Publications by authors named "Liam C Lee"

8 Publications

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Paediatric Burkitt lymphoma patient-derived xenografts capture disease characteristics over time and are a model for therapy.

Br J Haematol 2021 01 3;192(2):354-365. Epub 2020 Sep 3.

Division of Cellular and Molecular Pathology, Department of Pathology, University of Cambridge, Cambridge, UK.

Burkitt lymphoma (BL) accounts for almost two-thirds of all B-cell non-Hodgkin lymphoma (B-NHL) in children and adolescents and is characterised by a MYC translocation and rapid cell turnover. Intensive chemotherapeutic regimens have been developed in recent decades, including the lymphomes malins B (LMB) protocol, which have resulted in a survival rate in excess of 90%. Recent clinical trials have focused on immunochemotherapy, with the addition of rituximab to chemotherapeutic backbones, showing encouraging results. Despite these advances, relapse and refractory disease occurs in up to 10% of patients and salvage options for these carry a dismal prognosis. Efforts to better understand the molecular and functional characteristics driving relapse and refractory disease may help improve this prognosis. This study has established a paediatric BL patient-derived xenograft (PDX) resource which captures and maintains tumour heterogeneity, may be used to better characterise tumours and identify cell populations responsible for therapy resistance.
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http://dx.doi.org/10.1111/bjh.17043DOI Listing
January 2021

IL10RA modulates crizotinib sensitivity in NPM1-ALK+ anaplastic large cell lymphoma.

Blood 2020 10;136(14):1657-1669

Department of Pediatric Hematology and Oncology, University Hospital Hamburg-Eppendorf, Hamburg, Germany.

Anaplastic large cell lymphoma (ALCL) is a T-cell malignancy predominantly driven by a hyperactive anaplastic lymphoma kinase (ALK) fusion protein. ALK inhibitors, such as crizotinib, provide alternatives to standard chemotherapy with reduced toxicity and side effects. Children with lymphomas driven by nucleophosmin 1 (NPM1)-ALK fusion proteins achieved an objective response rate to ALK inhibition therapy of 54% to 90% in clinical trials; however, a subset of patients progressed within the first 3 months of treatment. The mechanism for the development of ALK inhibitor resistance is unknown. Through genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) activation and knockout screens in ALCL cell lines, combined with RNA sequencing data derived from ALK inhibitor-relapsed patient tumors, we show that resistance to ALK inhibition by crizotinib in ALCL can be driven by aberrant upregulation of interleukin 10 receptor subunit alpha (IL10RA). Elevated IL10RA expression rewires the STAT3 signaling pathway, bypassing otherwise critical phosphorylation by NPM1-ALK. IL-10RA expression does not correlate with response to standard chemotherapy in pediatric patients, suggesting that a combination of crizotinib and chemotherapy could prevent ALK inhibitor resistance-specific relapse.
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http://dx.doi.org/10.1182/blood.2019003793DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7530646PMC
October 2020

The targetable kinase PIM1 drives ALK inhibitor resistance in high-risk neuroblastoma independent of MYCN status.

Nat Commun 2019 11 28;10(1):5428. Epub 2019 Nov 28.

Division of Cellular and Molecular Pathology, Department of Pathology, University of Cambridge, Lab Block level 3, Box 231, Cambridge Biomedical Campus, Cambridge, CB2 0QQ, UK.

Resistance to anaplastic lymphoma kinase (ALK)-targeted therapy in ALK-positive non-small cell lung cancer has been reported, with the majority of acquired resistance mechanisms relying on bypass signaling. To proactively identify resistance mechanisms in ALK-positive neuroblastoma (NB), we herein employ genome-wide CRISPR activation screens of NB cell lines treated with brigatinib or ceritinib, identifying PIM1 as a putative resistance gene, whose high expression is associated with high-risk disease and poor survival. Knockdown of PIM1 sensitizes cells of differing MYCN status to ALK inhibitors, and in patient-derived xenografts of high-risk NB harboring ALK mutations, the combination of the ALK inhibitor ceritinib and PIM1 inhibitor AZD1208 shows significantly enhanced anti-tumor efficacy relative to single agents. These data confirm that PIM1 overexpression decreases sensitivity to ALK inhibitors in NB, and suggests that combined front-line inhibition of ALK and PIM1 is a viable strategy for the treatment of ALK-positive NB independent of MYCN status.
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http://dx.doi.org/10.1038/s41467-019-13315-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6883072PMC
November 2019

MAP kinase and autophagy pathways cooperate to maintain RAS mutant cancer cell survival.

Proc Natl Acad Sci U S A 2019 03 1;116(10):4508-4517. Epub 2019 Feb 1.

Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892;

Oncogenic mutations in the small GTPase KRAS are frequently found in human cancers, and, currently, there are no effective targeted therapies for these tumors. Using a combinatorial siRNA approach, we analyzed a panel of mutant colorectal and pancreatic cancer cell lines for their dependency on 28 gene nodes that represent canonical RAS effector pathways and selected stress response pathways. We found that RAF node knockdown best differentiated mutant and WT cancer cells, suggesting RAF kinases are key oncoeffectors for addiction. By analyzing all 376 pairwise combination of these gene nodes, we found that cotargeting the RAF, RAC, and autophagy pathways can improve the capture of dependency better than targeting RAF alone. In particular, codepletion of the oncoeffector kinases BRAF and CRAF, together with the autophagy E1 ligase ATG7, gives the best therapeutic window between mutant cells and normal, untransformed cells. Distinct patterns of RAS effector dependency were observed across mutant cell lines, indicative of heterogeneous utilization of effector and stress response pathways in supporting KRAS addiction. Our findings revealed previously unappreciated complexity in the signaling network downstream of the oncogene and suggest rational target combinations for more effective therapeutic intervention.
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http://dx.doi.org/10.1073/pnas.1817494116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6410784PMC
March 2019

One-step immortalization of primary human airway epithelial cells capable of oncogenic transformation.

Cell Biosci 2016 11;6:57. Epub 2016 Nov 11.

Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, NIH., Bethesda, MD USA.

Background: The ability to transform normal human cells into cancer cells with the introduction of defined genetic alterations is a valuable method for understanding the mechanisms of oncogenesis. Easy establishment of immortalized but non-transformed human cells from various tissues would facilitate these genetic analyses.

Results: We report here a simple, one-step immortalization method that involves retroviral vector mediated co-expression of the human telomerase protein and a shRNA targeting the gene locus. We demonstrate that this method could successfully immortalize human small airway epithelial cells while maintaining their chromosomal stability. We further showed that these cells retain p53 activity and can be transformed by the oncogene.

Conclusions: Our method simplifies the immortalization process and is broadly applicable for establishing immortalized epithelial cell lines from primary human tissues for cancer research.
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http://dx.doi.org/10.1186/s13578-016-0122-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5106784PMC
November 2016

Oncogenesis driven by the Ras/Raf pathway requires the SUMO E2 ligase Ubc9.

Proc Natl Acad Sci U S A 2015 Apr 24;112(14):E1724-33. Epub 2015 Mar 24.

Laboratory of Cancer Biology and Genetics, Center for Cancer Research,

The small GTPase KRAS is frequently mutated in human cancer and currently there are no targeted therapies for KRAS mutant tumors. Here, we show that the small ubiquitin-like modifier (SUMO) pathway is required for KRAS-driven transformation. RNAi depletion of the SUMO E2 ligase Ubc9 suppresses 3D growth of KRAS mutant colorectal cancer cells in vitro and attenuates tumor growth in vivo. In KRAS mutant cells, a subset of proteins exhibit elevated levels of SUMOylation. Among these proteins, KAP1, CHD1, and EIF3L collectively support anchorage-independent growth, and the SUMOylation of KAP1 is necessary for its activity in this context. Thus, the SUMO pathway critically contributes to the transformed phenotype of KRAS mutant cells and Ubc9 presents a potential target for the treatment of KRAS mutant colorectal cancer.
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http://dx.doi.org/10.1073/pnas.1415569112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4394293PMC
April 2015

Development of siRNA payloads to target KRAS-mutant cancer.

Cancer Discov 2014 Oct 6;4(10):1182-1197. Epub 2014 Aug 6.

Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.

Unlabelled: RNAi is a powerful tool for target identification and can lead to novel therapies for pharmacologically intractable targets such as KRAS. RNAi therapy must combine potent siRNA payloads with reliable in vivo delivery for efficient target inhibition. We used a functional "Sensor" assay to establish a library of potent siRNAs against RAS pathway genes and to show that they efficiently suppress their targets at low dose. This reduces off-target effects and enables combination gene knockdown. We administered Sensor siRNAs in vitro and in vivo and validated the delivery of KRAS siRNA alone and siRNA targeting the complete RAF effector node (A/B/CRAF) as promising strategies to treat KRAS-mutant colorectal cancer. We further demonstrate that improved therapeutic efficacy is achieved by formulating siRNA payloads that combine both single-gene siRNA and node-targeted siRNAs (KRAS + PIK3CA/B). The customizable nature of Sensor siRNA payloads offers a universal platform for the combination target identification and development of RNAi therapeutics.

Significance: To advance RNAi therapy for KRAS-mutant cancer, we developed a validated siRNA library against RAS pathway genes that enables combination gene silencing. Using an in vivo model for real-time siRNA delivery tracking, we show that siRNA-mediated inhibition of KRAS as well as RAF or PI3K combinations can impair KRAS-mutant colorectal cancer in xenograft models.
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http://dx.doi.org/10.1158/2159-8290.CD-13-0900DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4184972PMC
October 2014

CRIPTO1 expression in EGFR-mutant NSCLC elicits intrinsic EGFR-inhibitor resistance.

J Clin Invest 2014 Jul 9;124(7):3003-15. Epub 2014 Jun 9.

The majority of non-small cell lung cancer (NSCLC) patients harbor EGFR-activating mutations that can be therapeutically targeted by EGFR tyrosine kinase inhibitors (EGFR-TKI), such as erlotinib and gefitinib. Unfortunately, a subset of patients with EGFR mutations are refractory to EGFR-TKIs. Resistance to EGFR inhibitors reportedly involves SRC activation and induction of epithelial-to-mesenchymal transition (EMT). Here, we have demonstrated that overexpression of CRIPTO1, an EGF-CFC protein family member, renders EGFR-TKI-sensitive and EGFR-mutated NSCLC cells resistant to erlotinib in culture and in murine xenograft models. Furthermore, tumors from NSCLC patients with EGFR-activating mutations that were intrinsically resistant to EGFR-TKIs expressed higher levels of CRIPTO1 compared with tumors from patients that were sensitive to EGFR-TKIs. Primary NSCLC cells derived from a patient with EGFR-mutated NSCLC that was intrinsically erlotinib resistant were CRIPTO1 positive, but gained erlotinib sensitivity upon loss of CRIPTO1 expression during culture. CRIPTO1 activated SRC and ZEB1 to promote EMT via microRNA-205 (miR-205) downregulation. While miR-205 depletion induced erlotinib resistance, miR-205 overexpression inhibited CRIPTO1-dependent ZEB1 and SRC activation, restoring erlotinib sensitivity. CRIPTO1-induced erlotinib resistance was directly mediated through SRC but not ZEB1; therefore, cotargeting EGFR and SRC synergistically attenuated growth of erlotinib-resistant, CRIPTO1-positive, EGFR-mutated NSCLC cells in vitro and in vivo, suggesting that this combination may overcome intrinsic EGFR-inhibitor resistance in patients with CRIPTO1-positive, EGFR-mutated NSCLC.
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http://dx.doi.org/10.1172/JCI73048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4071378PMC
July 2014