Non-Hodgkin and Hodgkin Lymphomas Select for Overexpression of BCLW.

Purpose: B-cell lymphomas must acquire resistance to apoptosis during their development. We recently discovered BCLW, an antiapoptotic BCL2 family member thought only to contribute to spermatogenesis, was overexpressed in diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma. To gain insight into the contribution of BCLW to B-cell lymphomas and its potential to confer resistance to BCL2 inhibitors, we investigated the expression of BCLW and the other antiapoptotic BCL2 family members in six different B-cell lymphomas. Experimental Design: We performed a large-scale gene expression analysis of datasets comprising approximately 2,300 lymphoma patient samples, including non-Hodgkin and Hodgkin lymphomas as well as indolent and aggressive lymphomas. Data were validated experimentally with qRT-PCR and IHC. Results: We report BCLW is significantly overexpressed in aggressive and indolent lymphomas, including DLBCL, Burkitt, follicular, mantle cell, marginal zone, and Hodgkin lymphomas. Notably, BCLW was preferentially overexpressed over that of BCL2 and negatively correlated with BCL2 in specific lymphomas. Unexpectedly, BCLW was overexpressed as frequently as BCL2 in follicular lymphoma. Evaluation of all five antiapoptotic BCL2 family members in six types of B-cell lymphoma revealed that BCL2, BCLW, and BCLX were consistently overexpressed, whereas MCL1 and A1 were not. In addition, individual lymphomas frequently overexpressed more than one antiapoptotic BCL2 family member. Conclusions: Our comprehensive analysis indicates B-cell lymphomas commonly select for BCLW overexpression in combination with or instead of other antiapoptotic BCL2 family members. Our results suggest BCLW may be equally as important in lymphomagenesis as BCL2 and that targeting BCLW in lymphomas should be considered. ©2017 AACR

Bcl2 is an independent prognostic marker of triple negative breast cancer (TNBC) and predicts response to anthracycline combination (ATC) chemotherapy (CT) in adjuvant and neoadjuvant settings

Background: TNBC represents a heterogeneous subgroup of BC with poor prognosis and frequently resistant to CT. Material and methods: The relationship between Bcl2 immunohistochemical protein expression and clinicopathological outcomes was assessed in 736 TNBC-patients: 635 patients had early primary-TNBC (EP-TNBC) and 101 had primary locally advanced (PLA)-TNBC treated with neo-adjuvant- ATC-CT. Results: Negative Bcl2 (Bcl2-) was observed in 70% of EP-TNBC and was significantly associated with high proliferation, high levels of P-Cadherin, E-Cadherin and HER3 (P’s<0.01), while Bcl2+ was significantly associated with high levels of p27, MDM4 and SPAG5 (P<0.01). After controlling for chemotherapy and other prognostic factors, Bcl2- was associated with 2-fold increased risk of death (P=0.006) and recurrence (P=0.0004). Furthermore, the prognosis of EP-TNBC/Bcl2- patients had improved both BC-specific survival (P=0.002) and disease-free survival (P = 0.003), if they received adjuvant-ATC-CT. Moreover, Bcl2- expression was an independent predictor of pathological complete response of primary locally advanced triple negative breast cancer (PLA-TNBC) treated with neoadjuvant-ATC-CT (P=0.008). Conclusion: Adding Bcl2 to the panel of markers used in current clinical practice could provide both prognostic and predictive information in TNBC. TNBC/Bcl2- patients appear to benefit from ATC-CT, whereas Bcl2+ TNBC seems to be resistant to ATC-CT and may benefit from a trial of different type of chemotherapy with/without novel-targeted agents. Key words: anthracyclin chemotherapy, Bcl2, predictive marker, prognostic marker, theraputic targets, triple negative breast cancer

Is now the time for molecular driven therapy for diffuse large B-cell lymphoma?

INTRODUCTION: Recent genetic and molecular discoveries regarding alterations in diffuse large B-cell lymphoma (DLBCL) deeply changed the approach to this lymphoproliferative disorder. Novel additional predictors of outcomes and new therapeutic strategies are being introduced to improve outcomes. Areas covered: This review aims to analyse the recent molecular discoveries in DLBCL, the rationale of novel molecular driven treatments and their impact on DLBCL prognosis, especially in ABC-DLBCL and High Grade B Cell Lymphoma. Pre-clinical and clinical evidences are reviewed to critically evaluate the novel DLBCL management strategies. Expert commentary: New insights in DLBCL molecular characteristics should guide the therapeutic approach; the results of the current studies which are investigating safety and efficacy of novel 'X-RCHOP' will probably lead, in future, to a cell of origin (COO) based upfront therapy. Moreover, it is necessary to identify early patients with DLBCL who carried MYC, BCL2 and/or BCL6 rearrangements double hit lymphomas (DHL) because they should not receive standard R-CHOP but high intensity treatment as reported in many retrospective studies. New prospective trials are needed to investigate the more appropriate treatment of DHL

Overexpression of microRNA-16 declines cellular growth, proliferation and induces apoptosis in human breast cancer cells

MicroRNAs (miRNA) are a large family of small single-stranded RNA molecules found in all multicellular organisms. Early studies have been shown that miRNA are involved in cancer development and progression, and this role can be done by working as an oncogenes and tumor suppressor genes, so manipulation of this molecules can be a promising approach in cancer therapy, and experimental results represented that the modification in breast cancer phenotype is possible by miRNA expression alteration. miR-16, which is located in 13q14 chromosome, plays critical roles as a tumor suppressor by targeting several oncogenes which regulate cell cycle and apoptosis. Hence, in the present study, we investigated whether miR-16 could decline growth and survival of MCF-7 cell line as model of human breast cancer. MCF-7 cell line was infected with lentiviruses containing miR-16 precursor sequence. The effects of ectopic expression of miR-16 on breast cancer phenotype were examined by cell cycle analysis and apoptosis assays. miR-16 cytotoxicity effect was measured by the MTT assay. We showed that the miR-16 overexpression reduces Cyclin D1 and BCL2 at messenger RNA (mRNA) and protein levels in MCF-7 cell line. In addition, this is found that enforced expression of miR-16 decreases cell growth and proliferation and induces apoptosis in MCF-7 cells. In conclusion, our results revealed that upregulation of miR-16 would be a potential approach for breast cancer therapy. © 2015, The Society for In Vitro Biology

The RNA-binding protein LARP1 is a post-transcriptional regulator of survival and tumorigenesis in ovarian cancer

RNA-binding proteins (RBPs) are increasingly identified as post-transcriptional drivers of cancer progression. The RBP LARP1 is an mRNA stability regulator, and elevated expression of the protein in hepatocellular and lung cancers is correlated with adverse prognosis. LARP1 associates with an mRNA interactome that is enriched for oncogenic transcripts. Here we explore the role of LARP1 in epithelial ovarian cancer, a disease characterized by the rapid acquisition of resistance to chemotherapy through the induction of pro-survival signalling. We show, using ovarian cell lines and xenografts, that LARP1 is required for cancer cell survival and chemotherapy resistance. LARP1 promotes tumour formation in vivo and maintains cancer stem cell-like populations. Using transcriptomic analysis following LARP1 knockdown, cross-referenced against the LARP1 interactome, we identify BCL2 and BIK as LARP1 mRNA targets. We demonstrate that, through an interaction with the 3 untranslated regions (3 UTRs) of BCL2 and BIK, LARP1 stabilizes BCL2 but destabilizes BIK with the net effect of resisting apoptosis. Together, our data indicate that by differentially regulating the stability of a selection of mRNAs, LARP1 promotes ovarian cancer progression and chemotherapy resistance

JMJD3 promotes survival of diffuse large B-cell lymphoma subtypes via distinct mechanisms.

JMJD3 (Jumonji domain containing-3), a histone H3 Lys27 (H3K27) demethylase, has been reported to be involved in the antigen-driven differentiation of germinal center B-cells. However, insight into the mechanism of JMJD3 in DLBCL (Diffuse large B-cell lymphoma) progression remains poorly understood. In this study, we investigated the subtype-specific JMJD3-dependent survival effects in DLBCL. Our data showed that in the ABC subtype, silencing-down of JMJD3 inhibited interferon regulatory factor 4 (IRF4) expression in a demethylase activity-dependent fashion. IRF4 reciprocally stimulated expression of JMJD3, forming a positive feedback loop that promoted survival in these cells. Accordingly, IRF4 expression was sufficient to rescue the pro-apoptotic effect of JMJD3 suppression in the ABC, but not in the GCB subtype. In contrast, ectopic overexpression of BCL-2 completely offset JMJD3-mediated survival in the GCB DLBCL cells. In vivo, treatment with siRNA to JMJD3 reduced tumor volume concordant with increased apoptosis in either subtype. This suggests it is a common target, though the distinctive signaling axes regulating DCBCL survival offer different strategic options for treating DLBCL subtypes

The kinesin Eg5 inhibitor K858 induces apoptosis but also survivin-related chemoresistance in breast cancer cells

Inhibitors of kinesin spindle protein Eg5 are characterized by pronounced antitumor activity. Our group has recently synthesized and screened a library of 1,3,4-thiadiazoline analogues with the pharmacophoric structure of K858, an Eg5 inhibitor. We herein report the effects of K858 on four different breast cancer cell lines: MCF7 (luminal A), BT474 (luminal B), SKBR3 (HER2 like) and MDA-MB231 (basal like). We demonstrated that K858 displayed anti-proliferative activity on every analyzed breast cancer cell line by inducing apoptosis. However, at the same time, we showed that K858 up-regulated survivin, an anti-apoptotic molecule. We then performed a negative regulation of survivin expression, with the utilization of wortmannin, an AKT inhibitor, and obtained a significant increase of K858-dependent apoptosis. These data demonstrate that K858 is a potent inhibitor of replication and induces apoptosis in breast tumor cells, independently from the tumor phenotype. This anti-proliferative response of tumor cells to K858 can be limited by the contemporaneous over-expression of survivin; consequently, the reduction of survivin levels, obtained with AKT inhibitors, can sensitize tumor cells to K858-induced apoptosis

Protein kinase CK2 is widely expressed in follicular, Burkitt and diffuse large B-cell lymphomas and propels malignant B-cell growth.

Serine-threonine kinase CK2 is highly expressed and pivotal for survival and proliferation in multiple myeloma, chronic lymphocytic leukemia and mantle cell lymphoma. Here, we investigated the expression of \u3b1 catalytic and \u3b2 regulatory CK2 subunits by immunohistochemistry in 57 follicular (FL), 18 Burkitt (BL), 52 diffuse large B-cell (DLBCL) non-Hodgkin lymphomas (NHL) and in normal reactive follicles. In silico evaluation of available Gene Expression Profile (GEP) data sets from patients and Western blot (WB) analysis in NHL cell-lines were also performed. Moreover, the novel, clinical-grade, ATP-competitive CK2-inhibitor CX-4945 (Silmitasertib) was assayed on lymphoma cells. CK2 was detected in 98.4% of cases with a trend towards a stronger CK2\u3b1 immunostain in BL compared to FL and DLBCL. No significant differences were observed between Germinal Center B (GCB) and non-GCB DLBCL types. GEP data and WB confirmed elevated CK2 mRNA and protein levels as well as active phosphorylation of specific targets in NHL cells. CX-4945 caused a dose-dependent growth-arresting effect on GCB, non-GCB DLBCL and BL cell-lines and it efficiently shut off phosphorylation of NF-\u3baB RelA and CDC37 on CK2 target sites. Thus, CK2 is highly expressed and could represent a suitable therapeutic target in BL, FL and DLBCL NHL

Targeting translation initiation by synthetic rocaglates for treating MYC-driven lymphomas.

MYC-driven lymphomas, especially those with concurrent MYC and BCL2 dysregulation, are currently a challenge in clinical practice due to rapid disease progression, resistance to standard chemotherapy, and high risk of refractory disease. MYC plays a central role by coordinating hyperactive protein synthesis with upregulated transcription in order to support rapid proliferation of tumor cells. Translation initiation inhibitor rocaglates have been identified as the most potent drugs in MYC-driven lymphomas as they efficiently inhibit MYC expression and tumor cell viability. We found that this class of compounds can overcome eIF4A abundance by stabilizing target mRNA-eIF4A interaction that directly prevents translation. Proteome-wide quantification demonstrated selective repression of multiple critical oncoproteins in addition to MYC in B-cell lymphoma including NEK2, MCL1, AURKA, PLK1, and several transcription factors that are generally considered undruggable. Finally, (-)-SDS-1-021, the most promising synthetic rocaglate, was confirmed to be highly potent as a single agent, and displayed significant synergy with the BCL2 inhibitor ABT199 in inhibiting tumor growth and survival in primary lymphoma cells in vitro and in patient-derived xenograft mouse models. Overall, our findings support the strategy of using rocaglates to target oncoprotein synthesis in MYC-driven lymphomas.P30 CA036727 - NCI NIH HHS; R24 GM111625 - NIGMS NIH HHS; R35 GM118173 - NIGMS NIH HHS; LB506 - Nebraska Department of Health and Human Services (Nebraska DHHS)Accepted manuscriptSupporting documentatio

Notch/Delta signaling constrains reengineering of pro-T cells by PU.1

PU.1 is essential for early stages of mouse T cell development but antagonizes it if expressed constitutively. Two separable mechanisms are involved: attenuation and diversion. Dysregulated PU.1 expression inhibits pro-T cell survival, proliferation, and passage through β-selection by blocking essential T cell transcription factors, signaling molecules, and Rag gene expression, which expression of a rearranged T cell antigen receptor transgene cannot rescue. However, Bcl2 transgenic cells are protected from this attenuation and may even undergo β-selection, as shown by PU.1 transduction of defined subsets of Bcl2 transgenic fetal thymocytes with differentiation in OP9-DL1 and OP9 control cultures. The outcome of PU.1 expression in these cells depends on Notch/Delta signaling. PU.1 can efficiently divert thymocytes toward a myeloid-like state with multigene regulatory changes, but Notch/Delta signaling vetoes diversion. Gene expression analysis distinguishes sets of critical T lineage regulatory genes with different combinatorial responses to PU.1 and Notch/Delta signals, suggesting particular importance for inhibition of E proteins, Myb, and/or Gfi1 (growth factor independence 1) in diversion. However, Notch signaling only protects against diversion of cells that have undergone T lineage specification after Thy-1 and CD25 up-regulation. The results imply that in T cell precursors, Notch/Delta signaling normally acts to modulate and channel PU.1 transcriptional activities during the stages from T lineage specification until commitment