Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) is an aberrant fusion gene product with tyrosine kinase activity and is expressed in substantial subset of anaplastic large cell lymphomas (ALCL). It has been shown that NPM-ALK binds to and activates signal transducer and activator of transcription 3 (STAT3). Although NPM-ALK+ ALCL overall shows a better prognosis, there is a sub-group of patients who relapses and is resistant to conventional chemotherapeutic regimens. NPM-ALK is a potential target for small molecule kinase inhibitors. Crizotinib (PF-2341066) is a small, orally bioavailable molecule that inhibits growth of tumors with ALK activity as shown in a subgroup of non-small lung cancer patients with EML4-ALK expression. In this study, we have investigated the in vitro effects of Crizotinib in ALCL cell line with NPM-ALK fusion. Crizotinib induced marked downregulation of STAT3 phosphorylation, which was associated with significant apoptotic cell death. Apoptosis induction was attributed to caspase-3 cleavage and marked downregulation of the Bcl-2 family of proteins including MCL-1. These findings implicate that Crizotinib has excellent potential to treat patients with NPM-ALK+ ALCL through induction of apoptotic cell death and downregulation of major oncogenic proteins in this aggressive lymphoma.
Background: Hematogones (HG) are benign precursor B-cells that are seen in increased numbers in the bone marrow during childhood, following chemotherapy or bone marrow transplant, in certain immune deficiencies, and in autoimmune disorders. Flow cytometry typically shows expression of TdT, CD10, CD19, variable CD20, dim CD22, CD34 (early HG), and dim CD45. As this phenotype is also seen in patients with B lymphoblastic leukemia (B-ALL), it causes a significant problem in distinguishing leukemic blasts from HG, particularly in a regenerating marrow. Furthermore, hematogones are usually more numerous at baseline in younger patient populations, the same age group with a relatively higher incidence of B-ALL. Despite guidance by earlier studies using the above markers for differentiating HG from B-ALL, these markers are not always sufficient and may hinder correct interpretation. Previous studies demonstrate CD58 is commonly expressed in B-ALL but not in hematogones; however, CD58 as a single marker is somewhat limited when expressed at lower levels. In order to improve diagnostic accuracy, we generated a five color antibody panel including CD10, CD19, CD45, CD38, and CD58 to assess the utility of a single tube panel in distinguishing B-ALL from HG. Design: A total of 35 cases with immature B-cell populations, 16 B-ALL (diagnostic and residual/relapsed cases) and 19 HG, were analyzed by 5-color flow cytometry. 32/35 cases were bone marrow aspirates and 3/35 cases were peripheral blood. A single tube containing CD10 FITC/CD58 PE/CD19 ECD/CD38 PC5/CD45 PC7 was analyzed together with the standard acute leukemia panels. To eliminate technical and fluorochrome variability in expression level analysis, relative antigenic expression was determined through comparison with appropriate internal controls. Antigen expression, as measured by the geometric mean fluorescent intensity (MFI), was then compared between B-ALL and HG using the Mann-Whitney U-test to assess for significant difference. Results were correlated with the morphologic, immunohistochemical, cytogenetic, and molecular findings for precise diagnostic classification as HG or leukemia. Results: HG demonstrated significantly brighter expression of CD38 (p<0.01) than that seen in B-ALL. In contrast, B-ALL expressed significantly brighter CD58 (p <0.01) than HG, which showed dim to no expression of the antigen. HG also showed significantly bright expression of CD10 relative to internal control granulocytes; however, this level of expression was similar to that seen in B-ALL. Median antigen expression. Hematogones show bright CD38, but dim to no CD58. Conversely, B-ALL expresses very dim CD38 and variable CD58. CD10 expression, though, demonstrates overlap between the two populations. B-ALL = B lymphoblastic leukemia, MFI = mean fluorescent intensity Comparative antigenic expression levels in hematogones and B-ALL. Select representative histograms showing HG and B-ALL blasts for the antigens CD38, CD58, and CD10 were selected from various patients studied based on those with the closest relative MFI to the overall median detected for that population in the study. The far right column shows the distribution in MFI of relative antigen expression exhibited the populations studied. HG show significantly brighter CD38 expression than B-ALL does (p<0.01). While B-ALL generally expresses brighter CD58 relative to internal controls, expression levels are variable. HG, though, show significantly dimmer CD58 to essentially no CD58 expression, compared to B-ALL (p<0.01). Similar to CD38, HG demonstrate significantly bright CD10, while B-ALL shows overall bright CD10 but variable expression levels amongst studied cases. These expression levels for CD10 overlap between HG and B-ALL and show no real statistical difference. Conclusions: The combination of CD38 and CD58 in a single tube increases the diagnostic accuracy in differentiation of HG from B-ALL. Without utilizing both antigens together, certain cases would have been difficult to interpret. Based on this analysis, we recommend that these markers be utilized in the routine evaluation for acute lymphoblastic leukemia. This is especially critical in post treatment cases in order to avoid misdiagnosis. Furthermore, the use this single tube panel would cut costs while at the same time improve patient care. Table Table. Figure Figure. Disclosures No relevant conflicts of interest to declare.
Background: Overexpression of MYC and BCL2 or/and BCL6 due to genomic rearrangements is the key molecular feature of double hit lymphoma (DHL) or triple hit lymphoma (THL). Patients with DHL and THL show very aggressive disease course and poor survival because there is no effective treatment modality. The objective of this study is to assess the impact of MYC and BCL2 inhibition on established DHL and patient derived THL cells in vitro. Methods: Primary THL cell line named as CS-THL1 was established from an 85 year-old lymphoma patient. Immunohistochemistry (IHC) was conducted to determine the levels of MYC, BCL2, BCL6, or Ki-67 expression in formalin-fixed, paraffin-embedded tissues. MTS assays were employed to evaluate the anti-growth efficacy of MYC inhibitor 10058-F4 or JQ-1, and BCL-2 inhibitor ABT-199, a BH3 mimetic, in CS-THL1 and DoGKiT cells alone or together and with or without vincristine or doxorubicin. Apoptosis, cytochrome c release and intracellular expression of BCL2, BCL-xL, BIM, and MCL-1 protein in response to drugs were analyzed by flow cytometry. Results: Expression of MYC, BCL2, BCL6 and proliferation factor Ki-67 protein was increased in THL, as shown by IHC. Analysis of cell viability demonstrated that inhibition of MYC by 10058-F4 or JQ-1 or BCL2 by ABT-199 as a single agent significantly attenuated the growth of CS-THL1 and DoGKiT cells in dose- and time-dependent manners. Combination of 10058-F4 or JQ-1 and ABT-199 or together with vincristine or doxorubicin synergistically suppressed CS-THL1 and DoGKiT cell growth compared with single agent treatment. As demonstrated by multiple approaches, apoptosis due to inhibition of BCL2 by ABT-199 or inhibition of MYC with 10058-F4 or JQ-1 exposure was the underlying cause of the observed growth retardation in CS-THL1 and DoGKiT cells. Conclusion: These observations suggest that concurrent inhibition of MYC and BCL2 pathway signaling is a potential treatment modality for patients with aggressive DH and TH B-cell lymphomas. Citation Format: Munevver Cinar, Fred Rosenfelt, Sepehr Rokshar, Raju Pillai, Jean Lopategui, Melissa Cervania, Bekir Cinar, Serhan Alkan. Co-targeting of MYC and BCL2 signaling is a promising treatment strategy for double hit and triple hit B-cell lymphomas. [abstract]. In: Proceedings of the AACR Special Conference on Myc: From Biology to Therapy; Jan 7-10, 2015; La Jolla, CA. Philadelphia (PA): AACR; Mol Cancer Res 2015;13(10 Suppl):Abstract nr B12.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.