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Prolactin (PRL) is elevated in B-cell-mediated lymphoproliferative diseases and promotes B-cell survival. Whether PRL or PRL receptors drive the evolution of B-cell malignancies is unknown. We measure changes in B cells after knocking down the pro-proliferative, anti-apoptotic long isoform of the PRL receptor (LFPRLR) in vivo in systemic lupus erythematosus (SLE)- and B-cell lymphoma-prone mouse models, and the long plus intermediate isoforms (LF/IFPRLR) in human B-cell malignancies. To knockdown LF/IFPRLRs without suppressing expression of the counteractive short PRLR isoforms (SFPRLRs), we employ splice-modulating DNA oligomers. In SLE-prone mice, LFPRLR knockdown reduces numbers and proliferation of pathogenic B-cell subsets and lowers the risk of B-cell transformation by downregulating expression of activation-induced cytidine deaminase. LFPRLR knockdown in lymphoma-prone mice reduces B-cell numbers and their expression of BCL2 and TCL1. In overt human B-cell malignancies, LF/IFPRLR knockdown reduces B-cell viability and their MYC and BCL2 expression. Unlike normal B cells, human B-cell malignancies secrete autocrine PRL and often express no SFPRLRs. Neutralization of secreted PRL reduces the viability of B-cell malignancies. Knockdown of LF/IFPRLR reduces the growth of human B-cell malignancies in vitro and in vivo. Thus, LF/IFPRLR knockdown is a highly specific approach to block the evolution of B-cell neoplasms.

Studying mechanisms of malignant transformation of human pre-B cells, we found that acute activation of oncogenes induced immediate cell death in the vast majority of cells. Few surviving pre-B cell clones had acquired permissiveness to oncogenic signaling by strong activation of negative feedback regulation of Erk signaling. Studying negative feedback regulation of Erk in genetic experiments at three different levels, we found that Spry2, Dusp6, and Etv5 were essential for oncogenic transformation in mouse models for pre-B acute lymphoblastic leukemia (ALL). Interestingly, a small molecule inhibitor of DUSP6 selectively induced cell death in patient-derived pre-B ALL cells and overcame conventional mechanisms of drug-resistance.

The unfolded protein response (UPR) pathway, a stress-induced signaling cascade emanating from the endoplasmic reticulum (ER), regulates the expression and activity of molecules including BiP (HSPA5), IRE1 (ERN1), Blimp-1 (PRDM1), and X-box binding protein 1 (XBP1). These molecules are required for terminal differentiation of B cells into plasma cells and expressed at high levels in plasma cell-derived multiple myeloma. Although these molecules have no known role at early stages of B-cell development, here we show that their expression transiently peaks at the pre-B-cell receptor checkpoint. Inducible, Cre-mediated deletion of Hspa5, Prdm1, and Xbp1 consistently induces cellular stress and cell death in normal pre-B cells and in pre-B-cell acute lymphoblastic leukemia (ALL) driven by BCR-ABL1- and NRAS(G12D) oncogenes. Mechanistically, expression and activity of the UPR downstream effector XBP1 is regulated positively by STAT5 and negatively by the B-cell-specific transcriptional repressors BACH2 and BCL6. In two clinical trials for children and adults with ALL, high XBP1 mRNA levels at the time of diagnosis predicted poor outcome. A small molecule inhibitor of ERN1-mediated XBP1 activation induced selective cell death of patient-derived pre-B ALL cells in vitro and significantly prolonged survival of transplant recipient mice in vivo. Collectively, these studies reveal that pre-B ALL cells are uniquely vulnerable to ER stress and identify the UPR pathway and its downstream effector XBP1 as novel therapeutic targets to overcome drug resistance in pre-B ALL.

The reaction of substituted acylthiourea derivatives (L1–L4) with [(PPh3)2Cu(μ‐Cl)2Cu(PPh3)] in a proper stoichiometric proportion gave the complexes (1–4) in good yields. The structures of the compounds (ligands and complexes) were advocated through spectroscopic analyses and x‐ray diffraction (XRD) studies; the latter confirmed their molecular structures. The complexes (1–4) were assessed for their cytotoxic potential through MTT assay against A549, T24, and HepG2 cancer cells, and their toxicity was evaluated using Vero and MCF‐10a normal cells. The complexes (1–4) displayed better cytotoxic action toward HepG2 cancer cells than cisplatin. Complexes 3 and 4 significantly destroyed A549 cancer cells with the IC50 values of 16.25 and 4.90 μM, respectively, which were lower than that of cisplatin (IC50 = 17.73 μM). Furthermore, the higher IC50 values in normal cells showed that the complexes were less hazardous to the normal cells except complex 3. The biocompatibility study showed that the complexes had minimal impact on normal human dermal fibroblast (NHDF) cells. The mode of cell death of complexes 3 and 4 against cancer cells was evaluated using staining assays. Both compounds demonstrated a significant reduction in live cells, an increase in early apoptotic cells, and pronounced proapoptotic effects, as evidenced by fragmented nuclei. Flow cytometry revealed a distinctive subpopulation in the subG0 phase, highlighting the compounds' ability to induce apoptosis and inhibit cell cycle progression. [ABSTRACT FROM AUTHOR]

The B cell-specific transcription factor BACH2 is required for affinity maturation of B cells. Here we show that Bach2-mediated activation of p53 is required for stringent elimination of pre-B cells that failed to productively rearrange immunoglobulin VH-DJH gene segments. After productive VH-DJH gene rearrangement, pre-B cell receptor signaling ends BACH2-mediated negative selection through B cell lymphoma 6 (BCL6)-mediated repression of p53. In patients with pre-B acute lymphoblastic leukemia, the BACH2-mediated checkpoint control is compromised by deletions, rare somatic mutations and loss of its upstream activator, PAX5. Low levels of BACH2 expression in these patients represent a strong independent predictor of poor clinical outcome. In this study, we demonstrate that Bach2(+/+) pre-B cells resist leukemic transformation by Myc through Bach2-dependent upregulation of p53 and do not initiate fatal leukemia in transplant-recipient mice. Chromatin immunoprecipitation sequencing and gene expression analyses carried out by us revealed that BACH2 competes with BCL6 for promoter binding and reverses BCL6-mediated repression of p53 and other cell cycle checkpoint-control genes. These findings identify BACH2 as a crucial mediator of negative selection at the pre-B cell receptor checkpoint and a safeguard against leukemogenesis.

Tyrosine kinase inhibitors (TKIs) are widely used to treat patients with leukaemia driven by BCR-ABL1 (ref. 1) and other oncogenic tyrosine kinases. Recent efforts have focused on developing more potent TKIs that also inhibit mutant tyrosine kinases. However, even effective TKIs typically fail to eradicate leukaemia-initiating cells (LICs), which often cause recurrence of leukaemia after initially successful treatment. Here we report the discovery of a novel mechanism of drug resistance, which is based on protective feedback signalling of leukaemia cells in response to treatment with TKI. We identify BCL6 as a central component of this drug-resistance pathway and demonstrate that targeted inhibition of BCL6 leads to eradication of drug-resistant and leukaemia-initiating subclones.

BCL6 protects germinal center (GC) B cells against DNA damage-induced apoptosis during somatic hypermutation and class-switch recombination. Although expression of BCL6 was not found in early IL-7-dependent B cell precursors, we report that IL-7Ralpha-Stat5 signaling negatively regulates BCL6. Upon productive VH-DJH gene rearrangement and expression of a mu heavy chain, however, activation of pre-B cell receptor signaling strongly induces BCL6 expression, whereas IL-7Ralpha-Stat5 signaling is attenuated. At the transition from IL-7-dependent to -independent stages of B cell development, BCL6 is activated, reaches expression levels resembling those in GC B cells, and protects pre-B cells from DNA damage-induced apoptosis during immunoglobulin (Ig) light chain gene recombination. In the absence of BCL6, DNA breaks during Ig light chain gene rearrangement lead to excessive up-regulation of Arf and p53. As a consequence, the pool of new bone marrow immature B cells is markedly reduced in size and clonal diversity. We conclude that negative regulation of Arf by BCL6 is required for pre-B cell self-renewal and the formation of a diverse polyclonal B cell repertoire.

Three Pd(II) complexes (1–3) containing tridentate chromone-based thiosemicarbazones (SVSHL1–3) have been investigated as potential anticancer agents. The complexes of the type [Pd(PPh3)(ONS-(SVSHL1–3)-OCH3)] were synthesized by the reaction of [PdCl2(PPh3)2] with the corresponding TSC ligands under inert conditions; the formation of the complexes occurred with the ligands undergoing in situ Michael addition. The characterization of complexes was achieved with the aid of analytical and various spectroscopic techniques. The molecular structure of complex 3 was determined using a single crystal X-ray diffraction (XRD) method, confirming the Michael addition pathway as well. The complexes were screened against a panel of cancer and normal cell lines to evaluate their anticancer activity; complexes 2 and 3 with a phenyl or cyclohexyl substituent on the N-terminal of the ligands exhibited IC50 values of 2.87 and 4.01 μM against HeLa cancer cells, respectively. The apoptotic mode of cell death was confirmed using microscopic and flow cytometry studies. Interestingly, normal cell architecture was seen in the tissues of mice treated with the active Pd(II) complexes, in contrast with those treated with cisplatin which induced severe damage to the normal tissues in mice. [ABSTRACT FROM AUTHOR]

The research and development department continues searching for effective and definite anticancer medications that would help eradicate cancer. The thiosemicarbazone ligands with indole fragment were utilized to create four new copper(I) complexes. Several spectral examinations and elemental analyses validated the formation of the complexes (1–4). Single crystal X‐ray diffraction (XRD) also proved that complex 4 had the expected tetrahedral structure. Complexes 1–4 were assessed for their cytotoxic property on different cell lines such as immortalized human vascular endothelial (EAhy.926), hepatocellular carcinoma (HepG‐2), and bladder cancer (T24), and kidney epithelial normal cells extracted from an African green monkey (Vero). The complexes showed reduced toxicity towards healthy Vero cells while being active in cancer cell lines. While the standard cisplatin revealed IC50 values of 26.60 and 49.90 μM, the N‐phenyl substituted complex 3 displayed a superior cytotoxic effect with IC50 values of 13.82 and 24.93 μM towards EAhy.926 and HepG‐2 cells, respectively. [ABSTRACT FROM AUTHOR]

Memory B cells are a dynamic subset of the mature B cell population that in some cases can reenter germinal centers (GCs) in response to iterative infections. Such a reactivation can lead to accumulation of genetic lesions in these cells, potentially from repetitive activation of the B cell mutator enzyme AID. Normal memory B cells do not survive repeated reentries into GCs. In this issue, Sungalee et al. demonstrate that memory B cells harboring the oncogenic BCL2:IGH translocation, which results in constitutive BCL2 expression, survive multiple GC entries upon repetitive immunization. Through these multiple GC reentries, the hallmark BCL2:IGH translocation enables AID-induced hypermutation and propagates clonal evolution toward malignant follicular lymphoma., Clonal evolution of follicular lymphomas Follicular lymphoma (FL) is an indolent subtype of non-Hodgkin's lymphoma (1) that clonally evolves over decades (2) before presenting as overt disease. In almost all [...]

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that drives the generation of myeloid cell subsets including neutrophils, monocytes, macrophages, and dendritic cells in response to stress, infections, and cancers. By modulating the functions of innate immune cells that serve as a bridge to activate adaptive immune responses, GMCSF globally impacts host immune surveillance under pathologic conditions. As with other soluble mediators of immunity, too much or too little GM-CSF has been found to promote cancer aggressiveness. While too little GM-CSF prevents the appropriate production of innate immune cells and subsequent activation of adaptive anti-cancer immune responses, too much of GM-CSF can exhaust immune cells and promote cancer growth. The consequences of GM-CSF signaling in cancer progression are a function of the levels of GM-CSF, the cancer type, and the tumor microenvironment. In this review, we first discuss the secretion of GM-CSF, signaling downstream of the GM-CSF receptor, and GM-CSF's role in modulating myeloid cell homeostasis. We then outline GM-CSF's anti-tumorigenic and pro-tumorigenic effects both on the malignant cells and on the nonmalignant immune and other cells in the tumor microenvironment. We provide examples of current clinical and preclinical strategies that harness GM-CSF's anti-cancer potential while minimizing its deleterious effects. We describe the challenges in achieving the Goldilocks effect during administration of GM-CSF-based therapies to patients with cancer. Finally, we provide insights into how technologies that map the immune microenvironment spatially and temporally may be leveraged to intelligently harness GM-CSF for treatment of malignancies. [ABSTRACT FROM AUTHOR]

Denticity plays a prominent role in the cytotoxicity of metal complexes. The impact of ligand denticity of chromone/chromene thiosemicarbazones (TSCs) in the Ru(II)‐DMSO complexes on the cytotoxicity of cancer cells was evaluated. Ru(II)‐DMSO complex containing tridentate chromone TSC (D1) exhibited better in vitro anticancer activity than the one with bidentate chromene TSC (D2). Using the optimized structures of the complexes, the reason behind the same was attributed to the thermodynamically stable nature of the tridentate system. Hydrolysis study of the complexes showed the kinetically labile nature of the chloride ligand in the tridentate species with respect to the bidentate counterpart. The selective cytotoxicity of complex D1 against breast cancer cells and its less toxicity against normal cells demonstrated the effective anticancer potential of the complex. Further, the binding efficacy of the complexes against DNA and BSA, along with imaging and flow cytometry experiments, warranted the effective cell death induced by the complexes against breast cancer cells via apoptosis. [ABSTRACT FROM AUTHOR]

Six different acylthiourea ligands (L1–L6) and their corresponding Ru(II)-p-cymene complexes (P1–P6) were designed to explore the structure–activity relationship of the complexes upon aliphatic chain and aromatic conjugation on the C- and N-terminals, respectively. The compounds were synthesized and adequately characterized using various analytical and spectroscopic techniques. The structures of P2–P6, solved using single crystal X-ray diffraction (XRD), confirmed the neutral monodentate coordination of the S atoms of the acylthiourea ligands to Ru(II) ions. In silico studies showed an increase of lipophilicity for the ligands with an increase in alkyl chain length or aromatic conjugation at the C- or N-terminal, respectively. Subsequently, mitogen-activated protein kinases (MAPK) were predicted as one of the primary targets for the complexes, which showed good binding affinity towards extracellular signal-regulated kinases (ERK1, ERK2 and ERK5), c-Jun N-terminal kinase (JNK) and p38 of the MAPK pathway. Henceforth, the complexes were tested for their anticancer activity in lung carcinoma (A549) and cisplatin-resistant lung carcinoma (cisA549R) cells and human umbilical vein epithelial normal cells (HUVEC). Interestingly, an increase in chain length or aromatic conjugation led to an increase in the activity of the complexes, with P5 (7.73 and 13.04 μM) and P6 (6.52 and 14.45 μM) showing the highest activity in A549 and cisA549R cells, which is better than the positive control, cisplatin (8.72 and 44.28 μM). Remarkably, we report the highest activity yet observed for complexes of the type [(η6-p-cymene)RuIICl2(S-acylthiourea)] in the tested cell lines. Aqueous solution studies showed that complexes P5 and P6 are rapidly hydrolyzed to produce solely aquated species that remained stable for 24 h. Staining assays and flow cytometric analyses of P5 and P6 in A549 cells revealed that the complexes induced apoptosis and arrested the cell cycle predominantly in the S phase. In vivo studies demonstrated the higher toxicity of cisplatin and a comparatively higher survival rate of mice injected with the most active complex P6. Histological analyses revealed that treatment with P6 at high doses of up to 8 mg kg−1 did not cause any palpable damage to the tested organs. [ABSTRACT FROM AUTHOR]

Fourteen new RuII–arene (p‐cymene/benzene) complexes (C1–C14) have been synthesized by varying the N‐terminal substituent in the furoylthiourea ligand and satisfactorily characterized by using analytical and spectroscopic techniques. Electrostatic potential maps predicted that the electronic effect of the substituents was mostly localized, with some influence seen on the labile chloride ligands. The structure–activity relationships of the Ru–p‐cymene and Ru–benzene complexes showed opposite trends. All the complexes were found to be highly toxic towards IMR‐32 cancer cells, with C5 (Ru–p‐cymene complex containing C6H2(CH3)3 as N‐terminal substituent) and C13 (Ru–benzene complex containing C6H4(CF3) as N‐terminal substituent) showing the highest activity among each set of complexes, and hence they were chosen for further study. These complexes showed different behavior in aqueous solutions, and were also found to catalytically oxidize glutathione. They also promoted cell death by apoptosis and cell cycle arrest. Furthermore, the complexes showed good binding ability with the receptors Pim‐1 kinase and vascular endothelial growth factor receptor 2, commonly overexpressed in cancer cells. [ABSTRACT FROM AUTHOR]

The targeted inactivation of individual oncogenes can elicit regression of cancers through a phenomenon called oncogene addiction. Oncogene addiction is mediated by cell-autonomous and immune-dependent mechanisms. Therapeutic resistance to oncogene inactivation leads to recurrence but can be counteracted by immune surveillance. Predicting the timing of resistance will provide valuable insights in developing effective cancer treatments. To provide a quantitative understanding of cancer response to oncogene inactivation, we developed a new 3-compartment mathematical model of oncogene-driven tumor growth, regression and recurrence, and validated the model using a MYC-driven transgenic mouse model of T-cell acute lymphoblastic leukemia. Our mathematical model uses imaging-based measurements of tumor burden to predict the relative number of drug-sensitive and drug-resistant cancer cells in MYC-dependent states. We show natural killer (NK) cell adoptive therapy can delay cancer recurrence by reducing the net-growth rate of drug-resistant cells. Our studies provide a novel way to evaluate combination therapy for personalized cancer treatment. [ABSTRACT FROM AUTHOR]

The reactions of CuCl2·2H2O with chromone thiosemicarbazone ligands containing a –H or –CH3 substituent on terminal N yielded monometallic Cu(II) complexes [Cu(HL1)Cl2] (1) and [Cu(HL2)Cl2] (2), whereas bimetallic Cu(II) complexes [Cu(μ-Cl)(HL3)]2Cl2 (3), [Cu(μ-Cl)(HL4)]2Cl2 (4) and [Cu(μ-Cl)(L5)]2 (5) were obtained when a –C2H5, –C6H11 or –C6H5 substituent was present, respectively, in the ligands. The complexes were characterized using elemental analyses, UV-Vis, FT-IR, EPR, mass and TGA studies. The structures of neutral monometallic and dicationic bimetallic complexes were confirmed by single crystal X-ray diffraction, and they exhibited a distorted square pyramidal geometry around Cu(II) ions. The catecholase-mimicking activity of complexes 1–5 was examined spectrophotometrically, and the results revealed that all the complexes except 5 had the ability to oxidize 3,5-di-tert-butylcatechol (3,5-DTBC) to 3,5-di-tert-butylquinone (3,5-DTBQ) under aerobic conditions with moderate turnover numbers. In order to find the possible complex–substrate intermediates, a mass spectrometry study was carried out for complexes 1–4 in the presence of 3,5-DTBC. The phosphatase-like activity of 1–5 was also investigated using 4-nitrophenylphosphate (4-NPP) as a model substrate. All the complexes exhibited excellent phosphatase activity in DMF-H2O medium. The complexes displayed significant biomolecular interactions and antioxidant potential. Complex 3 showed good interaction with apoptotic CASP3 protein, VEGFR2 and PIM-1 kinase receptors as revealed by a molecular docking study. Complexes (3–5) exhibited promising cytotoxicity against HeLa-cervical cancer cells with IC50 values of 2.24 (3), 2.25 (4) and 3.77 (5) μM, respectively, and showed a two-fold higher activity than cisplatin. The active complex 3 showed complete inhibition of colony formation at 10 μM concentration. In addition, the acridine orange (AO)/ethidium bromide (EB) staining and real-time live cell imaging results confirmed that complex 3 induced cell death in HeLa cells. [ABSTRACT FROM AUTHOR]

Background The telomerase enzyme is a viable target for anti-cancer therapy given the innate differences in telomerase activity between tumour cells and normal somatic cells. However, the time lag between telomerase inhibition and telomeres becoming critically short to trigger cell death, allows cancer cells to acquire drug resistance. Inhibition of DNA repair pathways along with telomerase could be an alternative strategy to enhance anti-tumour effects and circumvent the possibility of drug resistance. Poly (ADP-Ribose) Polymerase-1 (PARP-1), an important DNA damage sensor and a DNA repair factor, has important roles in maintaining telomeres and chromosomal stability. In this study, the effects of combined inhibition of PARP-1 and telomerase in mouse embryonic fibroblasts (MEFs) following sodium arsenite exposure (a carcinogen and potent DNA damaging agent), were evaluated. Results Inhibition of PARP in telomerase deficient MEFs induced an increase in arsenite-induced DNA damage as compared to control cells. Combined inhibition also resulted in enhanced genomic instability, demonstrated by elevated micronuclei induction and chromosomal aberrations with decreased cell survival. In addition, telomerase inhibition in PARP-1 deficient MEFs led to greater telomere shortening and increased genomic instability. Conclusions Our study demonstrated that the co-inhibition of PARP-1 and telomerase in MEFs rendered cells more susceptible to DNA damaging agents. Hence, these results offer support for the use of combined inhibition of PARP-1 and telomerase as a strategy to minimise the problems associated with long-term telomerase inhibition in cancer therapeutics., Authors: Resham L Gurung [1]; Lakshmidevi Balakrishnan [1]; Rabindra N Bhattacharjee [2]; Jayapal Manikandan [1]; Srividya Swaminathan [1]; M Prakash Hande (corresponding author) [1] Background Telomeres are specialised dynamic structures [...]