RESUMEN
B-cell receptor (BCR) signals play a critical role in the pathogenesis of chronic lymphocytic leukemia (CLL), but their role in regulating CLL cell proliferation has still not been firmly established. Unlike normal B cells, CLL cells do not proliferate in vitro upon engagement of the BCR, suggesting that CLL cell proliferation is regulated by other signals from the microenvironment, such as those provided by Toll-like receptors or T cells. Here, we report that BCR engagement of human and murine CLL cells induces several positive regulators of the cell cycle, but simultaneously induces the negative regulators CDKN1A, CDKN2A, and CDKN2B, which block cell-cycle progression. We further show that introduction of genetic lesions that downregulate these cell-cycle inhibitors, such as inactivating lesions in CDKN2A, CDKN2B, and the CDKN1A regulator TP53, leads to more aggressive disease in a murine in vivo CLL model and spontaneous proliferation in vitro that is BCR dependent but independent of costimulatory signals. Importantly, inactivating lesions in CDKN2A, CDKN2B, and TP53 frequently co-occur in Richter syndrome (RS), and BCR stimulation of human RS cells with such lesions is sufficient to induce proliferation. We also show that tumor cells with combined TP53 and CDKN2A/2B abnormalities remain sensitive to BCR-inhibitor treatment and are synergistically sensitive to the combination of a BCR and cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitor both in vitro and in vivo. These data provide evidence that BCR signals are directly involved in driving CLL cell proliferation and reveal a novel mechanism of Richter transformation.
Asunto(s)
Transformación Celular Neoplásica , Inhibidor p15 de las Quinasas Dependientes de la Ciclina , Inhibidor p16 de la Quinasa Dependiente de Ciclina , Leucemia Linfocítica Crónica de Células B , Receptores de Antígenos de Linfocitos B , Transducción de Señal , Proteína p53 Supresora de Tumor , Animales , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/inmunología , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/inmunología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/inmunología , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/inmunología , Ratones , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/inmunología , Transducción de Señal/genética , Transducción de Señal/inmunología , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/inmunologíaRESUMEN
The BCL-2 inhibitor venetoclax has only limited activity in DLBCL despite frequent BCL-2 overexpression. Since constitutive activation of the B cell receptor (BCR) pathway has been reported in both ABC and GCB DLBCL, we investigated whether targeting SYK or BTK will increase sensitivity of DLBCL cells to venetoclax. We report that pharmacological inhibition of SYK or BTK synergistically enhances venetoclax sensitivity in BCL-2-positive DLBCL cell lines with an activated BCR pathway in vitro and in a xenograft model in vivo, despite the only modest direct cytotoxic effect. We further show that these sensitizing effects are associated with inhibition of the downstream PI3K/AKT pathway and changes in the expression of MCL-1, BIM, and HRK. In addition, we show that BCR-dependent GCB DLBCL cells are characterized by deficiency of the phosphatase SHP1, a key negative regulator of the BCR pathway. Re-expression of SHP1 in GCB DBLCL cells reduces SYK, BLNK, and GSK3 phosphorylation and induces corresponding changes in MCL1, BIM, and HRK expression. Together, these findings suggest that SHP1 deficiency is responsible for the constitutive activation of the BCR pathway in GCB DLBCL and identify SHP1 and BCL-2 as potential predictive markers for response to treatment with a venetoclax/BCR inhibitor combination.