IκB kinase-α coordinates BRD4 and JAK/STAT signaling to subvert DNA damage-based anticancer therapy.

Activation of the IκB kinase (IKK) complex has recurrently been linked to colorectal cancer (CRC) initiation and progression. However, identification of downstream effectors other than NF-κB has remained elusive. Here, analysis of IKK-dependent substrates in CRC cells after UV treatment revealed that phosphorylation of BRD4 by IKK-α is required for its chromatin-binding at target genes upon DNA damage. Moreover, IKK-α induces the NF-κB-dependent transcription of the cytokine LIF, leading to STAT3 activation, association with BRD4 and recruitment to specific target genes. IKK-α abrogation results in defective BRD4 and STAT3 functions and consequently irreparable DNA damage and apoptotic cell death upon different stimuli. Simultaneous inhibition of BRAF-dependent IKK-α activity, BRD4, and the JAK/STAT pathway enhanced the therapeutic potential of 5-fluorouracil combined with irinotecan in CRC cells and is curative in a chemotherapy-resistant xenograft model. Finally, coordinated expression of LIF and IKK-α is a poor prognosis marker for CRC patients. Our data uncover a functional link between IKK-α, BRD4, and JAK/STAT signaling with clinical relevance.

ß-Catenin activity induces an RNA biosynthesis program promoting therapy resistance in T-cell acute lymphoblastic leukemia.

Understanding the molecular mechanisms that contribute to the appearance of chemotherapy resistant cell populations is necessary to improve cancer treatment. We have now investigated the role of ß-catenin/CTNNB1 in the evolution of T-cell Acute Lymphoblastic Leukemia (T-ALL) patients and its involvement in therapy resistance. We have identified a specific gene signature that is directly regulated by ß-catenin, TCF/LEF factors and ZBTB33/Kaiso in T-ALL cell lines, which is highly and significantly represented in five out of six refractory patients from a cohort of 40 children with T-ALL. By subsequent refinement of this gene signature, we found that a subset of ß-catenin target genes involved with RNA-processing function are sufficient to segregate T-ALL refractory patients in three independent cohorts. We demonstrate the implication of ß-catenin in RNA and protein synthesis in T-ALL and provide in vitro and in vivo experimental evidence that ß-catenin is crucial for the cellular response to chemotherapy, mainly in the cellular recovery phase after treatment. We propose that combination treatments involving chemotherapy plus ß-catenin inhibitors will enhance chemotherapy response and prevent disease relapse in T-ALL patients.

Distinct roles for PARP-1 and PARP-2 in c-Myc-driven B-cell lymphoma in mice.

Dysregulation of the c-Myc oncogene occurs in a wide variety of hematologic malignancies, and its overexpression has been linked with aggressive tumor progression. Here, we show that poly (ADP-ribose) polymerase 1 (PARP-1) and PARP-2 exert opposing influences on progression of c-Myc-driven B-cell lymphoma. PARP-1 and PARP-2 catalyze the synthesis and transfer of ADP-ribose units onto amino acid residues of acceptor proteins in response to DNA strand breaks, playing a central role in the response to DNA damage. Accordingly, PARP inhibitors have emerged as promising new cancer therapeutics. However, the inhibitors currently available for clinical use are not able to discriminate between individual PARP proteins. We found that genetic deletion of PARP-2 prevents c-Myc-driven B-cell lymphoma, whereas PARP-1 deficiency accelerates lymphomagenesis in the Eµ-Myc mouse model of aggressive B-cell lymphoma. Loss of PARP-2 aggravates replication stress in preleukemic Eµ-Myc B cells, resulting in accumulation of DNA damage and concomitant cell death that restricts the c-Myc-driven expansion of B cells, thereby providing protection against B-cell lymphoma. In contrast, PARP-1 deficiency induces a proinflammatory response and an increase in regulatory T cells, likely contributing to immune escape of B-cell lymphoma, resulting in an acceleration of lymphomagenesis. These findings pinpoint specific functions for PARP-1 and PARP-2 in c-Myc-driven lymphomagenesis with antagonistic consequences that may help inform the design of new PARP-centered therapeutic strategies, with selective PARP-2 inhibition potentially representing a new therapeutic approach for the treatment of c-Myc-driven tumors.

The evolution of relapse of adult T cell acute lymphoblastic leukemia.

BACKGROUND: Adult T cell acute lymphoblastic leukemia (T-ALL) is a rare disease that affects less than 10 individuals in one million. It has been less studied than its cognate pediatric malignancy, which is more prevalent. A higher percentage of the adult patients relapse, compared to children. It is thus essential to study the mechanisms of relapse of adult T-ALL cases. RESULTS: We profile whole-genome somatic mutations of 19 primary T-ALLs from adult patients and the corresponding relapse malignancies and analyze their evolution upon treatment in comparison with 238 pediatric and young adult ALL cases. We compare the mutational processes and driver mutations active in primary and relapse adult T-ALLs with those of pediatric patients. A precise estimation of clock-like mutations in leukemic cells shows that the emergence of the relapse clone occurs several months before the diagnosis of the primary T-ALL. Specifically, through the doubling time of the leukemic population, we find that in at least 14 out of the 19 patients, the population of relapse leukemia present at the moment of diagnosis comprises more than one but fewer than 108 blasts. Using simulations, we show that in all patients the relapse appears to be driven by genetic mutations. CONCLUSIONS: The early appearance of a population of leukemic cells with genetic mechanisms of resistance across adult T-ALL cases constitutes a challenge for treatment. Improving early detection of the malignancy is thus key to prevent its relapse.

Inhibition of Xanthine Oxidoreductase Enhances the Potential of Tyrosine Kinase Inhibitors against Chronic Myeloid Leukemia.

Chronic myeloid leukemia (CML) is characterized by the expression of the oncogenic kinase BCR-ABL. Although tyrosine kinase inhibitors (TKIs) against BCR-ABL represent the standard therapeutic option for CML, resistances to TKIs can be a serious problem. Thus, the search for novel therapeutic approaches is still needed. CML cells show an increased ROS production, which is required for maintaining the BCR-ABL signaling cascade active. In line with that, reducing ROS levels could be an interesting therapeutic strategy for the clinical management of resistant CML. To analyze the therapeutic potential of xanthine oxidoreductase (XOR) in CML, we tested the effect of XOR inhibitor allopurinol. Here, we show for the first time the therapeutic potential of allopurinol against BCR-ABL-positive CML cells. Allopurinol reduces the proliferation and clonogenic ability of the CML model cell lines K562 and KCL22. More importantly, the combination of allopurinol with imatinib or nilotinib reduced cell proliferation in a synergistic manner. Moreover, the co-treatment arms hampered cell clonogenic capacity and induced cell death more strongly than each single-agent arm. The reduction of intracellular ROS levels and the attenuation of the BCR-ABL signaling cascade may explain these effects. Finally, the self-renewal potential of primary bone marrow cells from CML patients was also severely reduced especially by the combination of allopurinol with TKIs. In summary, here we show that XOR inhibition is an interesting therapeutic option for CML, which can enhance the effectiveness of the TKIs currently used in clinics.

Changes in the expression of LIMP-2 during cerulein-induced pancreatitis in rats: Effect of inhibition of leukocyte infiltration, cAMP and MAPKs early on in its development.

Lysosomal integral membrane protein-2 (LIMP-2) is an important protein in lysosomal biogenesis and function and also plays a role in the tissue inflammatory response. It is known that lysosomes play a central role in acute pancreatitis, with inflammatory cell infiltration triggering the disease early on. In this study we report increases in pancreatic LIMP-2 protein and mRNA levels as early events that occur during the development of cerulein (Cer)-induced acute pancreatitis (AP) in rats. GdCl3, a macrophage inhibitor, but not FK506, a T lymphocyte inhibitor, was able to reverse the increase in LIMP-2 expression after Cer treatment, although such reversion was abolished if the animals were depleted of neutrophils due to a vinblastine sulfate pre-treatment. Immunostaining revealed that the cellular source of LIMP-2 was mainly acinar cells. Additionally, pre-treatments with the MAPKs inhibitors SP600125 and PD98059, inhibitors of JNK and ERK½ activation, respectively, but not of rolipram, a type IV phosphodiesterase inhibitor, suppressed the increase in the expression of LIMP-2 after Cer administration. Together, these results indicate that neutrophils are able to drive a macrophage activation that would regulate the increase in LIMP-2 expression during the early phase of Cer-induced AP, with the stress kinases JNK and ERK½ also playing a coordinated role in the increase of LIMP-2 expression due to Cer.

Modulation in the expression of SHP-1, SHP-2 and PTP1B due to the inhibition of MAPKs, cAMP and neutrophils early on in the development of cerulein-induced acute pancreatitis in rats.

The protein tyrosine phosphatases (PTPs) SHP-1, SHP-2 and PTP1B are overexpressed early on during the development of cerulein -induced acute pancreatitis (AP) in rats, and their levels can be modulated by some species of mitogen-activated protein kinases (MAPKs), the intracellular levels of cAMP and by general leukocyte infiltration, the latter at least for SHP-2 and PTP1B. In this study we show that cerulein treatment activates extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) but not p38 MAPK during the early phase of cerulein-induced AP (2h after the first injection of cerulein). Therefore, by using the MAPK inhibitors SP600125 (a specific JNK inhibitor) and PD98059 (a specific ERK inhibitor), we have unmasked the particular MAPK that underlies the modulation of the expression levels of these PTPs. JNK would act by preventing SHP-1 protein expression from increasing beyond a certain level. ERK 1/2 was the main MAPK involved in the increase in SHP-2 protein expression due to cerulein. JNK negatively modulated the SH2-domain containing PTPs. Both MAPKs played a role in the increase in PTP1B protein expression due to cerulein. Finally, by using the white blood cell inhibitors vinblastine sulfate, gadolinium chloride and FK506 (tacrolimus), we show that the macrophage activity or T-lymphocytes does not modulate the expression of any of the PTPs, although neutrophil infiltration was found to be a regulator of SHP-2 and PTP1B protein expression due to cerulein.