The present studies examined viability and DNA damage levels in mammary carcinoma cells following PARP1 and CHK1 inhibitor drug combination exposure. PARP1 inhibitors [AZD2281 ; ABT888 ; NU1025 ; AG014699] interacted with CHK1 inhibitors [UCN-01 ; AZD7762 ; LY2603618] to kill mammary carcinoma cells. PARP1 and CHK1 inhibitors interacted to increase both single strand and double strand DNA breaks that correlated with increased γH2AX phosphorylation. Treatment of cells with CHK1 inhibitors increased the phosphorylation of CHK1 and ERK1/2. Knock down of ATM suppressed the drug-induced increases in CHK1 and ERK1/2 phosphorylation and enhanced tumor cell killing by PARP1 and CHK1 inhibitors. Expression of dominant negative MEK1 enhanced drug-induced DNA damage whereas expression of activated MEK1 suppressed both the DNA damage response and tumor cell killing. Collectively our data demonstrate that PARP1 and CHK1 inhibitors interact to kill mammary carcinoma cells and that increased DNA damage is a surrogate marker for the response of cells to this drug combination.
PARP1; CHK1; DNA damage; ATM; kinase; apoptosis; comet
Interactions between the Bruton tyrosine kinase (BTK) inhibitor PCI-32765 and the proteasome inhibitor (bortezomib) were examined in diffuse large-B cell lymphoma (DLBCL) and mantle cell lymphoma (MCL) cells, including those highly resistant to bortezomib. Co-administration of PCI-32765/bortezomib synergistically increased mitochondrial injury and apoptosis in germinal centre- or activated B-cell-like-DLBCL cells and in MCL cells. These events were accompanied by marked AKT and nuclear factor (NF)-κB (NFKB1) inactivation, down-regulation of Mcl-1 (MCL1), Bcl-xL (BCL2L1), and XIAP, and enhanced DNA damage (e.g., γH2A.X formation) and endoplasmic reticulum (ER) stress. Similar interactions were observed in highly bortezomib-resistant DLBCL and MCL cells, and in primary DLBCL cells. In contrast, PCI-32765/bortezomib regimens displayed minimal toxicity toward normal CD34+ bone marrow cells. Transfection of DLBCL cells with a constitutively active AKT construct attenuated AKT inactivation and significantly diminished cell death, whereas expression of an NF-κB “super-repressor” (IκBαser34/36) increased both PCI-32765 and bortezomib lethality. Moreover, cells in which the ER stress response was disabled by a dominant-negative eIF2α construct were resistant to this regimen. Finally, combined exposure to PCI-32765 and bortezomib resulted in more pronounced and sustained reactive oxygen species (ROS) generation, and ROS scavengers significantly diminished lethality. Given promising early clinical results for PCI-32765 in DLBCL and MCL, a strategy combining BTK/ proteasome inhibitor warrants attention in these malignancies.
PCI32765; bortezomib; BTK; DLBCL; mantle cell lymphoma
Programmed cell death is well-orchestrated process regulated by multiple pro-apoptotic and anti-apoptotic genes, particularly those of the Bcl-2 gene family. These genes are well documented in cancer with aberrant expression being strongly associated with resistance to chemotherapy and radiation.
This review focuses on the resistance induced by the Bcl-2 family of anti-apoptotic proteins and current therapeutic interventions currently in preclinical or clinical trials that target this pathway. Major resistance mechanisms that are regulated by Bcl-2 family proteins and potential strategies to circumvent resistance are also examined. Although antisense and gene therapy strategies are used to nullify Bcl-2 family proteins, recent approaches use small molecule inhibitors and peptides. Structural similarity of the Bcl-2 family of proteins greatly favors development of inhibitors that target the BH3 domain, called BH3 mimetics.
Strategies to specifically identify and inhibit critical determinants that promote therapy-resistance and tumor progression represent viable approaches for developing effective cancer therapies. From a clinical perspective, pretreatment with novel, potent Bcl-2 inhibitors either alone or in combination with conventional therapies hold significant promise for providing beneficial clinical outcomes. Identifying small molecule inhibitors with broader and higher affinities for inhibiting all of the Bcl-2 pro-survival proteins will facilitate development of superior cancer therapies.
BH3 domain; apoptosis; Mcl-1; radiation resistance; chemotherapy resistance
Melanoma differentiation associated gene-9 (mda-9/syntenin) encodes an adapter scaffold protein whose expression correlates with and mediates melanoma progression and metastasis. Tumor angiogenesis represents an integral component of cancer metastasis prompting us to investigate a possible role of mda-9/syntenin in inducing angiogenesis. Genetic (gain-of-function and loss-of-function) and pharmacological approaches were employed to modify mda-9/syntenin expression in normal immortal melanocytes, early radial growth phase melanoma and metastatic melanoma cells. The consequence of modifying mda-9/syntenin expression on angiogenesis was evaluated using both in vitro and in vivo assays, including tube formation assays using human vascular endothelial cells, CAM assays and xenograft tumor animal models. Gain-of-function and loss-of-function experiments confirm that MDA-9/syntenin induces angiogenesis by augmenting expression of several pro-angiogenic factors/genes. Experimental evidence is provided for a model of angiogenesis induction by MDA-9/syntenin in which MDA-9/syntenin interacts with the ECM activating Src and FAK resulting in activation by phosphorylation of Akt, which induces HIF-1α. The HIF-1α activates transcription of Insulin Growth Factor Binding Protein-2 (IGFBP-2), which is secreted thereby promoting angiogenesis and further induces endothelial cells to produce and secrete VEGF-A augmenting tumor angiogenesis. Our studies delineate an unanticipated cell non-autonomous function of MDA-9/syntenin in the context of angiogenesis, which may directly contribute to its metastasis-promoting properties. As a result, targeting MDA-9/syntenin or its downstream-regulated molecules may provide a means of simultaneously impeding metastasis by both directly inhibiting tumor cell transformed properties (autonomous) and indirectly by blocking angiogenesis (non-autonomous).
mda-9/syntenin; melanoma; angiogenesis; IGFBP-2; HuVECs; CAM assay
It is historically well known that signaling by the PI3K-AKT and MEK1/2-ERK1/2 pathways in a cell type-dependent fashion can collaborate to maintain cell viability.1-3 Signaling pathways can also crosstalk with each other wherein one pathway can signal to either enhance or suppress signaling by another.4 Signaling by the ERK1/2 pathway can also stimulate release of growth factors which can feed back onto tumor cells to re-energize signaling pathways.5 The studies described by Toulany et al. add to this knowledge base by examining the relationship between PI3K-AKT and MEK1/2-ERK1/2 pathway signaling, EGF receptor signaling, K-RAS function, and tumor cell survival.6
K-RAS; NSCLC; HNSCC; EGFR; PI3K/Akt; MAPK/ERK; erlotinib; PI-103
Prior studies demonstrated that resistance to the ERBB1/2 inhibitor Lapatinib in HCT116 cells was mediated by increased MCL-1 expression. We examined whether inhibition of BCL-2 family function could restore Lapatinib toxicity in Lapatinib adapted tumor cells and enhance Lapatinib toxicity in naive cells. The BCL-2 family antagonist Obatoclax (GX15-070), that inhibits BCL-2/BCL-Xl/MCL-1 function, enhanced Lapatinib toxicity in parental HCT116 and Lapatinib adapted HCT116 cells. In breast cancer lines, regardless of elevated ERBB1/2 expression, GX15-070 enhanced Lapatinib toxicity within 3–12 h.The promotion of Lapatinib toxicity neither correlated with cleavage of caspase 3 nor was blocked by inhibition caspases; and was not associated with changes in the activities of ERK1/2, JNK1/2 or p38 MAPK but with reduced AKT, mTOR and S6K1 phosphorylation. The promotion of Lapatinib toxicity by GX15-070 correlated with increased cytosolic levels of apoptosis inducing factor (AIF) and expression of ATG8 (LC3), and the formation of large vesicles that intensely stained for a transfected LC3-GFP construct. Knockdown of the autophagy regulatory proteins ATG5 or Beclin1 suppressed the induction of LC3-GFP vesicularization and significantly reduced cell killing, whereas knock down of MCL-1 and BCL-Xl enhanced the induction of LC3-GFP vesicularization and significantly enhanced cell killing. Knockdown of Beclin1 and AIF abolished cell killing. Collectively, our data demonstrate that Obatoclax mediated inhibition of MCL-1 rapidly enhances Lapatinib toxicity in tumor cells via a toxic form of autophagy and via AIF release from the mitochondrion.
lapatinib; obatoclax; autophagy; cell death; resistance
We have further defined mechanism(s) by which the drug OSU-03012 (OSU) kills brain cancer cells. OSU toxicity was enhanced by the HSP90 inhibitor 17-N-Allylamino-17-demethoxygeldanamycin (17AAG) that correlated with reduced expression of ERBB1 and ERBB2. Inhibition of the extrinsic apoptosis pathway blocked the interaction between 17AAG and OSU. OSU toxicity was enhanced by the inhibitor of ERBB1/2/4, lapatinib. Knock down of ERBB1/2/4 in a cell line specific fashion promoted OSU toxicity. Combined exposure of cells to lapatinib and OSU resulted in reduced AKT and ERK1/2 activity; expression of activated forms of AKT and to a lesser extent MEK1 protected cells from the lethal effects of the drug combination. Knock down of PTEN suppressed, and expression of PTEN enhanced, the lethal interaction between OSU and lapatinib. Downstream of PTEN, inhibition of mTOR recapitulated the effects of lapatinib. Knock down of CD95, NOXA, PUMA, BIK or AIF, suppressed lapatinib and OSU toxicity. Knock down of MCL-1 enhanced, and overexpression of MCL-1 suppressed, drug combination lethality. Lapatinib and OSU interacted in vivo to suppress the growth of established tumors. Collectively our data argue that the inhibition of ERBB receptor function represents a useful way to enhance OSU lethality in brain tumor cells.
glioblastoma; medulloblastoma; lapatinib; OSU-03012; apoptosis; autophagy; ERBB1; PTEN
Sorafenib (Nexavar) is a multi-kinase inhibitor that was developed as an inhibitor of RAF-1, in the ERK1/2 pathway, but which was subsequently shown to inhibit class III tyrosine kinase receptors.1 More recently regorafenib (Stivarga) has been developed, which is a further fluorinated version of sorafenib with greater bioavailability and similar inhibitory properties against RAF-1/class III RTKs.2 Some of the anti-tumor effects of sorafenib have been ascribed to anti-angiogenic actions of this agent on endothelial associated kinases such as VEGFR2. Other effects of sorafenib clearly have to be due to its effects on the inherent biology of the tumor cells themselves. For example, through various mechanisms sorafenib has been shown in the laboratory and the clinic to suppress expression of the protective protein MCL-1.3 Sorafenib has also been linked to inhibition of STAT3, NFκB, and activation of the death receptor CD95.4 Sorafenib is routinely dosed daily (400 mg BID) and 7 d after the start of dosing has a Cmax of ~21 μM with a nadir at 12 h of ~10 μM, and is a highly protein bound based on in vitro assays.5 Despite this in vitro binding data sorafenib has profound in vivo effects on tumor cells in renal carcinoma and hepatocellular carcinoma patients; cells which are not per se addicted to high activity oncogene signals that are targets of sorafenib/regorafenib. Thus the precise stable bioavailable level of sorafenib/regorafenib in patient plasma is not known.
sorafenib; ovarian clear cell carcinoma; progression-free survival
It has been known for many years that elevated signaling by the ERK1/2 pathway is frequently associated with the growth and survival of many tumor cell types under a variety of normal and stressful conditions, including the response of cells to other cancer interventional therapeutic strategies e.g., references 1–4. There is, however, a modest significant literature showing that enhanced ERK1/2 signaling can also cause tumor cell death e.g., references 5–8. The role of ERK1/2 signaling is clearly complex, for example as shown by the Koumenis group where inhibition of radiation-induced ERK1/2 signaling caused radiosensitization, whereas inhibition of curcumin-hyper-stimulated ERK1/2 signaling reduced radiosensitivity.7 Presumably this Janus-faced behavior of the ERK1/2 pathway in terms of cell survival regulation will depend upon the tumor cell type, the intensity of ERK1/2 stimulation, and the molecular intervention/drug being used.
ERK; MKP1; camptothecin; human colon cancer
In the present study we show that histone deacetylase inhibitors (HDACIs) enhance the anti-tumor effects of melanoma differentiation associated gene-7/interleukin 24 (mda-7/IL-24) in human renal carcinoma cells. Similar data were obtained in other GU tumor cells. Combination of these two agents resulted in increased autophagy that was dependent on expression of ceramide synthase 6, with HDACIs enhancing MDA-7/IL-24 toxicity by increasing generation of ROS and Ca2+. Knock down of CD95 protected cells from HDACI and MDA-7/IL-24 lethality. Sorafenib treatment further enhanced (HDACI + MDA-7/IL-24) lethality. Anoikis resistant renal carcinoma cells were more sensitive to MDA-7/IL-24 that correlated with elevated SRC activity and tyrosine phosphorylation of CD95. We employed a recently constructed serotype 5/3 adenovirus, which is more effective than a serotype 5 virus in delivering mda-7/IL-24 to renal carcinoma cells and which conditionally replicates (CR) in tumor cells expressing MDA-7/IL-24 by virtue of placing the adenoviral E1A gene under the control of the cancer-specific promoter progression elevated gene-3 (Ad.5/3-PEG-E1A-mda-7; CRAd.5/3-mda-7, Ad.5/3-CTV), to define efficacy in renal carcinoma cells. Ad.5/3-CTV decreased the growth of renal carcinoma tumors to a significantly greater extent than did a non-replicative virus Ad.5/3-mda-7. In contralateral uninfected renal carcinoma tumors Ad.5/3-CTV also decreased the growth of tumors to a greater extent than did Ad.5/3-mda-7. In summation, our data demonstrates that HDACIs enhance MDA-7/IL-24-mediated toxicity and tumor specific adenoviral delivery and viral replication of mda-7/IL-24 is an effective pre-clinical renal carcinoma therapeutic.
MDA-7/IL-24; HDACI; ceramide; apoptosis; bystander; cytokine; ROS; caspase; animal study
To many investigators PARP1 is simply a substrate for caspase 3, and whose cleavage is thought indicative of apoptosis. However, in reality PARP1 plays a major role in the biology of the cell cycle and DNA repair.1,2 PARP1 binds to damaged DNA where it becomes enzymatically activated and ADP ribosylates itself and other proteins. PARP facilitates DNA repair complex formation, e.g., with BRCA1/2, and the activation of the cell cycle regulatory enzymes ATM and ATR.2 PARP inhibitors as a single agent have only shown any degree of efficacy in breast and ovarian cancer patients who lack BRCA1/2 function.3,4 The present studies examined PARP1 inhibitor biology in a range of triple negative and non-triple negative breast cancer cell lines.
PARP inhibitors; olaparib; iniparib; breast cancer; triple-negative; CDK1; RO-3306
The present studies focused on defining the mechanisms by which anoikis-resistant (AR) mammary carcinoma cells can be reverted to a therapy-sensitive phenotype. AR mammary carcinoma cells had reduced expression of the toxic BH3 domain proteins BAX, BAK, NOXA, and PUMA. In AR cells expression of the protective BCL-2 family proteins BCL-XL and MCL-1 was increased. AR cells were resistant to cell killing by multiple anti-tumor cell therapies, including ERBB1/2 inhibitor + MCL-1 inhibitor treatment, and had a reduced autophagic flux response to these therapies, despite similarly exhibiting increased levels of LC3II processing. Knockdown of MCL-1 and BCL-XL caused necro-apoptosis in AR cells to a greater extent than in parental cells. Pre-treatment of anoikis-resistant cells with histone deacetylase inhibitors (HDACIs) for 24 h increased the levels of toxic BH3 domain proteins, reduced MCL-1 levels, and restored/re-sensitized the cell death response of AR tumor cells to multiple toxic therapies. In vivo, pre-treatment of AR breast tumors in the brain with valproate restored the chemo-sensitivity of the tumors and prolonged animal survival. These data argue that one mechanism to enhance the anti-tumor effect of chemotherapy could be HDACI pre-treatment.
autophagy; anoikis; BH3 domain; MCL-1; ERBB1; tumor; signaling; necrosis; BAK; NOXA
The nomenclature for the serine/threonine protein phosphatases was established by Professor Sir Philip Cohen over 30 years ago.1 At that time protein phosphatase 1 was known to have two small inhibitory proteins (I-1 and I-2) and be regulated by sub-cellular location whereas no protein inhibitor had yet been discovered for the related multi-subunit phosphatase PP2A. That paradigm subsequently changed, and several PP2A protein inhibitors have been discovered.2 The protein I2PP2A (SET) is considered to be oncogenic, i.e., PP2A is a tumor suppressor, and is overexpressed in many tumor cell types (ref. 3, and refs. therein). I2PP2A also has other targets besides PP2A, e.g., DNA exonucleases and modification of histone acetylation.4 PP2A activity is known to be regulated by the bioactive lipid ceramide, and this occurs through both I2PP2A inhibition and PP2A de-repression and through ceramide actions on subunits of the PP2A enzyme complex.5,6 In the present manuscript the authors examined the expression of I2PP2A in prostate cancer and prostate epithelial cells. They determined whether ceramide could decrease accumulation of the oncogene c-Myc through inhibition of I2PP2A and activation of PP2A. As I2PP2A is also an inhibitor of histone acetylation they determined whether ceramide could block the epigenetic action of I2PP2A.
c-Myc; cell signaling; ceramide; histone acetyl transferase; inhibitor 2 of protein phosphatase 2A; protein phosphatase 2A; tumor suppressor lipid
It is now well recognized that in the vast majority of tumor types, for the approach of “kinase inhibition” to exhibit a significant effect, whether the data are from an in vitro assay, an animal model or the clinic, requires that multiple complementary kinases be simultaneously inhibited. This combined inhibition is not only kill the tumor cell but also to suppress and kill tumor cells that seek to avoid the initial induction of death processes via compensatory survival signaling mechanisms.1 Even within the broad brush definition of carcinomas from a particular organ, there are a range of mutations which present that will profoundly or sometimes more subtly change the paradigm for therapeutic intervention using multiple kinase inhibitor combinations. For example, in colorectal cancer the K-RAS oncogene frequently has an activating mutation implying that inhibition of RAF-MEK1/2-ERK1/2 signaling, but not an initiating receptor upstream of K-RAS, could have a therapeutic effect; however, some colon cell lines with the K-RAS mutation are still noted to be sensitive to upstream ERBB1 inhibitors.2,3 Also, compensatory feedback survival signaling loops can cause, after inhibition of a mutant active intracellular oncogenic kinase such as B-RAF V600E, a survival activation of growth factor receptors in a tumor cell.4 The clinical studies in the manuscript by Al-Marrawi et al. describe the rational combination of signaling inhibitors in a colon cancer patient whose tumor cells express a mutant active B-RAF V600E protein that signals into the MEK1/2-ERK1/2 pathway downstream of K-RAS; this is a particularly aggressive form of colon cancer for which few rational therapeutic interventions have been available until recent times.5,6
BAY43-9006; RAF inhibitor; cetuximab; colorectal cancer therapy; combined therapy; sorafenib; sorafenib and cetuximab
It has been known for a number of years that mutated “inactive” p53 proteins still capable of binding to DNA per se, can bind to DNA sequences that are non-canonical for p53, with for example, a resultant increase in the transcription and expression of growth factor receptors such as ERBB1,1,2 i.e., mutation of p53 not merely results in “no p53 function” but in fact results in “oncogenic p53 function”. And in agreement with this postulate transduction of p53 null cells with mutant p53 can cause transformation.3 In prior studies the authors of the present manuscript had demonstrated that expression of p53 (R175H) and ERBB1 could transform immortalized primary esophageal cells, in parallel with increased migratory ability.4 These present studies have defined why those transformed cells became invasive: increased c-Met activity.5
c-Met; esophageal cancer; p53 mutation; tumor invasion
BH3 mimetic drugs induce cell death by antagonizing the activity of anti-apoptotic Bcl-2 family proteins. Cyclin-dependent kinase (CDK) inhibitors that function as transcriptional repressors down-regulate the Bcl-2 family member Mcl-1 and increase the activity of selective BH3-mimetics that fail to target this protein. In this study, we determined whether CDK inhibitors potentiate the activity of pan-BH3 mimetics by directly neutralizing Mcl-1. Specifically, we evaluated interactions between the prototypical pan-CDK inhibitor flavopiridol and the pan-BH3-mimetic obatoclax in multiple myeloma (MM) cells in which Mcl-1 is critical for survival. Co-administration of flavopiridol and obatoclax synergistically triggered apoptosis in both drug-naive and drug-resistant MM cells. Mechanistic investigations revealed that flavopiridol inhibited Mcl-1 transcription but increased transcription of Bim and its binding to Bcl-2/Bcl-xL. Obatoclax prevented Mcl-1 recovery and potentiated release of Bim from Bcl-2/Bcl-xL and Mcl-1, accompanied by activation of Bax/Bak. Whether administered singly or in combination with obatoclax, flavopiridol also induced up-regulation of multiple BH3-only proteins, including BimEL, BimL, Noxa, and Bik/NBK. Notably, shRNA knock-down of Bim or Noxa abrogated lethality triggered by the flavopiridol/obatoclax combination in vitro and in vivo. Together, our findings demonstrate that CDK inhibition potentiates pan-BH3-mimetic activity through a cooperative mechanism involving up-regulation of BH3-only proteins with coordinate down-regulation of their anti-apoptotic counterparts. These findings have immediate implications for the clinical trial design of BH3 mimetic-based therapies that are presently being studied intensively for the treatment of diverse hematopoietic malignancies, including lethal multiple myeloma.
BH3-only protein; Bim; Cdk inhibitor; BH3-mimetic; myeloma
A major problem in the treatment of cancer and prolongation of patient survival is the dissemination of cells from a defined tumor site into a loco-regional disease and ultimately to full metastatic spread into distant organs. In the manuscript by Ierano et al. multiple chemically diverse histone deacetylase inhibitors (HDACIs) in tumor cell types of many diverse origins were shown to increase expression of the receptor CXCR4; a receptor whose expression promotes metastatic spread of tumor cells and that is correlated with a stage independent poor prognosis.1,2 The ligand of CXCR4, CXCL12, also called stromal cell-derived factor (SDF1), stimulates signaling through multiple pathways downstream of the CXCR4 receptor including SRC kinases, ERK1/2, and STAT3. Inhibition of SRC, ERK, or STAT3 can all suppress tumor cell migration and reduce the threshold at which tumor cells undergo apoptosis.3-8 The authors noted that despite increased CXCR4 expression following HDACI treatment, exogenous CXCL12 ligand had a reduced ability to stimulate cell signaling processes, with the phosphorylation of both SRC and STAT3 at activating sites declining. This resulted in less induced migration of HDACI-treated tumor cells. No studies were undertaken to determine whether HDACI-treated cells transduced to express activated forms of SRC or STAT3 or retained their invasive phenotype; however a loss of SRC and STAT3 signaling would predict for a less invasive phenotype.
CXCR4; romidepsin; histone deacetylase inhibitor; CXCL12; apicidin; vorinostat; entinostat
It has been known for many years that for a "normal" un-transformed cell to become immortal and subsequently tumorigenic requires multiple pro-oncogenic changes in the levels of protein expression and function. Genes most commonly associated with the process of oncogenesis include: p53 inactivating mutation; hDM2 overexpression; p16 reduced expression; K-/H-RAS activating mutation; PTEN inactivating mutation/deletion; EGFR activating mutation and overexpression; retinoblastoma inactivating mutation and deletion; Cyclin proteins overexpression; CD95 reduced expression; protective BCL-2 proteins overexpression; to name but just a few of such molecules.1-5 That the minimally required specific proteins for oncogenesis are not known for many specific tumor types remains a challenge for the rational design of molecular targeted therapies.
basal-like breast cancer; EGFR; PTEN; p53; MCF10A; mammary cells; oncogene; tumor suppressor; transformation; soft agar
It has been known for many years that the protein Fas-associated death domain (FADD) is an essential protein forming the apical portion of the extrinsic apoptosis pathway that permits association of death receptors, e.g., CD95, DR4, DR5 with pro-caspases 8 and 10, thereby facilitating caspase activation (e.g., ref. 1, and references therein). It is also known that FADD can recruit other proteins to regulate NFκB and MAPK pathways which in turn can promote proliferation and cell cycle progression. In NSCLC high expression of FADD has been associated with shorter survival times and lymph node metastasis or oral cancer and worse survival, and the present manuscript in head and neck cancer demonstrates similar findings with respect to lymph node metastasis and survival.2,3
FADD; prognosis; metastasis; head and neck cancer; DR5; caspase-8; immunohistochemistry
It has been known for many years that arsenic trioxide (As2O3; ATO) is an effective therapy for acute promyelocytic leukemia but has little activity against other forms of the disease. ATO has diverse modes of action, but is well known to generate high levels of reactive oxygen species in cells which are believed to be causal in many of its biologic actions.1 ROS can both activate and suppress signaling through multiple intracellular pathways based on the amount and duration of ROS production.2 As the basal activity of the MEK1/2-ERK1/2 pathway is often high in acute myeloid leukemias, and that ATO is known to stimulate MEK1/2-ERK1/2 signaling in leukemia, the authors investigated whether knock down of the downstream effector of ERK1/2, RSK1, could enhance the anti-leukemic activity of ATO.3,4
RSK signaling; arsenic trioxide; kinase; leukemia; mRNA translation; mTOR
Kupffer cells are the resident liver macrophages of the liver; other tissues also have resident immune cells e.g., microglia in the brain.1 These cells have a distinct embryonic lineage when compared with circulating myeloid cells. In both the liver and brain micro-environments activation of their resident immune cells results in the synthesis of multiple growth factors and cytokines which stimulate tumor growth and that in part provide the permissive “soil” in which the tumor “seed” grows.2,3 In the manuscript by Wen et al., studies defined whether liver localized Kupffer cells supported or inhibited the growth of colorectal tumor metastases in an immune competent animal model.4 The authors also determined whether the most important changes in the biology of metastatic tumors were associated with the numbers of CD3-positive T cells and the numbers of VEGF and iNOS expressing cells.4
Kupffer cell; colorectal cancer; liver metastases; macrophage; tumor-associated macrophage
Cardiac toxicity is a major dose-limiting factor for the anthracycline drug doxorubicin. The reasons why doxorubicin causes heart damage are not fully understood, and the manuscript by Wong et al. postulates that inflammatory cytokines released from macrophages or other cell types may play a significant role in the damage process in response to doxorubicin and possibly other chemotherapeutic agents.1 Expression of many cytokines requires activation of both the p38 MAPK and JNK pathways and, additionally, doxorubicin toxicity can be blocked by combined inhibition of both pathways.2,3 The MAP3K responsible for doxorubicin-induced p38 MAPK and JNK activation in keratinocytes was previously shown by these authors to be ZAK.4 ZAK is of note because it can be targeted by FDA approved agents such as nilotinib and sorafenib.4-7
MAPK; ZAK; cytokines; doxorubicin; nilotinib; ponatinib; sorafenib
The present studies sought to further understand how the anti-folate pemetrexed and the multi-kinase inhibitor sorafenib interact to kill tumor cells. Sorafenib activated SRC, and via SRC the drug combination activated ERK1/2. Expression of dominant negative SRC or dominant negative MEK1 abolished drug-induced ERK1/2 activation, together with drug-induced autophagy, acidic lysosome formation, and tumor cell killing. Protein phosphatase 2A is an important regulator of the ERK1/2 pathway. Fulvestrant resistant MCF7 cells expressed higher levels of the PP2A inhibitor SET/I2PP2A, had lower endogenous PP2A activity, and had elevated basal ERK1/2 activity compared with their estrogen dependent counterparts. Overexpression of I2PP2A blocked drug-induced activation of ERK1/2 and tumor cell killing. PP2A can be directly activated by ceramide and SET/I2PP2A can be inhibited by ceramide. Inhibition of the de novo ceramide synthase pathway blocked drug-induced ceramide generation, PP2A activation and tumor cell killing. Collectively these findings demonstrate that ERK1/2 plays an essential role downstream of SRC in pemetrexed and sorafenib lethality and that PP2A plays an important role in regulating this process.
ERK; I2PP2A; PP2A; SRC; autophagy; ceramide; pemetrexed; sorafenib
Adenovirus (Ad)-based gene therapy represents a potentially viable strategy for treating colorectal cancer. The infectivity of serotype 5 adenovirus (Ad.5), routinely used as a transgene delivery vector, is dependent on Coxsackie-adenovirus receptors (CAR). CAR expression is downregulated in many cancers thus preventing optimum therapeutic efficiency of Ad.5-based therapies. To overcome the low CAR problem, a serotype chimerism approach was used to generate a recombinant Ad (Ad.5/3) that is capable of infecting cancer cells via Ad.3 receptors in a CAR-independent manner. We evaluated the improved transgene delivery and efficacy of Ad.5/3 recombinant virus expressing melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24), an effective wide-spectrum cancer-selective therapeutic. In low CAR human colorectal cancer cells RKO, wild-type Ad.5 virus expressing mda-7/IL-24 (Ad.5-mda-7) failed to infect efficiently resulting in lack of expression of MDA-7/IL-24 or induction of apoptosis. However, a recombinant Ad.5/3 virus expressing mda-7/IL-24 (Ad.5/3-mda-7) efficiently infected RKO cells resulting in higher MDA-7/IL-24 expression and inhibition of cell growth both in vitro and in nude mice xenograft models. Addition of the novel Bcl-2 family pharmacological inhibitor Apogossypol derivative BI-97C1 (Sabutoclax) significantly augmented the efficacy of Ad.5/3-mda-7. A combination regimen of suboptimal doses of Ad.5/3-mda-7 and BI-97C1 profoundly enhanced cytotoxicity in RKO cells both in vitro and in vivo. Considering the fact that Ad.5-mda-7 has demonstrated significant objective responses in a Phase I clinical trial for advanced solid tumors, Ad.5/3-mda-7 alone or in combination with BI-97C1 would be predicted to exert significantly improved therapeutic efficacy in colorectal cancer patients.
Viral gene therapy; Mcl-1 inhibition; apoptosis induction; anti-tumor activity
Melanoma differentiation-associated gene-7/interleukin-24 (mda-7/IL-24), a unique member of the IL-10 gene family, displays a broad range of antitumor properties including cancer-specific induction of apoptosis, inhibition of tumor angiogenesis, and modulation of anti-tumor immune responses. Here we identify clusterin (CLU) as a MDA-7/IL-24 interacting protein in DU-145 cells and investigate the role of MDA-7/IL-24 in regulating CLU expression and mediating the antitumor properties of mda-7/IL-24 in prostate cancer. Ad.mda-7 decreased expression of soluble CLU (sCLU) and increased expression of nuclear CLU (nCLU). In the initial phase of Ad.mda-7 infection sCLU expression increased and CLU interacted with MDA-7/IL-24 producing a cytoprotective effect. Infection of stable clones of DU-145 prostate cancer cells expressing sCLU with Ad.mda-7 resulted in generation of nCLU that correlated with decreased cell viability and increased apoptosis. In the presence of mda-7/IL-24, sCLU-DU-145 cells displayed G2/M phase arrest followed by apoptosis. Similarly, Ad.mda-7 infection decreased cell migration by altering cytoskeleton in sCLU-DU-145 cells. Ad.mda-7-treated sCLU-DU-145 cells displayed a significant reduction in tumor growth in mouse xenograft models and reduced angiogenesis when compared to the vector control group. Tumor tissue lysates demonstrated enhanced nCLU generated from sCLU with increased apoptosis in the presence of MDA-7/IL-24. Our findings reveal novel aspects relative to the role of sCLU/nCLU in regulating the anticancer properties of MDA-7/IL-24 that may be exploited for developing enhanced therapies for prostate cancer.
MDA-7/IL-24; soluble clusterin; nuclear clusterin; G2/M arrest; apoptosis