PO-145 ERK5 pathway inhibitors inhibit the maintenance of chronic myeloid leukaemia stem cells

Introduction Chronic myeloid leukaemia (CML) is a hematopoietic stem cell (HSC)-driven neoplasia characterised by the expression of the constitutively active tyrosine kinase BCR/ABL. CML therapy based on tyrosine kinase inhibitors (TKi) is highly effective in inducing remission but not in targeting leukaemia stem cells (LSC), which sustain the minimal residual disease and are responsible for CML relapse following discontinuation of treatment. Our aim was to address the effects of the inhibition of the ERK5 pathway on the maintenance of CML LSC. Material and methods KCL22 and K562 CML cell lines, patient-derived CML cells or CD34 +peripheral blood cells from healthy donors (informed consent) were incubated in normoxic or hypoxic (0.1% O 2 ) primary cultures (LC1) in the presence or the absence of drugs. At the end of incubation (day 7), cells were analysed on a flow cytometer to determine the expression of stem cell markers or transferred to drug-free normoxic secondary cultures (LC2) to measure LC2 repopulation as a read-out of progenitor/stem cell potential (CRA assay). In the serial Colony Formation Ability (CFA) assay colonies were scored on day 7 of each passage (III passages). In the Long-Term Culture-Initiating Cells (LTC-IC) assay the number of colonies was scored after 14 days. Compounds: XMD8-92 (ERK5 inhibitor) and BIX02189 (MEK5 inhibitor); imatinib and dasatinib (BCR/ABL inhibitors). Results and discussions In CML patient-derived cells and cell lines, we found that the MEK5/ERK5 pathway is active and necessary for optimal proliferation in low oxygen, a condition typical of normal hematopoietic and leukemic stem cell niches. Treatment of primary CML cells with XMD8-92 or BIX02189, but not with TKi, strikingly reduced Culture Repopulation Ability (CRA), serial Colony Formation Ability and Long-Term Culture-Initiating Cells (LTC-IC). Importantly, inhibition of MEK5/ERK5 was effective on CML cells regardless of the presence or absence of imatinib (IM), and did not reduce CRA or LTC-IC of normal CD34 +cells. Interestingly, in hypoxia, combined treatment XMD8-92/IM decreased the expression of genes relevant for stem cell maintenance such as c-MYC, SOX2 and NANOG and the expression of CD26, a CML LSC marker. Conclusion We propose ERK5 pathway inhibitors as a novel therapeutic approach to prevent CML relapse and, in combination with TKi, enhance induction of remission

The Role of Alpha 6 Integrin in Prostate Cancer Migration and Bone Pain in a Novel Xenograft Model

Of the estimated 565,650 people in the U.S. who will die of cancer in 2008, almost all will have metastasis. Breast, prostate, kidney, thyroid and lung cancers metastasize to the bone. Tumor cells reside within the bone using integrin type cell adhesion receptors and elicit incapacitating bone pain and fractures. In particular, metastatic human prostate tumors express and cleave the integrin A6, a receptor for extracellular matrix components of the bone, i.e., laminin 332 and laminin 511. More than 50% of all prostate cancer patients develop severe bone pain during their remaining lifetime. One major goal is to prevent or delay cancer induced bone pain. We used a novel xenograft mouse model to directly determine if bone pain could be prevented by blocking the known cleavage of the A6 integrin adhesion receptor. Human tumor cells expressing either the wildtype or mutated A6 integrin were placed within the living bone matrix and 21 days later, integrin expression was confirmed by RT-PCR, radiographs were collected and behavioral measurements of spontaneous and evoked pain performed. All animals independent of integrin status had indistinguishable tumor burden and developed bone loss 21 days after surgery. A comparison of animals containing the wild type or mutated integrin revealed that tumor cells expressing the mutated integrin resulted in a dramatic decrease in bone loss, unicortical or bicortical fractures and a decrease in the ability of tumor cells to reach the epiphyseal plate of the bone. Further, tumor cells within the bone expressing the integrin mutation prevented cancer induced spontaneous flinching, tactile allodynia, and movement evoked pain. Preventing A6 integrin cleavage on the prostate tumor cell surface decreased the migration of tumor cells within the bone and the onset and degree of bone pain and fractures. These results suggest that strategies for blocking the cleavage of the adhesion receptors on the tumor cell surface can significantly prevent cancer induced bone pain and slow disease progression within the bone. Since integrin cleavage is mediated by Urokinase-type Plasminogen Activator (uPA), further work is warranted to test the efficacy of uPA inhibitors for prevention or delay of cancer induced bone pain