Chemotherapy of vascularised tumours: Role of vessel density and the effect of vascular “pruning”

In this work we propose to model chemotherapy taking into account the mutual interaction between tumour growth and the development of tumour vasculature. By adopting a simple model for this interaction, and assuming that the efficacy of a drug can be modulated by the vessel density, we study the constant continuous therapy, the periodic bolus-based therapy, and combined therapy in which a chemotherapic drug is associated with an anti-angiogenic agent. The model allows to represent the vessel-disrupting activity of some standard chemotherapic drugs, and shows, in the case of constant continuous drug administration, the possibility of multiple stable equilibria. The multistability suggests an explanation for some sudden losses of control observed during therapy, and for the beneficial effect of vascular “pruning” exerted by anti-angiogenic agents in combined therapy. Moreover, in case of periodic therapies in which the drug amount administered per unit time is constant (“metronomic” delivery), the model predicts a response, as a function of the bolus frequency, significantly influenced by the extent of the anti-angiogenic activity of the chemotherapic drug and by the dependence of the drug efficacy on the vessel density.     

Understanding the role of the tumour vasculature in the transport of drugs to solid cancer tumours

OBJECTIVES: The vasculature of tumours imposes certain barriers that transport of anti-cancer drugs must overcome. Here follows an account of development of a general computational model that describes the mechanisms of drug transport to a solid tumour, with an emphasis on modelling the vasculature using solute transport concepts. MATERIALS AND METHODS: Investigation into the biological parameters that enhance/prevent anticancer drug transport to the tumour provides a means to evaluate the effects of these parameters on the treatment process. Sensitivity analysis of these provides useful insights concerning anticancer drug transport mechanisms from the vasculature to the solid tumour for a non-specified drug and non-specified solid tumour by revealing the conditions that promote or prevent effective drug transport. The effect of the vasculature on transport efficiency is studied using a parametric analysis of some of the transport and biological parameters. Understanding the various transport mechanisms provides a basis to evaluate the effectiveness of the drug treatment a priori. RESULTS: It was found that increases in the capillary hydrostatic pressure, diffusive permeability coefficient and hydraulic conductivity all result in a decrease in tumour size. Similarly, decreases in the interstitium hydrostatic pressure and filtration constant result in a decrease in tumour size. Dependence of the change in the tumour size to changes in these parameters is non-linear. These results demonstrate the potential of the integrated computational model of the tumour and its vasculature to estimate efficacy of a particular treatment process. Regardless of the dependency of the outcome on the assumed model parameters and the assumed kinetics, mathematical models of this type can provide more explanation on the issues related to the transport barriers, the efficacy of the treatment, and the development of effective anticancer drugs. A case study also is presented to demonstrate the model's flexibility to accommodate a two-cell-glioma population.     

Mathematical model for chemotherapeutic drug efficacy in arresting tumour growth based on the cancer stem cell hypothesis

OBJECTIVE: Cancer stem cells have been identified as the growth root for various malignant tumours and are thought to be responsible for cancer recurrence following treatment. MATERIALS AND METHODS: Here, a predictive mathematical model for the cancer stem cell hypothesis is used to understand tumour responses to chemotherapeutic drugs and judge the efficacy of treatments in arresting tumour growth. The impact of varying drug efficacies on different abnormal cell populations is investigated through the kinetics associated with their decline in response to therapy. RESULTS AND CONCLUSIONS: The model predicts the clinically established 'dandelion phenomenon' and suggests that the best response to chemotherapy occurs when a drug targets abnormal stem cells. We compare continuous and periodic drug infusion. For the latter, we examine the relative importance of the drug cell-kill rate and the mean time between successive therapies, to identify the key attributes for successful treatment.     

Tumours can adapt to anti-angiogenic therapy depending on the stromal context: lessons from endothelial cell biology

It has long been recognized that interference with the blood supply of a tumour is an effective way to halt tumour progression, and even induce tumour regression. This can be accomplished by anti-angiogenic treatment which prevents the formation of a tumour neovasculature, or anti-vascular treatment, which aims at destruction of existent tumour vessels. The latter has received relatively little attention because there is a lack of specific tumour-endothelial markers. Instead, the current detailed knowledge on the factors and mechanisms, involved in angiogenesis, has enabled the development of a variety of angiogenesis inhibitors, especially those that target cellular signalling by vascular endothelial growth factor-A (VEGF-A), the most potent angiogenic factor known. These inhibitors have received lots of attention because they effectively inhibit tumour growth in pre-clinical models. However, in clinical trials these same inhibitors showed very poor anti-tumour activity. In this review we discuss this discrepancy, and we show that the tumour microenvironment is crucial to the sensitivity of tumours to anti-angiogenic therapy.     

Coupled modelling of tumour angiogenesis, tumour growth and blood perfusion

We propose a mathematical modelling system to investigate the dynamic process of tumour cell proliferation, death and tumour angiogenesis by fully coupling the vessel growth, tumour growth and blood perfusion. Tumour growth and angiogenesis are coupled by the chemical microenvironment and the cell-matrix interaction. The haemodynamic calculation is carried out on the updated vasculature. The domains of intravascular, transcapillary and interstitial fluid flow were coupled in the model to provide a comprehensive solution of blood perfusion variables. An estimation of vessel collapse is made according to the wall shear stress criterion to provide feedback on vasculature remodelling. The simulation can show the process of tumour angiogenesis and the spatial distribution of tumour cells for periods of up to 24 days. It can show the major features of tumour and tumour microvasculature during the period such as the formation of a large necrotic core in the tumour centre with few functional vessels passing through, and a well circulated tumour periphery regions in which the microvascular density is high and associated with more aggressive proliferating cells of the growing tumour which are all consistent with physiological observations. The study also demonstrated that the simulation results are not dependent on the initial tumour and networks, which further confirms the application of the coupled model feedback mechanisms. The model enables us to examine the interactions between angiogenesis and tumour growth, and to study the dynamic response of a solid tumour to the changes in the microenvironment. This simulation framework can be a foundation for further applications such as drug delivery and anti-angiogenic therapies.     

Periostin: a putative mediator involved in tumour resistance to anti‐angiogenic therapy?

Despite advances in the development of anti-angiogenic agents for cancer treatment, the increase in the survival duration of cancer patients is still rather modest. One major obstacle in anti-angiogenic therapy is the emergence of drug resistance. Understanding the molecular mechanisms that enable a tumour to evade anti-angiogenic treatment is valuable to improve therapeutic efficacy. Targeting blood supply usually causes hypoxic responses of tumours that trigger a series of adaptive changes leading to a resistant phenotype. Periostin, a secreted ECM (extracellular matrix) protein, is mainly produced by CAFs (cancer-associated fibroblasts) on hypoxic stress. As CAFs have been casually linked to tumour resistance to angiogenesis blockade and periostin can influence many aspects of tumour biology, we hypothesized that periostin might be a crucial mediator involved anti-angiogenic resistance in cancer treatment. This hypothesis is indirectly supported by the following facts: (a) high levels of periostin promote tumour angiogenesis; (b) periostin improves cancer cell survival under hypoxic conditions; and (c) genetic modulation of periostin induces EMT (epithelial-mesenchymal transition) and enhances cancer cell invasion and metastasis, which represents an escape mechanism from anticancer treatment. Testing and confirmation of this hypothesis will give more insight into the resistance mechanisms and provide the rationale for improvement of therapeutic outcome of anti-angiogenic therapy.     

Nanoparticle-mediated gene delivery to tumour neovasculature

The alpha(v)beta(3) integrin is a potential pharmacological target for anti-angiogenic therapy. A recent report describes the use of alpha(v)beta(3)-targeted nanoparticles to deliver a gene to tumour vasculature selectively. This resulted in substantial tumour regression in several experimental mouse tumour models. Hence, this approach has great potential for the treatment of human cancer.     

Anti-Dll4 therapy: can we block tumour growth by increasing angiogenesis?

Since the early 1970s, the dogma postulating that blocking tumour angiogenesis can inhibit tumour growth has been accepted widely and has resulted in the generation of a variety of successful anti-angiogenic therapies. More recently, new signalling pathways, such as the Dll4-Notch signalling pathway, have been shown to regulate angiogenesis during development. In pathological conditions, such as cancer, Dll4 is up-regulated strongly in the tumour vasculature. Based on this expression pattern, different molecules have been generated to block Dll4 signalling. Unexpectedly, these blocking agents inhibited tumour growth in vivo by triggering excessive but nonfunctional angiogenesis. Altogether, these molecules constitute a new category of pro-angiogenic yet anticancer agents and offer an exciting alternative to previously described vascular targeting molecules.     

Manipulation of the tumour-associated vasculature to improve tumour therapy

Inhibition of tumour vascular growth, destruction of the tumour associated vasculature (TAV), and manipulation of the endothelial lining of the TAV provide powerful tools for anti-tumour therapy. We previously demonstrated that addition of TNF to chemotherapy improved tumour response. The major effect of TNF is an increased permeability of the tumour vascular bed resulting in augmented accumulation of co-administered drug in the tumour. As the TAV is recognised as a major candidate in tumour therapy it is becoming important to understand anti-vascular effects better. In our laboratory we examine the effect of immunotherapy on the TAV, and the effect of anti tumour-vascular therapy on tumours. This is studied in animal models, which exhibit similarities with the clinical setting, such as tumour perfusion treatment.     

MANIPULATION OF THE TUMOUR-ASSOCIATED VASCULATURE TO IMPROVE TUMOUR THERAPY

ABSTRACT

Inhibition of tumour vascular growth, destruction of the tumour associated vasculature (TAV), and manipulation of the endothelial lining of the TAV provide powerful tools for anti-tumour therapy. We previously demonstrated that addition of TNF to chemotherapy improved tumour response. The major effect of TNF is an increased permeability of the tumour vascular bed resulting in augmented accumulation of co-administered drug in the tumour. As the TAV is recognised as a major candidate in tumour therapy it is becoming important to understand anti-vascular effects better. In our laboratory we examine the effect of immunotherapy on the TAV, and the effect of anti tumour-vascular therapy on tumours. This is studied in animal models, which exhibit similarities with the clinical setting, such as tumour perfusion treatment.     

Anti-metastatic effects of viral and non-viral mediated Nk4 delivery to tumours

The most common cause of death of cancer sufferers is through the occurrence of metastases. The metastatic behaviour of tumour cells is regulated by extracellular growth factors such as hepatocyte growth factor (HGF), a ligand for the c-Met receptor tyrosine kinase, and aberrant expression/activation of the c-Met receptor is closely associated with metastatic progression. Nk4 (also known as Interleukin (IL)32b) is a competitive antagonist of the HGF c-Met system and inhibits c-Met signalling and tumour metastasis. Nk4 has an additional anti-angiogenic activity independent of its HGF-antagonist function. Angiogenesis-inhibitory as well as cancer-specific apoptosis inducing effects make the Nk4 sequence an attractive candidate for gene therapy of cancer. This study investigates the inhibition of tumour metasasis by gene therapy mediated production of Nk4 by the primary tumour. Optimal delivery of anti-cancer genes is vital in order to achieve the highest therapeutic responses. Non-viral plasmid delivery methods have the advantage of safety and ease of production, providing immediate transgene expression, albeit short-lived in most tumours. Sustained presence of anti-angiogenic molecules is preferable with anti-angiogenic therapies, and the long-term expression mediated by Adeno-associated Virus (AAV) might represent a more appropriate delivery in this respect. However, the incubation time required by AAV vectors to reach appropriate gene expression levels hampers efficacy in many fast-growing murine tumour models. Here, we describe murine trials assessing the effects of Nk4 on the spontaneously metastatic Lewis Lung Carcinoma (LLC) model when delivered to primary tumour via plasmid lipofection or AAV2 vector. Intratumoural AAV-Nk4 administration produced the highest therapeutic response with significant reduction in both primary tumour growth and incidence of lung metastases. Plasmid-mediated therapy also significantly reduced metastatic growth, but with moderate reduction in primary subcutaneous tumour growth. Overall, this study demonstrates the potential for Nk4 gene therapy of metastatic tumours, when delivered by AAV or non-viral methods.     

PEDF: a potential molecular therapeutic target with multiple anti-cancer activities

Pigment epithelium-derived factor (PEDF) is an endogenously produced protein that is widely expressed throughout the human body, and exhibits multiple and varied biological activities. Already established as a potent anti-angiogenic molecule, PEDF has recently shown promise as a potential anti-tumour agent, causing both direct and indirect tumour suppression. Here, we explore the unique anti-tumour properties of PEDF and discuss its role as an effective anti-angiogenic, anti-proliferative and pro-differentiation factor. We also discuss the prospects for PEDF therapy and the need for a closer evaluation of issues such as delivery, stability and potential toxicity.     

Combination of vasostatin gene therapy with cyclophosphamide inhibits growth of B16(F10) melanoma tumours

Angiogenesis, i.e. formation of new blood vessels out of pre-existing capillaries, is essential to the development of tumour vasculature. The discovery of specific antiangiogenic inhibitors has important therapeutic implications for the development of novel cancer treatments. Vasostatin, the N-terminal domain of calreticulin, is a potent endogenous inhibitor of angiogenesis and tumour growth. In our study, using B16(F10) murine melanoma model and electroporation we attempted intramuscular transfer of human vasostatin gene. The gene therapy was combined with antiangiogenic drug dosing schedule of a known chemotherapeutic (cyclophosphamide). The combination of vasostatin gene therapy and cyclophosphamide administration improved therapeutic effects in melanoma tumours. We observed both significant inhibition of tumour growth and extended survival of treated mice. To our knowledge, this is one of the first reports showing antitumour efficacy of electroporation-mediated vasostatin gene therapy combined with antiangiogenic chemotherapy.     

Antibody drug-conjugates targeting the tumor vasculature: Current and future developments

Reducing the blood supply of tumors is one modality to combat cancer. Monoclonal antibodies are now established as a key therapeutic approach for a range of diseases. Owing to the ability of antibodies to selectively target endothelial cells within the tumor vasculature, vascular targeting programs have become a mainstay in oncology drug development. However, the antitumor activity of single agent administration of conventional anti-angiogenic compounds is limited and the improvements in patient survival are most prominent in combinations with chemotherapy. Furthermore, prolonged treatment with conventional anti-angiogenic drugs is associated with toxicity and drug resistance. These circumstances provide a strong rationale for novel approaches to enhance the efficacy of mAbs targeting tumor vasculature such as antibody-drug conjugates (ADCs). Here, we review trends in the development of ADCs targeting tumor vasculature with the aim of informing future research and development of this class of therapeutics.Key words: tumor, vasculature, immunotherapy, antibody-drug conjugates, monoclonal antibody, cancer, angiogenesis     

Predictive Modeling of Neuroblastoma Growth Dynamics in Xenograft Model After Bevacizumab Anti-VEGF Therapy

Neuroblastoma is the leading cause of cancer death in young children. Although treatment for neuroblastoma has improved, the 5-year survival rate of patients still remains less than half. Recent studies have indicated that bevacizumab, an anti-VEGF drug used in treatment of several other cancer types, may be effective for treating neuroblastoma as well. However, its effect on neuroblastoma has not been well characterized. While traditional experiments are costly and time-consuming, mathematical models are capable of simulating complex systems quickly and inexpensively. In this study, we present a model of vascular tumor growth of neuroblastoma IMR-32 that is complex enough to replicate experimental data across a range of tumor cell properties measured in a suite of in vitro and in vivo experiments. The model provides quantitative insight into tumor vasculature, predicting a linear relationship between vasculature and tumor volume. The tumor growth model was coupled with known pharmacokinetics and pharmacodynamics of the VEGF blocker bevacizumab to study its effect on neuroblastoma growth dynamics. The results of our model suggest that total administered bevacizumab concentration per week, as opposed to dosage regimen, is the major determining factor in tumor suppression. Our model also establishes an exponentially decreasing relationship between administered bevacizumab concentration and tumor growth rate.     

Inhibitory effects of novel integrin-targeting peptides on angiogenesis activity in HUVEC cells in vitro

Integrins are critically involved in many tumour-promoting activities. The development of inhibitors against integrins may suppress tumour growth by inhibiting tumour angiogenic signalling. In this study, we investigated the effects of two novel peptides containing the integrin binding arginine-glycine-aspartic acid-motif on inhibiting diverse cell behaviours, including cell adhesion, motility, invasion, tube formation and cell cytoskeleton. Cell adhesion and motility assays demonstrated that cyclopeptides c-Gly and c-Lys might inhibit the adhesive and motile activity at the concentration of 25 μM. There was no significant effect on cell invasion, indicating the importance of extracellular matrix degradation in modulating the anti-invasive effect of human umbilical vein endothelial cells (HUVECs). More importantly, the tubular network formation of HUVECs was significantly inhibited by cyclopeptide c-Lys besides causing a remarkable inhibition of cytoskeletal organization, disrupting the focal adhesion and actin stress fibres formation. In conclusion, this study results indicated that the novel peptide c-Lys has the ability to inhibit diverse cell behaviours of HUVECs, and the effects may be mediated at different levels of the tumour growth. Therefore, c-Lys is perhaps proposed to be a potent anti-angiogenic drug candidate.     

Clinical pharmacokinetics of 5-fluorouracil with consideration of chronopharmacokinetics

Even though 5-fluorouracil (FU) is one of the oldest anticancer drugs, its use in cancer chemotherapy continues to increase. Fluorouracil is a pro-drug that requires intracellular activation to exert its effects. This makes it difficult to associate blood drug concentration with cell toxicity directly, although data from the literature show the existence of such a relationship. The relationship between FU pharmacokinetics and patient response has been explored extensively and reports attest a link between systemic drug exposure and response and survival. This has led to the concept of maximal tolerated exposure, and strategies to achieve this rely on pharmacokinetic follow-up and individual dose adjustment. More than 80% of the administered FU dose is eliminated by catabolism through dihydropyrimidine dehydrogenase (DPD), the rate-limiting enzyme. Dihydropyrimidine dehydrogenase activity is found in most tissues but is highest in the liver. Peripheral blood mononuclear cells (PBMC) are used to monitor clinically DPD activity. A significant, but weak correlation between PBMC and liver DPD activity has been observed. The relationship between PBMC-DPD activity and FU systemic clearance is weak (r²=0.10); thus, simply determining PBMC-DPD is not sufficient to predict accurately FU clearance. Population pharmacokinetic analysis identified patient co-variables that influence FU clearance; drug kinetics is significantly reduced by increased age, high serum alkaline phosphatase, length of drug infusion, and low PBMC-DPD. Autoregulation of FU metabolism also is suggested; inhibition of DPD activity was observed after FU administration in both colorectal cancer patients and an animal model. Circadian rhythmicity in DPD activity is suggested from both human and animal investigations. In patients receiving protracted low dose 5-FU infusion, the circadian rhythm in FU plasma concentration peaks at 11:00h and is lowest at 23:00h, on average. The inverse relationship observed between the circadian profile of FU plasma concentration and PBMC-DP activity in these same patients suggests a link between DPD activity and FU pharmacokinetics. The impact of the biological time of drug administration was also studied with short venous infusions; clearance was 70% greater at 13:00h than at 01:00h. Similarly, peak drug concentration occurred in the first half of the night in patients receiving constant rate 5-FU infusion for 2-5 d. Several studies describe wide interindividual variation in the timing of the peak and trough of the 24h rhythm in DPD activity. The rational for FU chronomodulated therapy has been the circadian rhythm in host drug tolerance, which is greatest during the night time when the proliferation of normal target tissue is least. A randomized study of chronomodulated FU therapy with maximal delivery rate at 04:00h was shown clearly to be significantly more effective and less toxic than control flat FU therapy. Future research must focus on easy-to-obtain markers of specific rhythms to individualize the chronomodulated FU delivery.     

Tumour susceptibility to innate and adaptive immunotherapy changes during tumour maturation

Immunotherapy of tumours using T cells expanded in vitro has met with mixed clinical success suggesting that a greater understanding of tumour/T-cell interaction is required. We used a HPV16E7 oncoprotein-based mouse tumour model to study this further. In this study, we demonstrate that a HPV16E7 tumour passes through at least three stages of immune susceptibility over time. At the earliest time point, infusion of intravenous immune cells fails to control tumour growth although the same cells given subcutaneously at the tumour site are effective. In a second stage, the tumour becomes resistant to subcutaneous infusion of cells but is now susceptible to both adjuvant activated and HPV16E7-specific immune cells transferred intravenously. In the last phase, the tumour is susceptible to intravenous transfer of HPV16E7-specific cells, but not adjuvant-activated immune cells. The requirement for IFN-gamma and perforin also changes with each stage of tumour development. Our data suggest that effective adoptive T-cell therapy of tumour will need to be matched with the stage of tumour development.