferative capacity, homing and migrations of chronic lymphocytic leukemia (CLL) cells, and cause their redistribution from

In chronic lymphocytic leukemia (CLL), stabilizing mutations of NOTCH1, affecting up to 10–15% of cases, have been associated to poor prognosis, disease progression and refractoriness to chemotherapy. NOTCH1 mutations are significantly overrepresented in trisomy 12 CLL, a disease subset frequently expressing CD49d, the α4 chain of the very-late-activation-4 integrin, a well-known key regulator of microenviromental interactions, and negative prognosticator in CLL. In the present study, by analysing a wide cohort of 1180 CLL, we observed a very strong association between the presence of NOTCH1 mutations and the expression of CD49d (PNOTCH1 gene configuration, we provide evidence that triggering of the NOTCH1 pathway resulted in a positive CD49d expression regulation, which was driven by a NOTCH1-dependent activation of nuclear factot-κB (NF-κB). Consistently, pharmacological inhibition of the NOTCH1 and/or of the NF-κB pathways resulted in impaired NF-κB nuclear translocation with consequent down-modulation of CD49d expression. Altogether, our data link for the first time NOTCH1 mutations to CD49d expression regulation through the involvement of the NF-κB pathway in CLL.

Trisomy 12 is a recurrent cytogenetic abnormality that occurs in 15–20% of Chronic Lymphocytic Leukemia (CLL).[1][1],[2][2] Within the hierarchical model proposed by Döhner et al .,[3][3] trisomy 12 CLL (tris12 CLL) carry an intermediate prognostic risk, with median overall survival (OS) and time

In chronic lymphocytic leukemia (CLL), mutations in the NOTCH1 coding region (coding NOTCH1 mutations) have been associated with impaired degradation of NOTCH1 protein[1][1],[2][2] and, clinically, with shorter time to first treatment, shorter overall survival,[1][1]–[3][3] and resistance to anti-

In chronic lymphocytic leukemia (CLL), the mechanisms controlling cell growth and proliferation in the presence of NOTCH1 mutations remain largely unexplored. By performing a gene expression profile of NOTCH1-mutated (NOTCH1-mut) versus NOTCH1 wild-type CLL, we identified a gene signature of NOTCH1-mut CLL characterized by the upregulation of genes related to ribosome biogenesis, such as nucleophosmin 1 (NPM1) and ribosomal proteins (RNPs). Activation of NOTCH1 signaling by ethylenediaminetetraacetic acid or by coculture with JAGGED1-expressing stromal cells increased NPM1 expression, and inhibition of NOTCH1 signaling by either NOTCH1-specific small interfering RNA (siRNA) or γ-secretase inhibitor reduced NPM1 expression. Bioinformatic analyses and in vitro activation/inhibition of NOTCH1 signaling suggested a role of MYC as a mediator of NOTCH1 effects over NPM1 and RNP expression in NOTCH1-mut CLL. Chromatin immunoprecipitation experiments performed on NOTCH1 intracellular domain (NICD)-transfected CLL-like cells showed the direct binding of NOTCH1 to the MYC promoter, and transfection with MYC-specific siRNA reduced NPM1 expression. In turn, NPM1 determined a proliferation advantage of CLL-like cells, as demonstrated by NPM1-specific siRNA transfection. In conclusion, NOTCH1 mutations in CLL are associated with the overexpression of MYC and MYC-related genes involved in protein biosynthesis including NPM1, which are allegedly responsible for cell growth and/or proliferation advantages of NOTCH1-mut CLL.