Decreased apoptotic priming and loss of BCL-2 dependence are functional hallmarks of Richter's syndrome.

Richter's syndrome (RS) is the transformation of chronic lymphocytic leukemia (CLL) into a high-grade B-cell malignancy. Molecular and functional studies have pointed out that CLL cells are close to the apoptotic threshold and dependent on BCL-2 for survival. However, it remains undefined how evasion from apoptosis evolves during disease transformation. Here, we employed functional and static approaches to compare the regulation of mitochondrial apoptosis in CLL and RS. BH3 profiling of 17 CLL and 9 RS samples demonstrated that RS cells had reduced apoptotic priming and lower BCL-2 dependence than CLL cells. While a subset of RS was dependent on alternative anti-apoptotic proteins and was sensitive to specific BH3 mimetics, other RS cases harbored no specific anti-apoptotic addiction. Transcriptomics of paired CLL/RS samples revealed downregulation of pro-apoptotic sensitizers during disease transformation. Albeit expressed, effector and activator members were less likely to colocalize with mitochondria in RS compared to CLL. Electron microscopy highlighted reduced cristae width in RS mitochondria, a condition further promoting apoptosis resistance. Collectively, our data suggest that RS cells evolve multiple mechanisms that lower the apoptotic priming and shift the anti-apoptotic dependencies away from BCL-2, making direct targeting of mitochondrial apoptosis more challenging after disease transformation.

Novel Spirocyclic Dimer, SpiD3, Targets Chronic Lymphocytic Leukemia Survival Pathways with Potent Preclinical Effects.

Chronic lymphocytic leukemia (CLL) cell survival and growth is fueled by the induction of B-cell receptor (BCR) signaling within the tumor microenvironment (TME) driving activation of NFκB signaling and the unfolded protein response (UPR). Malignant cells have higher basal levels of UPR posing a unique therapeutic window to combat CLL cell growth using pharmacologic agents that induce accumulation of misfolded proteins. Frontline CLL therapeutics that directly target BCR signaling such as Bruton tyrosine kinase (BTK) inhibitors (e.g., ibrutinib) have enhanced patient survival. However, resistance mechanisms wherein tumor cells bypass BTK inhibition through acquired BTK mutations, and/or activation of alternative survival mechanisms have rendered ibrutinib ineffective, imposing the need for novel therapeutics. We evaluated SpiD3, a novel spirocyclic dimer, in CLL cell lines, patient-derived CLL samples, ibrutinib-resistant CLL cells, and in the Eµ-TCL1 mouse model. Our integrated multi-omics and functional analyses revealed BCR signaling, NFκB signaling, and endoplasmic reticulum stress among the top pathways modulated by SpiD3. This was accompanied by marked upregulation of the UPR and inhibition of global protein synthesis in CLL cell lines and patient-derived CLL cells. In ibrutinib-resistant CLL cells, SpiD3 retained its antileukemic effects, mirrored in reduced activation of key proliferative pathways (e.g., PRAS, ERK, MYC). Translationally, we observed reduced tumor burden in SpiD3-treated Eµ-TCL1 mice. Our findings reveal that SpiD3 exploits critical vulnerabilities in CLL cells including NFκB signaling and the UPR, culminating in profound antitumor properties independent of TME stimuli. SIGNIFICANCE: SpiD3 demonstrates cytotoxicity in CLL partially through inhibition of NFκB signaling independent of tumor-supportive stimuli. By inducing the accumulation of unfolded proteins, SpiD3 activates the UPR and hinders protein synthesis in CLL cells. Overall, SpiD3 exploits critical CLL vulnerabilities (i.e., the NFκB pathway and UPR) highlighting its use in drug-resistant CLL.

A novel potent class I HDAC inhibitor reverses the STAT4/p66Shc apoptotic defect in B cells from chronic lymphocytic leukemia patients.

Chronic Lymphocytic Leukemia (CLL) patients have a defective expression of the proapoptotic protein p66Shc and of its transcriptional factor STAT4, which evoke molecular abnormalities, impairing apoptosis and worsening disease prognosis and severity. p66Shc expression is epigenetically controlled and transcriptionally modulated by STAT4; epigenetic modifiers are deregulated in CLL cells and specific histone deacetylases (HDACs) like HDAC1, are overexpressed. Reactivation of STAT4/p66Shc expression may represent an attractive and challenging strategy to reverse CLL apoptosis defects. New selective class I HDAC inhibitors (HDACis, 6a-g) were developed with increased potency over existing agents and preferentially interfering with the CLL-relevant isoform HDAC1, to unveil the role of class I HDACs in the upregulation of STAT4 expression, which upregulates p66Shc expression and hence normalizes CLL cell apoptosis. 6c (chlopynostat) was identified as a potent HDAC1i with a superior profile over entinostat. 6c induces marked apoptosis of CLL cells compared with SAHA, which was associated with an upregulation of STAT4/p66Shc protein expression. The role of HDAC1, but not HDAC3, in the epigenetic upregulation of STAT4/p66Shc was demonstrated for the first time in CLL cells and was validated in siRNA-induced HDAC1/HDAC3 knock-down EBV-B cells. To sum up, HDAC1 inhibition is necessary to reactivate STAT4/p66Shc expression in patients with CLL. 6c is one of the most potent HDAC1is known to date and represents a novel pharmacological tool for reversing the impairment of the STAT4/p66Shc apoptotic machinery.

Linc01588 deletion inhibits the malignant biological characteristics of hydroquinone-induced leukemic cells via miR-9-5p/SIRT1.

Leukemia caused by environmental chemical pollutants has attracted great attention, the malignant leukemic transformation model of TK6 cells induced by hydroquinone (HQ) has been previously found in our team. However, the type of leukemia corresponding to this malignant transformed cell line model needs further study and interpretation. Furthermore, the molecular mechanism of malignant proliferation of leukemic cells induced by HQ remains unclear. This study is the first to reveal the expression of aberrant genes in leukemic cells of HQ-induced malignant transformation, which may correspond to chronic lymphocytic leukemia (CLL). The expression of Linc01588, a long non-coding RNA (lncRNA), was significantly up-regulated in CLL patients and leukemic cell line model which previously described. After gain-of-function assays and loss-of-function assays, feeble cell viability, severe apoptotic phenotype and the increased secretion of TNF-α were easily observed in malignant leukemic TK6 cells with Linc01588 deletion after HQ intervention. The tumors derived from malignant TK6 cells with Linc01588 deletion inoculated subcutaneously in nude mice were smaller than controls. In CLL and its cell line model, the expression of Linc01588 and miR-9-5p, miR-9-5p and SIRT1 were negative correlation respectively in CLL and cell line model, while the expression of Linc01588 and SIRT1 were positive correlation. The dual-luciferase reporter assay showed that Linc01588 & miR-9-5p, miR-9-5p & SIRT1 could bind directly, respectively. Furthermore, knockdown of miR-9-5p successfully rescued the severe apoptotic phenotype and the increased secretion of TNF-α caused by the Linc01588 deletion, the deletion of Linc01588 in human CLL cell line MEC-2 could also inhibit malignant biological characteristics, and the phenotype caused by the deletion of Linc01588 could also be rescued after overexpression of SIRT1. Moreover, the regulation of SIRT1 expression in HQ19 cells by Linc01588 and miR-9-5 P may be related to the Akt/NF-κB pathway. In brief, Linc01588 deletion inhibits the malignant biological characteristics of HQ-induced leukemic cells via miR-9-5p/SIRT1, and it is a novel and hopeful clue for the clinical targeted therapy of CLL.

Diverse B-cell tumors associated with t(14;19)(q32;q13)/IGH::BCL3 identified by G-banding and fluorescence in situ hybridization.

We characterized 5 B-cell tumors carrying t(14;19)(q32;q13) that creates the IGH::BCL3 fusion gene. The patients' ages ranged between 55 and 88 years. Two patients presented with progression or recurrence of B-cell chronic lymphocytic leukemia (B-CLL)/small lymphocytic lymphoma (SLL), two with diffuse large B-cell lymphoma (DLBCL) of non-germinal center B-like phenotype, and the remaining one with composite angioimmunoblastic T-cell lymphoma and Epstein-Barr virus-positive DLBCL. The presence of t(14;19)(q32;q13) was confirmed by fluorescence in situ hybridization (FISH), showing colocalization of 3' IGH and 3' BCL3 probes on der(14)t(14;19) and 5' BCL3 and 5' IGH probes on der(19)t(14;19). One B-CLL case had t(2;14)(p13;q32)/IGH::BCL11A, and 2 DLBCL cases had t(8;14)(q24;q32) or t(8;11;14)(q24;q11;q32), both of which generated IGH::MYC by FISH, and showed nuclear expression of MYC and BCL3 by immunohistochemistry. The IGH::BCL3 fusion gene was amplified by long-distance polymerase chain reaction in 2 B-CLL/SLL cases and the breakpoints occurred immediately 5' of BCL3 exon 1 and within the switch region associated with IGHA1. The 5 cases shared IGHV preferentially used in B-CLL cells, but the genes were unmutated in 2 B-CLL/SLL cases and significantly mutated in the remaining 3. B-cell tumors with t(14;19)(q32;q13) can be divided into B-CLL/SLL and DLBCL groups, and the anatomy of IGH::BCL3 in the latter may be different from that of the former.

An unappreciated cell survival-independent role for BAFF initiating chronic lymphocytic leukemia.

Background: Chronic Lymphocytic Leukemia (CLL) is characterized by the expansion of CD19+ CD5+ B cells but its origin remains debated. Mutated CLL may originate from post-germinal center B cells and unmutated CLL from CD5+ mature B cell precursors. Irrespective of precursor types, events initiating CLL remain unknown. The cytokines BAFF and APRIL each play a significant role in CLL cell survival and accumulation, but their involvement in disease initiation remains unclear. Methods: We generated novel CLL models lacking BAFF or APRIL. In vivo experiments were conducted to explore the impact of BAFF or APRIL loss on leukemia initiation, progression, and dissemination. Additionally, RNA-seq and quantitative real-time PCR were performed to unveil the transcriptomic signature influenced by BAFF in CLL. The direct role of BAFF in controlling the expression of tumor-promoting genes was further assessed in patient-derived primary CLL cells ex-vivo. Results: Our findings demonstrate a crucial role for BAFF, but not APRIL, in the initiation and dissemination of CLL cells. In the absence of BAFF or its receptor BAFF-R, the TCL1 transgene only increases CLL cell numbers in the peritoneal cavity, without dissemination into the periphery. While BAFF binding to BAFF-R is dispensable for peritoneal CLL cell survival, it is necessary to activate a tumor-promoting gene program, potentially linked to CLL initiation and progression. This direct role of BAFF in controlling the expression of tumor-promoting genes was confirmed in patient-derived primary CLL cells ex-vivo. Conclusions: Our study, involving both mouse and human CLL cells, suggests that BAFF might initiate CLL through mechanisms independent of cell survival. Combining current CLL therapies with BAFF inhibition could offer a dual benefit by reducing peripheral tumor burden and suppressing transformed CLL cell output.

Targeting BTK in B Cell Malignancies: From Mode of Action to Resistance Mechanisms.

The B cell receptor (BCR) signaling pathway plays a crucial role in B cell development and contributes to the pathogenesis of B cell neoplasms. In B cell malignancies, the BCR is constitutively active through both ligand-dependent and ligand-independent mechanisms, resulting in continuous Bruton tyrosine kinase (BTK) signaling activation, which provides a survival and proliferation advantage to the neoplastic clone. Among B cell malignancies, those in which the most significant results were obtained by treatment with BTK inhibitors (BTKi) include chronic lymphocytic leukemia, mantle cell lymphoma, lymphoplasmacytic lymphoma, and diffuse large B cell lymphoma. Covalent BTKi (namely ibrutinib, acalabrutinib, and zanubrutinib) functions by irreversibly blocking BTK through covalent binding to the cysteine residue 481 (Cys-481) in the ATP-binding domain. Despite the high efficacy and safety of BTKi treatment, a significant fraction of patients affected by B cell malignancies who are treated with these drugs experience disease relapse. Several mechanisms of resistance to covalent BTKi, including Cys-481 mutations of BTK, have been investigated in B cell malignancies. Non-covalent BTKi, such as pirtobrutinib, have been developed and proven effective in patients carrying both Cys-481-mutated and unmutated BTK. Moreover, targeting BTK with proteolysis-targeting chimeras (PROTACs) represents a promising strategy to overcome resistance to BTKi in B cell neoplasms.

SOLUBLE CD150 ISOFORM LEVEL IN PLASMA OF CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS.

BACKGROUND: SLAMF1/CD150 is an active player in B cell signaling networks in chronic lymphocytic leukemia (CLL). CD150-mediated signaling initiates through a homophilic CD150 binding, which spans the adjacent cells, or the interaction with the soluble CD150 isoform (sCD150). The expression of sCD150 isoform at the mRNA and protein levels ex vivo was confirmed. However, it is unclear whether sCD150 isoform present in the blood plasma of CLL patients is a factor in the constitutive activation of CD150+ cells. The aim of this study was to develop an ELISA assay for the specific sCD150 evaluation and assess the sCD150 levels in the blood plasma of CLL patients with different CD150 expression on B cells. MATERIALS AND METHODS: Blood plasma samples and peripheral blood mononuclear cells from 40 previously untreated CLL patients were analyzed. An ELISA method, ex vivo drug sensitivity assay, and a cell viability assay were used. RESULTS: The sCD150 isoform was found in all studied plasma samples of CLL patients at different levels regardless of the cell surface CD150 expression status of B cells and sCD150 mRNA expression. CLL cases with low levels of the cell surface CD150 expression in B cells are characterized by high levels of sCD150 in blood plasma in contrast to the CLL cases with high cell surface CD150 expression on B cells. The elevated levels of sCD150 in blood plasma are associated with a better sensitivity of malignant B cells to cyclophosphamide and bendamustine. CONCLUSIONS: The sCD150 isoform is actively secreted by CLL B cells with its accumulation in blood plasma, which may be regarded as an additional factor in the CLL clinicopathologic variability.

Genetically predicted telomere length and the risk of 11 hematological diseases: a Mendelian randomization study.

OBJECTIVE: Previous studies have demonstrated that various hematologic diseases (HDs) induce alterations in telomere length (TL). The aim of this study is to investigate whether genetically predicted changes in TL have an impact on the risk of developing HDs. METHODS: GWAS data for TL and 11 HDs were extracted from the database. The R software package "TwoSampleMR" was employed to conduct a two-sample Mendelian randomization (MR) analysis, in order to estimate the influence of TL changes on the risk of developing the 11 HDs. RESULTS: We examined the effect of TL changes on the risk of developing the 11 HDs. The IVW results revealed a significant causal association between genetically predicted longer TL and the risk of developing acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MANTLE), and hodgkin lymphoma (HODGKIN). However, there was no significant causal relationship observed between TL changes and the risk of developing chronic myeloid leukemia (CML), diffuse large b-cell lymphoma (DLBCL), marginal zone b-cell lymphoma (MARGINAL), follicular lymphoma (FOLLICULAR), monocytic leukemia (MONOCYTIC), and mature T/NK-cell lymphomas (TNK). CONCLUSIONS: The MR analysis revealed a positive association between genetically predicted longer TL and an increased risk of developing ALL, AML, CLL, MANTLE, and HODGKIN. This study further supports the notion that cells with longer TL have greater proliferative and mutational potential, leading to an increased risk of certain HDs.

A tower of babel of acronyms? The shadowlands of MGUS/MBL/CHIP/TCUS.

With the advent of outperforming and massive laboratory tools, such as multiparameter flow cytometry and next-generation sequencing, hematopoietic cell clones with putative abnormalities for a variety of blood malignancies have been appreciated in otherwise healthy individuals. These conditions do not fulfill the criteria of their presumed cancer counterparts, and thus have been recognized as their precursor states. This is the case of monoclonal gammopathy of unknown significance (MGUS), the first blood premalignancy state described, preceding multiple myeloma (MM) or Waldenström macroglobulinemia (WM). However, in the last 2 decades, an increasing list of clonopathies has been recognized, including monoclonal B cell lymphocytosis (MBL), which antecedes chronic lymphocytic leukemia (CLL), clonal hematopoiesis of indeterminate potential (CHIP) for myeloid neoplasms (MN), and T-cell clones of uncertain significance (TCUS) for T-cell large chronic lymphocytic leukemia (LGLL). While for some of these entities diagnostic boundaries are precisely set, for others these are yet to be fully defined. Moreover, despite mostly considered of "uncertain significance," they have not only appeared to predispose to malignancy, but also to be capable of provoking set of immunological and cardiovascular complications that may require specialized management. The clinical implications of the aberrant clones, together with the extensive knowledge generated on the pathogenetic events driving their evolution, raises the question whether earlier interventions may alter the natural history of the disease. Herein, we review this Tower of Babel of acronyms pinpointing diagnostic definitions, differential diagnosis, and the role of genomic profiling of these precursor states, as well as potential interventional strategies.

Impact of Binet Staging versus Tumour Bulk on Treatment Outcome in Chronic Lymphocytic Leukaemia.

BACKGROUND: Most of the predictive tools put up to prognosticate treatment outcomes in patients with chronic lymphocytic leukaemia (CLL) are not easily available and affordable in our resource-constrained environment. AIM: The aim of this study was to evaluate the impact of staging and some tumour bulk on treatment outcomes of persons with CLL, Enugu, Nigeria. PATIENTS AND METHODS: This is a 10-year review of the CLL data from the haemato-oncology unit of a Nigerian tertiary hospital to evaluate the impact of staging and tumour bulk indicators. Data were retrieved from the case notes of 102 patients with CLL receiving care at the facility. Data of interest include basic demographic variables, clinical features including spleen size and disease staging and blood counts. Statistical analysis was done using SPSS version 22. RESULTS: The median absolute lymphocyte count (ALC) was 108.05 (confidence interval [CI] = 50.8-201.3, interquartile range [IQR] = 124.4) ×109/L, and duration of survival for the study cohort was 5.5 (CI = 3.5-31.9, IQR = 27) months. Majority (69, 79.3%) were in Stage C. The Binet stage showed a significant association with the ALC (r = 0.338; P = 0.002) but not with spleen size (r = 0.198; P = 0.056). The duration of survival only showed a significant inverse relationship with the ALC (r = 0.35, P = 0.006) but with neither the Binet stage (r = 0.103, P = 0.431) nor spleen size (r = 0.184, P = 0.116). CONCLUSION: In CLL patients, ALC at presentation correlates with the duration of survival. We recommend that the ALC at presentation be used as a prognostic marker in our clime.

Treatment of Double-Refractory Chronic Lymphocytic Leukemia-An Unmet Clinical Need.

Recent years have seen significant improvement in chronic lymphocytic leukemia (CLL) management. Targeting B-cell lymphoma (BCL-2) and Bruton's kinase (BTK) have become the main strategies to restrain CLL activity. These agents are generally well tolerated, but the discontinuation of these therapies happens due to resistance, adverse effects, and Richter's transformation. A growing population of patients who have previously used both BTK inhibitors and BCL2 suffer from the constriction of the following regimens. This review explores the resistance mechanisms for both ibrutinib and venetoclax. Moreover, we present innovative approaches evaluated for treating double-refractory CLL.

Optical genomic mapping is a helpful tool for detecting CCND1 rearrangements in CD5-negative small B-cell lymphoma: Two cases of leukemic non-nodal mantle cell lymphoma.

Optical genome mapping (OGM) is a new DNA-based technology which provides comprehensive examination of the entire genome. We report two patients who presented with splenomegaly and leukocytosis with lymphocytosis including villous lymphocytes. Neither patient had lymphadenopathy. Bone marrow evaluation showed involvement by small B-cell lymphoma in a sinusoidal and interstitial distribution, and immunophenotypic analysis showed that the neoplastic cells were positive for B-cell markers and cyclin D1 but were negative for SOX11 and CD5. Initially, the clinicopathologic features in both patients were thought to be suspicious for hairy cell leukemia variant or splenic marginal zone lymphoma. However, OGM detected CCND1 rearrangement: t(2;11)/IGK::CCND1 in one case and t(11;14)/IGH::CCND1 in the other case. These cases illustrate the valuable role OGM can play in establishing the diagnosis of MCL. Case 1 also contributes to the paucity of literature on the rare occurrence of IGK::CCND1 in MCL.

Leukemic cell-secreted interleukin-9 suppresses cytotoxic T cell-mediated killing in chronic lymphocytic leukemia.

The tumor microenvironment (TME) plays a central role in the pathogenesis of chronic lymphocytic leukemia (CLL), contributing to disease progression and chemoresistance. Leukemic cells shape the TME into a pro-survival and immunosuppressive niche through contact-dependent and contact-independent interactions with the cellular components of the TME. Immune synapse (IS) formation is defective in CLL. Here we asked whether soluble factors released by CLL cells contribute to their protection from cytotoxic T cell (CTL)-mediated killing by interfering with this process. We found that healthy CTLs cultured in media conditioned by leukemic cells from CLL patients or Eµ-TCL1 mice upregulate the exhaustion marker PD-1 and become unable to form functional ISs and kill target cells. These defects were more pronounced when media were conditioned by leukemic cells lacking p66Shc, a proapoptotic adapter whose deficiency has been implicated in disease aggressiveness both in CLL and in the Eµ-TCL1 mouse model. Multiplex ELISA assays showed that leukemic cells from Eµ-TCL1 mice secrete abnormally elevated amounts of CCL22, CCL24, IL-9 and IL-10, which are further upregulated in the absence of p66Shc. Among these, IL-9 and IL-10 were also overexpressed in leukemic cells from CLL patients, where they inversely correlated with residual p66Shc. Using neutralizing antibodies or the recombinant cytokines we show that IL-9, but not IL-10, mediates both the enhancement in PD-1 expression and the suppression of effector functions in healthy CTLs. Our results demonstrate that IL-9 secreted by leukemic cells negatively modulates the anti-tumor immune abilities of CTLs, highlighting a new suppressive mechanism and a novel potential therapeutical target in CLL.

Cyclic Fasting-Mimicking Diet Plus Bortezomib and Rituximab Is an Effective Treatment for Chronic Lymphocytic Leukemia.

Cyclic fasting-mimicking diet (FMD) is an experimental nutritional intervention with potent antitumor activity in preclinical models of solid malignancies. FMD cycles are also safe and active metabolically and immunologically in cancer patients. Here, we reported on the outcome of FMD cycles in two patients with chronic lymphocytic leukemia (CLL) and investigated the effects of fasting and FMD cycles in preclinical CLL models. Fasting-mimicking conditions in murine CLL models had mild cytotoxic effects, which resulted in apoptosis activation mediated in part by lowered insulin and IGF1 concentrations. In CLL cells, fasting conditions promoted an increase in proteasome activity that served as a starvation escape pathway. Pharmacologic inhibition of this escape mechanism with the proteasome inhibitor bortezomib resulted in a strong enhancement of the proapoptotic effects of starvation conditions in vitro. In mouse CLL models, combining cyclic fasting/FMD with bortezomib and rituximab, an anti-CD20 antibody, delayed CLL progression and resulted in significant prolongation of mouse survival. Overall, the effect of proteasome inhibition in combination with FMD cycles in promoting CLL death supports the targeting of starvation escape pathways as an effective treatment strategy that should be tested in clinical trials. SIGNIFICANCE: Chronic lymphocytic leukemia cells resist fasting-mimicking diet by inducing proteasome activation to escape starvation, which can be targeted using proteasome inhibition by bortezomib treatment to impede leukemia progression and prolong survival.

ROR1 expression in mature B lymphoid neoplasms by flow cytometry.

Immunophenotyping by flow cytometry is an integral part of the diagnosis and classification of leukemias/lymphomas. The expression of ROR1 associated with chronic B lymphocytic leukemia (CLL) is well described in the literature, both in its diagnosis and in the follow-up of minimal residual disease (MRD) research, however, there are few studies regarding the expression pattern of ROR1 in other subtypes of mature B lymphoid neoplasms. With the aim of evaluating the expression of ROR1 and associating it with the expression of other important markers for the differentiation of mature B lymphoid neoplasms (MBLN), 767 samples of cases that entered our laboratory for immunophenotyping with clinical suspicion of MBLN were studied. ROR1 expression is predominant in CD5+/CD10- neoplasms. Overall, positive ROR1 expression was observed in 461 (60.1%) cases. The CD5+/CD10- group had a significantly higher proportion of ROR1 positive samples (89.9%) and more brightly expressed ROR1 than the other groups. Our results highlight the importance of evaluating ROR1 expression in the diagnosis of MBLN to contribute to the differential diagnosis, and possibly therapy of mainly CLL, and indicate that this marker could be considered as a useful addition to immunophenotypic panels, particularly for more challenging cases.

The CIpP activator, TR-57, is highly effective as a single agent and in combination with venetoclax against CLL cells in vitro.

Despite advances in treatment, a significant proportion of patients with chronic lymphocytic leukemia (CLL) will relapse with drug-resistant disease. The imipridones, ONC-201 and ONC-212, are effective against a range of different cancers, including acute myeloid leukemia (AML) and tumors of the brain, breast, and prostate. These drugs induce cell death through activation of the mitochondrial protease, caseinolytic protease (CIpP), and the unfolded protein response (UPR). Here we demonstrate that the novel imipridone analog, TR-57, has efficacy as a single agent and synergises with venetoclax against CLL cells under in vitro conditions that mimic the tumor microenvironment. Changes in protein expression suggest TR-57 activates the UPR, inhibits the AKT and ERK1/2 pathways and induces pro-apoptotic changes in the expression of proteins of the BCL-2 family. The study suggests that TR-57, as a single agent and in combination with venetoclax, may represent an effective treatment option for CLL.