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Nanoparticles are suitable platforms for cancer targeting and diagnostic applications. Typically, less than 10% of all systemically administered nanoparticles accumulate in the tumour. Here we explore the interactions of blood components with nanoparticles and describe how these interactions influence solid tumour targeting. In the blood, serum proteins adsorb onto nanoparticles to form a protein corona in a manner dependent on nanoparticle physicochemical properties. These serum proteins can block nanoparticle tumour targeting ligands from binding to tumour cell receptors. Additionally, serum proteins can also encourage nanoparticle uptake by macrophages, which decreases nanoparticle availability in the blood and limits tumour accumulation. The formation of this protein corona will also increase the nanoparticle hydrodynamic size or induce aggregation, which makes nanoparticles too large to enter into the tumour through pores of the leaky vessels, and prevents their deep penetration into tumours for cell targeting. Recent studies have focused on developing new chemical strategies to reduce or eliminate serum protein adsorption, and rescue the targeting potential of nanoparticles to tumour cells. An in-depth and complete understanding of nanoparticle-blood interactions is key to designing nanoparticles with optimal physicochemical properties with high tumour accumulation. The purpose of this review article is to describe how the protein corona alters the targeting of nanoparticles to solid tumours and explains current solutions to solve this problem.

Activatable cell-penetrating peptides (aCPPs) allow non-viral, low cytotoxic and selective delivery of compounds into target cells for cancer therapy. In tumour cells, up-regulation of human telomerase reverse transcriptase (hTERT) frequently occurs and is being considered as a target in cancer diagnosis and treatment. siRNA sequence that target hTERT mRNA can silence the gene and reduce hTERT protein expression to reduce cell proliferation and inhibit cell growth. In our study, we tested a matrix metalloproteinase-2 (MPP2) aCPP in delivering hTERT siRNA into hepatocellular carcinoma cells (SMMC-7721) to silence the hTERT gene. Cultured SMMC-7721 cells were transfected with a complex of aCPPs and hTERT-specific siRNA-encoding or control plasmids. Compared with cells treated with the complex of control plasmid–CPPs, cells treated with the hTERT-specific siRNA-encoding plasmid–CPP complex had a prolonged G1-phase, but a shorter G2/S-phase, indicating a G1-arrest. Treatment with the hTERT-specific siRNA resulted in a significant decrease (by 26%; P, In the present study, we delivered human telomerase reverse transcriptase (hTERT) siRNA into SMMC-7721 hepatoma cells using a matrix metalloproteinase-2 (MMP2)-activatable cell-penetrating peptide (aCPP). The siRNA subsequently induced down-regulation of the hTERT gene and G1-arrest, implicating the utility of this delivery system in cancer therapy.

Targeted radionuclide therapy with high-energy beta-emitters is generally considered suboptimal to cure small tumours (300 mg). Tumour targeting of the CCK2 receptor-binding minigastrin analogue PP-F11 was determined in a tumour-bearing mouse model at increasing peptide amounts. The optimal therapy was analysed for PP-F11 labelled with (90)Y, (177)Lu or (213)Bi, accounting for the radionuclide specific activities (SAs), the tumour absorbed doses and tumour (radio) biology.Tumour uptake of (111)In-PP-F11 was determined in nude mice bearing CCK2 receptor-transfected A431 xenografts at 1 and 4 h post-injection for escalating peptide masses of 0.03 to 15 nmol/mouse. The absorbed tumour dose was estimated, assuming comparable biodistributions of the (90)Y, (177)Lu or (213)Bi radiolabelled peptides. The linear-quadratic (LQ) model was used to calculate the tumour control probabilities (TCP) as a function of tumour mass and growth.Practically achievable maximum SAs for PP-F11 labelled with (90)Y and (177)Lu were 400 MBq (90)Y/nmol and 120 MBq(177)Lu/nmol. Both the large elution volume from the 220 MBq (225)Ac generator used and reaction kinetics diminished the maximum achieved (213)Bi SA in practice: 40 MBq (213)Bi/nmol. Tumour uptakes decreased rapidly with increasing peptide amounts, following a logarithmic curve with ED50 = 0.5 nmol. At 0.03 nmol peptide, the (300 mg) tumour dose was 9 Gy after 12 MBq (90)Y-PP-F11, and for (111)In and (177)Lu, this was 1 Gy. A curative dose of 60 Gy could be achieved with a single administration of 111 MBq (90)Y labelled to 0.28 nmol PP-F11 or with 4 × 17 MBq (213)Bi (0.41 nmol) when its α-radiation relative biological effectiveness (RBE) was assumed to be 3.4. Repeated dosing is preferable to avoid complete tumour receptor saturation. Tumours larger than 200 mg are curable with (90)Y-PP-F11; the other radionuclides perform better in smaller tumours. Furthermore, (177)Lu is not optimal for curing fast-growing tumours.Receptor saturation, specific radiopharmaceutical activities and absorbed doses in the tumour together favour therapy with the CCK2 receptor-binding peptide PP-F11 labelled with (90)Y, despite its longer β-particle range in tissue, certainly for tumours larger than 300 mg. The predicted TCPs are of theoretical nature and need to be compared with the outcome of targeted radionuclide experiments.

Promising results have come from attempts to direct drugs, nucleic acids and diagnostic agents to tumours by using polymeric nanoparticles. Such carriers are versatile; their encapsulation capacity, drug release profile, and biological performance vary with their chemical makeup, morphology, and size. Polymeric nanoparticles may therefore be engineered for therapeutic and diagnostic purposes in accordance with the type, developmental stage and location of the cancer as well as the required route of administration. This article examines recent developments in design and engineering of polymeric nanoparticles and related platforms to include supramolecular systems such as nanocapsules and nanoparticlebased hydrogels, and assesses their potential diagnostic and therapeutic applications in experimental and clinical oncology.

Contains fulltext : 48915.pdf (Publisher’s version ) (Closed access) PURPOSE: An effective pretargeting strategy was developed for renal cell carcinoma (RCC) based on a biologically produced bispecific monoclonal antibody: anti-RCCxanti-DTPA(In) (bsMAb: G250xDTIn-1). Tumour uptake of a (111)In-labelled bivalent peptide after pretargeting with bsMAb G250xDTIn-1 was relatively high compared with that in other pretargeting systems using chemically coupled F(ab')(2) fragments. Here, we investigated the effect of the bsMAb form in the pretargeting strategy. METHODS: To determine the optimal interval between the administration of each of the bsMAb forms and the (111)In-labelled bivalent peptide, the biodistribution of the radioiodinated bsMAb forms was studied in athymic mice with subcutaneous SK-RC-1 RCC tumours. Since tumour targeting of the radiolabelled peptide depends on the bsMAb form and dose, a bsMAb dose escalation study was carried out for both bsMAb forms. Under optimised conditions, the biodistribution of the (111)In label in mice with pretargeted RCC was determined from 4 h up to 7 days p.i. RESULTS: The optimal interval between the two administrations was 72 h for the bsMAb IgG and 4 h for the bsMAb F(ab')(2). The optimal bsMAb dose for intact IgG was 67 pmol and the optimal bsMAb F(ab')(2) dose was 200 pmol. Targeting of the pretargeted RCC with 4 pmol (111)In-labelled bivalent peptide revealed high tumour uptake with both bsMAb forms. CONCLUSION: With the pretargeting strategy, using either bsMAb IgG or bsMAb F(ab')(2), very efficient peptide targeting of the tumour was obtained. Uptake and retention of the radiolabel in the tumour with the pretargeting approach are not affected by the bsMAb form used.

Two neuropeptides, bombesin (BBS) and neurotensin (NT) and their radiolabeled analogues, have great potential for tumour targeting, either for diagnosis (e.g., with 99mTc) or therapy (e.g., with 90Y or 188Re). In this study, we investigated NT(8-13) and BBS(7-14) analogues with Nalpha-histidinyl acetate linked to the N-terminus of the peptide. This His-derivative forms a stable and inert tridentate complex with the 99mTc(CO)3 and the 188Re(CO)3 moieties. The stability of 99mTc-labeled neurotensin and bombesin analogues was tested in human plasma samples and in tumour cell cultures in the presence and absence of specific enzyme inhibitors. The inhibitor of ACE (angiotensin converting enzyme) was the most effective in inhibiting the peptide cleavage of both NT(8-13) and BBS(7-14). In agreement with this finding, the replacement of Ile12 by tert-leucine (NT) and Leu13 by cyclohexylalanin (BBS) brought about a better stability. With NT(8-13) analogues, higher tumour to nontarget (t/nt) ratios and the same affinity to the receptor was observed, but with BBS(7-14) derivatives the affinity was lower and the t/nt ratio was not significantly improved. Toxicity tests showed no effect in mice of up to a five-hundred-fold higher dose than planned for patient application, which started successfully with NT(8-13) analogues.

Specific targeting of radionuclides is a promising approach to improve diagnosis and treatment of tumors. Targeting vectors may be monoclonal antibodies directed toward tumour-specific antigens or regulatory peptides binding to receptors overexpressed on or by malignant cells. Depending on the aim of the procedure and the biokinetics of the targeting vectors, radionuclides with different nuclear properties (decay scheme, half-life, etc.) must be applied. Halogen radioisotopes are attractive since they exhibit a variety of nuclear properties suitable for various applications. At the same time, their chemistry shows great similarities, which enables the use of similar labelling procedures for different nuclides. A problem in using radiohalogens for labelling of tumour-targeting proteins and peptides is that the commonly used radiohalogenation methods provide labels, which, after internalisation and lysosomal digestion, rapidly "leak" from malignant cells as radiohalogenated degradation products. The main reason for such leakage is free diffusion of the radiometabolites through lysosomal and cellular membranes. This review describes current approaches in molecular design to improve cellular retention of radiohalogen labels. These approaches include the use of prosthetic groups for the attachment of radiohalogens to targeting vectors of bulky hydrophilic non-charged molecules, molecules positively charged at lysosomal pH and negatively charged molecules. The emphasis in this paper is on labelling chemistry and the results of the biological testing of labelled compounds.

Live bacteria were first actively used in the treatment of cancer nearly 150 years ago, work that ultimately led to the study of immunomodulation. Today, with the discovery of bacterial strains that specifically target tumours, and aided by genomic sequencing and genetic engineering, there is new interest in the use of bacteria as tumour vectors. Bifodobacterium, Clostridium, and Salmonella have all been shown to preferentially replicate within solid tumours when injected from a distal site, and all three types of bacteria have been used to transport and amplify genes encoding factors such as prodrug-converting enzymes, toxins, angiogenesis inhibitors, and cytokines. In this review we provide a historical discussion of this area, and describe the development of the bacteria, which are currently being prepared for use in clinical trials in patients with cancer.

We prepared small unilamellar liposomes derivatised with single chain antibody fragments specific for the ED-B domain of B-fibronectin. This extracellular matrix associated protein is expressed around newly forming blood vessels in the vicinity of many types of tumours. The single chain antibody fragments were functionalised by introduction of C-terminal cysteines and linked to liposomes via maleimide groups located at the terminal ends of poly(ethylene glycol) modified phospholipids. The properties of these anti-ED-B single chain antibody fragments-liposomes were analysed in vitro on ED-B fibronectin expressing Caco-2 cells and in vivo by studying their biodistribution and their therapeutic potential in mice bearing subcutanous F9 teratocarcinoma tumours. Radioactively labelled (114mIndium) single chain antibody fragments-liposomes accumulated in the tumours at 2–3-fold higher concentrations during the first 2 h after i.v. injection compared to unmodified liposomes. After 6–24 h both liposome types were found in similar amounts (8–10% injected dose g−1) in the tumours. Animals treated i.v. with single chain antibody fragments-liposomes containing the new cytotoxic agent 2′-deoxy-5-fluorouridylyl-N4-octadecyl-1-β-D-arabinofuranosylcytosine (30 mg kg-1 per dose, five times every 24 h) showed a reduction of tumour growth by 62–90% determined on days 5 and 8, respectively, compared to animals receiving control liposomes. Histological analysis revealed a marked reduction of F9 tumour cells and excessive deposition of fibronectin in the extracellular matrix after treatment with single chain antibody fragments-2-dioxy-5-fluorouridylyl-N4-octadecyl-1-β-D-arabinofuranosylcytosine-liposomes. Single chain antibody fragments-liposomes targeted to ED-B fibronectin positive tumours therefore represent a promising and versatile novel drug delivery system for the treatment of tumours. British Journal of Cancer (2002) 87, 106–112. doi:10.1038/sj.bjc.6600423 www.bjcancer.com © 2002 Cancer Research UK

Polyhedral boron clusters (PBC) are three-dimensional inorganic aromatic systems. Some of them can easily be halogenated, and the halogen-boron bond in such systems is very strong. We consider the use of PBC as linkers for attachment of radioactive halogen isotopes to tumor-targeting proteins and peptides. In this review the major preconditions for such applications, such as biological considerations, knowledge concerning coupling chemistry and radiolabeling of PBC, are described. A review with 90 references.

Despite improvements in our understanding of cancer and the concept of personalised medicine, cancer is still a major cause of death. It is established that solid tumours are highly heterogeneous, with a complex tumour microenvironment. Indeed, the tumour microenvironment is made up of a collection of immune cells, cancer-activated fibroblasts, and endothelial cells and in some cases a dense extracellular matrix. Accumulating evidence shows that the tumour microenvironment is a major barrier for the effective delivery of therapeutic drugs to tumour cells. Importantly, nanotechnology has come to the forefront as highly effective delivery vehicles for therapeutic agents. This perspective will discuss how nanomedicine can be used to target and deliver therapeutic drugs specifically to tumour cells. Moreover, emerging opportunities to modulate the tumour microenvironment and increase the delivery and efficacy of chemotherapy agents to solid tumours will be highlighted. From the Clinical Editor Improving drug delivery to treatment resistant tumors is a major target of many nanomedicine-based applications. This comprehensive review discusses the currently available and emerging opportunities, in addition to discussing tumor microenvironment modulation to facilitate efficient delivery.

A porous silicon (PSi) that is a biodegradable semiconductor material with nanoporous holes has been extensively developed for biomedical applications over the past few decades. In particular, researchers have attempted to use the PSi material for the detection and visualization of tumours, since it has excellent biocompatibility and intrinsic photoluminescence. This chapter will summarize examples of PSi materials that have been utilized to target and image tumours and discuss their impact on cancer diagnosis. The PSi microparticles or nanoparticles that can be systemically introduced into the biological system will be discussed with regard to their impact on tumour targeting and imaging.

Microtubule-targeting agents (MTAs) are amongst the most successful chemotherapeutic drugs commonly used in the clinic for the treatment of human cancers. Although originally administered at or close to the maximum tolerated dose once every 3 weeks, the discovery of their potent antiangiogenic properties at the end of the 1990s has led to the re-evaluation of treatment protocols. Nowadays, MTAs are often administered at lower doses either weekly or even more frequently following a metronomic schedule, thus leading to increased efficacy and decreased toxicity. In this chapter, we present an overview of the in vitro and in vivo studies that have contributed to the development of MTA-based metronomic chemotherapy protocols and increased our understanding of their mechanisms of action. First, we discuss the complex cellular and molecular mechanisms involved in the antiangiogenic activity of MTAs. We also present their effects on the immune system, which may contribute to the antitumour efficacy of MTA-based metronomic chemotherapy. Then, we review the results obtained with this type of therapeutic approach in preclinical models of human cancer, focusing on the most promising combination treatments. Finally, we oversee the future developments in this field in terms of new MTAs and novel formulations currently in development with the aims to improve efficacy and bioavailability while increasing tumour targeting and specificity.

The contribution of nuclear medicine to the diagnosis and treatment of endocrine malignancy is increasing. Advances in molecular biology offer new opportunities for tumour targeting via surface receptor recognition and tumour-specific metabolic markers. Imaging the biodistribution of these markers allows quantitative, in vivo characterization of tumour function. There is growing interest in the therapeutic potential of nuclear medicine targeting, substituting therapeutic beta-emitting radionuclides for the gamma-emitters used in diagnostic imaging. Limited clinical experience supports the rationale of this approach in patients with inoperable or disseminated disease and controlled trials are in progress. This chapter outlines the place of nuclear medicine techniques in the routine management of endocrine malignancy and explores areas for further development.

Immunoliposomes (antibody-coupled liposomes) have been regarded as very attractive drug-targeting systems for chemotherapeutic cancer treatment. Fundamental problems regarding immunoliposome preparation and application such as antibody coupling and immunoliposome stability and pharmacokinetics have been overcome during the last decade. Therefore, several promising studies on tumour targeting have been described in recent years. Adding to existing reviews on liposomal drug delivery, this article focuses on immunoliposome tumour targeting and summarises various experiments of immunoliposome application in vitro and in vivo with respect to structural liposomal parameters, therapeutic potential and the requirements of the target sites. New therapeutic trends related to immunoliposomes are also considered. Remaining problems in immunoliposome application, especially immunological aspects, are discussed, as are strategies that might help to overcome these obstacles.

Polymer-directed enzyme prodrug therapy (PDEPT) is a novel two-step antitumour approach using a combination of a polymeric prodrug and polymer-enzyme conjugate to generate cytotoxic drug selectively at the tumour site. In this study the polymeric prodrug N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer-Gly-Phe-Leu-Gly-doxorubicin conjugate PK1 (currently under Phase II clinical evaluation) was selected as the model prodrug, and HPMA copolymer-cathepsin B as a model for the activating enzyme conjugate. Following polymer conjugation (yield of 30–35%) HPMA copolymer-cathepsin B retained ~20–25% enzymatic activity in vitro. To investigate pharmacokinetics in vivo,125I-labelled HPMA copolymer-cathepsin B was administered intravenously (i.v.) to B16F10 tumour-bearing mice. HPMA copolymer-cathespin B exhibited a longer plasma half-life (free cathepsin B t1/2α= 2.8 h; bound cathepsin B t1/2α= 3.2 h) and a 4.2-fold increase in tumour accumulation compared to the free enzyme. When PK1 (10 mg kg−1dox-equiv.) was injected i.v. into C57 mice bearing subcutaneously (s.c.) palpable B16F10 tumours followed after 5 h by HPMA copolymer-cathepsin B there was a rapid increase in the rate of dox release within the tumour (3.6-fold increase in the AUC compared to that seen for PK1 alone). When PK1 and the PDEPT combination were used to treat established B16F10 melanoma tumour (single dose; 10 mg kg−1dox-equiv.), the antitumour activity (T/C%) seen for the combination PDEPT was 168% compared to 152% seen for PK1 alone, and 144% for free dox. Also, the PDEPT combination showed activity against a COR-L23 xenograft whereas PK1 did not. PDEPT has certain advantages compared to ADEPT and GDEPT. The relatively short plasma residence time of the polymeric prodrug allows subsequent administration of polymer-enzyme without fear of prodrug activation in the circulation and polymer-enzyme conjugates have reduced immunogenicity. This study proves the concept of PDEPT and further optimisation is warranted. © 2001 Cancer Research Campaign http://www.bjcancer.com

One way to improve the selectivity of therapeutic molecules in clinical oncology would be to target them on the tumour site, thereby sparing normal tissues. The development of targeted therapeutic methodologies relies in most cases on the availability of binding molecules specific for tumour-associated markers. The display of repertoires of polypeptides on the surface of filamentous phage, together with the efficient selection-amplification of the desired binding specificities using affinity capture, represents an efficient route towards the isolation of specific peptides and proteins that could act as vehicles for tumour targeting applications. Most investigations in this area of research have so far been performed with phage derived recombinant antibodies, which have been shown to selectively target tumour-associated markers both in preclinical animal models and in the clinic. However, future developments with other classes of polypeptides (small constrained peptides, small globular proteins) promise to be important for the selective delivery of therapeutic agents to the tumour site.

The relationship between tumour size and uptake of111In-DTPA-labelled pegylated liposomes has been examined in a human head and neck cancer xenograft model in nude mice. The mean tumour uptake of111In-labelled pegylated liposomes at 24 hours was 7.2 ± 6.6% ID/g. Liposome uptake for tumours < 0.1 g, 0.1–1.0 g and > 1.0 g was 15.1 ± 10.8, 5.9 ± 2.2 and 3.0 ± 1.3% ID/g, respectively. An inverse correlation between tumour weight and liposome uptake was observed by both Spearman’s rank correlation test (r s= – 0.573, P< 0.001) and Pearson’s correlation coefficient (r s= – 0.555, P< 0.001). For 18 tumours with macroscopic central necrosis, the ratio of uptake in the tumour rim relative to the necrotic tumour core was 11.2 ± 6.4. Measurement of tumour vascular volume for tumours of various sizes revealed an inverse correlation between tumour weight and tumour vascular volume (Spearman’s rank correlation test, r s= – 0.598, P< 0.001), consistent with poor or heterogeneous vascularization of larger tumours. These data have important implications for the clinical application of pegylated liposome targeted strategies for solid cancers which are discussed in detail. © 2000 Cancer Research Campaign

Tumour capillaries are frequently hyperpermeable compared with normal vasculature, and thus they offer a much sought-after gateway for targeted delivery of cancer chemotherapy. Phase I clinical trials reported recently describe the first synthetic polymer–drug conjugate to be tested in man. N-(2-Hydroxypropyl)methacrylamide copolymer-doxorubicin (PK1, FCE 28068) displayed antitumour activity in chemotherapy-refractory patients, considerably reduced toxicity compared with doxorubicin, and evidence of tumour-selective targeting. With increasing understanding of the vector- and tumour-related factors that govern vascular permeability, non-viral vectors are being designed for tumour-selective targeting and subsequent intracytoplasmic delivery of macromolecular medicines such as genes, antisense oligonucleotides, proteins and peptides.

Recently, we developed a powerful cytotoxic analogue of bombesin AN-215, in which the bombesin-like carrier peptide Gln–Trp–Ala–Val–Gly–His–Leu–Ψ(CH2-NH)–Leu–NH2 (RC-3094) is conjugated to a potent derivative of doxorubicin, 2-pyrrolinodoxorubicin (AN-201). Small-cell lung carcinomas (SCLCs) are known to express high levels of bombesin receptors. We evaluated whether these receptors could be used for targeting cytotoxic bombesin analogue to H-69 SCLC cells. H-69 cells were xenografted into male nude mice, which then received an intravenous injection of AN-215, cytotoxic radical AN-201, the carrier peptide RC-3094 alone or unconjugated mixture of RC-3094 and AN-201. The levels of mRNA for bombesin receptor subtypes were evaluated by reverse transcription-polymerase chain reaction. In vitro, both the analogue AN-215 and the radical AN-201 showed strong antiproliferative effects on H-69 cells, AN-215 requiring more time to exert its action at 10–8M concentration than AN-201. In vivo, the growth of H-69 SCLC tumours was significantly inhibited by the treatment with 200 nmol kg–1 of AN-215, while equimolar doses of the cytotoxic radical AN-201 or the mixture of AN-201 and the carrier peptide were toxic and produced only a minor tumour inhibition as compared with control groups. mRNA for bombesin receptor subtypes 2 (BRS-2) and 3 (BRS-3) was detected in H-69 tumours. The mRNA levels for BRS-3, but not for BRS-2, were lower in the AN-215-treated tumours as compared with controls. Our results demonstrate that the cytotoxic bombesin analogue AN-215 could be considered for targeted therapy of tumours, such as SCLC, that express bombesin receptors. © 1999 Cancer Research Campaign

Flagellated bacteria are a promising and powerful source for the development of reliable and efficient micro-robots. These bio-actuated devices have a wide variety of application in medical field including targeted drug delivery into the tumour. As of now drugs are delivered into the tumour through the blood vessels on the outside but it's hard to infiltrate the tumour itself. Our goal is to develop bacterial driven micro-beads which would deliver the drugs inside the tumour and degrade it from inside out. For this purpose we have developed 10 μm size polyethylene glycol (PEG) beads using chemical cross linking and incorporated inside it nanoparticles carrying the anticancer drug. For the providing the propulsion force and tumour targeting capability to the microbeads we have attached the genetically engineered Salmonella Typhimurium bacteria onto the surface of beads. These bacterial driven microbeads were found to be highly mobile and have high tumour targeting capability.

Tumour targeting nanoparticles (NPs) have demonstrated great potential for enhancing anticancer drug delivery to tumour sites and for reducing the side effects of chemotherapy. However, many nanoparticulate delivery systems still lack efficient tumour accumulation. In this work, we present a porous silicon (PSi) nanovector functionalized with a tumour-homing peptide, which targets the mammary-derived growth inhibitor (MDGI) expressing cancer cells both in vitro and in vivo, thereby enhancing the accumulation of the NPs in the tumours. We demonstrated that the tumour homing peptide (herein designated as CooP) functionalized thermally hydrocarbonized PSi (THCPSi) NPs homed specifically to the subcutaneous MDGI-expressing xenograft tumours. The THCPSi-CooP NPs were stable in human plasma and their uptake by MDGI-expressing cancer cells measured by confocal microscopy and flow cytometry was significantly increased compared to the non-functionalized THCPSi NPs. After intravenous injections into nude mice bearing MDGI-expressing tumours, effective targeting was detected and THCPSi-CooP NPs showed ∼9-fold higher accumulation in the tumour site compared to the control THCPSi NPs. Accumulation of both NPs in the vital organs was negligible.

Background: Administration of certain chemotherapy drugs at the maximum tolerated dose, vascular-disrupting agents (VDAs) and irradiation can induce mobilisation and tumour homing of proangiogenic bone marrow-derived cells (BMDCs). Increases in cytokines and chemokines contribute to such mobilisation that eventually promotes tumour (re)growth. NGR-TNF is a vascular-targeting agent in advanced clinical development, coupling the CNGRCG angiogenic vessel-homing peptide with tumour necrosis factor-alpha (TNF). We investigated whether NGR-TNF mobilises host BMDCs and growth factors. Methods: Blood was obtained from Lewis lung carcinoma and 4T1 tumour-bearing mice at different time points following NGR-TNF, VDA or anti-VEGFR2/flk-1 antibody treatment. Levels of circulating growth factors were assessed by ELISAs. BMDCs were characterised by FACS. Circulating endothelial progenitor cells were defined as CD45−/CD13+/flk-1+/CD117+/7AAD−, Tie2-expressing monocytes as CD45+/CD11b+/Tie2+ and myeloid-derived suppressor cells as CD45+/CD11b+/Gr1+ cells. Results: NGR-TNF decreases tumour blood vessel density-inducing apoptosis of tumour and tumour endothelial cells. Unlike VDAs, or high-dose NGR-TNF, lower doses of NGR-TNF, comparable to those used in clinical trials, neither mobilise nor recruit to the tumour site proangiogenic BMDCs or induce host growth factors. Conclusion: Low-dose NGR-TNF exerts antitumour activity without inducing proangiogenic host responses, conceivably important for preventing/overcoming resistance and designing combination therapeutic strategies.

Over the past two decades, fluorescent quantum dots (QDs) have been highly attractive for a myriad of bioapplications due to their unique optical properties. For bioimaging applications, QD-based in vivo specific tumour targeting is vitally important in the biological and biomedical fields. Aqueous synthesized QDs (aqQDs) exhibit excellent aqueous dispersibility without requiring any post-treatment and have small hydrodynamic diameters (generally

We report the synthesis, in vitro and in vivo investigation of folate-targeted, biocompatible, biodegradable self-assembled nanoparticles radiolabelled with 99mTc, as potential new SPECT or SPECT/CT imaging agent. Nanoparticles with hydrodynamic size in the range of 75–200 nm were prepared by self-assembly of chitosan and folated poly-γ-glutamic acid, and then radiolabelled with 99mTc. The nanoparticles target tumour cells overexpressing folate receptors and internalize specifically into them to realize early tumour diagnosis detected by SPECT and SPECT/CT modalities. Rat hepatocellular carcinoma cells were used as model system. Cell specificity and tumour targeting efficacy of these nanosystems were investigated in vitro, and in vivo using SPECT and fusion nanoSPECT/CT imaging. In vitro results showed that the radiolabeled nanosystem was efficiently internalized by tumour cells. Whole-body biodistribution of the new radiolabelled, folate-targeted nanoparticles revealed higher uptake in the tumorous kidney compared to the non-tumorous contralateral side. Uptake by the lungs and thyroids was negligible, which confirmed the stability of the nanoparticles in vivo. In vivo SPECT and SPECT/CT imaging visually reinforced the uptake results and were in accordance with the biodistribution data: the new nanoparticles as a targeted contrast agent improve tumour targeting and are able to detect folate-receptor-overexpressing tumours in animal models with enhanced contrast.

Nuclear-receptors are often overexpressed in tumours and can thereby be used as targets when designing novel selective chemotherapeutic agents. To date, many conjugates incorporating an estrogen receptor (ER) ligand have been synthesised in order to direct chemical agents to tissue sites containing ERs. A series of ER ligand conjugates were synthesised incorporating an antagonistic ER ligand scaffold based on endoxifen, covalently-bound via an amide linkage to a variety of combretastatin-based analogues, which may act as antimitotic agents. These novel endoxifen-combretastatin hybrid scaffold analogues were biochemically evaluated in order to determine their antiproliferative and cytotoxicity effects in both the ER-positive MCF-7 and the ER-negative MDA-MB-231 human breast cancer cell lines. ER competitive binding assays were carried out to assess the binding affinity of the lead conjugate 28 towards both the ER alpha and ER beta isoforms. In results from the NCI 60-cell line screen, the lead conjugate 28 displayed potent and highly selective antiproliferative activity towards the MCF-7 human cancer cell line (IC50 = 5 nM). In the ER-binding assays, the lead conjugate 28 demonstrated potent ER competitive binding in ER alpha (IC50 value: 0.9 nM) and ER beta (IC50 value: 4.7 nM). Preliminary biochemical results also demonstrate that the lead conjugate 28 may exhibit pure antagonism. This series makes an important addition to the class of ER antagonists and may have potential applications in anticancer therapy. Refereed/Peer-reviewed

Nanobodies are antigen specific, recombinant, single domain, variable fragments of Camelide heavy chain only antibodies, which are able to bind specific antigen. They are considered as smallest naturally derived antigen-antibody fragment. The supremacy of nanobodies over antibodies is their minimal size, great stability, reversible refolding and high solubility in aqueous solution. Nanobodies, because of small size might be considered as "magic bullets" of molecular imaging in the near future, especially oncologic imaging. Construction of nanobodies involves stages such as immunisation, isolation and cloning and genetic engineering. Nanobodies are mainly used for the molecular imaging application. Apart from molecular imaging it will also used for development of biological sensors and clinical applications in viral infections, atherosclerosis etc. Therapeutic applications of nanobodies includes; inhibition of tumour cell proliferation, binding and intracellular routing of nanobody albumin nanoparticles, nanobullets-nanobody coupled liposomes for treating angiogenesis and tumour, nanobodies against infection, nanobodies against pathogens and cancer. Drug loaded nanobodies are also used as a novel drug delivery system. In the future more targets and their corresponding nanobodies would be identified. Furthermore, future clinical research should investigate the application value of nanobodies. Properties of nanobodies like high resistance to pH and heat, refolding capacity, high solubility, high stability etc make them more prominent than antibodies. Considering all these aspects, nanobodies appear to be a promising method of tumour targeting therapy and diagnosis.

A recombinant fusion protein of colon carcinoma binding A33 single chain antibody with cytosine deaminase displayed specific antigen binding and enzyme activity in surface plasmon resonance and is catalytic activity assay. In vitro, it selectively increased the toxicity of 5-FC to A33 antigen-positive cells by 300-fold, demonstrating the potency of this ADEPT strategy.

A new acid-sensitive drug-delivery nanocarrier has been developed for tumour targeting. The self-assembling co-polymer stearoyl-PEG-poly-sulfadimethoxine methacrylate (stearoyl-PEG-polySDM) was prepared to obtain micelles with responsive behaviour in the physiopathologic pH range. Stearoyl-PEG-polySDM was synthesised using a multi-step procedure that includes pH-sensitive sulfadimethoxine methacrylate polymerisation by AGET-ATRP at the amino terminal side of stearoyl-PEG-NH2. Chemical analysis showed that the stearoyl-PEG-polySDM co-polymer contained a mean of seven methacryloyl sulfadimethoxines per molecule. Potentiometric and turbidimetric analyses showed that stearoyl-PEG-polySDM has an apparent pKa of 7.2 and a cloud point at pH 7.0. In water at pH 7.4, the co-polymer assembled spontaneously into 13.2±3.1 nm micelles with a critical micelle concentration (CMC) of 36 μM. Cell-culture studies showed that the material was more biocompatible with respect to the control Brij-700®. The paclitaxel loading capacity of the micelles was 3.25±0.25% (w/w, %). The colloidal formulations were stable at pH 7.4 for several hours, while at pH 6.5, they rapidly rearranged and aggregated. Fluorescence spectroscopic and cytofluorimetric studies showed that the incubation of MCF-7 tumour cells with fluorescein-labelled stearoyl-PEG-polySDM at pH 6.5 resulted in massive time-dependent cell association, while the incubation at pH 7.4 showed significantly lower cell interaction. Confocal microscopy confirmed that at pH 6.5, the micelles are taken up by cells and that the fluorescein-labelled stearoyl-PEG-polySDM is distributed into the cytosol. At pH 6.5, paclitaxel-loaded stearoyl-PEG-polySDM micelles had a higher cytotoxic effect than the micelles incubated at pH 7.4. The former displayed similar cytotoxic activity to free paclitaxel.

Objectives rh-IFNα2a-NGR is a promising anti-tumor candidate. The aim of present study was to compare pharmacokinetics of rh-IFNα2a-NGR with rh-IFNα2a. Methods Pharmacokinetics and elimination were investigated after intravenous administration to mice and rats. Compared tumor and tissue distribution profiles between rh-IFNα2a-NGR and rh-IFNα2a were illustrated in the tumor transplanted mice of SP2/0 myeloma. Double antibody sandwich ELISA method was used to assess the level of both rh-IFNα2a-NGR and rh-IFNα2a in serum, tissue, bile and urine. Key findings After a single intravenous administration, the pharmacokinetic characters of rh-IFNα2a-NGR and rh-IFNα2a were described using a two-compartment model. No significant differences were observed between the two drugs in pharmacokinetic and elimination data. However, the concentration of rh-IFNα2a-NGR in tumor was 5.34 times and 1.52 times as high as that of rh-IFNα2a at 0.5 h (P < 0.01) and 1 h. In addition, immunohistochemical stain displayed rh-IFNα2a-NGR was predominantly located in tumor vascular tissues. Conclusions rh-IFNα2a-NGR could be an agent for tumor vascular-targeting therapy and these findings provided references for further clinical study.

Ziel: Zur Darstellung von Integrin αvβ3 von Tumoren oder Tumorneugefäßen wählten wir das zyklische Bindungspeptid Arg-Gly-Asp-D-Tyr-Lys (RGDyK) gekoppelt an NODAGA zur Markierung mit 68Ga. Methoden: NODAGA-RGDyK (ABX) wurde mit dem 68Ga-Eluat des 68Ge-Generators IGG100 markiert mit der Prozessoreinheit PharmTracer. Bioverteilungen wurden in weiblichen Hsd-Mäusen 10, 30, 60 und 90 min nach i.v.-Injektion von 68Ga-NODAGA-RGDyK gemessen und als OLINDA-Dosimetrie auf den Menschen projiziert. In SCID-Mäusen, die A431 und andere Tumoren trugen, wurde die Tumoraufnahme mit microPET und Bioverteilungen studiert. Ergebnisse: Die effektive Halbwertszeit von 68Ga-NODAGA-RGDyK war ~25 min im Ganzkörper und den meisten Organen außer Leber und Milz, für die eine stabile Aufnahme gemessen wurde. Die für Frauen und Männer berechnete effektive Dosis (ED) war 0.012 bzw. 0.016 mSv/MBq mit der Annahme eines Blasenleerungsintervalls von 0.5 h. Mit dynamischem PET wurde eine rasche A431-Tumoraufnahme innerhalb von 10 Minuten beobachtet, gefolgt von einem langsamen Abfluss. Bioverteilungen zeigten eine A431-Tumoraufnahme von 68Ga-NODAGA-RGDyK von 3.4±0.4 und 2.7±0.3%ID/g nach 1 bzw. 2 Stunden. Ähnliche Tumoranreicherungen in der Maus wurden in einem Maus- und Humanbrustkrebs gemessen sowie in humanem Ovarialkrebs. Co-Injektion mit einer Überdosis (5 mg/kg) unmarkiertem NODAGA-RGDyK mit dem Radiopharmazeutikum reduzierte die Tumoraufnahme auf 0.23±0.01%ID/g nach 1 Stunde, jedoch wurde eine ähnlich reduzierte Aufnahme auch in andern normalen Organen beobachtet. Wenn unmarkiertes Peptid 15 min nach 68Ga-NODAGA-RGDyK injiziert wurde, verminderte sich die Aufnahme besonders in Tumor und Nebennieren. Diese Beobachtung kann im Vergleich zu anderen Normalgeweben als unterschiedliches Bindungsverhalten gedeutet werden. Schlussfolgerung: NODAGA-RGDyK konnte zuverlässig mit 68Ga markiert werden und zeigte für Menschen eine vorhergesagte ED von 0.014 mSv/MBq. Die Tumoranreicherung war schnell und signifikant. Sie wurde aus Tumor und Nebennieren durch nachgespritztes, unmarkiertes NODAGA-RGDyK in ähnlicher Weise verdrängt. Aim: To visualize neovasculature and/or tumour integrin αvβ3 we selected the binding moiety Arg-Gly-Asp-D-Tyr-Lys (RGDyK) coupled to NODAGA for labeling with 68Ga. Methods: NODAGA-RGDyK (ABX) was labeled with the 68Ga eluate from the 68Ge generator IGG100 using the processor unit PharmTracer. Biodistribution was measured in female Hsd mice sacrificed 10, 30, 60 and 90 min after i.v. injection of 68Ga-NODAGA-RGDyK for OLINDA dosimetry extrapolated to humans. Tumour targeting was studied in SCID mice bearing A431 and other tumour transplants using microPET and biodistribution measurements. Results: Effective half-life of 68Ga-NODAGA-RGDyK was ~25 min for total body and most organs except liver and spleen that showed stable activity retention. With a bladder voiding interval of 0.5 h the calculated effective dose (ED) was 0.012 and 0.016 mSv/MBq for males and females, respectively. Rapid uptake within 10 min was observed in A431 tumours with dynamic PET followed by a slow release. Biodistribution measurements showed a 68Ga-NODAGA-RGDyK uptake in A431 tumours of 3.4±0.4 and 2.7±0.3%ID/g at 1 and 2 h, respectively. Similar uptakes were observed in a mouse and human breast and ovarian cancer xenografts. Co-injection of excess (5 mg/kg) unlabeled NODAGA-RGDyK with the radiotracer reduced tumour uptake at one hour to 0.23±0.01%ID/g, but similarly decreased uptake in normal organs as well. When unlabeled peptide was injected 15 min after 68Ga-NODAGA-RGDyK, uptake diminished particularly in tumour and adrenals, suggestive of a different binding mode compared with other normal tissues. Conclusion: NODAGA-RGDyK was reliably labeled with 68Ga and revealed a predicted ED of 0.014 mSv/MBq. Tumour uptake was rapid and significant and was chased with unlabeled RGDyK in a similar manner as adrenal uptake.

Solid tumour accounts for 90% of all cancers. The current treatment approach for most solid tumours is surgery, however it is limited to early stage tumours. Other treatment options such as chemotherapy and radiotherapy are non-selective, thus causing damage to both healthy and cancerous tissue. Past research has focused on understanding tumour cells themselves, and conventional wisdom has aimed at targeting these cells directly. Recent research has shifted towards understanding the tumour microenvironment and it’s differences from that of healthy cells/tissues in the body and then to exploit these differences for treatmeat of the tumour. One such approach is utilizing anaerobic bacteria. Several strains of bacteria have been shown to selectively colonize in solid tumours, making them valuable tools for selective tumour targeting and destruction. Amongst them, the anaerobicClostridiumhas shown great potential in penetration and colonization of the hypoxic and necrotic areas of the tumour microenvironment, causing significant oncolysis as well as enabling the delivery of therapeutics directly to the tumourin situ. Various strategies utilizingClostridiumare currently being investigated, and represent a novel area of emerging cancer therapy. This review provides an update review of tumour microenvironment as well as summary of the progresses and current status of Clostridial spore-based cancer therapies.

Despite aggressive treatments the prognosis of high-grade gliomas remains poor. Meningiomas are generally benign but for high-grade histotypes or partially resected tumours, recurrence is fairly common. Since a curative option is not currently available, new therapeutic strategies are required to extend the progression-free interval and the overall survival of brain tumour and meningiomas patients. Radioimmunotherapy (RIT) and peptide receptor radionuclide therapy (PRRT) represents two examples of targeted radiotherapy which offer exciting prospect of increasing the specificity of tumour cell irradiation using radionuclides. Although these treatments have been principally experimented for systemic approach, RIT and PRRT may also be exploited topically, in order to reduce systemic activity and enhance tumour targeting. In this chapter the authors report on intracavitary use of radionuclides in HGG and their experience concerning peptide receptor radionuclide therapy in meningiomas.

The introduction of multifunctionalities for tumour targeting is becoming a popular strategy toward the development of new therapeutic agents. In particular, the multifaceted potential of ruthenium(II)-arene complexes show great promise as chemotherapeutics. An ever-increasing number of papers dealing with the integration of ruthenium complexes with biologically active molecules to derive bioorganometallic molecules of chemotherapeutic significance have been published in recent years. This perspective review presents a short overview of multifunctional ruthenium-based drugs, especially those containing arene ruthenium complexes, with the emphasis on the combination of photosensitizers with ruthenium complexes for the preparation of novel multifunctional photodynamic therapy agents.

Hodgkin lymphoma (HL) represents a category of lymphoid neoplasms with unique features, notably the usual scarcity of tumour cells in involved tissues. The most common subtype of classical HL, nodular sclerosis HL, characteristically comprises abundant fibrous tissue stroma. Little information is available about the protein composition of the stromal environment from HL. Moreover, the identification of valid protein targets, specifically and abundantly expressed in HL, would be of utmost importance for targeted therapies and imaging, yet the biomarkers must necessarily be accessible from the bloodstream. To characterize HL stroma and to identify potentially accessible proteins, we used a chemical proteomic approach, consisting in the labelling of accessible proteins and their subsequent purification and identification by mass spectrometry. We performed an analysis of potentially accessible proteins in lymph node biopsies from HL and reactive lymphoid tissues, and in total, more than 1400 proteins were identified in 7 samples. We have identified several extracellular matrix proteins overexpressed in HL, such as versican, fibulin-1, periostin, and other proteins such as S100-A8. These proteins were validated by immunohistochemistry on a larger series of biopsy samples, and bear the potential to become targets for antibody-based anti-cancer therapies.

Nanocarriers can penetrate the tumour vasculature through its leaky endothelium and, in this way, accumulate in several solid tumours. This is called the enhanced permeation and retention (EPR) effect. Together with nanocarriers whose surface is tailored for prolonged blood circulation times, the concept is referred to as passive targeting. Targeting ligands, which bind to specific receptors on the tumour cells and endothelium, can be attached on the nanocarrier surface. This active targeting increases the selectivity of the delivery of drugs. Passive and active drug targeting with nanocarriers to tumours reduce toxic side-effects, increase efficacy, and enhance delivery of poorly soluble or sensitive therapeutic molecules. In this review, currently studied and used passive and active targeting strategies in cancer therapy are presented.

The human epidermal growth-factor like receptor (HER) family of receptor tyrosine kinases are important targets for cancer therapy. The family consists of four members - EGFR, HER2, HER3 and HER4 - that normally transfer stimulatory signals from extracellular growth factors to the intracellular signalling network. Over-activation of these receptors leads to uncontrolled cell proliferation and is seen in several types of tumours. The aim of the studies reported in this thesis was to study the uptake and effects of affibody molecules against EGFR, HER2 and HER3 in cultured cells. Affibody molecules are affinity proteins originally derived from one of the domains of protein A, and their small size and robust structure make them suitable agents for tumour targeting and therapy. Papers I and II of this thesis concern EGFR-specific affibody molecules, which were shown to be more similar to the antibody cetuximab than the natural ligand EGF in terms of cellular uptake, binding site and internalisation rate. In addition, fluorescence-based methods for the quantification of internalisation were evaluated. In the studies reported in papers III and IV, HER2-specific affibody molecules were utilised as carriers of radionuclides. Paper III reports that different cell lines exhibit different radiosensitivities to 211At-labelled affibody molecules; radiosensitivity was found to correlate with cell geometry and the rate of internalisation. Paper IV discusses the use of 17-AAG, an agent that induces HER2 internalisation and degradation, to force the internalisation of 211At- and 111In-labelled affibody molecules. Papers V and VI describe the selection and maturation of HER3-specific affibody molecules, which were found to compete with the receptor’s natural ligand, heregulin, for receptor binding. These affibody molecules were demonstrated to inhibit heregulin-induced HER3 activation and cell proliferation. The studies summarised in this paper will hopefully contribute to a better understanding of these affibody molecules and bring them one step closer to being helpful tools in the diagnosis and treatment of cancer.