Group Leaders: Assoc. Prof. Timothy P Hughes; Dr A. Bruce Lyons
Staff: Deb White, Verity Saunders
Students: Andrea Dewar, Pongtep Viboonjuntra, Sarah Moore
The focus of the Melissa White Laboratory is the cell and molecular biology of Chronic Myeloid Leukaemia, with specific emphasis on responses to the new therapeutic agent imatinib (Glivec), and the development of mutations which cause resistance to this drug. By furthering our understanding of leukaemic cell responses to imatinib, we hope to gain insight into how therapy can be improved.
1. Cytokine signalling may protect Chronic Myeloid Leukaemia progenitors from the action of imatinib. (with Prof. AF Lopez, Human Immunology)
As a number of cytokine signalling pathways share similarities with the pathways used by Bcr-Abl, the oncogenic fusion protein of CML, we wished to examine if cytokines could replace the siganlling of Bcr-Abl when its activity was blocked by imatinib. Our findings indicate that GM-CSF can protect CML progenitors from the anti-proliferative and anti-survival effects of this new drug. Using specific cytokine antagonists, we can demonstrate that blocking cytokine activity is a potentially useful combination therapy which may enhance the efficacy of imatinib.
2. The development of muations engendering resistance to imatinib. (with S Branford, Molecular Pathology)
Although imatinib has been extremely effective clinically, there has been significant experience of mutation in the Bcr-Abl gene which leads to resistance to the drug. We are currently examining development of mutations to gain insight into this process, including if imatinib selects from a pre-existing pool of mutations. One unexpected finding was that mutations in the P- loop region of the Bcr-Abl tyrosine kinase was associated with very poor prognosis.
3. Does initial responsiveness to imatinib have prognostic significance?
We have been examining intrinsic differences in the response of leukaemic cells to the drug at presentation, using Western Blotting to examine the inhibition of the tyrosine kinase activity of Bcr-ABL, which is its key leukaemogenic property. Surprisingly, we see a relatively wide variation in the IC50 (the drug concentration required to halve the enzymic activity of Bcr-Abl) of the drug for individual cases of CML. Our preliminary studies suggest that the IC50 is predictive of the rate of the response of patients to imatinib. We are currently gathering data to determine if this has long term prognostic value, which may provide a means for identifying patients requiring more intensive therapy.
4. Imatinib (Glivec) inhibits the growth and differentiation of normal human monocytes.
Even though imatinib has specificity for the causative tyrosine kinase of chronic myeloid leukaemia, there is potential for this agent to affect normal cells by inhibiting other kinases. Our recent studies show that imatinib inhibits growth of monocyte/macrophage colonies from normal bone marrow progenitors, and prevents monocytes from peripheral blood maturing into macrophages. These effects are not thought to be due to inhibition of known tyrosine kinases. These findings suggest that imatinib or closely related derivatives may be useful for modifying immune responses or in the treatment of conditions having a significant role for macrophages, such as rheumatoid arthritis.
2004 : $30,000 "The role of DNA repair genes in genetic instability and progression of CML"
IMVS Contributing Haematologists Committee,
S Branford, TP Hughes, AB Lyons
2003 : $19,914 "Genetic instability in Chronic Myeloid Leukaemia cells may predict early loss of response to chemotherapeutic agents"
RAH/IMVS Research Committee,
AB Lyons, TP Hughes
2003-4 : $122,796 "Analysis of the biology of mutant forms of BCR/ABL resistant to the tyrosine kinase inhibitor imatinib (Glivec)"
Cancer Council of South Australia,
AB Lyons, TP Hughes
2002 : $51,606 "Dissecting the aberrant differentiation program of leukaemic CD34+ progenitor cells"
Anti-Cancer Foundation of South Australia,
AB Lyons
2002 : $87,687 "Potential enhancement of the activity of STI571 on CML progenitors, using the cytokine receptor antagonist E21R"
British Biotech.
AB Lyons, AF Lopez, TP Hughes
2002 : $99,000 "Assessing the effect of PI-88 on Human haemopoietic engraftment into sub-lethally irradiated NOD/SCID mice"
Progen Industries.
I Lewis, TP Hughes, AB Lyons
2000-1 : $96,000 "Enhancing the activity of the ABL-specific tyrosine kinase inhibitor STI571 on the leukaemic stem cells in CML"
Anti-Cancer Foundation of South Australia.
TP Hughes, AB Lyons (as associate)
Lewis ID, McDiarmid LA, Samels LM, To LB, Hughes TP. (1998) Establishment of a reproducible model of chronic phase chronic myeloid leukemia in NOD/SCID mice using blood derived mononuclear or CD34 positive cells. Blood 91: 630-640.
Moore S, Haylock DN, Levesque J-P, McDiarmid L, Samels L, To LB, Simmons PJ, Hughes TP. (1998) Stem cell factor as a single agent induces marked proliferation of the Philadelphia chromosome positive fraction of CML CD34+ cells. Blood 92: 2461-2470
AB Lyons (1999) Divided we stand: tracking cell proliferation with CFSE. Immunol. Cell Biol. 77, 509-515
Branford S, Hughes TP, Rudzki Z. (1999) Monitoring chronic myeloid leukaemia therapy by real time quantitative PCR in blood is a reliable alternative to bone marrow cytogenetics. Br J Haematol 107: 587-599.
Grigg AP, Szer J, Beresford J, Dodds A, Bradstock K, Durrant S, Schwarer AP, Hughes TP, Herrmann R, Gibson J, Arthur C, Mathews J. Factors affecting the outcome of allogeneic bone marrow transplantation for adult patients with refractory or relapsed acute leukaemia. (1999) Br J Haematol, 107: 409-18
AB Lyons (2000) Analysing lymphocyte cell division using CFSE dye dilution. J. Immunol. Meths. 243, 147-154
Moore S, McDiarmid L, Hughes TP. (2000) Stem cell factor and chronic myeloid leukaemia. Leukaemia and Lymphoma 38:211-220
Branford S, Rudzki Z, Hughes TP. (2000) A novel BCR-ABL transcript (e8a2) with the inverted sequence of ABL intron 1b in a patient with Philadelphia-positive chronic myeloid leukaemia. (2000) Br J Haematol, 109: 635-637.
Szabo F, Horvath N, Seimon S, Hughes TP. (2000) Inappropriate antidiuretic hormone secretion, abdominal pain and disseminated varicella-zoster virus infection: an unusual triad in a patient 6 months post mini-allogeneic peripheral stem cell transplant for chronic myeloid leukaemia. Bone Marrow Transplant, 26:231-3
Craddock C, Bardy P, Kreiter S, Johnston R, Apperley J, Marks D, Huber C, Kolbe K, Goulding R, Lawler M, Goldman J, Hughes TP, Derigs G. (2000) Engraftment of T-depleted allogeneic haematopoietic stem cells using a reduced intensity conditioning regimen. British Journal of Haematology 111, 797-800
PD Hodgkin and AB Lyons (2000) Tracking cell division using CFSE. Today’s Life Science 12, 36-38
Hui CH, Bardy P, Hughes TP, Horvath N, To LB. (2001) Successful salvage of RAEB/AML relapsing early post allograft with FLAG-Ida conditioned mini-allograft: a report of two cases. Clinics in Laboratory Haematology 23, 135-138.
Lutwyche JK, Keough RA, Hughes TP, Gonda TJ. (2001) Mutation screening of the c-MYB negative regulatory domain in acute and chronic myeloid leukaemia. British Journal of Haematology 114:632-634, 2001.
A Dewar, KV Doherty, GM Woods, AB Lyons and HK Muller (2001) Acquisition of immune function by neonatal Langerhans cells. Immunology 103, 61-69
AB Lyons, J Hasbold and PD Hodgkin (2001) Flow cytometric analysis of cell division history using dilution of CFSE, a stably integrated fluorescent probe. Methods Cell Biol. 63, 375-398.
Branford S, Hughes TP, Rudzki Z. (2002) Proposed mechanism for transcription of both b2a2 and b3a2 BCR-ABL in chronic myeloid leukemia involving polymorphism within the BCR gene. British Journal of Haematology 117:875-877.
Branford S, Rudzki Z, Walsh S, Grigg A, Arthur C, Taylor K, Herrmann R, Lynch KP, Hughes TP (2002). High Frequency of Point Mutations Clustered Within the ATP Binding Region of BCR/ABL in CML and Ph-positive ALL Patients who Develop Imatinib (STI571) Resistance. Blood 99: 3472-3475
Mullighan CG, Heatley S, Doherty K, Szabo F, Grigg A, Hughes TP, Schwarer AP, Szer J, Tait BD, Bik To L, Bardy PG. (2002) Mannose-binding lectin gene polymorphisms are associated with major infection following allogeneic hemopoietic stem cell transplantation. Blood 99:3524-9
Frost MJ, Ferrao PT, Hughes TP, Ashman LK. (2002) Juxtamemberane mutant V560Gkit is more sensitive to imatinib (STI571) compared to wild-type c-KIT whereas the kinase domain mutant D816Vkit is resistant. Molecular Cancer Therapeutics 1: 1115-1124.
R Wilkinson, AB Lyons, D Roberts, MX Wong, PA Bartley and DE Jackson (2002) PECAM-1 is essential in regulating B-cell development, B-cell antigen receptor (BCR)-mediated activation and autoimmune disease. Blood 100, 184-193
O’Brien S G, Guilhot F, Larson R A, Gathmann I, Baccarani M, Cervantes F, Cornelissen J, Fischer T, Hochhaus A, Hughes TP, Lechner K, Nielsen J, Rousselot P, Reiffers J, Saglio G, Shepherd J, Simonsson B, Gratwohl A, Goldman J, Kantarjian, Taylor K, Verhoef G, Bolton A, Capdeville R, Druker B. (2003) Imatinib compared with Interferon and Low-Dose Cytarabine for Newly Diagnosed Chronic-Phase Chronic Myeloid Leukaemia. New England Journal of Medicine: 348: 994-1004.
Hughes TP and Branford S. (2003) Molecular Monitoring of CML. Semin Haematol 40, number 2 (suppl 2) 62-68.
Hughes TP. (2003) Comments on the Hammersmith Policy Semin Haematol 40, number 2 (suppl 2) 111-112.
Branford S, Rudzki Z, Walsh S, Parkinson I, Grigg A, Szer J, Taylor K, Herrmann R, Seymour J, Arthur C, Joske D, Lynch K, Hughes TP. (2003) The detection of BCR-ABL mutations in imatinib-treated CML patients is virtually always accompanied by clinical resistance, and mutations in the ATP phosphate-binding loop (P-loop) are associated with a poor prognosis. Blood 102: 276-283.
Roberts MM, Dyson P, Rawling C, Thorp D, Rawling T, White D, Horvath N, Bardy P, Hui CH, Dart GW, To LB, Hughes TP. Selected CD34 blood cell allografts for older patients: low transplant-related mortality, graft failure and acute GvHD. Cytotherapy (2003) Vol 5, No 6, 534-541.
Ross DM, Hughes TP. (2003) Cancer treatment with kinase inhibitors: what have we learnt from imatinib? British Journal of Cancer 90: 12-19.
CH Hui, KY Goh, D White, S Branford, A Grigg, JF Seymour, YL Kwan, S Walsh, R Hoyt, A Trickett, B Rudzki, DDF Ma, LB To, TP Hughes.(2003) Successful peripheral blood stem cell mobilisation with filgrastim in patients with chronic myeloid leukaemia achieving complete cytogenetic response with imatinib, without increasing disease burden as measured by quantitative real-time PCR. Leukemia 17:821-828.
Hui CH, Hughes TP (2003) Strategies for the treatment of imatinib-resistant chronic myeloid leukaemia. Clinical Advances in Hematology and Oncology 1: 538-545.
Hughes TP. (2003) Molecular monitoring in chronic myeloid leukemia in Hematology 2003 American Society of Hematology Education Program Book.
Hughes TP, Kaeda J, Branford S, Rudzki Z, Hochhaus A, Henslee ML, Gathmann I, Bolton AE, van Hoomissen IC, Goldman JM and Radich J. (2003) Frequency of major molecular responses to imatinib or interferon alfa plus cytarabine in newly diagnosed patients with chronic myeloid leukemia. New England Journal of Medicine 2003;349:1421-30
AL Dewar, RM Domaschenz, KV Doherty, TP Hughes and AB Lyons. (2003) Imatinib inhibits the in vitro development of the monoctye/macrophage lineage from normal human bone marrow progenitors. Leukemia 17, 1713-1721
RN Morrison, AB Lyons, Nowak, B.F., Hayball, J.D., (2004). Snapper (Pagrus auratus) leucocyte proliferation is synergistically enhanced by simultaneous stimulation with LPS and PHA. Fish & Shellfish Immunology 16, 307-319.
AB Lyons and KV Doherty (2004) Flow cytometric analysis of cell division by dye dilution. In; Current protocols in cytometry, 9.11.1-9.11.10. Eds. JP Robinson et al (John Wiley and sons, New York)
Faderl S, Hochhaus A, Hughes TP (2004) Monitoring of minimal residual disease in CML North American Clinics in Hematology and Oncology in press.
Hochhaus A, Hughes TP Clinical resistance to imatinib: mechanisms and implications. (2004) North American Clinics in Hematology and Oncology in press.
Branford S, Rudzki Z, Harper A, Grigg A, Taylor K, Durrant S, Arthur C, Browett P, Schwarer AP, Ma D, Seymour JF, Bradstock K, Joske D, Lynch K, Gathmann I, Hughes TP. Imatinib produces significantly superior molecular responses compared to interferon alfa plus cytarabine in patients with newly diagnosed chronic myeloid leukemia in chronic phase. Leukemia 17; 2401-9