Search

Abstract Most cancer researchers regularly practice the responsible conduct of research (RCR) without consciously considering it. As professional scientists, we simply do what we are trained to do. However, as we train a new generation of cancer researchers in our laboratories, we must be vigilant against undue complacency. In an age when misconduct in research is receiving more media attention than ever before, we should periodically take a moment of pause and reflect upon the meaning and practice of responsibly conducting research. Rather than meeting minimum standards in a compliance-driven manner, we should practice forethought and periodically consider how we can improve. We, as leaders in cancer research, must then push our peers to do the same. By embedding RCR into the culture of cancer research through a multilayer approach, including regular assessment at the levels of individual research groups, departmentally, and institutionally, we will become a model discipline in the responsible conduct of research.

Abstract Background Protein used in medicine, e.g. interferon, are immunogenic and quickly broken down by the body. Pegylation is a recognized way of preserving their integrity and reducing immune reactions, and works well with enzymes used to degrade amino acids, a recent focus of attention in controlling cancer growth. Of the two arginine-degrading enzymes being explored clinically, arginine deiminase is a decidedly foreign mycoplasm-derived enzyme, whereas human arginase 1 is a native liver enzyme. Both have been pegylated, the former with adjuncts of 20 kD, the latter with 5 kD PEG. Pegylation is done by several different methods, not all of which are satisfactory or desirable. Methods The preparation of novel polyethylene glycol (PEG) derivatives for modifying proteins is described, but directed specifically at pegylation of recombinant human arginase 1 (rhArg1). rhArg1 expressed in Escherichia coli was purified and coupled in various ways with 5 different PEG molecules to compare their protective properties and the residual enzyme activity, using hepatocellular cell lines both in vitro and in vivo. Results Methoxypolyethylene glycol-succinimidyl propionate (mPEG-SPA 5,000) coupled with very high affinity under mild conditions. The resulting pegylated enzyme (rhArg1-peg5,000 mw) had up to 6 PEG chains of 5K length which not only protected it from degradation and any residual immunogenicity, but most importantly let it retain >90% of its native catalytic activity. It remained efficacious in depleting arginine in rats after a single ip injection of 1,500 U of the conjugate as the native enzyme, plasma arginine falling to >0.05 μM from ~170 μM within 20 min and lasting 6 days. The conjugate had almost the same efficacy as unpegylated rhArg1 on 2 cultured human liver cancer (HCC) cell lines. It was considerably more effective than 4 other pegylated conjugates prepared. Conclusion Valuable data on the optimization of the pegylation procedure and choice of ligand that best stabilizes the enzyme arginase 1 are presented, a protocol that should equally fit many other enzymes and proteins. It is a long lasting arginine-depleting enzyme in vivo which will greatly improve its use in anti-cancer therapy.

Abstract Background Lovastatin and other statins may reduce the development of melanomas. The effects on melanoma cells and their ability to enhance angiogenesis in a co-culture system presented an opportunity to assess whether Lovastatin act on melanoma cells, HUVEC or both types of cells. Results Direct effects of co-culturing two different malignant melanoma cells (A375 and G361) on the process of angiogenesis in vitro was studied with our angiogenesis model1, based on human dermal fibroblasts and human umbilical vein endothelial cells (HUVEC). Co-cultures were set up using "sland" and "dispersed seeding" techniques. A statistically significant increase in tubule formation in both cases was observed compared to controls. The effects of doses equivalent to therapeutic concentrations of Lovastatin were analysed. The drug inhibited the growth of all cell types, induced apoptosis, and markedly reduced the formation of tubules in the angiogenesis model at low concentrations. Its action was successfully reversed by the introduction of geranylgeranyl pyrophosphate. Conclusion Lovastatin can reduce both tumour (melanoma) cell growth, and the angiogenic activity of these cells in co-cultures using an established 2-dimensional model angiogenesis system beyond that which would be seen by reduced proliferation alone.

Abstract Clinicians will argue that cancer can only really receive the treatment that is needed through thorough understanding of medicine. However, even empirical approaches to therapy result in experimental analysis of the agencies involved on test cells, usually in culture. From the obverse perspective, cell biologists will argue that until we fully understand cell cycle regulation, tumour management will be too imprecise to make the best advances. A forum is needed whereby the fundamental studies on cells prior to, during and after transformation in vitro can be freely reported (open access) and discussed. The action of anticancer agents and cancer preventative substances can more easily be studied in vitro before the often excessive complexity of making similar studies in experimental and human cancers is tackled. Cancer Cell International is committed to providing such a forum. Ironically within a few months of launching this open access journal, Elsevier had much the same idea, and there one has to pay for the privilege of downloading vital papers in this biomedical field.

Abstract Background The issue remains unresolved as to whether low frequency magnetic fields can affect cell behaviour, with the possibility that they may be in part responsible for the increased incidence of leukaemia in parts of the population exposed to them. Methods Combined treatment of HeLa cells with gamma-irradiation (1, 3 and 5 Grays) and extra low frequency magnetic fields of ~50 Hz was carried out under rigorously controlled conditions. Results Synchronised cells progressing from S-phase arrived at mitosis on average marginally ahead of irradiation controls not exposed to ELF. In no instance out of a total of twenty separate experiments did this "double-insult" further delay entry of cells into mitosis, as had been anticipated. Conclusion This apparently "non-genotoxic" agent (ELF) appears to be capable of affecting cells that would normally arrest for longer in G2, suggesting a weakening of the stringency of the late cycle (G2) checkpoint.

We adopt the position that metabolism originated at (or near) mineral surfaces prior to the origin of the first cells. Based on current views of the organization of contemporary animal cells we speculate that the metabolism of the immediate ancestors of eukaryotic cells required these non-biological surfaces, but that the latter were subsequently replaced by membranes, and nuclear and cytoplasmic matrix proteins which, we argue, remain as required participants in the intermediary metabolism of contemporary eukaryotic cells. The idea that such an intracellular organization could have provided a fundamental means by which to control metabolic rate at the level of the intact animal is considered next. In the case of vertebrates we suggest that the organismic level of control might operate through the rate of capillary blood flow, as proposed in the Flow Theory of Coulson (1986): by controlling the rate at which the organized enzyme arrays within the cells are perfused with substrate, cellular metabolic rates could be set throughout the organism in an integrated fashion. Although there are problems with this linkage the interesting possibility arises that the metabolic rate of individual cells may be subservient to the organism, being driven not so much by the well known intracellular controls of concentration-based-biochemistry as by the flow of nutrients through the cells.

Background: Protein used in medicine, e.g. interferon, are immunogenic and quickly broken down by the body. Pegylation is a recognized way of preserving their integrity and reducing immune reactions, and works well with enzymes used to degrade amino acids, a recent focus of attention in controlling cancer growth. Of the two arginine-degrading enzymes being explored clinically, arginine deiminase is a decidedly foreign mycoplasm-derived enzyme, whereas human arginase 1 is a native liver enzyme. Both have been pegylated, the former with adjuncts of 20 kD, the latter with 5 kD PEG. Pegylation is done by several different methods, not all of which are satisfactory or desirable. Methods: The preparation of novel polyethylene glycol (PEG) derivatives for modifying proteins is described, but directed specifically at pegylation of recombinant human arginase 1 (rhArg1). rhArg1 expressed in Escherichia coli was purified and coupled in various ways with 5 different PEG molecules to compare their protective properties and the residual enzyme activity, using hepatocellular cell lines both in vitro and in vivo. Results: Methoxypolyethylene glycol-succinimidyl propionate (mPEG-SPA 5,000) coupled with very high affinity under mild conditions. The resulting pegylated enzyme (rhArg1-peg5,000 mw) had up to 6 PEG chains of 5K length which not only protected it from degradation and any residual immunogenicity, but most importantly let it retain >90% of its native catalytic activity. It remained efficacious in depleting arginine in rats after a single ip injection of 1,500 U of the conjugate as the native enzyme, plasma arginine falling to >0.05 µM from ~170 µM within 20 min and lasting 6 days. The conjugate had almost the same efficacy as unpegylated rhArg1 on 2 cultured human liver cancer (HCC) cell lines. It was considerably more effective than 4 other pegylated conjugates prepared. Conclusion: Valuable data on the optimization of the pegylation procedure and choice of ligand that best stabilizes the enzyme arginase 1 are presented, a protocol that should equally fit many other enzymes and proteins. It is a long lasting arginine-depleting enzyme in vivo which will greatly improve its use in anti-cancer therapy. [ABSTRACT FROM AUTHOR]

Background: The relationship between body mass (M) and standard metabolic rate (B) among living organisms remains controversial, though it is widely accepted that in many cases B is approximately proportional to the three-quarters power of M. Results: The biological significance of the straight-line plots obtained over wide ranges of species when B is plotted against log M remains a matter of debate. In this article we review the values ascribed to the gradients of such graphs (typically 0.75, according to the majority view), and we assess various attempts to explain the allometric power-law phenomenon, placing emphasis on the most recent publications. Conclusion: Although many of the models that have been advanced have significant attractions, none can be accepted without serious reservations, and the possibility that no one model can fit all cases has to be more seriously entertained. [ABSTRACT FROM AUTHOR]

The thesis is presented that only within very small microdomains of the cell internum might diffusion operate in the sorting of molecular affinities. Much of cell metabolism is guided and controlled in rate by the speed with which molecules that have to interact encounter one another. What is clear, however, is that the cell does not have a single 'modus operandi' but has the choice of many different strategies, each of which can contribute in different proportion to the rate of ongoing activity. It is probably our own desire to simplify things and use the most (or more) probable strategy that confines our appreciation of the overall robustness of the cell as a 'survival machine'. The main operative process at any given time (perfusion, diffusion or whatever) has always to be considered very carefully in relation to the organisational structure of the cell, which can be transient and fickle but nevertheless has been seen as involving an extensive cytomatrix, a ground substance, within an aqueous environment in which the degree of water structuring is even more fickle. [ABSTRACT FROM AUTHOR]

The development of vessel-like structures in vitro to mimic as well as to realize the possibility of tissue-engineered small vascular networks presents a major challenge to cell biologists and biotechnologists. We aimed to establish a three-dimensional (3-D) culture system with an endothelial network that does not require artificial substrates or ECM compounds. By using human skin fibroblasts and endothelial cells (ECs) from the human umbilical vein (HUVECs) in diverse spheroid coculture strategies, we verified that fibroblast support and modulate EC migration, viability, and network formation in a 3-D tissue-like stromal environment. In mixed spheroid cultures consisting of human ECs and fibroblasts, a complex 3-D network with EC tubular structures, lumen formation, pinocytotic activity, and tight junction complexes has been identified on the basis of immunohistochemical and transmission electron microscopic imaging. Tubular networks with extensions up to 400 mum were achieved. When EC suspensions were used, EC migration and network formation were critically affected by the status of the fibroblast. However, the absence of EC migration into the center of 14-day, but not 3-day, precultured fibroblast spheroids could not be attributed to loss of F viability. In parallel, it was also confirmed that migrated ECs that entered cluster-like formations became apoptotic, whereas the majority of those forming vessel-like structures remained viable for >8 days. Our protocols allow us to study the nature of tubule formation in a manner more closely related to the in vivo situation as well as to understand the basis for the integration of capillary networks in tissue grafts and develop methods of quantifying the amount of angiogenesis in spheroids using fibroblast and other cells isolated from the same patient, along with ECs.

BACKGROUND: The issue remains unresolved as to whether low frequency magnetic fields can affect cell behaviour, with the possibility that they may be in part responsible for the increased incidence of leukaemia in parts of the population exposed to them. METHODS: Combined treatment of HeLa cells with gamma-irradiation (1, 3 and 5 Grays) and extra low frequency magnetic fields of ~50 Hz was carried out under rigorously controlled conditions. RESULTS: Synchronised cells progressing from S-phase arrived at mitosis on average marginally ahead of irradiation controls not exposed to ELF. In no instance out of a total of twenty separate experiments did this "double-insult" further delay entry of cells into mitosis, as had been anticipated. CONCLUSION: This apparently "non-genotoxic" agent (ELF) appears to be capable of affecting cells that would normally arrest for longer in G2, suggesting a weakening of the stringency of the late cycle (G2) checkpoint.