ENMD-981693 Is an Orally-Active Kinase Inhibitor with Activity towards Human Hematologic Cancers In Vitro and In Vivo.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1377-1377
Author(s):  
Mark R. Bray ◽  
Graham C. Fletcher ◽  
Trisha A. Denny ◽  
John Xu ◽  
Xiaoru Chen ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Rapid Development ◽  
Xenograft Model ◽  
Myelogenous Leukemia ◽  
Aurora B ◽  
Ic50 Value ◽  
Ic50 Values ◽  
Orally Active

Abstract The clinical success of Gleevec (imatinib) for chronic myelogenous leukemia has demonstrated that small-molecule inhibitors of specific kinases can be developed into effective oncology therapies. However, the rapid development of resistance in leukemic cells to drugs such as Gleevec and the limited therapeutic indications addressed with these molecules suggests that considerable opportunity exists for new inhibitors with a distinctive spectrum of activities against multiple kinase targets. ENMD-981693 is a novel, orally-active molecule that was discovered through a screening effort directed towards Aurora kinases, a family of serine/threonine kinases that are essential for mitotic progression. ENMD-981693 is selective for the Aurora A isoform, with an IC50 value of 25 nM, compared to an IC50 value of ~700 nM for Aurora B. The activity of ENMD-981693 was evaluated against a panel of 100 recombinant kinases, and the compound was shown to inhibit a broad range of tyrosine kinase targets including Flt3, CSF1R, Lck, JAK2, and c-Kit. ENMD-981693 inhibited the in vitro growth of human hematopoietic cancer lines including MV4;11, K562, THP-1, Jurkat, TF-1, U937, and HL-60 with IC50 values ranging from 0.04 – 21 μM. ENMD-981693 was shown to induce G2/M cell cycle arrest followed by apoptosis in U937 cells, without induction of the endo-reduplication phenotype (≥4N DNA content) associated with Aurora B-acting inhibitors such as MK-0457 (VX-680) and AZD1152. Primary cells derived from AML patients were sensitive to treatment with ENMD-981693 in vitro, resulting in IC50 values from 0.2 – 6.0 μM. Sensitivity of primary CML samples was more variable in in vitro cytotoxicity assays, with IC50s in the range of 0.1 – 40 μM. ENMD-981693 shows significant antitumor activity and is well tolerated in xenograft studies, with no weight loss or morbidity observed in administration schedules of up to 100 mg/kg bid or 200 mg/kg qd, given continuously for more than 30 days. Results from in vivo efficacy studies with ENMD-981693 using the MV4;11 xenograft model will be described. In conclusion, ENMD-981693 is a novel kinase inhibitor with potent activity towards a number of targets important in hematologic cancers.

In Vitro and In Vivo Activity of the Src/Abl Kinase Inhibitor AP23464 and Analogs Against Activation Loop Mutants of KIT.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 793-793 ◽  
Author(s):  
Amie S. Corbin ◽  
Shadmehr Demehri ◽  
Ian J. Griswold ◽  
Chester A. Metcalf ◽  
William C. Shakespeare ◽  
...  
Keyword(s):  
Cell Lines ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Myelogenous Leukemia ◽  
Activation Loop ◽  
Wild Type ◽  
Abl Kinase ◽  
Ic50 Values

Abstract Oncogenic mutations of the KIT receptor tyrosine kinase have been identified in several malignancies including gastrointestinal stromal tumors (GIST), systemic mastocytosis (SM), seminomas/dysgerminomas and acute myelogenous leukemia (AML). Mutations in the regulatory juxtamembrane domain are common in GIST, while mutations in the activation loop of the kinase (most commonly D816V) occur predominantly in SM and at low frequency in AML. Several ATP-competitive kinase inhibitors, including imatinib, are effective against juxtamembrane KIT mutants, however, the D816V mutant is largely resistant to inhibition. We analyzed the sensitivities of cell lines expressing wild type KIT, juxtamembrane mutant KIT (V560G) and activation loop mutant KIT (D816V,F,Y and murine D814Y) to a potent Src/Abl kinase inhibitor, AP23464, and analogs. IC50 values for inhibition of cellular KIT phosphorylation by AP23464 were 5–11 nM for activation loop mutants, 70 nM for the juxtamembrane mutant and 85 nM for wild type KIT. Consistent with this, IC50 values in cell proliferation assays were 3–20 nM for activation loop mutants and 100 nM for wild type KIT and the juxtmembrane mutant. In activation loop mutant-expressing cell lines, AP23464, at concentrations ≤50 nM, induced apoptosis, arrested the cell cycle in G0/G1 and down-regulated phosphorylation of Akt and STAT3, signaling pathways critical for the transforming capacity of mutant KIT. In contrast, 500 nM AP23464 was required to induce equivalent effects in wild-type KIT and juxtamembrane mutant-expressing cell lines. These data demonstrate that activation loop KIT mutants are considerably more sensitive to inhibition by AP23464 than wild type or juxtamembrane mutant KIT. Non-specific toxicity in parental cells occurred only at concentrations above 2 μM. Additionally, at concentrations below 100 nM, AP23464 did not inhibit formation of granulocyte/macrophage and erythrocyte colonies from normal bone marrow, suggesting that therapeutic drug levels would not impact normal hematopoiesis. We also examined in vivo target inhibition in a mouse model. Mice were subcutaneously injected with D814Y-expressing (D816V homologous) murine mastocytoma cells. Once tumors were established, compound was administered three-times daily by oral gavage. One hour post treatment we observed >90% inhibition of KIT phosphorylation in tumor tissue. Following a three-day treatment regimen, there was a statistically significant difference in tumor size compared to controls. Thus, AP23464 analogs effectively target D816-mutant KIT both in vitro and in vivo and inhibit activation loop KIT mutants more potently than the wild type protein. These data provide evidence that this class of kinase inhibitors may have therapeutic potential for D816V-expressing malignancies such as SM or AML.

scholarly journals Targeting USP47 overcomes tyrosine kinase inhibitor resistance and eradicates leukemia stem/progenitor cells in chronic myelogenous leukemia

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Hu Lei ◽  
Han-Zhang Xu ◽  
Hui-Zhuang Shan ◽  
Meng Liu ◽  
Ying Lu ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Progenitor Cells ◽  
Chronic Myelogenous Leukemia ◽  
Drug Targets ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Myelogenous Leukemia ◽  
Tki Resistance

AbstractIdentifying novel drug targets to overcome resistance to tyrosine kinase inhibitors (TKIs) and eradicating leukemia stem/progenitor cells are required for the treatment of chronic myelogenous leukemia (CML). Here, we show that ubiquitin-specific peptidase 47 (USP47) is a potential target to overcome TKI resistance. Functional analysis shows that USP47 knockdown represses proliferation of CML cells sensitive or resistant to imatinib in vitro and in vivo. The knockout of Usp47 significantly inhibits BCR-ABL and BCR-ABLT315I-induced CML in mice with the reduction of Lin−Sca1+c-Kit+ CML stem/progenitor cells. Mechanistic studies show that stabilizing Y-box binding protein 1 contributes to USP47-mediated DNA damage repair in CML cells. Inhibiting USP47 by P22077 exerts cytotoxicity to CML cells with or without TKI resistance in vitro and in vivo. Moreover, P22077 eliminates leukemia stem/progenitor cells in CML mice. Together, targeting USP47 is a promising strategy to overcome TKI resistance and eradicate leukemia stem/progenitor cells in CML.

scholarly journals Identification of Hsp90 inhibitors as potential drugs for the treatment of TSC1/TSC2 deficient cancer

2021 ◽  
Author(s):  
Evelyn M. Mrozek ◽  
Vineeta Bajaj ◽  
Yanan Guo ◽  
Izabela Malinowska ◽  
Jianming Zhang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Xenograft Model ◽  
Growth Delay ◽  
Hsp90 Inhibitors ◽  
Null Cell ◽  
Standard Dosage

Inactivating mutations in either TSC1 or TSC2 cause Tuberous Sclerosis Complex, an autosomal dominant disorder, characterized by multi-system tumor and hamartoma development. Mutation and loss of function of TSC1 and/or TSC2 also occur in a variety of sporadic cancers, and rapamycin and related drugs show highly variable treatment benefit in patients with such cancers. The TSC1 and TSC2 proteins function in a complex that inhibits mTORC1, a key regulator of cell growth, which acts to enhance anabolic biosynthetic pathways. In this study, we identified and validated five cancer cell lines with TSC1 or TSC2 mutations and performed a kinase inhibitor drug screen with 197 compounds. The five cell lines were sensitive to several mTOR inhibitors, and cell cycle kinase and HSP90 kinase inhibitors. The IC50 for Torin1 and INK128, both mTOR kinase inhibitors, was significantly increased in three TSC2 null cell lines in which TSC2 expression was restored.  Rapamycin was significantly more effective than either INK128 or ganetespib (an HSP90 inhibitor) in reducing the growth of TSC2 null SNU-398 cells in a xenograft model. Combination ganetespib-rapamycin showed no significant enhancement of growth suppression over rapamycin. Hence, although HSP90 inhibitors show strong inhibition of TSC1/TSC2 null cell line growth in vitro, ganetespib showed little benefit at standard dosage in vivo. In contrast, rapamycin which showed very modest growth inhibition in vitro was the best agent for in vivo treatment, but did not cause tumor regression, only growth delay.

The Natural Product Borrelidin Induces UPR and Apoptosis in AML Cell Lines

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4260-4260
Author(s):  
Leah Jackson ◽  
Shelby Bechler ◽  
Justin Miller ◽  
Amy Brownell ◽  
Danielle Garshott ◽  
...  
Keyword(s):  
Cell Death ◽  
Er Stress ◽  
Natural Product ◽  
Cell Lines ◽  
Time Course ◽  
Conflicts Of Interest ◽  
Xenograft Model ◽  
Myelogenous Leukemia ◽  
Ic50 Values

Abstract Abstract 4260 Acute Myelogenous Leukemia (AML) is the most common form of leukemia. Current therapies are intense and even those fortunate enough to achieve remission often relapse extending extremely poor prognoses to these patient. The most commonly used therapeutics, namely cytarabine aribinoside, the anthracyclines and etoposide, are decades old and target ubiquitous cellular processes. We have previously reported that small molecules and natural products that activate and exacerbate the unfolded protein response (UPR) can effectively and selectively induce cell death in a wide variety of solid tumor cells. We hypothesized that the UPR might be a viable new therapeutic target in AML and sought to determine whether or not the novel UPR-inducing natural product borrelidin might be used as such an agent. A luminescent proliferation assay performed with panel of four AML cell lines treated with the ER stress-inducing antibiotic tunicamycin (Tm) revealed that three of the cell lines displayed IC50 values between 0.47–2.5μ M, doses of Tm which are known to induce a low to moderate level of ER stress. We then repeated the experiment with the more general UPR-inducing natural product borrelidin, which has been shown to have potent anti-inflammatory properties in several murine assays in vivo. All four cell lines were sensitive to borrelidin, displaying IC50 values between 0.032–0.29 μ M. Time course assays performed with borrelidin revealed 4–20 fold increases in active caspase 3 and 7 indicating borrelidin-induced AML decreases in cell proliferation might be the result of apoptosis. Quantitative reverse-transcription real time PCR performed with mRNA isolated from two AML cell lines revealed an increase in the UPR-related transcripts CHOP, ATF4, and GADD34 and the cell death genes Noxa, Puma, DR5 and Bim confirming that borrelidin could induce the UPR and apoptosis in AML cells. Studies currently underway in our laboratory will determine the ability of borrelidin and other UPR-inducing agents to reduce leukemic burden in an in vivo xenograft model. Disclosures: No relevant conflicts of interest to declare.

Rapid rational drug targeting of Merkel cell polyomavirus (MCV)-positive Merkel cell carcinoma (MCC) using the survivin inhibitor YM155.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. 8577-8577
Author(s):  
Reety Arora ◽  
Masahiro Shuda ◽  
Anna Guastafierro ◽  
Tuna Toptan ◽  
Yanis Tolstov ◽  
...  
Keyword(s):  
Xenograft Model ◽  
Survivin Expression ◽  
T Antigen ◽  
Merkel Cell Polyomavirus ◽  
Myelogenous Leukemia ◽  
Merkel Cell ◽  
Rational Drug ◽  
Mouse Xenograft Model

8577 Background: MCC is an aggressive, chemoresistant skin cancer causing more deaths each year than chronic myelogenous leukemia. We discovered a new virus, Merkel cell polyomavirus (MCV), clonally integrated into ~80% of primary and metastatic MCC in 2008. To find therapeutic targets for this cancer, we examined cellular genes perturbed by MCV infection. Methods: Digital transcriptome subtraction was used to discover MCV and also to reveal survivin gene (BIRC5) upregulation in virus-positive tumors. MCV T antigen knockdown studies in seven MCC lines and large T (LT) transduction into BJ fibroblasts were used to confirm this. Drug screening was performed in vitro using Cell-Titer Glo assays in a two stage analysis. In vivo screening used an MKL-1 (MCV+) MCC NOD-SCIDg mouse xenograft model with a single three-week treatment round. Results: MCV large T oncoprotein induces survivin transcription through retinoblastoma protein sequestration by the LT LXCXE motif. MCV T antigen knockdown results in nonapoptotic MCC cell death and loss of survivin expression. YM155, a phase II survivin transcription inhibitor, causes MCV+ MCC cell necroptosis associated with autophagy at 1-12 nM EC50. Of 1359 other drugs from LOPAC and NCI Oncology Set II libraries, only bortezomib had in vitro potency comparable to YM155. In MKL-1 xenograft studies, mice were treated with saline, bortezomib or YM155 for three weeks using standard dosings. Bortezomib did not significantly improve mouse survival (33%) over saline (24%) during treatment. In contrast, all YM155-treated mice survived (100%, p<0.001) the 3 week treatment period. Tumors resumed growth once YM155 treatment was stopped suggesting that YM155 is cytostatic in vivo rather than cytotoxic. Conclusions: Survivin expression is induced by MCV LT and is critical to MCV+ MCC survival. A survivin inhibitor, YM155 was nontoxic to mice and cytostatic for MCV+ MCC xenografts. Using genomic technologies, in less than four years, the primary viral cause for most MCC was discovered, new diagnostic tests developed and a promising rational drug candidate identified. A cooperative group trial (E1611) for YM155 and bortezomib in MCC patients is currently planned.

Abstract 117: Ponatinib Mediated Cardiotoxicity Is Driven By Pro-inflammatory S100A8/A9-NLRP3-IL-1β Signaling Circuit

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Anand P Singh ◽  
Sultan Tousif ◽  
Prachi Umbarkar ◽  
Cristi L Galindo ◽  
Nicholas Wheeler ◽  
...  
Keyword(s):  
Cardiac Function ◽  
High Fat Diet ◽  
Kinase Inhibitor ◽  
Clinical Investigation ◽  
Pressure Overload ◽  
Immunosuppressive Agent ◽  
Myelogenous Leukemia ◽  
High Fat

Background: Ponatinib is a third-generation tyrosine kinase inhibitor (TKI) for chronic myelogenous leukemia (CML) treatment. Of note, ponatinib is the only treatment option for CML patients with T315I (gatekeeper) mutation. Unexpected clinical cardiotoxicity, including fatal myocardial infarction and congestive heart failure, has hampered its clinical use. Herein, we aimed to investigate the cardiotoxic mechanism of ponatinib and strategies to prevent the cardiotoxic manifestations. Methods: We employed wild-type C57BL/6, cardiovascular (CV) comorbidity models e.g., transverse aortic constriction (TAC)-pressure overload (cardiac comorbidity) and high-fat diet fed ApoE -/- (vascular comorbidity), to investigate the cardiotoxic mechanism of ponatinib. Echocardiography was performed to assess cardiac function. Comprehensive immune profiling was performed to identify ponatinib-induced immune dynamics using flow cytometry analysis. Results: Echocardiographic assessment of ponatinib treated high-fat diet fed ApoE -/- and pressure overload (PO) murine model showed significant decline in cardiac function, suggesting the key role of CV-comorbidities in ponatinib-induced cardiomyopathy. An unbiased RNA-Seq analysis identified the enrichment of dysregulated inflammatory genes, including a multi-fold upregulation of alarmins S100A8/A9 as a top hit in ponatinib-treated hearts. A combination of in vitro and in vivo mechanistic analysis, identified that ponatinib activates the S100A8/9-TLR4-NLRP3-IL-1β signaling pathway in cardiac and systemic myeloid cells (monocytes and neutrophils), thereby leading to excessive myocardial and systemic inflammation. Finally, we demonstrate that ponatinib-induced excessive inflammation is central to the cardiac pathology because a broad immunosuppressive agent dexamethasone abolished the adverse cardiac remodeling and dysfunction of ponatinib treated hearts. Conclusions: These findings uncover a novel mechanism of ponatinib-induced cardiac inflammation leading to cardiac dysfunction. Our results provide critical preclinical data and rationale for clinical investigation into immunosuppressive interventions to mitigate ponatinib-induced cardiotoxicity.

Multi-Targeted Receptor Tyrosine Kinase Inhibitor, ABT-869, Induces Apoptosis and Suppresses Proliferation of Ba/F3 FLT-3 ITD Mutant Cells in Vitro and in Vivo through Inhibition of FLT3 and AKT.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1938-1938
Author(s):  
Jenny E Hernandez ◽  
Joan Zape ◽  
Keith Glaser ◽  
Elliot Landaw ◽  
Cecilia Fu ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Cell Lines ◽  
Receptor Tyrosine Kinase ◽  
Kinase Inhibitor ◽  
Mutant Cell ◽  
Vehicle Control ◽  
Ic50 Value ◽  
Mutant Cells

Abstract FLT3 is a receptor tyrosine kinase of the subclass III family that plays a vital role in the regulation of differentiation, proliferation and survival of normal hematopoietic cells. FLT3 mutations are often found in patients with acute myelogenous leukemia (AML) and confer a poor prognosis. Of these mutations, 15–35% are FLT3 ITD (internal tandem duplication) mutations and 5–7% are point mutations in the FLT3 kinase activation loop, e.g. D835V. We are studying the signaling pathways associated with a small molecule multi-targeted receptor tyrosine kinase inhibitor (RTKI), ABT-869. To determine the effects of ABT-869 in vitro and in vivo, a Ba/F3 mouse pro-B lymphocytic cell line harboring the FLT-3 ITD or FLT-3 D835V mutation was used as an isolated FLT-3 mutant model system. In vitro, ABT-869 is effective in inhibiting the proliferation of Ba/F3 Flt-3 ITD mutant cells (IC50 value of 1 nM) when compared to Ba/F3 Flt-3 D835V mutant (IC50 value between 1 and 10 μM) and Ba/F3 Flt-3 wildtype (WT) cells (IC50 value of 10 μM). Annexin V and propidium iodide staining of cells revealed that an increase in apoptosis occurred in Ba/F3 Flt-3 ITD mutant cells treated with 1μM ABT-869 for 24 hours (42.8%) when compared to untreated (4.7%) or vehicle control (4.0%) cells. Ba/F3 Flt-3 D835V mutant cell lines demonstrated a 12.5% rate of apoptosis at 1μM, compared to untreated (1.99%) and vehicle control (2.1%) cell lines. Propidium iodide staining of treated Ba/F3 Flt-3 WT cell lines revealed no difference in apoptosis when compared to untreated Ba/F3 Flt-3 WT cells or DMSO controls. PARP cleavage was observed in Ba/F3 FLT-3 ITD mutant cells, following 6 hours of treatment with 1 to 100 nM ABT-869, whereas no cleavage was observed in Ba/F3 WT cells treated with ABT-869. To study the effects of ABT-869 in vivo, we treated SCID mice injected with Ba/F3 Flt-3 ITD, Ba/F3 Flt-3 D835V, or Ba/F3 Flt-3 WT cells and monitored disease progression using bioluminescence imaging. The mice injected with the Ba/F3 FLT-3 ITD mutant cells and treated with vehicle control developed metastases and had a median survival time of 2 weeks. In contrast, the ABT-869 treated group had slower disease progression with median survival of 6.2 weeks (P&lt;0.008). Both control and treated mice injected with Ba/F3 FLT-3 D835V mutant cell lines developed metastases and had similar survival (median 1.7 and 1.9 weeks, respectively). Survival times of control and treated mice injected with Ba/F3 FLT-3 WT cells were also similar (median 8.4 and 8.1 weeks, respectively). Previous work identified that ABT-869 induced apoptosis of acute myeloid leukemia cells through inhibition of FLT-3 reception phosphorylation, which is observed as early as 3 hours after treatment. In Ba/F3 cells expressing FLT-3 ITD, ABT-869 also inhibited phosphorylation of AKT, which is upstream of the pro-apoptotic protein Bad. Therefore, our preclinical data suggest that ABT-869 induces apoptosis of FLT-3 ITD mutant cells both in vitro and in vivo. These studies provide rationale for the treatment of acute leukemia patients harboring the FLT3-ITD mutation with ABT-869 and the potential benefit of combining small molecule inhibitors that target both RTKs and AKT.

scholarly journals Broad anti-coronaviral activity of FDA approved drugs against SARS-CoV-2 in vitro and SARS-CoV in vivo

2020 ◽  
Author(s):  
Stuart Weston ◽  
Christopher M. Coleman ◽  
Rob Haupt ◽  
James Logue ◽  
Krystal Matthews ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Rapid Development ◽  
Drug Repurposing ◽  
Ic50 Values ◽  
Human Use ◽  
Approved Drugs ◽  
Fda Approved Drugs

AbstractSARS-CoV-2 emerged in China at the end of 2019 and has rapidly become a pandemic with roughly 2.7 million recorded COVID-19 cases and greater than 189,000 recorded deaths by April 23rd, 2020 (www.WHO.org). There are no FDA approved antivirals or vaccines for any coronavirus, including SARS-CoV-2. Current treatments for COVID-19 are limited to supportive therapies and off-label use of FDA approved drugs. Rapid development and human testing of potential antivirals is greatly needed. A quick way to test compounds with potential antiviral activity is through drug repurposing. Numerous drugs are already approved for human use and subsequently there is a good understanding of their safety profiles and potential side effects, making them easier to fast-track to clinical studies in COVID-19 patients. Here, we present data on the antiviral activity of 20 FDA approved drugs against SARS-CoV-2 that also inhibit SARS-CoV and MERS-CoV. We found that 17 of these inhibit SARS-CoV-2 at a range of IC50 values at non-cytotoxic concentrations. We directly follow up with seven of these to demonstrate all are capable of inhibiting infectious SARS-CoV-2 production. Moreover, we have evaluated two of these, chloroquine and chlorpromazine, in vivo using a mouse-adapted SARS-CoV model and found both drugs protect mice from clinical disease.

scholarly journals Evaluation of Hsp90 and mTOR inhibitors as potential drugs for the treatment of TSC1/TSC2 deficient cancer

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0248380
Author(s):  
Evelyn M. Mrozek ◽  
Vineeta Bajaj ◽  
Yanan Guo ◽  
Izabela A. Malinowska ◽  
Jianming Zhang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Xenograft Model ◽  
Mtor Inhibitors ◽  
Growth Delay ◽  
Null Cell ◽  
Standard Dosage

Inactivating mutations in either TSC1 or TSC2 cause Tuberous Sclerosis Complex, an autosomal dominant disorder, characterized by multi-system tumor and hamartoma development. Mutation and loss of function of TSC1 and/or TSC2 also occur in a variety of sporadic cancers, and rapamycin and related drugs show highly variable treatment benefit in patients with such cancers. The TSC1 and TSC2 proteins function in a complex that inhibits mTORC1, a key regulator of cell growth, which acts to enhance anabolic biosynthetic pathways. In this study, we identified and validated five cancer cell lines with TSC1 or TSC2 mutations and performed a kinase inhibitor drug screen with 197 compounds. The five cell lines were sensitive to several mTOR inhibitors, and cell cycle kinase and HSP90 kinase inhibitors. The IC50 for Torin1 and INK128, both mTOR kinase inhibitors, was significantly increased in three TSC2 null cell lines in which TSC2 expression was restored. Rapamycin was significantly more effective than either INK128 or ganetespib (an HSP90 inhibitor) in reducing the growth of TSC2 null SNU-398 cells in a xenograft model. Combination ganetespib-rapamycin showed no significant enhancement of growth suppression over rapamycin. Hence, although HSP90 inhibitors show strong inhibition of TSC1/TSC2 null cell line growth in vitro, ganetespib showed little benefit at standard dosage in vivo. In contrast, rapamycin which showed very modest growth inhibition in vitro was the best agent for in vivo treatment, but did not cause tumor regression, only growth delay.

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