Proceedings of the Third Global Workshop on Chronic Myeloid Leukemia.

Tyrosine kinase inhibitor (TKI) therapy has resulted in unprecedented responses and survival rates in Philadelphia chromosome-positive chronic myelogenous leukemia (CML) that are durable for years. However, a third of patients either fail to respond or respond suboptimally to imatinib therapy, while some others are intolerant to imatinib. Increased understanding of the molecular basis of imatinib resistance has led to rational development of second-generation TKIs as effective second-line treatment options for imatinib-resistant patients. However, persistence of minimal residual disease (MRD) and development of resistance against TKI therapy are proving to be significant challenges. Treatment options are evolving for patients with CML and the promising results with several novel agents showing activity in CML, including in patients with the T315I mutation, are encouraging advancements in the field. Recently, a panel of global experts in CML deliberated on the imminent approval of second-generation TKIs in the frontline setting, ways to improve on frontline therapy, integration of new agents in current treatment algorithms, and design of future clinical trials; the proceedings of the discussion are summarized in this article.

Treatment of patients with acute promyelocytic leukemia: a consensus statement on risk-adapted approaches to therapy.

The integration of all-trans-retinoic acid (ATRA) with anthracycline chemotherapy into up-front regimens for acute promyelocytic leukemia (APL) has produced striking improvements in clinical outcomes, making APL one of the most curable forms of hematologic malignancies in adults. In addition, arsenic trioxide has proven efficacy in relapse and when combined with ATRA in frontline regimens has demonstrated high efficacy in a number of trials and the potential to replace chemotherapy, which would thereby reduce chemotherapy-related adverse events. Cure in APL is also largely dependent on timely and effective supportive care measures that counteract such life-threatening emergencies as coagulopathy and APL differentiation syndrome. The risk of mortality during induction and the risk of relapse following frontline therapy are related to the individual's initial clinical presentation and relapse-risk score at diagnosis; however, these risks are now quite low even in high-risk patients as long as appropriate therapy is initiated expeditiously. Multiple European and North American trials have investigated risk-adapted approaches to the treatment of APL and have reported high success rates. Further refinement of risk-adapted strategies is ongoing and will likely contribute to better patient care. A roundtable conference was conducted to discuss risk-adapted therapy for APL and to develop a consensus statement and approach for clinical oncologists. Expert opinions and evidence-based strategies presented during the roundtable are summarized herein.

Risk-adapted approaches to therapy for high-risk acute promyelocytic leukemia.

In the past decade, risk stratification based on white blood cell (WBC) and platelet counts at initial clinical presentation in patients with acute promyelocytic leukemia (APL) has allowed adjustments in intensity and type of up-front agents used in the treatment of different risk groups, thereby optimizing clinical outcomes. This review summarizes the key issues, as currently understood, in the management of high-risk (WBC count > 10,000/µL) APL. Multiple European and North American trials have successfully attempted incorporation of risk-adapted approaches into the consolidation phase of frontline treatment; clinical data from the integration of all-trans-retinoic acid and cytarabine into therapeutic regimens for high-risk disease are discussed. Another agent that has gained enormous interest in APL management in recent years is arsenic trioxide (ATO). A positive impact of adding ATO to consolidation regimens was reported for all risk groups of APL in the North American Leukemia Intergroup C9710 trial; recent results from that trial are also reviewed. In addition to therapeutic strategies, optimal management of APL, especially high-risk disease, also involves using effective supportive care measures, monitoring response to therapy, and preventing relapse. Efforts to better understand these aspects and further refine risk-tailored therapy are under way.

Reconstitution of the nuclear transport of the MAP kinase ERK2.

The nuclear-cytoplasmic distribution of ERK2 is regulated in response to various stimuli and changes in cell context. Furthermore, the nuclear flux of ERK2 occurs by several energy- and carrier-dependent and -independent mechanisms. ERK2 has been shown to translocate into and out of the nucleus by facilitated diffusion through the nuclear pore, interacting directly with proteins within the nuclear pore complex, as well as by karyopherin-mediated transport. Nuclear export has been suggested to be CRM1- and MEK1/2-dependent. Here, we describe a general nuclear import assay of wild-type ERK2 that can be employed to identify different mechanisms governing nuclear entry of the protein kinase, adapted to evaluate ERK2 mutants that impair nuclear entry to dissect energy- and carrier-dependent and -independent mechanisms, and extended to characterize export mechanisms.

Mutations in ERK2 binding sites affect nuclear entry.

The MAPK ERK2 can enter and exit the nucleus by an energy-independent process that is facilitated by direct interactions with nuclear pore proteins. Several studies also suggest that the localization of ERK2 can be influenced by carrier proteins. Using import reconstitution assays, we examined a group of ERK2 mutants defective in known protein interactions to determine structural properties of ERK2 that contribute to its nuclear entry. ERK2 mutants defective in binding to substrates near the active site or to basic/hydrophobic docking (D) motifs were imported normally. Several ERK2 mutants defective in interactions with FXF motifs displayed slowed rates of nuclear import. The import-impaired mutants also showed reduced binding to a recombinant C-terminal fragment of nucleoporin 153 that is rich in FXF motifs. Despite the deficit revealed in some mutants via reconstitution assays, all but one of the ERK2 mutants accumulated in nuclei of stimulated cells in a manner comparable with the wild type protein; the mutant most defective in import remained in the cytoplasm. These results further support the idea that direct interactions with nucleoporins are involved in ERK2 nuclear entry and that multiple events contribute to the ligand-dependent relocalization of these protein kinases.

Randomized phase III trial results of panitumumab, a fully human anti-epidermal growth factor receptor monoclonal antibody, in metastatic colorectal cancer.

Monoclonal antibodies against the epidermal growth factor receptor have proven efficacy as monotherapy and in combination with chemotherapy in patients with metastatic colorectal cancer (CRC: mCRC). Initial clinical trials in CRC used the human-murine chimeric monoclonal antibody cetuximab. Ongoing studies are being conducted to evaluate the efficacy and safety of the fully human anti-epiderman growth factor receptor monoclonal antibody panitumumab. The results of a phase III trial which compared panitumumab as a single agent to best supportive care in patients with previously treated metastatic CRC have recently been reported Pantitumumab therapy resulted in a 46% reduction in the risk of tumor progression and a partial response rate of 8%. Rash was reported in 90% of patients with increased severity significantly correlated with improved medium overall survival (OS). Further clinical studies re ongoing and planned to test panitumumab in combination with chemotherapy in first-line therapy of advanced-stage CRC and adjuvant treatment of colon cancer.

The MAP kinase ERK5 binds to and phosphorylates p90 RSK.

We showed previously that p90 RSK was activated in cells expressing an activated mutant of MEK5, the activator of the MAP kinase ERK5. Based on the following evidence, we suggest that ERK5 can directly activate RSK in cells. ERK5 binds to RSK in vitro and co-immunoprecipitates from cell extracts; activation of ERK5 weakens its binding to RSK, suggesting that RSK is released upon activation. Phosphorylation of RSK by ERK5 in vitro causes its activation, indicating that RSK is a substrate of ERK5. In cells activation of ERK5 but not p38 or the c-Jun N-terminal kinase is associated with RSK activation. The large C-terminal domain of ERK5 is not required for binding or activation of RSK by ERK5; however, the common docking or CD domain of ERK5 and the docking or D domain of RSK are important for their association.

The nuclear localization of ERK2 occurs by mechanisms both independent of and dependent on energy.

The mitogen-activated protein (MAP) kinases ERK1 and ERK2 often accumulate in the nuclei of stimulated cells to mediate changes in transcription. The mechanisms underlying stimulus-dependent redistribution of these kinases remain unclear. We have used a permeabilized cell reconstitution assay in HeLa cells and human foreskin fibroblasts to explore the processes by which ERK2 enters and exits the nucleus. We previously reported that entry of unphosphorylated ERK2 into the nucleus occurs by facilitated diffusion not requiring cytosolic transport factors. We find that export, like import, can occur by an energy- and carrier-independent mechanism. An energy-dependent mechanism of ERK2 export can also be distinguished, mediated at least in part through the exportin CRM1. We have also examined import and export of thiophosphorylated, active ERK2. Import of active ERK2 is significantly enhanced by the addition of exogenous transport factors and an energy regeneration system. These studies support a model in which multiple constitutive and regulated processes control the subcellular distribution of ERK2.