Vascular Targeting Agent Research Articles

Abstract 3216: CKD-516, a novel vascular disrupting agent, enhances anticancer activity of anti-PD-1 antibody in SMAD4-deficient colon cancer model

Abstract CKD-516 is a potent vascular disrupting agent (VDA), selectively acting on tumor vessels. A clinical study of CKD-516 combined with irinotecan is undergoing in colorectal cancer. There have been exponential gains in immune-oncology (I-O) in recent times through the development of immune checkpoint inhibitors (ICIs). ICIs demonstrated durable response and some patients achieved disease control for several years. However, there are still critical unmet medical needs for the combination therapies due to limited response rate of ICIs. The first step of the cancer-immunity cycle is dendritic cells (DCs) maturation which is necessary for the anticancer immunity. It is well known that mature DCs play critical roles in priming immune responses in cancer patients. We found that CKD-516 is able to induce DC maturation through Rho signaling pathway in DCs. A tumor suppressor gene, SMAD4 is associated with several cancers including stomach cancer, colorectal cancer, and pancreas cancer. The loss of SMAD4 is a poor prognostic biomarker in these cancer types and its mutation is related to poor clinical outcome on immunotherapies. A SMAD4-deficient mouse colorectal cancer cell line (3349LM) was primary cultured from a spontaneous intestinal adenocarcinoma formed in a Villin-Cre;Smad4(F/F);Trp53(F/F) mouse. CKD-516 showed a synergistic effect with anti-PD-1 antibody in 3349LM syngeneic model. In addition, 3349LM is also a MSS type cancer. Thus, CKD-516 displayed a therapeutic potential for several cancer patients who do not respond to immunotherapy. In summary, CKD-516 is a novel VDA with immune boosting effect. In SMAD4 deficient cancer model, CKD-516 has shown synergistic effects in combination with PD-1 antibody. Therefore, these data suggest that CKD-516 potentiates the anticancer activity of immunotherapy. Citation Format: Soo Jin Kim, Hark Kyun Kim, Keun Ho Ryu, Chung Il Hong. CKD-516, a novel vascular disrupting agent, enhances anticancer activity of anti-PD-1 antibody in SMAD4-deficient colon cancer model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3216.

Abstract 350: Investigating the mechanisms of cellular uptake and metabolism of ICT2588, an MT-MMP-activated prodrug

Abstract ICT2588 is a non-toxic MT1-MMP-activated peptide prodrug of the potent vascular disrupting agent (VDA) azademethylcolchicine [1]. ICT2588 produces significant tumor-selective delivery (≥10-fold) of the toxic and potent azademethylcolchicine, as compared to the administration of azademethylcolchicine alone, in murine models of human colorectal, breast, lung and prostate cancers. In addition, ICT2588 exhibited reduced potential for toxicity, and cardiotoxicity in particular (a phenomenon associated with most VDAs) as a result of its metabolic stability in plasma and normal tissues [2]. ICT2588 is progressing towards clinical evaluation in the UK in 2019. In this study we aim to further investigate the cellular uptake and metabolism of ICT2588 and related compounds in MT1-MMP positive (HT1080) and MT1-MMP negative (MCF-7) cells in vitro. ICT2588 and related compounds were synthesized in-house and purified by preparative HPLC in excellent yield and purity. Activation of ICT2588 by recombinant MT1-MMP and its preferential metabolism in MT1-MMP positive tumor xenograft homogenates has been shown previously [1]. We are currently studying the mechanism of cellular uptake and metabolism of these peptide prodrugs by incubating these compounds in HT1080 and MCF-7 cell cultures. Cellular concentrations of ICT2588 and all metabolites identified were determined using RP-HPLC, mass spectrometry and immunofluorescent microscopy. We describe the first data on the cellular mechanism of uptake and metabolism of ICT2588 and related peptide prodrugs in cancer cells. Our findings have important implications in the future design of effective peptide-based tumor-targeted prodrugs. Reference: [1] Atkinson, J.M et al “Development of a tumor-targeted vascular disrupting agent activated by Membrane-type Matrix Metalloproteinases (MT-MMPs).” Cancer Research, 2010, 70, 6902-12. [2] Gill, J.H et al “Tumor-targeted prodrug ICT2588 demonstrates therapeutic activity against solid tumors and reduced potential for cardiovascular toxicity”. Mol Pharm. 2014,11:1294-300. Citation Format: Francis Mprah Barnieh, Amanda D. Race, Steven D. Shnyder, Paul M. Loadman, Robert A. Falconer. Investigating the mechanisms of cellular uptake and metabolism of ICT2588, an MT-MMP-activated prodrug [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 350.

Abstract CT067: A Phase I study of Fosbretabulin in combination with everolimus in neuroendocrine tumors (Grades 1-3) that have progressed after at least one prior regimen for metastatic disease

Abstract Background: Based on the SEER database, the incidence of gastroenteropancreatic neuroendocrine tumors (GEPNETs) has risen six-fold in the last four decades. At present, more than 150,000 GEPNET patients exist in the United States. Fosbretabulin, a synthetic, water-soluble, phosphorylated prodrug of the natural product combretastatin A4 (CA4P), initially isolated from the bark of the South African bush willow, Combretum caffrum, is the lead compound in a class of agents termed vascular disrupting agents (VDAs) and has shown activity as a single agent in ovarian cancer and GEPNETs. Everolimus, an mTOR inhibitor, is FDA approved for the management of well-differentiated NETs. A Phase I trial combining fosbretabulin and everolimus to determine the recommended Phase II trial dose (RP2D) and safety data in metastatic GEPNET patients was conducted. Method: An investigator-initiated, single center, open-label, phase I study involving GEPNETs incorporated partial order continual reassessment method (PO-CRM) to define the dose escalation. The primary objective was to establish the maximum tolerated dose (MTD) of the combination of everolimus and fosbretabulin in NETs that have progressed after at least one prior regimen for metastatic disease. Secondary objective included identifying the safety profile of the combination using NCI CTCAE4 reporting criteria. Patients received daily oral everolimus (2.5 mg, 5 mg, 7.5 mg, and 10 mg). Fosbretabulin was administered IV 60 mg/m2 either q3 weekly or q weekly based on PO-CRM. Patients were treated for 12 weeks with all combinations. Results: Of the 17 patients enrolled, 16 completed the 12-week trial. One patient was not evaluable due to noncompliance with the treatment regimen. No DLTs were observed at day 21. The highest dose of 10 mg daily oral everolimus in combination with weekly 60mg/m2 IV fosbretabulin is the RP2D. No grade 4 or 5 toxicities were noted. Grade 3 toxicities were seen in 5 patients that include increased abdominal pain and hyperglycemia (not related to study drug), fatigue (possibly related), decreased lymphocyte count and anemia (related). Several patients had delay in treatment due to grade 2 AE’s (GI symptoms, rash, thrombocytopenia) and one patient was unable to complete treatment due to pneumonitis. Only one patient had radiologic progression at the first q 3 monthly CT scan of chest, abdomen, and pelvis. A detailed table with all grade toxicities will be presented at the meeting. Conclusion: Ten mg PO daily everolimus plus 60 mg/m2 fosbretabulin IV weekly is the RP2D. ClinicalTrials.gov Identifier: NCT0301429 Citation Format: Aman Chauhan, Susanne Arnold, John Wu, Stacey Slone, Emily Dressler, Heather Flynn, Val Adams, Heidi Weiss, Lowell Anthony. A Phase I study of Fosbretabulin in combination with everolimus in neuroendocrine tumors (Grades 1-3) that have progressed after at least one prior regimen for metastatic disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT067.

Anti-angiogenesis Therapy in Cancer Treatment

Anti-angiogenesis Therapy in Cancer Treatment

Vascular Disrupting Agents in cancer treatment: Cardiovascular toxicity and implications for co-administration with other cancer chemotherapeutics.

Destruction of the established tumour vasculature by a class of compound termed Vascular Disrupting Agents (VDAs) is showing considerable promise as a viable approach for the management of solid tumours. VDAs induce a rapid shutdown and collapse of tumour blood vessels, leading to ischaemia and consequent necrosis of the tumour mass. Their efficacy is hindered by the persistence of a viable rim of tumour cells, supported by the peripheral normal vasculature, necessitating their co-administration with additional chemotherapeutics for maximal therapeutic benefit. However, a major limitation for the use of many cancer therapeutics is the development of life-threatening cardiovascular toxicities, with significant consequences for treatment response and the patient's quality of life. The aim of this review is to outline VDAs as a cancer therapeutic approach and define the mechanistic basis of cardiovascular toxicities of current chemotherapeutics, with the overall objective of discussing whether VDA combinations with specific chemotherapeutic classes would be good or bad in terms of cardiovascular toxicity.

Clinical efficacy and toxicity data on phase I study of fosbretabulin in combination with everolimus in neuroendocrine tumors.

4114 Background: Fosbretabulin, a synthetic, water-soluble, phosphorylated prodrug of the natural product combretastatin A4 (CA4P), initially isolated from the bark of the South African bush willow, Combretum caffrum, is the lead compound in a class of agents termed vascular disrupting agents (VDAs). Everolimus, an mTOR inhibitor, is FDA approved for the management of well-differentiated NETs. A Phase I trial combining fosbretabulin and everolimus to determine the recommended Phase II trial dose (RP2D), safety data and early clinical efficacy in metastatic GEPNET patients was conducted. Methods: An investigator-initiated, single center, open-label, phase I study involving GEPNETs incorporated partial order continual reassessment method (PO-CRM) to define the dose escalation. The primary objective was to establish the maximum tolerated dose (MTD) of the combination of everolimus and fosbretabulin in NETs that have progressed after at least one prior regimen for metastatic disease. Secondary objective included identifying the safety profile of the combination using NCI CTCAE4 reporting criteria. Patients received daily oral everolimus (2.5 mg, 5 mg, 7.5 mg, and 10 mg). Fosbretabulin was administered IV 60 mg/m2 either q3 weekly or q weekly based on PO-CRM. Patients were treated for 12 weeks with all combinations. RECIST 1.1 was used to evaluate radiological responses at 3 month. Results: Of the 17 patients enrolled, 16 completed the 12-week trial. One patient was not evaluable due to noncompliance. No DLTs were observed at day 21. The highest dose of 10 mg daily oral everolimus in combination with weekly 60mg/m2 IV fosbretabulin is the RP2D. No grade 4 or 5 toxicities were noted. Grade 3 toxicities were seen in 5 patients; abdominal pain and hyperglycemia (not related to study drug), fatigue (possibly related), decreased lymphocyte count and anemia (related). Several patients had delay in treatment due to grade 2 AE’s (GI symptoms, rash, thrombocytopenia) and one patient was unable to complete treatment due to pneumonitis. All evaluable patients except one had stable disease at 3 months. One patient showed SD but non target lesion demonstrated PD. One patient had > 30% decrease in tumor size but overall sum of lesions showed SD. A detailed table with all grade toxicities and waterfall plot of RR will be presented at the meeting. Conclusions: Ten mg PO daily everolimus plus 60 mg/m2 fosbretabulin IV weekly is the RP2D. Early clinical data suggests clinical activity and stable disease in all but one patient at 3 months. Clinical trial information: NCT0301429.

Role of Angiopoietin-2 in Vascular Physiology and Pathophysiology.

Angiopoietins 1–4 (Ang1–4) represent an important family of growth factors, whose activities are mediated through the tyrosine kinase receptors, Tie1 and Tie2. The best characterized are angiopoietin-1 (Ang1) and angiopoietin-2 (Ang2). Ang1 is a potent angiogenic growth factor signaling through Tie2, whereas Ang2 was initially identified as a vascular disruptive agent with antagonistic activity through the same receptor. Recent data demonstrates that Ang2 has context-dependent agonist activities. Ang2 plays important roles in physiological processes and the deregulation of its expression is characteristic of several diseases. In this review, we summarize the activity of Ang2 on blood and lymphatic endothelial cells, its significance in human physiology and disease, and provide a current view of the molecular signaling pathways regulated by Ang2 in endothelial cells.

Structure Guided Design, Synthesis, and Biological Evaluation of Novel Benzosuberene Analogues as Inhibitors of Tubulin Polymerization.

A promising design paradigm for small-molecule inhibitors of tubulin polymerization that bind to the colchicine site draws structural inspiration from the natural products colchicine and combretastatin A-4 (CA4). Our previous studies with benzocycloalkenyl and heteroaromatic ring systems yielded promising inhibitors with dihydronaphthalene and benzosuberene analogues featuring phenolic (KGP03 and KGP18) and aniline (KGP05 and KGP156) congeners emerging as lead agents. These molecules demonstrated dual mechanism of action, functioning both as potent vascular disrupting agents (VDAs) and as highly cytotoxic anticancer agents. A further series of analogues was designed to extend functional group diversity and investigate regioisomeric tolerance. Ten new molecules were effective inhibitors of tubulin polymerization (IC50 < 5 μM) with seven of these exhibiting highly potent activity comparable to CA4, KGP18, and KGP03. For one of the most effective agents, dose-dependent vascular shutdown was demonstrated using dynamic bioluminescence imaging in a human prostate tumor xenograft growing in a rat.

PI3Kgamma Inhibitor Attenuates Immunosuppressive Effect of Poly(l-Glutamic Acid)-Combretastatin A4 Conjugate in Metastatic Breast Cancer.

Vascular disrupting agents (VDAs) have great potential for cancer treatment. Poly(l‐glutamic acid)‐combretastatin A4 conjugate (PLG‐CA4) is a novel class of VDAs. Though it has notable antitumor activity, it can induce host immune responses that promote tumor growth. Here, PLG‐CA4 induces the polarization of tumor‐associated macrophages (TAMs) toward the M2‐like phenotype in 4T1 metastatic breast cancer (Control 30% vs PLG‐CA4 53%; p < 0.05). Compared to the monotherapy of PLG‐CA4, inhibition of phosphoinositide 3‐kinase gamma (PI3Kγ) attenuates the immunosuppressive effect of PLG‐CA4 treatment by decreasing the number of M2‐like TAMs (2.0 × 104 to 1.5 × 104 per tumor) and potential enhancement of cytotoxic T lymphocyte (3.0 × 104 to 5.7 × 104 per tumor). Importantly, PI3Kγ inhibitor synergizing with PLG‐CA4 significantly extends the mean survival time from 52 days in monotherapy‐treated mice to 61.8 days. Additionally, the combination of PLG‐CA4 and PI3Kγ inhibitor improves the tumor therapeutic effect of NLG919, an inhibitor of immune checkpoint indoleamine 2,3‐dioxygenase (IDO). As far as it is known, this is the first demonstrated study that VDAs induce the reshaping of macrophages to the M2‐like phenotype. The findings also indicate a potential therapeutic strategy of the combination VDAs with an accurate immune modifier in the tumor to reverse the immune resistance.

Cytotoxic Stilbenes and Derivatives as Promising Antimitotic Leads for Cancer Therapy

The growing incidence of cancer, the toxic side-effects associated with conventional chemotherapeutic agents and the development of multidrug resistance (MDR) drive the search for novel and more effective drugs with multi-target activity and selectivity towards cancer cells. Stilbenes are a group of naturally occurring phenolic compounds of plant origin derived from the phenylpropanoid pathway that may exist as cis- or trans-isomers. Although the trans-isomer is the more common and stable configuration, resveratrol being a representative compound, cis-stilbenes are potent cytotoxic agents that bind to and inhibit tubulin polymerization, destabilizing microtubules. This review summarizes the chemistry and biological evaluation of cytotoxic stilbenes and their synthetic derivatives as promising antimitotic leads for cancer therapy, focusing on the most potent compounds, the combretastatins. Combretastatins isolated from the South African bushwillow Combretum caffrum are among the most potent antimitotic and vascular disrupting agents (VDAs) of natural origin. Preclinical studies have demonstrated their potent antitumor effects in a wide variety of tumors, both in vitro and in vivo, being currently under evaluation in phase 2 and phase 3 clinical trials for several types of solid tumors. Topics covered herein include synthetic medicinal chemistry, modes of action, structure-activity relationships (SAR), preclinical and clinical studies as VDAs in cancer therapy, either as single agents or in combination with cytotoxic anticancer drugs, antiangiogenic agents, or radiation therapy, and development of appropriate formulations based on nanocarriers (e.g., liposomes, nanoemulsions, polymeric, lipid and ceramic nanoparticles, carbon nanotubes) for improved bioavailability and targeted delivery of combretastatins to the tumor vasculature.

Selectively Potentiating Hypoxia Levels by Combretastatin A4 Nanomedicine: Toward Highly Enhanced Hypoxia-Activated Prodrug Tirapazamine Therapy for Metastatic Tumors.

Hypoxia-activated prodrugs (HAPs) have the potential to selectively kill hypoxic cells and convert tumor hypoxia from a problem to a selective treatment advantage. However, HAPs are unsuccessful in most clinical trials owing to inadequate hypoxia within the treated tumors, as implied by a further substudy of a phase II clinical trial. Here, a novel strategy for the combination of HAPs plus vascular disrupting agent (VDA) nanomedicine for efficacious solid tumor therapy is developed. An effective VDA nanomedicine of poly(l-glutamic acid)-graft-methoxy poly(ethylene glycol)/combretastatin A4 (CA4-NPs) is prepared and can selectively enhance tumor hypoxia and boost a typical HAP tirapazamine (TPZ) therapy against metastatic 4T1 breast tumors. After treatment with the combination of TPZ plus CA4-NPs, complete tumor reduction is observed in 4T1 xenograft mice (initial tumor volume is 180 mm3 ), and significant tumor shrinkage and antimetastatic effects are observed in challenging large tumors with initial volume of 500 mm3 . The report here highlights the potential of using a combination of HAPs plus VDA nanomedicine in solid tumor therapy.

Safety and Tolerability of Anti-Angiogenic Protein Kinase Inhibitors and Vascular-Disrupting Agents in Cancer: Focus on Gastrointestinal Malignancies.

Angiogenesis is an essential process for tumor growth and metastasis. Inhibition of angiogenesis as an anticancer strategy has shown significant results in a plethora of tumors. Anti-angiogenic agents are currently part of many standard-of-care options for several metastatic gastrointestinal cancers. Bevacizumab, aflibercept, ramucirumab, and regorafenib have significantly improved both progression-free and overall survival in different lines of treatment in metastatic colorectal cancer. Second-line ramucirumab and third-line apatinib are effective anti-angiogenic treatments for patients with metastatic gastric cancer. Unfortunately, the anti-angiogenic strategy has major practical limitations: resistance inevitably develops through redundancy of signaling pathways and selection for subclonal populations adapted for hypoxic conditions. Anti-angiogenic agents may be more effective in combination therapies, with not only cytotoxics but also other emerging compounds in the anti-angiogenic class or in the separate class of the so-called vascular-disrupting agents. This review aims to provide an overview of the approved and "under development" anti-angiogenic compounds as well as the vascular-disrupting agents in the treatment of gastrointestinal cancers, focusing on the actual body of knowledge available on therapy challenges, pharmacodynamic and pharmacokinetic mechanisms, safety profiles, promising predictive biomarkers, and future perspectives.

Synthesis and biological evaluation of structurally diverse α-conformationally restricted chalcones and related analogues.

Numerous members of the combretastatin and chalcone families of natural products function as inhibitors of tubulin polymerization through a binding interaction at the colchicine site on β-tubulin. These molecular scaffolds inspired the development of many structurally modified derivatives and analogues as promising anticancer agents. A productive design blueprint that involved molecular hybridization of the pharmacophore moieties of combretastatin A-4 (CA4) and the chalcones led to the discovery of two promising lead molecules referred to as KGP413 and SD400. The corresponding water-soluble phosphate prodrug salts of KGP413 and SD400 selectively damaged tumor-associated vasculature, thus highlighting the potential development of these molecules as vascular disrupting agents (VDAs). These previous studies prompted our current investigation of conformationally restricted chalcones. Herein, we report the synthesis of cyclic chalcones and related analogues that incorporate structural motifs of CA4, and evaluation of their cytotoxicity against human cancer cell lines [NCI-H460 (lung), DU-145 (prostate), and SK-OV-3 (ovarian)]. While these molecules proved inactive as inhibitors of tubulin polymerization (IC50 > 20 μM), eight molecules demonstrated good antiproliferative activity (GI50 < 20 μM) against all three cancer cell lines, and compounds 2j and 2l demonstrated sub-micromolar cytotoxicity. To the best of our knowledge these molecules represent the most potent (based on GI50) cyclic chalcones known to date, and are promising lead molecules for continued investigation.

An eximious and affordable GSH stimulus-responsive poly(α-lipoic acid) nanocarrier bonding combretastatin A4 for tumor therapy.

Due to poor penetration of cytotoxic-drug-loaded-nanomedicines, more and more attention has been paid to nanodrugs of vascular disrupting agents (VDAs). However, traditional VDA nanodrugs lack tumor-selectivity, so new nano-carriers for tumor-selective CA4 delivery are urgently needed. Here, a novel PEGylated poly(alpha-lipoic acid) graft combretastatin A4 (PALA-g-mPEG/CA4) nanoparticle with glutathione (GSH) stimulus responsive ability was prepared from alpha-lipoic acid in a simple approach, which can accumulate and release CA4 selectively in a tumor site. Furthermore, this simple system has potential application value for tumor-targeting delivery and GSH sensitive release of other drugs.

Efficient synthetic methodology for the construction of dihydronaphthalene and benzosuberene molecular frameworks

Benzosuberene analogues (1 and 2) and dihydronaphthalene analogues (3 and 4) function as potent inhibitors of tubulin polymerization, demonstrate pronounced cytotoxicity (low nM to pM range) against human cancer cell lines, and are promising vascular disrupting agents (VDAs). As such, these compounds represent lead anticancer agents with potential translatability towards the clinic. Methodology previously established by us (and others) facilitated synthetic access to a variety of structural and functional group modifications necessary to explore structure activity relationship considerations directed towards the development of these (and related) molecules as potential therapeutic agents. During the course of these studies it became apparent that the availability of synthetic methodology to facilitate direct conversion of the phenolic-based compounds to their corresponding aniline congeners would be beneficial. Accordingly, modified synthetic routes toward these target phenols (benzosuberene 1 and dihydronaphthalene 3) were developed in order to improve scalability and overall yield [45-57% (1) and 32% (3)]. Moreover, benzosuberene-based phenolic analogue 1 and separately dihydronaphthalene-based phenolic analogue 3 were successfully converted into their corresponding aniline analogues 2 and 4 in good yield (>60% over three steps) using a palladium catalyzed amination reaction.

1,1-Diheterocyclic Ethylenes Derived from Quinaldine and Carbazole as New Tubulin-Polymerization Inhibitors: Synthesis, Metabolism, and Biological Evaluation.

We report the synthesis and metabolic and biological evaluation of a series of 17 novel heterocyclic derivatives of isocombretastatin-A4 (iso-CA-4) and their structure-activity relationships. Among these derivatives, the most active compound, 4f, inhibited the growth of a panel of seven cancer cell lines with an IC50 in the low nanomolar range. In addition, 4f showed interesting activity against CA-4-resistant colon-carcinoma cells and multidrug-resistant leukemia cells. It also induced G2/M cell-cycle arrest. Structural data indicated binding of 4f to the colchicine site of tubulin, likely preventing the curved-to-straight tubulin structural changes that occur during microtubule assembly. Also, 4f disrupted the blood-vessel-like assembly formed by human umbilical-vein endothelial cells in vitro, suggesting its function as a vascular-disrupting agent. An in vitro metabolism study of 4f showed its high human-microsomal stability in comparison with that of iso-CA-4. The physicochemical properties of 4f may be conducive to CNS permeability, suggesting that this compound may be a possible candidate for the treatment of glioblastoma.

New naphthopyran analogues of LY290181 as potential tumor vascular-disrupting agents

A series of 19 analogues of the antiproliferative naphthopyran LY290181 were prepared for structure–activity relationship studies. We found the best activities for test compounds bearing small substituents at the meta position of the phenyl ring. The mode of action of LY290181 and eight new analogues was studied in detail. The compounds were highly anti-proliferative with IC50 values in the sub-nanomolar to triple-digit nanomolar range. The new analogues led to G2/M arrest due to interruption of the microtubule dynamics. In 518A2 melanoma cells they caused a mitotic catastrophe which eventually led to apoptosis. The naphthopyrans also induced a disruption of the vasculature in the chorioallantoic membrane (CAM) of fertilized chicken eggs as well as in xenograft tumors in mice. In a preliminary therapy trial, the difluoro derivative 2b retarded the growth of resistant xenograft tumors in mice.

Gadofosveset-enhanced MRI as simple surrogate parameter for real-time evaluation of the initial tumour vessel infarction by retargeted tissue factor tTF-NGR.

Truncated tissue factor (tTF)-NGR consists of the extracellular domain of the human TF and the binding motif NGR. tTF-NGR activates blood coagulation within the tumour vasculature following binding to CD13, and is overexpressed in the endothelial cells of tumour vessels, resulting in tumour vessel infarction and subsequent retardation/regression of tumour growth. The aim of the present study was to investigate gadofosveset-based real-time dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in evaluating the initial therapeutic effects of the anti-vascular tTF-NGR approach. DCE-MRI (3.0 T) was performed in human U87-glioblastoma tumour-bearing nude mice. During a dynamic T1w GE-sequence, a gadolinium-based blood pool contrast agent (gadofosveset) was injected via a tail vein catheter. Following the maximum contrast intensity inside the tumour being obtained, tTF-NGR was injected (controls received NaCl) and the contrast behaviour of the tumour was monitored by ROI analysis. The slope difference of signal intensities between controls and the tTF-NGR group was investigated, as well as the differences between the average area under the curve (AUC) of the two groups. The association between intensity, group (control vs. tTF-NGR group) and time was analysed by fitting a linear mixed model. Following the injection of tTF-NGR, the signal intensity inside the tumours exhibited a statistically significantly stronger average slope decrease compared with the signal intensity of the tumours in the NaCl group. Furthermore, the initial average AUC values of mice treated with tTF-NGR were 5.7% lower than the average AUC of the control animals (P<0.05). Gadofosveset-enhanced MRI enables the visualization of the initial tumour response to anti-vascular treatment in real-time. Considering the clinical application of tTF-NGR, this method may provide a simple alternative parameter for monitoring the tumour response to vascular disrupting agents and certain vascular targeting agents in humans.

Noninvasive Anatomical and Functional Imaging of Orthotopic Glioblastoma Development and Therapy using Multispectral Optoacoustic Tomography

PURPOSE: Here we demonstrate the potential of multispectral optoacoustic tomography (MSOT), a new non-invasive structural and functional imaging modality, to track the growth and changes in blood oxygen saturation (sO2) in orthotopic glioblastoma (GBMs) and the surrounding brain tissues upon administration of a vascular disruptive agent (VDA). METHODS: Nude mice injected with U87MG tumor cells were longitudinally monitored for the development of orthotopic GBMs up to 15 days and observed for changes in sO2 upon administration of combretastatin A4 phosphate (CA4P, 30 mg/kg), an FDA approved VDA for treating solid tumors. We employed a newly-developed non-negative constrained approach for combined MSOT image reconstruction and unmixing in order to quantitatively map sO2 in whole mouse brains. RESULTS: Upon longitudinal monitoring, tumors could be detected in mouse brains using single-wavelength data as early as 6 days post tumor cell inoculation. Fifteen days post-inoculation, tumors had higher sO2 of 63 ± 11% (n = 5, P < .05) against 48 ± 7% in the corresponding contralateral brain, indicating their hyperoxic status. In a different set of animals, 42 days post-inoculation, tumors had lower sO2 of 42 ± 5% against 49 ± 4% (n = 3, P < .05) in the contralateral side, indicating their hypoxic status. Upon CA4P administration, sO2 in 15 days post-inoculation tumors dropped from 61 ± 9% to 36 ± 1% (n = 4, P < .01) within one hour, then reverted to pre CA4P treatment values (63 ± 6%) and remained constant until the last observation time point of 6 hours. CONCLUSION: With the help of advanced post processing algorithms, MSOT was capable of monitoring the tumor growth and assessing hemodynamic changes upon administration of VDAs in orthotopic GBMs.

Abstract 3533: Elevated expression of heme related proteins in viable tumor cells treated with vascular disrupting agent, combretastatin A-4 phosphate in xenograft models of NSCLC

Abstract Tumors cannot grow without a supply of oxygen and nutrients. Hence, targeting tumor vasculature with Vascular Disrupting Agents (VDAs) represents a promising therapeutic strategy, especially for the treatment of solid tumors. However, VDAs cause central necrosis, leaving a rim of viable tumor cells that often eventually lead to tumor recurrence and development of resistance. Here, we aim to better understand the underlying molecular mechanisms leading to the acquisition of VDA-resistance and its correlation with the extent of hypoxia and respiratory functions. A well-known VDA, Combretastatin A-4 phosphate (CA4P), was used for the study. First, a NSCLC cell line expressing luciferase was implanted into the right flanks of female SCID mice. Next, CA4P was administered intraperitoneally. The effect was monitored 3 and 24 hours post injection of CA4P through BLI (Perkin Elmer's IVIS Lumina III) after subcutaneous injection of luciferin. After sacrificing the mice, the tumors were harvested, formalin-fixed, and paraffin-embedded for Immunohistochemistry (IHC). Our BLI data indicated significant decrease in radiance 3 hour post treatment (total flux in photons per second) which remained so at 24 hours, when luciferin was re-administered. Our IHC data indicated enhanced levels of enzymes involved in heme biosynthesis, uptake and degradation, and hemoproteins involved in oxidative phosphorylation. Furthermore, we examined the effect of a mitochondrial targeting agent (MTA) that undermined the expression of these proteins, on the progression of subcutaneous tumor xenografts. Citation Format: Sanchareeka Dey, Sharda Kumari, Sarada Preeta Kalainayakan, Poorva Ghosh, Li Zhang, Li Liu. Elevated expression of heme related proteins in viable tumor cells treated with vascular disrupting agent, combretastatin A-4 phosphate in xenograft models of NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3533.