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PDF file - 65 KB, Acute dosing with GDC-0941 leads to an increase in apoptotic signaling in in sensitive tumors.

PDF file - 75 KB, Uptake of 18F-FDG decreases significantly after ~18h of GDC-0941 treatment in U87, but not HCT116, tumors.

PDF file - 74 KB, GDC-0941 inhibits hypoxic signaling in drug sensitive U87 tumors.

PDF file - 46 KB, Chronic dosing with GDC-0941 leads to an increase in MAP-kinase pathway signaling in sensitive tumors.

The phosphoinositide 3-kinase (PI3K) pathway is deregulated in a range of cancers, and several targeted inhibitors are entering the clinic. This study aimed to investigate whether the positron emission tomography tracer 3′-deoxy-3′-[18F]fluorothymidine ([18F]-FLT) is suitable to mark the effect of the novel PI3K inhibitor GDC-0941, which has entered phase II clinical trial. CBA nude mice bearing U87 glioma and HCT116 colorectal xenografts were imaged at baseline with [18F]-FLT and at acute (18 hours) and chronic (186 hours) time points after twice-daily administration of GDC-0941 (50 mg/kg) or vehicle. Tumor uptake normalized to blood pool was calculated, and tissue was analyzed at sacrifice for PI3K pathway inhibition and thymidine kinase (TK1) expression. Uptake of [18F]-FLT was also assessed in tumors inducibly overexpressing a dominant-negative form of the PI3K p85 subunit p85α, as well as HCT116 liver metastases after GDC-0941 therapy. GDC-0941 treatment induced tumor stasis in U87 xenografts, whereas inhibition of HCT116 tumors was more variable. Tumor uptake of [18F]-FLT was significantly reduced following GDC-0941 dosing in responsive tumors at the acute time point and correlated with pharmacodynamic markers of PI3K signaling inhibition and significant reduction in TK1 expression in U87, but not HCT116, tumors. Reduction of PI3K signaling via expression of Δp85α significantly reduced tumor growth and [18F]-FLT uptake, as did treatment of HCT116 liver metastases with GDC-0941. These results indicate that [18F]-FLT is a strong candidate for the noninvasive measurement of GDC-0941 action. Mol Cancer Ther; 12(5); 819–28. ©2013 AACR.

PDF file - 51 KB

Figures S1-S9

From microarray analysis, genes from VGP melanocyte neoplasia whose transcripts are significantly up- or down-regulated compared to wild-type tissue that are associated with lipid metabolism and are not differentially expressed also in BRAFV600E or V12RAS RGP samples.

Alterations in lipid metabolism in cancer cells impact cell structure, signaling, and energy metabolism, making lipid metabolism a potential diagnostic marker and therapeutic target. In this study, we combined PET, desorption electrospray ionization-mass spectrometry (DESI-MS), nonimaging MS, and transcriptomic analyses to interrogate changes in lipid metabolism in a transgenic zebrafish model of oncogenic RAS-driven melanocyte neoplasia progression. Exogenous fatty acid uptake was detected in melanoma tumor nodules by PET using the palmitic acid surrogate tracer 14(R,S)-18F-fluoro-6-thia-heptadecanoic acid ([18F]-FTHA), consistent with upregulation of genes associated with fatty acid uptake found through microarray analysis. DESI-MS imaging revealed that FTHA uptake in tumors was heterogeneous. Transcriptome and lipidome analyses further highlighted dysregulation of glycerophospholipid pathways in melanoma tumor nodules, including increased abundance of phosphatidyl ethanolamine and phosphatidyl choline species, corroborated by DESI-MS, which again revealed heterogeneous phospholipid composition in tumors. Overexpression of the gene encoding lipoprotein lipase (LPL), which was upregulated in zebrafish melanocyte tumor nodules and expressed in the majority of human melanomas, accelerated progression of oncogenic RAS-driven melanocyte neoplasia in zebrafish. Depletion or antagonism of LPL suppressed human melanoma cell growth; this required simultaneous fatty acid synthase (FASN) inhibition when FASN expression was also elevated. Collectively, our findings implicate fatty acid acquisition as a possible therapeutic target in melanoma, and the methods we developed for monitoring fatty acid uptake have potential for diagnosis, patient stratification, and monitoring pharmacologic response.Significance:These findings demonstrate the translational potential of monitoring fatty acid uptake and identify lipoprotein lipase as a potential therapeutic target in melanoma.

PARP inhibitors (PARPi) enhance radiation sensitivity in multiple cancer models, both in vitro and in vivo. Our observation that the radiosensitizing properties of PARPi are most pronounced in rapidly proliferating cells is reflected in early phase clinical trial data showing exacerbation of acute radiation toxicity in rapidly proliferating tissues such as oropharyngeal and esophageal mucosa. Lack of radiosensitization in late responding, slowly proliferating normal tissues indicates that PARPi may be more effectively combined with radiation therapy (RT) in patients with brain tumors. We are therefore evaluating the oral PARPi olaparib in combination with RT and/or temozolomide (TMZ) in the treatment of glioblastoma (GBM), the most prevalent and most aggressive primary brain tumor. Patients with GBM experience very poor outcomes in terms of median survival (c.1 year) and neurocognitive decline caused primarily by RT. Olaparib was initially evaluated in combination with daily low-dose TMZ in patients with recurrent GBM in the OPARATIC trial. Pharmacokinetic studies revealed that olaparib penetrates both core and margin regions of GBM, indicating that the BBB is significantly disrupted throughout these tumors. Olaparib could be safely combined with daily TMZ (75 mg/m2), but intermittent olaparib dosing (150 mg three days per week) was required to avoid dose-limiting hematological toxicity. Early phase testing of the olaparib-radiotherapy combination is now underway in three populations of patients with newly diagnosed GBM. Patients aged >65 with MGMT unmethylated GBM are being recruited to a randomized, placebo-controlled phase II study (PARADIGM) after a phase I dose escalation study showed that olaparib (200 mg twice daily) was extremely well tolerated when combined with brain irradiation (40 Gray in 15#). Good performance status patients aged Citation Format: Anthony J. Chalmers, Rodrigo Gutierrez-Quintana, David J. Walker, Karin Williams, Duncan Forster, Mark R. Jackson, Sarah Derby, Jon Stobo, Lorna Sweeting, Caroline Kelly, Stephen Durant, Kaye J. Williams. Enhancing the therapeutic ratio for glioblastoma by combining radiation therapy with PARP inhibitors [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr IA-006.

Alterations in lipid metabolism in cancer cells impact cell structure, signaling, and energy metabolism, making lipid metabolism a potential diagnostic marker and therapeutic target. In this study, we combined PET, desorption electrospray ionization-mass spectrometry (DESI-MS), nonimaging MS, and transcriptomic analyses to interrogate changes in lipid metabolism in a transgenic zebrafish model of oncogenic RAS-driven melanocyte neoplasia progression. Exogenous fatty acid uptake was detected in melanoma tumor nodules by PET using the palmitic acid surrogate tracer 14(R,S)-18F-fluoro-6-thia-heptadecanoic acid ([18F]-FTHA), consistent with upregulation of genes associated with fatty acid uptake found through microarray analysis. DESI-MS imaging revealed that FTHA uptake in tumors was heterogeneous. Transcriptome and lipidome analyses further highlighted dysregulation of glycerophospholipid pathways in melanoma tumor nodules, including increased abundance of phosphatidyl ethanolamine and phosphatidyl choline species, corroborated by DESI-MS, which again revealed heterogeneous phospholipid composition in tumors. Overexpression of the gene encoding lipoprotein lipase (LPL), which was upregulated in zebrafish melanocyte tumor nodules and expressed in the majority of human melanomas, accelerated progression of oncogenic RAS-driven melanocyte neoplasia in zebrafish. Depletion or antagonism of LPL suppressed human melanoma cell growth; this required simultaneous fatty acid synthase (FASN) inhibition when FASN expression was also elevated. Collectively, our findings implicate fatty acid acquisition as a possible therapeutic target in melanoma, and the methods we developed for monitoring fatty acid uptake have potential for diagnosis, patient stratification, and monitoring pharmacologic response. Significance: These findings demonstrate the translational potential of monitoring fatty acid uptake and identify lipoprotein lipase as a potential therapeutic target in melanoma.

Nonpeptidic Arg-Gly-Asp (RGD)-mimic ligands were designed and synthesized by click chemistry between an arginine-azide mimic and an aspartic acid-alkyne mimic. Some of these molecules combine excellent in vitro properties (high α

Hypoxia was identified as a microenvironmental component of solid tumours over 60 years ago and was immediately recognised as a potential barrier to therapy through the reliance of radiotherapy on oxygen to elicit maximal cytotoxicity. Over the last two decades both clinical and experimental studies have markedly enhanced our understanding of how hypoxia influences cellular behaviour and therapy response. Furthermore, they have confirmed early assumptions that low oxygenation status in tumours is an exploitable target in cancer therapy. Generally such approaches will be more beneficial to patients with hypoxic tumours, necessitating the use of biomarkers that reflect oxygenation status. Tissue biomarkers have shown utility in many studies. Further significant advances have been made in the non-invasive measurement of tumour hypoxia with positron emission tomography, magnetic resonance imaging and other imaging modalities. Here, we describe the complexities of defining and measuring tumour hypoxia and highlight the therapeutic approaches to combat it.

The phosphoinositide 3-kinase (PI3K) pathway is deregulated in a range of cancers, and several targeted inhibitors are entering the clinic. This study aimed to investigate whether the positron emission tomography tracer 3′-deoxy-3′-[18F]fluorothymidine ([18F]-FLT) is suitable to mark the effect of the novel PI3K inhibitor GDC-0941, which has entered phase II clinical trial. CBA nude mice bearing U87 glioma and HCT116 colorectal xenografts were imaged at baseline with [18F]-FLT and at acute (18 hours) and chronic (186 hours) time points after twice-daily administration of GDC-0941 (50 mg/kg) or vehicle. Tumor uptake normalized to blood pool was calculated, and tissue was analyzed at sacrifice for PI3K pathway inhibition and thymidine kinase (TK1) expression. Uptake of [18F]-FLT was also assessed in tumors inducibly overexpressing a dominant-negative form of the PI3K p85 subunit p85α, as well as HCT116 liver metastases after GDC-0941 therapy. GDC-0941 treatment induced tumor stasis in U87 xenografts, whereas inhibition of HCT116 tumors was more variable. Tumor uptake of [18F]-FLT was significantly reduced following GDC-0941 dosing in responsive tumors at the acute time point and correlated with pharmacodynamic markers of PI3K signaling inhibition and significant reduction in TK1 expression in U87, but not HCT116, tumors. Reduction of PI3K signaling via expression of Δp85α significantly reduced tumor growth and [18F]-FLT uptake, as did treatment of HCT116 liver metastases with GDC-0941. These results indicate that [18F]-FLT is a strong candidate for the noninvasive measurement of GDC-0941 action. Mol Cancer Ther; 12(5); 819–28. ©2013 AACR.

Alzheimer's disease (AD) is the most common form of dementia in the elderly. Due to ongoing advances in our understanding of the underlying pathology of AD, many potential new targets for therapeutics are becoming available. Transgenic mouse models of AD have helped in furthering our understanding of AD and also provide a vehicle for preclinical testing of new, putative disease-modifying therapeutics, which may have potential for translation to use in clinical trials. To identify possible translational biomarkers, we have studied the longitudinal cerebral metabolic pattern of the TASTPM transgenic AD mouse, a double transgenic mouse overexpressing human mutant amyloid precursor protein (hAPP695swe) and presenilin-1 (M146V) by (1) H magnetic resonance spectroscopy, along with concurrent brain T(1) /T(2) mapping and behavioral testing. We found significant differences in creatine, glutamate, N-acetylaspartate, choline-containing compounds, and myo-inositol between TASTPM and wild-type mice. In the case of N-acetylaspartate and myo-inositol, there were similarities to differences detected in human AD. T(1) /T(2) values were shorter overall in TASTPM mice, indicating possible differences in water content between TASTPM and wild-type mice. In older TASTPM mice, exploratory behavior became more random, indicating a possible memory deficiency. The decrease in behavioral performance correlated in the transgenic group with higher expression of myo-inositol. Magn Reson Med, 2012. (c) 2012 Wiley Periodicals, Inc.

Radiation therapy (RT) is an important anti-cancer treatment. Understanding the molecular pathways involved in the response of tumors to RT has led to interest in developing drugs which target these; one such target is Akt. Previous studies support the hypothesis that Akt inhibition might enhance the radiosensitivity of tumors through modification of cellular and micro-environmental factors. AZD5363 is a novel, orally bio-available, ATP competitive inhibitor of Akt and here we have evaluated whether AZD5363 will increase the efficacy of RT in preclinical models of head and neck (H&N) cancer. The ability of AZD5363 to inhibit the Akt pathway was investigated in vitro in 5 H&N cell lines and in vivo (FaDu xenografts) using western blot and ELISA techniques. The effect of AZD5363 on the radiosensitivity in vitro was assessed using clonogenic assays. The FaDu xenograft model was used to test the effect of AZD5363 on tumor growth when used as a sequential treatment (for 7 days before RT) or adjuvant therapy (continuously after RT). RT was given as a 6Gy single dose. The effect of these treatments on the tumor microenvironment prior to and after irradiation was also assessed using FAZA PET/CT, immunohistochemistry and ELISA techniques. AZD5363 effectively inhibited the Akt pathway in in vitro and in vivo. There was no effect on radiosensitivity in vitro when drug was administered in a variety of sequences and durations around RT. In vivo, there was no improvement in tumor control when AZD5363 was given for 7 days before RT. But survival was improved (p = 0.006) when AZD5363 was administered continuously after RT with reduced tumor growth (p = 0.0035, 7 days after RT). There was no effect of drug alone on tumor growth and no difference in the level of tumor hypoxia (measured using FAZA PET/CT or pimonidazole binding) after 7 days AZD5363. Human VEGF and HIF1α levels were reduced (p = 0.0104 and p = 0.002, respectively) after 7 days AZD5363. There was no difference in microvessel density (MVD) prior to RT, but 7 days after, MVD was significantly reduced if adjuvant AZD5363 had been given (p = 0.0317). AZD5363 was also found to effect the rate of proliferation of human umbilical vein endothelial (HUVEC) cells in vitro. AZD5363 effectively improves long term control of FaDu tumors when combined with RT. However, this only occurs when the drug is used in the adjuvant setting. AZD5363 has no effect on the radiosensitivity of H&N cells in vitro, which suggests that the in vivo effects are dependent on the tumor microenvironment. AZD5363 reduces tumor VEGF and HIF1α in an oxygen independent manner and inhibits the rate of proliferation of vascular endothelial cells, effects that would reduce vascular recovery after RT. This is the first time a specific inhibitor of Akt has been reported to have this sequence dependent ability to enhance the effects of RT and has implications for how AZD5363 and potentially other Akt inhibitors are used in combination with RT in the clinic. Citation Format: Emma J. Searle, Brian Telfer, Duncan Forster, Kaye J. Williams, Barry Davies, Timothy Illidge, Ian Stratford. The novel Akt inhibitor AZD5363 following radiotherapy improves tumor control in mouse models of head and neck cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3042.

Alzheimer's disease (AD) is the most common form of dementia in the elderly. Due to ongoing advances in our understanding of the underlying pathology of AD, many potential new targets for therapeutics are becoming available. Transgenic mouse models of AD have helped in furthering our understanding of AD and also provide a vehicle for preclinical testing of new, putative disease-modifying therapeutics, which may have potential for translation to use in clinical trials. To identify possible translational biomarkers, we have studied the longitudinal cerebral metabolic pattern of the TASTPM transgenic AD mouse, a double transgenic mouse overexpressing human mutant amyloid precursor protein (hAPP695swe) and presenilin-1 (M146V) by (1) H magnetic resonance spectroscopy, along with concurrent brain T1 /T2 mapping and behavioral testing. We found significant differences in creatine, glutamate, N-acetylaspartate, choline-containing compounds, and myo-inositol between TASTPM and wild-type mice. In the case of N-acetylaspartate and myo-inositol, there were similarities to differences detected in human AD. T1 /T2 values were shorter overall in TASTPM mice, indicating possible differences in water content between TASTPM and wild-type mice. In older TASTPM mice, exploratory behavior became more random, indicating a possible memory deficiency. The decrease in behavioral performance correlated in the transgenic group with higher expression of myo-inositol.