Chronic lymphocytic leukemia (CLL) is a B lymphoid malignancy highly dependent on the microenvironment. Despite new targeted therapies such as ibrutinib and venetoclax, disease progression and ...relapse remain an issue. CLL cell interactions with the supportive tissue microenvironment play a critical role in disease pathogenesis. We used a platform for drug discovery based on systems biology and artificial intelligence, to identify drugs targeting key proteins described to have a role in the microenvironment. The selected compounds were screened in CLL cell lines in the presence of stromal cells to mimic the microenvironment and validated the best candidates in primary CLL cells. Our results showed that the commercial drug simvastatin was the most effective and selective out of the tested compounds. Simvastatin decreased CLL cell survival and proliferation as well as cell adhesion. Importantly, this drug enhanced the antitumor effect of venetoclax and ibrutinib. We proposed that systems biology approaches combined with pharmacological screening could help to find new drugs for CLL treatment and to predict new combinations with current therapies. Our results highlight the possibility of repurposing widely used drugs such as statins to target the microenvironment and to improve the efficacy of ibrutinib or venetoclax in CLL cells.
Animal bioprosthetic heart valves (BHV) are used to replace defective valves in patients with valvular heart disease. Especially young BHV recipients may experience a structural valve deterioration ...caused by an immune reaction in which alpha-Gal and Neu5Gc are potential target antigens. The expression of these and other carbohydrate antigens in animal tissues used for production of BHV was explored. Protein lysates of porcine aortic and pulmonary valves, and porcine, bovine and equine pericardia were analyzed by Western blotting using anti-carbohydrate antibodies and lectins.N-glycans were released by PNGase F digestion andO-glycans by beta-elimination. Released oligosaccharides were analyzed by liquid chromatography - tandem mass spectrometry. In total, 102N-glycans and 40O-glycans were identified in animal heart tissue lysates. TheN- andO-glycan patterns were different between species. alpha-Gal and Neu5Gc were identified on bothN- andO-linked glycans,N,N '-diacetyllactosamine (LacdiNAc) onN-glycans only and sulfatedO-glycans. The relative amounts of alpha-Gal-containingN-glycans were higher in bovine compared to equine and porcine pericardia. In contrast to the restricted number of proteins carrying alpha-Gal and LacdiNAc, the distribution of proteins carrying Neu5Gc-determinants varied between species and between different tissues of the same species. Porcine pericardium carried the highest level of Neu5Gc-sialylatedO-glycans, and bovine pericardium the highest level of Neu5Gc-sialylatedN-glycans. The identifiedN-andO-linked glycans, some of which may be immunogenic and remain in BHVs manufactured for clinical use, could direct future genetic engineering to prevent glycan expression rendering the donor tissues less immunogenic in humans.
Animal bioprosthetic heart valves (BHV) are used to replace defective valves in patients with valvular heart disease. Especially young BHV recipients may experience a structural valve deterioration ...caused by an immune reaction in which α-Gal and Neu5Gc are potential target antigens. The expression of these and other carbohydrate antigens in animal tissues used for production of BHV was explored. Protein lysates of porcine aortic and pulmonary valves, and porcine, bovine and equine pericardia were analyzed by Western blotting using anti-carbohydrate antibodies and lectins.
N
-glycans were released by PNGase F digestion and
O
-glycans by β-elimination. Released oligosaccharides were analyzed by liquid chromatography – tandem mass spectrometry. In total, 102
N
-glycans and 40
O
-glycans were identified in animal heart tissue lysates. The
N
- and
O
-glycan patterns were different between species. α-Gal and Neu5Gc were identified on both
N
- and
O
-linked glycans,
N,N´
-diacetyllactosamine (LacdiNAc) on
N
-glycans only and sulfated
O
-glycans. The relative amounts of α-Gal-containing
N
-glycans were higher in bovine compared to equine and porcine pericardia. In contrast to the restricted number of proteins carrying α-Gal and LacdiNAc, the distribution of proteins carrying Neu5Gc-determinants varied between species and between different tissues of the same species. Porcine pericardium carried the highest level of Neu5Gc-sialylated
O
-glycans, and bovine pericardium the highest level of Neu5Gc-sialylated
N
-glycans. The identified
N-
and
O-
linked glycans, some of which may be immunogenic and remain in BHVs manufactured for clinical use, could direct future genetic engineering to prevent glycan expression rendering the donor tissues less immunogenic in humans.
Splicing factor 3B subunit 1 (SF3B1) is involved in pre-mRNA branch site recognition and is the target of antitumor-splicing inhibitors. Mutations in
are observed in 15% of patients with chronic ...lymphocytic leukemia (CLL) and are associated with poor prognosis, but their pathogenic mechanisms remain poorly understood. Using deep RNA-sequencing data from 298 CLL tumor samples and isogenic
WT and K700E-mutated CLL cell lines, we characterize targets and pre-mRNA sequence features associated with the selection of cryptic 3' splice sites upon
mutation, including an event in the
gene relevant for activation of NF-κB signaling. Using the H3B-8800 splicing modulator, we show, for the first time in CLL, cytotoxic effects in vitro in primary CLL samples and in
-mutated isogenic CLL cell lines, accompanied by major splicing changes and delayed leukemic infiltration in a CLL xenotransplant mouse model. H3B-8800 displayed preferential lethality towards
-mutated cells and synergism with the BCL2 inhibitor venetoclax, supporting the potential use of SF3B1 inhibitors as a novel therapeutic strategy in CLL.
Chronic lymphocytic leukemia (CLL) is characterized by a high degree of genetic variability and interpatient heterogeneity. In the last decade, novel alterations have been described. Some of them ...impact on the prognosis and evolution of patients. The approval of BTK inhibitors, PI3K inhibitors and Bcl-2 inhibitors has drastically changed the treatment of patients with CLL. The effect of these new targeted therapies has been widely analyzed in
-mutated cases, but few data exist about the response of patients carrying other recurrent mutations. In this review, we describe the biological pathways recurrently altered in CLL that might have an impact on the response to these new therapies together with the possibility to use new actionable targets to optimize treatment responses.
Mantle cell lymphoma (MCL), a rare and aggressive B-cell non-Hodgkin lymphoma, mainly develops in the lymph node (LN) and creates a protective and immunosuppressive niche that facilitates tumor ...survival, proliferation and chemoresistance. To capture disease heterogeneity and tumor microenvironment (TME) cues, we have developed the first patient-derived MCL spheroids (MCL-PDLS) that recapitulate tumor oncogenic pathways and immune microenvironment in a multiplexed system that allows easy drug screening, including immunotherapies. MCL spheroids, integrated by tumor B cells, monocytes and autologous T-cells self-organize in disc-shaped structures, where B and T-cells maintain viability and proliferate, and monocytes differentiate into M2-like macrophages. RNA-seq analysis demonstrated that tumor cells recapitulate hallmarks of MCL-LN (proliferation, NF-kB and BCR), with T cells exhibiting an exhaustion profile (PD1, TIM-3 and TIGIT). MCL-PDLS reproduces in vivo responses to ibrutinib and demonstrates that combination of ibrutinib with nivolumab (anti-PD1) may be effective in ibrutinib-resistant cases by engaging an immune response with increased interferon gamma and granzyme B release. In conclusion, MCL-PDLS recapitulates specific MCL-LN features and in vivo responses to ibrutinib, representing a robust tool to study MCL interaction with the immune TME and to perform drug screening in a patient-derived system, advancing toward personalized therapeutic approaches.
Chronic lymphocytic leukemia (CLL) is a B-cell neoplasm with a heterogeneous clinical behavior. In 5-10% of patients the disease transforms into a diffuse large-B cell lymphoma known as Richter ...transformation (RT), which is associated with dismal prognosis. Here, we aimed to establish patient-derived xenograft (PDX) models to study the molecular features and evolution of CLL and RT. We generated two PDXs by injecting CLL (PDX12) and RT (PDX19) cells into immunocompromised NSG mice. Both PDXs were morphologically and phenotypically similar to RT. Whole-genome sequencing analysis at different time points of the PDX evolution revealed a genomic landscape similar to RT tumors from both patients and uncovered an unprecedented RT subclonal heterogeneity and clonal evolution during PDX generation. In PDX12, the transformed cells expanded from a very small subclone already present at the CLL stage. Transcriptomic analysis of PDXs showed a high oxidative phosphorylation (OXPHOS) and low B-cell receptor (BCR) signaling similar to the RT in the patients. IACS-010759, an OXPHOS inhibitor, reduced proliferation, and circumvented resistance to venetoclax. In summary, we have generated new RT-PDX models, one of them from CLL cells that mimicked the evolution of CLL to RT uncovering intrinsic features of RT cells of therapeutical value.
PDF
