Alan K. Ikeda

Targeting CREB for Cancer Therapy: Friend or Foe

The cyclic-AMP response element-binding protein (CREB) is a nuclear transcription factor activated by phosphorylation at Ser133 by multiple serine/threonine (Ser/Thr) kinases. Upon phosphorylation, CREB binds the transcriptional co-activator, CBP (CREB-binding protein), to initiate CREB-dependent gene transcription. CREB is a critical regulator of cell differentiation, proliferation and survival in the nervous system. Recent studies have shown that CREB is involved tumor initiation, progression and metastasis, supporting its role as a proto-oncogene. Overexpression and over-activation of CREB were observed in cancer tissues from patients with prostate cancer, breast cancer, non-small-cell lung cancer and acute leukemia while down-regulation of CREB in several distinct cancer cell lines resulted in inhibition of cell proliferation and induction of apoptosis, suggesting that CREB may be a promising target for cancer therapy. Although CREB, as a transcription factor, is a challenging target for small molecules, various small molecules have been discovered to inhibit CREB phosphorylation, CREB-DNA, or CREB-CBP interaction. These results suggest that CREB is a suitable transcription factor for drug targeting and therefore targeting CREB could represent a novel strategy for cancer therapy.

FMS‐Like Tyrosine Kinase 3 in Normal Hematopoiesis and Acute Myeloid Leukemia

Ligand‐mediated activation of the FMS‐like tyrosine kinase 3 (FLT3) receptor is important for normal proliferation of primitive hematopoietic cells. However, activating mutations in FLT3 induce ligand‐independent downstream signaling that promotes oncogenesis through pathways involved in proliferation, differentiation, and survival. FLT3 mutations are identified as the most frequent genetic abnormality in acute myeloid leukemia and are also observed in other leukemias. Multiple small‐molecule inhibitors are under development to target aberrant FLT3 activity that confers a poor prognosis in patients.

The aggresome pathway as a target for therapy in hematologic malignancies

Misfolded or unfolded proteins are often refolded with the help of chaperones or degraded by the 26S proteasome. An alternative fate of these proteins is the aggresome pathway. The microtubule-organizing center (MTOC) transports unfolded proteins to lysosomes and are degraded through autophagy. Histone deacetylase 6 (HDAC6) deacetylates alpha-tubulin, which is thought to be a component of the MTOC. Recently, two small molecule inhibitors of the aggresome pathway and HDAC6 have been described. One inhibitor, tubacin, prevents deacetylation of alpha-tubulin and produces accumulation of polyubiquitinated proteins and apoptosis. Tubacin acts synergistically with the proteasome inhibitor, bortezomib, to induce cytotoxicity in one type of hematologic malignancy, multiple myeloma. The other, LBH589, is a pan HDAC inhibitor and hydroxamic acid derivative that induces apoptosis of multiple myeloma cells resistant to conventional therapies. In this review, we summarize recent reports on targeting the aggresome pathway and HDAC6 in hematologic malignancies.

Tubacin suppresses proliferation and induces apoptosis of acute lymphoblastic leukemia cells

Over the past decade, histone deacetylase inhibitors have increasingly been used to treat various malignancies. Tubacin (tubulin acetylation inducer) is a small molecule that inhibits histone deacetylase 6 (HDAC6) and induces acetylation of α-tubulin. We observed a higher antiproliferative effect of tubacin in acute lymphoblastic leukemia (ALL) cells than in normal hematopoietic cells. Treatment with tubacin led to the induction of apoptotic pathways in both pre-B and T cell ALL cells at a 50% inhibitory concentration (IC50) of low micromolar concentrations. Acetylation of α-tubulin increases within the first 30 min following treatment of ALL cells with tubacin. We also observed an accumulation of polyubiquitinated proteins and poly(ADP-ribose) polymerase (PARP) cleavage. Furthermore, the signaling pathways activated by tubacin appear to be distinct from those observed in multiple myeloma. In this article, we demonstrate that tubacin enhances the effects of chemotherapy to treat primary ALL cells in vitro and in vivo. These results suggest that targeting HDAC6 alone or in combination with chemotherapy could provide a novel approach to treat ALL.

Clinical efficacy of gene-modified stem cells in adenosine deaminase–deficient immunodeficiency

BACKGROUND. Autologous hematopoietic stem cell transplantation (HSCT) of gene-modified cells is an alternative to enzyme replacement therapy (ERT) and allogeneic HSCT that has shown clinical benefit for adenosine deaminase–deficient (ADA-deficient) SCID when combined with reduced intensity conditioning (RIC) and ERT cessation. Clinical safety and therapeutic efficacy were evaluated in a phase II study. METHODS. Ten subjects with confirmed ADA-deficient SCID and no available matched sibling or family donor were enrolled between 2009 and 2012 and received transplantation with autologous hematopoietic CD34+ cells that were modified with the human ADA cDNA (MND-ADA) &ggr;-retroviral vector after conditioning with busulfan (90 mg/m2) and ERT cessation. Subjects were followed from 33 to 84 months at the time of data analysis. Safety of the procedure was assessed by recording the number of adverse events. Efficacy was assessed by measuring engraftment of gene-modified hematopoietic stem/progenitor cells, ADA gene expression, and immune reconstitution. RESULTS. With the exception of the oldest subject (15 years old at enrollment), all subjects remained off ERT with normalized peripheral blood mononuclear cell (PBMC) ADA activity, improved lymphocyte numbers, and normal proliferative responses to mitogens. Three of nine subjects were able to discontinue intravenous immunoglobulin replacement therapy. The MND-ADA vector was persistently detected in PBMCs (vector copy number [VCN] = 0.1–2.6) and granulocytes (VCN = 0.01–0.3) through the most recent visits at the time of this writing. No patient has developed a leukoproliferative disorder or other vector-related clinical complication since transplant. CONCLUSION. These results demonstrate clinical therapeutic efficacy from gene therapy for ADA-deficient SCID, with an excellent clinical safety profile. TRIAL REGISTRATION. ClinicalTrials.gov NCT00794508. FUNDING. Food and Drug Administration Office of Orphan Product Development award, RO1 FD003005; NHLBI awards, PO1 HL73104 and Z01 HG000122; UCLA Clinical and Translational Science Institute awards, UL1RR033176 and UL1TR000124.

ABT-869 Inhibits the Proliferation of Ewing Sarcoma Cells and Suppresses Platelet-Derived Growth Factor Receptor β and c-KIT Signaling Pathways

The Ewing Sarcoma (EWS) family of tumors is one of the most common tumors diagnosed in children and adolescents and is characterized by a translocation involving the EWS gene. Despite advances in chemotherapy, the prognosis of metastatic EWS is poor with an overall survival of <30% after 5 years. EWS tumor cells express the receptor tyrosine kinases, platelet-derived growth factor receptor (PDGFR) and c-KIT. ABT-869 is a multitargeted small-molecule inhibitor that targets Fms-like tyrosine kinase-3, c-KIT, vascular endothelial growth receptors, and PDGFRs. To determine the potential therapeutic benefit of ABT-869 in EWS cells, we examined the effects of ABT-869 on EWS cell lines and xenograft mouse models. ABT-869 inhibited the proliferation of two EWS cell lines, A4573 and TC71, at an IC50 of 1.25 and 2 μmol/L after 72 h of treatment, respectively. The phosphorylation of PDGFRβ, c-KIT, and extracellular signal-regulated kinases was also inhibited. To examine the effects of ABT-869 in vivo, the drug was given to mice injected with EWS cells. We observed inhibition of growth of EWS tumor cells in a xenograft mouse model and prolonged survival in a metastatic mouse model of EWS. Therefore, our in vitro and in vivo studies show that ABT-869 inhibits proliferation of EWS cells through inhibition of PDGFRβ and c-KIT pathways. Mol Cancer Ther; 9(3); 653–60

Evaluating pharmacokinetics and pharmacodynamics of intravenous busulfan in pediatric patients receiving bone marrow transplantation

Abstract:  BU is a commonly used conditioning agent in BMT. However, it is a narrow therapeutic index drug which shows a strong correlation between AUC and both efficacy and toxicity. Studies in pediatric patients have suggested that children less than four yr of age have a greater clearance and thus lower AUC at standard adult doses. The goal of this retrospective analysis was to evaluate any age‐related pharmacokinetic and pharmacodynamic differences in pediatric patients who received BU as a conditioning agent. From 2003 to 2006, 21/77 pediatric patients who received BMT were reviewed. There were 15 males and six females with a mean age of six yr old. Diagnoses of leukemia (n = 11), Hodgkin’s lymphoma (n = 3), myelodysplastic syndrome (n = 2), and other (n = 5) were included. Sixteen patients received BU + cyclophosphamide while five patients received BU + another agent. There were 20 allogeneic and one autologous transplants among which 16 were human leukocyte antigen matched and five were mismatched. Average BU clearance in patients younger than four yr old (n = 8) was 4.1 ± 1.0 mL/min/kg vs. 3.1 ± 0.7 mL/min/kg in patients older than four yr old (n = 13) (p = 0.02). The corresponding averages for AUC were 998 ± 226 μm·min vs. 1155 ± 183 μm·min (p = 0.12). No patients younger than four yr old developed VOD while five of the older patients did (p = 0.044). There were no significant differences in terms of engraftment and acute GvHD. There were significant age‐related pharmacokinetic differences in pediatric patients less than four yr of age receiving BU for conditioning prior to BMT. There was a decrease in drug toxicity seen in these patients.

Invasive fungal infections in pediatric hematopoietic stem cell transplant patients

Abstract Background: Pediatric hematopoietic stem cell transplant (HSCT) recipients are at high risk of invasive fungal infections (IFIs). Methods: To characterize IFIs and changes in fungal organisms over time in pediatric HSCT patients, we performed a retrospective cohort study of all HSCTs performed in pediatric patients at UCLA between 1991 and 2006. Results: In all, 318 patients underwent 324 HSCT transplants over the 15-year period and 69 unique fungal infections were identified in 47 transplant patients. The overall incidence of fungal infections in HSCT recipients was 14.5%, with predominant organisms including Candida species (51%) and Aspergillus species (26%), with Candida albicans accounting for 18.8% of all fungal species. The distribution of organisms over time demonstrated a strong trend towards an increase in rare molds in more recent years. The respiratory tract was the main site of infection (52.6%), with urine and blood also noted as significant sites. Of all deaths in the patients with IFIs, fungal-related mortality accounted for 67.6% of deaths. Conclusions: HSCT patients have a much higher risk of fungal infections with rarer organisms becoming more prevalent, a finding likely linked to evolving antifungal practices over time. This emphasizes the need for the development and implementation of improved diagnostic, prophylactic, and therapeutic strategies to improve patient survival.

Dramatic response to rituximab in a child with severe cold autoimmune hemolytic anemia arising after allogeneic hematopoietic SCT.

Dramatic response to rituximab in a child with severe cold autoimmune hemolytic anemia arising after allogeneic hematopoietic SCT

Inappropriate sinus tachycardia after hematopoietic stem cell transplantation.

SUMMARY Survivors of childhood cancer and hematopoietic stem cell transplantation are subject to a range of cardiac late effects including conduction abnormalities. We present, the case of a 12-year-old girl diagnosed with inappropriate sinus tachycardia after a history of multiply relapsed acute lymphoblastic leukemia and an exposure history including anthracycline-based chemotherapy followed by the total body irradiation in the context of a hematopoietic stem cell transplant. The clinical importance of this late cardiac effect along with the potential options for therapy is discussed.