The desmoplastic reaction of pancreas cancer may begin as a wound healing response to the nascent neoplasm, but it soon creates an insidious shelter that can sustain the growing tumor and rebuff ...therapy. Among the many cell types subverted by transformed epithelial cells, fibroblasts are recruited and activated to lay a foundation of extracellular matrix proteins and glycosaminoglycans that alter tumor biophysics and signaling. Their near-universal presence in pancreas cancer and ostensible support of disease progression make fibroblasts attractive therapeutic targets. More recently, however, it has also become apparent that diverse subpopulations of fibroblasts with distinct phenotypes and secretomes inhabit the stroma, and that targeted depletion of particular fibroblast subsets could either provide substantial therapeutic benefit or accelerate disease progression. An improved characterization of these fibroblast subtypes, along with their potential relationships to tumor subtypes and mutational repertoires, is needed in order to make anti-fibroblast therapies clinically viable.
Kinase inhibitors have limited success in cancer treatment because tumors circumvent their action. Using a quantitative proteomics approach, we assessed kinome activity in response to MEK inhibition ...in triple-negative breast cancer (TNBC) cells and genetically engineered mice (GEMMs). MEK inhibition caused acute ERK activity loss, resulting in rapid c-Myc degradation that induced expression and activation of several receptor tyrosine kinases (RTKs). RNAi knockdown of ERK or c-Myc mimicked RTK induction by MEK inhibitors, and prevention of proteasomal c-Myc degradation blocked kinome reprogramming. MEK inhibitor-induced RTK stimulation overcame MEK2 inhibition, but not MEK1 inhibition, reactivating ERK and producing drug resistance. The C3Tag GEMM for TNBC similarly induced RTKs in response to MEK inhibition. The inhibitor-induced RTK profile suggested a kinase inhibitor combination therapy that produced GEMM tumor apoptosis and regression where single agents were ineffective. This approach defines mechanisms of drug resistance, allowing rational design of combination therapies for cancer.
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► Inhibition of the MEK-ERK pathway rapidly reprograms kinome activity in tumors ► c-Myc degradation induces expression and activation of receptor tyrosine kinases ► Receptor tyrosine kinase activation overcomes MEK inhibition, causing resistance ► Kinome inhibitor response profiling rationally predicts combination therapies
Chemical proteomics reveals the dynamic rewiring of kinase networks in response to treatment with MEK inhibitors, uncovering how resistance to these inhibitors emerges in tumors and highlighting combination therapy approaches for bypassing drug resistance.
For the majority of patients with pancreas cancer, the high metastatic proclivity is life limiting. Some patients, however, present with and succumb to locally destructive disease. A molecular ...understanding of these distinct disease manifestations can critically inform patient management. Using genetically engineered mouse models, we show that heterozygous mutation of Dpc4/Smad4 attenuates the metastatic potential of KrasG12D/+;Trp53R172H/+ pancreatic ductal adenocarcinomas while increasing their proliferation. Subsequent loss of heterozygosity of Dpc4 restores metastatic competency while further unleashing proliferation, creating a highly lethal combination. Expression levels of Runx3 respond to and combine with Dpc4 status to coordinately regulate the balance between cancer cell division and dissemination. Thus, Runx3 serves as both a tumor suppressor and promoter in slowing proliferation while orchestrating a metastatic program to stimulate cell migration, invasion, and secretion of proteins that favor distant colonization. These findings suggest a model to anticipate likely disease behaviors in patients and tailor treatment strategies accordingly.
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•DPC4 dosage regulates RUNX3 levels in a biphasic manner•RUNX3 controls the balance between local growth and dissemination in PDA•RUNX3 functions as both tumor suppressor and tumor promoter in PDA•DPC4 and RUNX3 levels can jointly inform clinical decision making
RUNX3 regulates the balance between proliferation and dissemination of pancreas cancer cells in response to gene dosage of DPC4/SMAD4. Evaluating the levels of DPC4 and RUNX3 can help predict patterns of relapse and response to therapies in patients, informing clinical disease management.
The dynamic nature of the kinome Graves, Lee M; Duncan, James S; Whittle, Martin C ...
Biochemical journal,
02/2013, Letnik:
450, Številka:
1
Journal Article
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Odprti dostop
Recent advances in proteomics have facilitated the analysis of the kinome 'en masse'. What these studies have revealed is a surprisingly dynamic network of kinase responses to highly selective kinase ...inhibitors, thereby illustrating the complex biological responses to these small molecules. Moreover these studies have identified key transcription factors, such as c-Myc and FOXO (forkhead box O), that play pivotal roles in kinome reprogramming in cancer cells. Since many kinase inhibitors fail despite a high efficacy of blocking their intended targets, elucidating kinome changes at a more global level will be essential to understanding the mechanisms of kinase inhibitor pharmacology. The development of technologies to study the kinome, as well as examples of kinome resilience and reprogramming, will be discussed in the present review.
The RUNX family transcription factors are critical regulators of development and frequently dysregulated in cancer. RUNX3, the least well characterized of the three family members, has been variously ...described as a tumor promoter or suppressor, sometimes with conflicting results and opinions in the same cancer and likely reflecting a complex role in oncogenesis. We recently identified RUNX3 expression as a crucial determinant of the predilection for pancreatic ductal adenocarcinoma (PDA) cells to proliferate locally or promulgate throughout the body. High RUNX3 expression induces the production and secretion of soluble factors that support metastatic niche construction and stimulates PDA cells to migrate and invade, while simultaneously suppressing proliferation through increased expression of cell cycle regulators such as CDKN1A/p21
. RUNX3 expression and function are coordinated by numerous transcriptional and post-translational inputs, and interactions with diverse cofactors influence whether the resulting RUNX3 complexes enact tumor suppressive or tumor promoting programs. Understanding these exquisitely context-dependent tumor cell behaviors has the potential to inform clinical decision-making including the most appropriate timing and sequencing of local vs. systemic therapies.
Protein kinases play key roles in oncogenic signaling and are a major focus in the development of targeted cancer therapies. Imatinib, a BCR-Abl tyrosine kinase inhibitor, is a successful front-line ...treatment for chronic myelogenous leukemia (CML). However, resistance to imatinib may be acquired by BCR-Abl mutations or hyperactivation of Src family kinases such as Lyn. We have used multiplexed kinase inhibitor beads (MIBs) and quantitative mass spectrometry (MS) to compare kinase expression and activity in an imatinib-resistant (MYL-R) and -sensitive (MYL) cell model of CML. Using MIB/MS, expression and activity changes of over 150 kinases were quantitatively measured from various protein kinase families. Statistical analysis of experimental replicates assigned significance to 35 of these kinases, referred to as the MYL-R kinome profile. MIB/MS and immunoblotting confirmed the over-expression and activation of Lyn in MYL-R cells and identified additional kinases with increased (MEK, ERK, IKKα, PKCβ, NEK9) or decreased (Abl, Kit, JNK, ATM, Yes) abundance or activity. Inhibiting Lyn with dasatinib or by shRNA-mediated knockdown reduced the phosphorylation of MEK and IKKα. Because MYL-R cells showed elevated NF-κB signaling relative to MYL cells, as demonstrated by increased IκBα and IL-6 mRNA expression, we tested the effects of an IKK inhibitor (BAY 65-1942). MIB/MS and immunoblotting revealed that BAY 65-1942 increased MEK/ERK signaling and that this increase was prevented by co-treatment with a MEK inhibitor (AZD6244). Furthermore, the combined inhibition of MEK and IKKα resulted in reduced IL-6 mRNA expression, synergistic loss of cell viability and increased apoptosis. Thus, MIB/MS analysis identified MEK and IKKα as important downstream targets of Lyn, suggesting that co-targeting these kinases may provide a unique strategy to inhibit Lyn-dependent imatinib-resistant CML. These results demonstrate the utility of MIB/MS as a tool to identify dysregulated kinases and to interrogate kinome dynamics as cells respond to targeted kinase inhibition.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Targeting the dysregulated BRAF-MEK-ERK pathway in cancer has increasingly emerged in clinical trial design. Despite clinical responses in specific cancers using inhibitors targeting BRAF and MEK, ...resistance develops often involving nongenomic adaptive bypass mechanisms. Inhibition of MEK1/2 by trametinib in patients with triple-negative breast cancer (TNBC) induced dramatic transcriptional responses, including upregulation of receptor tyrosine kinases (RTK) comparing tumor samples before and after one week of treatment. In preclinical models, MEK inhibition induced genome-wide enhancer formation involving the seeding of BRD4, MED1, H3K27 acetylation, and p300 that drives transcriptional adaptation. Inhibition of the P-TEFb-associated proteins BRD4 and CBP/p300 arrested enhancer seeding and RTK upregulation. BRD4 bromodomain inhibitors overcame trametinib resistance, producing sustained growth inhibition in cells, xenografts, and syngeneic mouse TNBC models. Pharmacologic targeting of P-TEFb members in conjunction with MEK inhibition by trametinib is an effective strategy to durably inhibit epigenomic remodeling required for adaptive resistance.
Widespread transcriptional adaptation to pharmacologic MEK inhibition was observed in TNBC patient tumors. In preclinical models, MEK inhibition induces dramatic genome-wide modulation of chromatin, in the form of
enhancer formation and enhancer remodeling. Pharmacologic targeting of P-TEFb complex members at enhancers is an effective strategy to durably inhibit such adaptation.
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Anticancer drug development is inefficient, but genetically engineered murine models (GEMM) and orthotopic, syngeneic transplants (OST) of cancer may offer advantages to in vitro and xenograft ...systems.
We assessed the activity of 16 treatment regimens in a RAS-driven, Ink4a/Arf-deficient melanoma GEMM. In addition, we tested a subset of treatment regimens in three breast cancer models representing distinct breast cancer subtypes: claudin-low (T11 OST), basal-like (C3-TAg GEMM), and luminal B (MMTV-Neu GEMM).
Like human RAS-mutant melanoma, the melanoma GEMM was refractory to chemotherapy and single-agent small molecule therapies. Combined treatment with AZD6244 mitogen-activated protein-extracellular signal-regulated kinase kinase (MEK) inhibitor and BEZ235 dual phosphoinositide-3 kinase (PI3K)/mammalian target of rapamycin (mTOR) inhibitor was the only treatment regimen to exhibit significant antitumor activity, showed by marked tumor regression and improved survival. Given the surprising activity of the "AZD/BEZ" combination in the melanoma GEMM, we next tested this regimen in the "claudin-low" breast cancer model that shares gene expression features with melanoma. The AZD/BEZ regimen also exhibited significant activity in this model, leading us to testing in even more diverse GEMMs of basal-like and luminal breast cancer. The AZD/BEZ combination was highly active in these distinct breast cancer models, showing equal or greater efficacy compared with any other regimen tested in studies of over 700 tumor-bearing mice. This regimen even exhibited activity in lapatinib-resistant HER2(+) tumors.
These results show the use of credentialed murine models for large-scale efficacy testing of diverse anticancer regimens and predict that combinations of PI3K/mTOR and MEK inhibitors will show antitumor activity in a wide range of human malignancies.
TP53 and the TGFβ pathway are major mediators of pancreatic cancer metastasis. The mechanisms by which they cause hematogenous metastasis have not been fully explored.
(
;
;
) mice were generated, ...and the frequency and morphology of organ-specific hematogenous metastases compared with that seen in
and
littermates (
). Key findings were validated in primary cells from each genotype and samples of human pancreatic cancer liver metastases.
The frequency of hematogenous metastasis in
mice was significantly lower than for
mice (41% vs. 68%,
< 0.05), largely due to a reduction in liver metastases. No differences were found between
and
lung metastases, whereas liver metastases in
mice showed a profound extravasation deficiency characterized by sinusoidal growth and lack of desmoplastic stroma. Analogous findings were confirmed in liver samples from patients indicating their clinical relevance. Portal vein colonization as a direct mode of access to the liver was observed in both mice and humans. Secretome analyses of
cells revealed an abundance of secreted prometastatic mediators including Col6A1 and Lcn2 that promoted early steps of metastatic colonization. These mediators were overexpressed in primary tumors but not metastases, suggesting that the ability to colonize is, in part, developed within the primary site, a phenomenon we refer to as the "Cinderella effect."
These findings establish a novel paradigm for understanding pancreatic cancer metastasis and the observed clinical latencies of liver versus lung metastases specifically.
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