Colon cancer metastasis in mouse liver is not affected by hypercoagulability due to Factor V Leiden mutation

Clinical trials have shown life-prolonging effects of antithrombotics in cancer patients, but the molecular mechanisms remain unknown due to the multitude of their effects. We investigated in a mouse model whether one of the targets of antithrombotic therapy, fibrin deposition, stimulates tumour development. Fibrin may provide either protection of cancer cells in the circulation against mechanical stress and the immune system, or form a matrix for tumours and/or angiogenesis in tumours to develop. Mice homozygous for Factor V Leiden (FVL), a mutation in one of the coagulation factors that facilitates fibrin formation, were used to investigate whether hypercoagulability affects tumour development in an experimental metastasis model. Liver metastases of colon cancer were induced in mice with the FVL mutation and wild-type littermates. At day 21, number and size of tumours at the liver surface, fibrin/fibrinogen distribution, vessel density and the presence of newly formed vessels in tumours were analysed. Number and size of tumours did not differ between mice with and without the FVL mutation. Fibrin/fibrinogen was found in the cytoplasm of hepatocytes and cancer cells, in blood vessels in liver and tumour tissue and diffusely distributed outside vessels in tumours, indicating leaky vessels. Vessel density and angiogenesis varied widely between tumours, but a pre-dominance for vessel-rich or vessel-poor tumours or vessel formation could not be found in either genotype. In conclusion, the FVL mutation has no effect on the development of secondary tumours of colon cancer in livers of mice. Fibrin deposition and thus inhibition of fibrin formation by anticoagulants do not seem to affect tumour development in this model.     

The Laser Treatment of Experimental Malignant Tumours

Some of the results of experiments performed during the past two years to assess effects of laser energy on experimental malignant tumours are reviewed. Twenty types of malignant tumours (most in the cheek pouch and 11 of human origin) were treated in over 700 Syrian hamsters. Results of laser treatment of malignant melanomas and thyroidal carcinomas are presented. A human patient with malignant melanoma treated by laser energy is described. Investigation of thermal effect revealed that the laser-treated tumour remained warm for about one minute, while the cautery-treated tumour cooled to normal temperature in five seconds. Direct action of laser on superficial tumours is possible; deeper lesions must be exposed surgically. Laser energy has a selective effect on certain malignant tumours, resulting in their progressive regression and ultimate dissolution. All hamsters with implanted malignant melanomas and carcinomas of human origin, after completion of a course of laser treatment, showed no gross or histologic evidence of tumour up to the date of last observation.     

Accessible hyaluronan receptors identical to ICAM-1 in mouse mast-cell tumours

Immunohistochemical studies of the hyaluronan (HA)-receptor (R), originally found on liver endothelial cells (LEC) and related to the intercellular adhesion molecule 1 (ICAM-1), showed that polyclonal antibodies against HARLEC (HA receptor on LEC) also stain structures in mouse mastocytomas, mainly vessels. To test if intravenously administered HA might target the tumour receptorsin vivo, mice carrying an inoculated mastocytoma in one hind leg muscle were injected in the tail vein with¹²⁵I-tyrosine (T)-labelled HA and killed 75 min after injection when organs and tissues were checked for radioactivity. When doses exceeding the binding capacity of the liver were injected, a significant increase in radioactivity (up to five-fold) within the tumour tissue was found. The weight adjusted difference between control and tumour tissue was greater for smaller tumours, probably due to necrosis in the larger. HA-staining of tumours from animals receiving¹²⁵I-T-HA, showed HA in areas that also stained weakly for ICAM-1 using monoclonal antibodies. ICAM-1 staining was dramatically increased after hyaluronidase treatment of the sections, indicating that the HA is bound to these receptors and thereby blocks antibody recognition.Abbreviations ICAM-1 intercellular adhesion molecule 1 - HA hyaluronan - HARLEC hyaluronan receptor on liver endothelial cells - MW molecular weight     

Delivery of mengovirus-derived RNA replicons into tumoural liver enhances the anti-tumour efficacy of a peripheral peptide-based vaccine

Hepatocellular carcinoma is a deadly cancer with growing incidence for which immunotherapy is one of the most promising therapeutic approach. Peptide-based vaccines designed to induce strong, sustained CD8+ T cell responses are effective in animal models and cancer patients. We demonstrated the efficacy of curative peptide-based immunisation against a unique epitope of SV40 tumour antigen, through the induction of a strong CD8+ T cell-specific response, in our liver tumour model. However, as in human clinical trials, most tumour antigen epitopes did not induce a therapeutic effect, despite inducing strong CD8+ T cell responses. We therefore modified the tumour environment to enhance peptide-based vaccine efficacy by delivering mengovirus (MV)-derived RNA autoreplicating sequences (MV-RNA replicons) into the liver. The injection of replication-competent RNA replicons into the liver converted partial tumour regression into tumour eradication, whereas non-replicating RNA had no such effect. Replicating RNA replicon injection induced local recruitment of innate immunity effectors (NK and NKT) to the tumour and did not affect specific CD8+ T cell populations or other myelolymphoid subsets. The local delivery of such RNA replicons into tumour stroma is therefore a promising strategy complementary to the use of peripheral peptide-based vaccines for treating liver tumours.     

Image-guided navigation for the control interstitial laser therapy of vascular malformations in the head and neck region]

Laser-induced interstitial thermal therapy (LITT) is a proven minimally invasive surgical technique for the treatment of haemangiomas and vascular malformations, and various tumours. The intra-lesion application of thermal energy destroys regional tissue. The percutaneous placement of the laser fibre for photocoagulation is done without the benefit of direct visual control. Image-data-based LITT was performed in patients (five procedures) with extensive venous malformations in the maxillofacial area. The system comprised a specially developed Nd:YAG laser fibre introducer set used in conjunction with fused high-resolution computed tomography, and magnetic resonance image-data--based surgical navigation. In all cases, follow-up examination clearly showed a diminishment in tumour volume, and all patients reported significant subjective improvement. The results suggest that navigation-guided LITT can be performed safely, preserving vital structures from collateral thermal damage.     

Pathogenetic mechanisms of nuclear pleomorphism of tumour cells based on the mutator phenotype theory of carcinogenesis

The nuclei of the cells of most solid tumours in histopathologic preparations vary in size, shape and chromatin pattern, both from normal nuclei and from each other. These features have not been explained in terms of conventional concepts of nuclear structure and theories of carcinogenesis. In recent years, the unfolded chromosomes have been shown to occupy "domains" in the nucleus during interphase, providing a relatively uniform density of fine chromatin fibres throughout the nucleus in the living state. This is in contrast to the appearances of interphase chromatin existing as coarse clumps and fibres (heterochromatin and euchromatin respectively) as are seen in histologic preparations. Additionally, the binding of chromatin to nuclear membrane, the possible existence of a nuclear matrix, the functions of nuclear pores, and the attachments of cytoskeletal structures to the outer nuclear membrane are now recognised. Studies of genetic instability of cancer cells (many random mutations are present in the genome, which vary from nucleus-to-nucleus in individual tumours) have shown that this phenomenon occurs early in tumour formation, can be present in morphologically-normal cells adjacent to tumours, and can result in thousands of genomic events per tumour cell. These observations form the basis for the mutator phenotype/clonal selection theory of carcinogenesis, which proposes that genetic instability is an essential early part of carcinogenesis. Genetic instability has been used to explain significant cell-to-cell variability of behaviour (tumour cell heterogeneity) among cells of individual tumours. This paper proposes that a high incidence of nucleus-to-nucleus-variable mutation of the genes for factors controlling nuclear morphology in tumours can explain nucleus-to-nucleus variations of histopathologic appearance of these nuclei when some additional effects of histological processing are taken into account.     

Tumour bed effect: hypoxic fraction of tumours growing in preirradiated beds

The reduction in tumour growth rate seen when tumours are implanted into preirradiated sites, the tumour bed effect (TBE), is believed to be due to radiation damage to vascular stroma, leading to defective angiogenesis in the tumour. The present work examined whether or not the functional inadequacy of irradiated stroma was accompanied by an increased hypoxic fraction in tumours growing in irradiated beds. Mouse flank skin was given 0 or 20 Gy X-rays and RIF-1 fibrosarcoma cells were implanted i.d. into the centre of the treatment field one week later. Tumours of 200 mm3 were irradiated under clamped or unclamped conditions and the hypoxic fraction measured from the displacement of the corresponding survival curves, assayed in vitro. Results indicated a small increase in the hypoxic fraction. Averaging values from three independent experiments, the percentage of hypoxic cells increased from 2.5 per cent for cells in tumours growing in unirradiated beds to 4.6 per cent for those from tumours in beds given 20 Gy. Thus an irradiated vascular bed is still to some extent able to maintain the proportion of oxic: hypoxic tumour cells found in tumours growing in unirradiated beds, despite manifest changes in tumour necrosis and growth rate.     

Large hepatocellular cancers: hepatic resection or liver transplantation?

Thirteen patients with primary hepatocellular cancer were studied to outline criteria for resectability in patients with large liver tumours. The mean age was 34 and the mean tumour diameter 13 cm (range 7-18 cm). Five of the tumours had a diameter of 15 cm or more. Extensive radiological investigations showed that seven of the patients had tumours of both right and left lobes of the liver, 10 had evidence of vascular invasion, and three had evidence of extrahepatic spread. Only two of the patients underwent a classically described formal hepatic resection, the rest having extensive resections crossing major anatomical planes. In no instance did the vascular invasion preclude resection, and extrahepatic spread could be verified in only one patient. The traditional criteria of resectability--that is, tumours located to one main lobe of the liver without vascular invasion and extrahepatic spread--can and should be extended. Resection may be preferable to transplantation even in patients with large primary liver tumours.     

Detection and molecular characterisation of disseminated tumour cells: Implications for anti-cancer therapy

Haematogenous distant metastasis is the leading cause of cancer-related death in solid tumours. By applying sensitive immunocytochemical and molecular assays, disseminated tumour cells (DTC) in bone marrow (BM) can be detected in 20–40% of cancer patients without any clinical or even histopathological signs of metastasis, and the presence of these DTC at primary diagnosis predicts the subsequent occurrence of overt metastases in bone and other organs. The detection and characterisation of DTC in BM may lead to a better understanding of the biology initiating metastatic spread in cancer patients and will eventually contribute to the development of more effective strategies to eliminate DTC. In this review, we will therefore discuss the detection and characterisation of DTC in the light of new therapeutic strategies targeting tumour-associated molecules and signalling pathways.     

Intratumoural light distribution in an experimental mouse tumour irradiated by a diffuse-light irradiator compared with unilateral helium-neon light for photodynamic therapy

Investigations on dosimetry in photodynamic therapy (PDT) of experimental mouse tumours transplanted into the right hind leg revealed a significant variability in the fluence rate reaching tumour cells in different parts of the tumour when irradiated by a 50 mW collimated He-Ne laser. Based on intratumoural fibre-optical light measurements, a new irradiation source was constructed, in which variability in the radiant energy fluence rate between different parts of the solid tumour was reduced. The new diffuse-light irradiator was constructed basically from two concentric water chambers surrounded by three linear 3000 W xenon flash lamps. The outer chamber was an optical band filter, and the inner chamber contained a light-dispersing solution of lipid droplets which created an isotropic light field in which the tumours were submerged for PDT. Compared with unilateral He-Ne laser irradiation, an enhancement factor of 7.3 in radiance was obtained for the diffuse-light irradiator measured in the tumour. The new apparatus provides a nearly isotropic light field for in vivo experimental PDT.     

Long-term Culture of Human Breast Cancer Specimens and Their Analysis Using Optical Projection Tomography

Breast cancer is a leading cause of mortality in the Western world. It is well established that the spread of breast cancer, first locally and later distally, is a major factor in patient prognosis. Experimental systems of breast cancer rely on cell lines usually derived from primary tumours or pleural effusions. Two major obstacles hinder this research: (i) some known sub-types of breast cancers (notably poor prognosis luminal B tumours) are not represented within current line collections; (ii) the influence of the tumour microenvironment is not usually taken into account.We demonstrate a technique to culture primary breast cancer specimens of all sub-types. This is achieved by using three-dimensional (3D) culture system in which small pieces of tumour are embedded in soft rat collagen I cushions. Within 2-3 weeks, the tumour cells spread into the collagen and form various structures similar to those observed in human tumours1. Viable adipocytes, epithelial cells and fibroblasts within the original core were evident on histology. Malignant epithelial cells with squamoid morphology were demonstrated invading into the surrounding collagen. Nuclear pleomorphism was evident within these cells, along with mitotic figures and apoptotic bodies.We have employed Optical Projection Tomography (OPT), a 3D imaging technology, in order to quantify the extent of tumour spread in culture. We have used OPT to measure the bulk volume of the tumour culture, a parameter routinely measured during the neo-adjuvant treatment of breast cancer patients to assess response to drug therapy. Here, we present an opportunity to culture human breast tumours without sub-type bias and quantify the spread of those ex vivo. This method could be used in the future to quantify drug sensitivity in original tumour. This may provide a more predictive model than currently used cell lines.     

Renal cryoablation: Multidisciplinary,collaborative and perspective approach

Renal cryoablation is becoming an established treatment option for small renal masses. It allows preservation of renal function without compromising cancer control. The technique has evolved considerably since it was first reported using liquid nitrogen over 20 years ago. We describe the modern technique for both laparoscopic and image guided renal cryoablation.Renal cryoablation is performed either laparoscopically or percutaneously depending on tumour characteristics. Common features include biopsy of the mass, protection of adjacent organs, and the use of compressed argon gas for freezing and helium for thawing. Dynamic monitoring is used to ensure adequate treatment. The shape of the iceball can be modified by adding extra needles or changing their positions. A double freeze/thaw is necessary for confident ablation of all cancer cells. The laparoscopic approach includes exposure of the tumour and may involve extensive mobilisation of the kidney. Laparoscopic ultrasound is essential for correct localisation of the tumour, needle placement, and monitoring the treatment. A Temperature probe is placed at the edge of the tumour to record treatment temperature. The percutaneous approach is typically performed with CT guidance. Adjacent organs can be protected by injecting saline or carbon dioxide. Early imaging is helpful to detect or rule out incomplete treatment. Post-operative follow-up is structured at specific intervals (e.g. 3, 6, 12 months then annually) and perhaps tailored or modified based on the degree of suspicion of inadequate treatment.     

Improved radioimmunodetection of tumours using liposome-entrapped antibody

The discrimination of radioimmunodetection of tumours is reduced by the presence of circulating radiolabelled antibody (primary antibody). We have prepared liposomes containing an antibody to the primary antibody (secondary antibody), with the intention of complexing and delivering to the liver primary antibody which is not associated with the tumour. In mice bearing xenografts of human tumours which secrete the marker carcinoembryonic antigen (CEA), liposomally entrapped secondary antibody was able to reduce the blood levels of 125I-labelled anti-CEA within 2 h, without reducing the amount of anti-CEA bound to the tumour. We therefore suggest that the use of liposomally entrapped secondary antibody would improve the diagnostic potential of radioimmunodetection of tumours and their metastases.     

Análisis de biopsias líquidas para el diagnóstico del cáncer: revisión sistemática

The incidence of cancer has increased in recent years, especially in those over 65 years of age, posing a major health problem. Many tumours have a poor prognosis because they are diagnosed at very advanced stages. It is therefore especially important to incorporate liquid biopsy into clinical practice as a method for detecting tumours at very early stages. A systematic review was conducted, with the main objective of analysing the available literature on the use of liquid biopsy in the early diagnosis of cancer, and as a secondary objective, to determine the types of tumours that can be diagnosed early by liquid biopsy and the available biomarkers. The results indicate a lack of agreement with the biomarkers detected and the technologies applied. This highlights the need for multicentre studies to look at large cohorts and to establish protocols of action, as well as to increase analytical validity and the possibility of using a screening test for each type of tumour. This could be a very important step forward, as it could improve the management of cancer patients to a great extent.     

A novel approach to improve the efficacy of tumour ablation during cryosurgery

Freezing tumours and ablating it using cryosurgery is becoming a popular surgical procedure for treatment of carcinomas. In order to improve the efficiency of the cryosurgical procedure different approaches have been implemented till now, e.g., injecting high thermal conductivity fluid inside the tumour, low latent heat fluids inside the tumour prior to cryosurgery etc. These techniques improve the cryosurgical process to some extent but lack in minimising the damage to the surrounding healthy tissues. In this study, a novel concept is proposed which advocates the use of solutions with specific thermophysical properties around the interface of tumour. Numerical modelling has been done to determine the location of the ice fronts in the presence of this solution around the boundary of the tumour. It is noticed that in the presence of solution layer, owing to its distinct thermophysical properties like low thermal conductivity, not only the cellular destruction is enhanced but also the damage to the surrounding healthy tissue is minimised. Further, results indicate that this strategy leads to a faster ablation rate reducing the surgical time immensely. Also, an optimal offset, the minimum distance between the tip of cryoprobe and the boundary of the tumour, is identified for a given tumour radius with a given active length which gives maximum tumour necrosis in less time. This optimal offset which has been identified for each case will help the surgeons in proper planning of cryosurgery and improving the effectiveness of this technique greatly, making it a better treatment modality than its counterparts in many ways. It is also observed that for a 2 mm increase in activelength of the cryoprobe, the decrease in optimal offset is approximately 1 mm, i.e. optimal offset decreases linearly with an increase in the activelength for a given radius of the tumour. Also, for tumour with different radii, ranging between 10 mm to 15 mm, with same active length, the time taken for complete ablation by the larger tumour is nearly 2.7 times the time taken by the smaller one for every 2.5 mm increase in the tumour radius.     

The use of clostridial spores for cancer treatment

Hypoxic/necrotic regions, absent in normal tissues, can be exploited to target tumours in cancer therapy using nonpathogenic strains of the bacterial genus Clostridium. Following administration of Clostridium spores to tumour-bearing organisms, these spores can only germinate within the hypoxic/necrotic regions of solid tumours, proving their exquisite selectivity. Low oxygen tension is a common feature of solid tumours, which may arise from the unique physiological environment, generated to a large extent by the abnormal tumour vasculature, and provides as such a niche for anaerobic bacteria. Some clostridia tested clearly showed innate oncolytic activity, but they could not completely eradicate the tumour. Recombinant clostridia producing prodrug-converting enzymes or cytokines resulted in the production of such proteins solely within the tumour, and where applicable, could convert the prodrug in a toxic compound. Moreover, in some cases, tumour eradication or tumour control could be observed. This review brings an overview of the relative successes and failures of the Clostridium-directed tumour therapy with both wild-type strains and strains producing proteins useful in antitumour therapy.     

Current issues in the utility of 19F nuclear magnetic resonance methodologies for the assessment of tumour hypoxia

It is now well established that uncontrolled proliferation of tumour cells together with the chaotic and poorly regulated blood supply of solid tumours result in tissue hypoxia, and that hypoxic regions of tumours are resistant to radiotherapy and chemotherapy. The development and application of non-invasive methods to rapidly determine the degree and extent of tumour hypoxia in an individual tumour would clearly enhance cancer treatment strategies. This review describes the current status of two (19)F nuclear magnetic resonance (NMR) methodologies that have been exploited to investigate tumour hypoxia, namely: (i) (19)F NMR oximetry following administration of perfluorocarbons, from which tumour p(O)(2) measurements can be made; and (ii) (19)F NMR measurements of the tumour retention of fluorinated 2-nitroimidazoles.     

Immunosuppressive cells and tumour microenvironment: focus on mesenchymal stem cells and myeloid derived suppressor cells

Tumours have been compared to unhealed wounds that produce large amounts of inflammatory mediators, including cytokines, chemokines, and growth factors. These molecules participate in the formation of a rich and heterogeneous microenvironment by attracting non malignant cells that promote tumour progression and dissemination. Tumour infiltrating cells include macrophages, myeloid-derived suppressor cells (MDSCs), mesenchymal stromal cells (MSCs) and TIE2-expressing monocytes. Most of them are bone marrow-derived, although MSC are present in virtually every tissue. This review focuses on MDSCs and MSCs, both of which can exert pro-tumorigenic effects through negative regulation of immune responses. MDSCs represent a heterogeneous population of cells of myeloid origin that are expanded and activated in response to growth factors and cytokines released by tumours. Once MDSCs are activated, they accumulate in lymphoid organs and tumours where they exert T cell immunosuppression. Like MDSCs, MSCs can be mobilized from the bone marrow into the bloodstream and home in the tumour stroma, where they either help or hinder tumour growth. Here, we will discuss the origin, the functions and the mechanisms of action of MSCs and MDSCs, as well as the strategies to target these cells for the therapeutic benefit of cancer patients.     

Roles of p38 MAPKs in invasion and metastasis

Cells from primary tumours need to go through several steps to become fully metastatic. During this process, cancer cells acquire the ability to invade, migrate across the surrounding tissue, enter into the circulation and colonize distant organs. In the present paper, we review recent progress in understanding how the p38 MAPK (mitogen-activated protein kinase) signalling pathway participates in the different steps of metastasis. Experimental evidence suggests that tumour cells need to modulate p38 MAPK activity levels to successfully metastasize.