Fibroblast activation and senescence in oral cancer.

There is now compelling evidence that the tumour stroma plays an important role in the pathogenesis of cancers of epithelial origin. The pre-eminent cell type of the stroma is carcinoma-associated fibroblasts. These cells demonstrate remarkable heterogeneity with activation and senescence being common stress responses. In this review, we summarise the part that these cells play in cancer, particularly oral cancer, and present evidence to show that activation and senescence reflect a unified programme of fibroblast differentiation. We report advances concerning the senescent fibroblast metabolome, mechanisms of gene regulation in these cells and ways in which epithelial cell adhesion is dysregulated by the fibroblast secretome. We suggest that the identification of fibroblast stress responses may be a valuable diagnostic tool in the determination of tumour behaviour and patient outcome. Further, the fact that stromal fibroblasts are a genetically stable diploid cell population suggests that they may be ideal therapeutic targets and early work in this context is encouraging.

The opposing roles of NOTCH signalling in head and neck cancer: a mini review.

NOTCH signalling can exert oncogenic or tumour suppressive effects in both solid and haematological malignancies. Similar to T-cell acute lymphoblastic leukaemia (T-ALL), early studies suggested a pro-tumorigenic role of NOTCH in head and neck squamous cell carcinoma (HNSCC), mainly based on the increased expression levels of the genes within the pathway. Recently, data from exome sequencing analyses unexpectedly pointed to a tumour suppressor role for NOTCH in HNSCC by identifying loss-of-function mutations in the NOTCH1 gene in a significant proportion of patients. These data have questioned the accepted role of NOTCH in HNSCC and the possible rationale of targeting NOTCH in this disease. This review summarises the current information on NOTCH signalling in HNSCC and discusses how this pathway can apparently exert opposing effects within the same disease.

Lysophosphatidic acid and calcitriol co-operate to promote human osteoblastogenesis: requirement of albumin-bound LPA.

Lysophosphatidic acid (LPA), a pleiotropic signalling lipid is assuming growing significance in osteoblast biology. Although committed osteoblasts from several mammalian species are receptive to LPA far less is known about the potential for LPA to influence osteoblast formation from their mesenchymal progenitors. An essential factor for both bone development and post-natal bone growth and homeostasis is the active metabolite of vitamin D3, calcitriol (D3). Previously we reported how a combination of LPA and D3 synergistically co-operated to enhance the differentiation of immature human osteoblasts. Herein we provide evidence for the formation of human osteoblasts from multiple, primary human bone marrow derived stromal (stem) cells (hBMSCs). Importantly osteoblast development from hBMSCs only occurred when LPA was administered as a complex with albumin, its natural carrier. Collectively our findings support a co-operative role of LPA and D3 in osteoblastogenesis, findings which may aid the development of novel treatment strategies for bone repair.

Molecular events including p53 and k-ras alterations in the in vitro progression of a human colorectal adenoma cell line to an adenocarcinoma.

The aim of the current study was to identify genetic abnormalities in human colorectal adenoma and carcinoma derived cell lines, and to determine whether the genetic changes which occur in vitro are relevant to the in vivo situation. Loss of 1p(33-35) region was shown to be the most common chromosome 1 abnormality and loss of heterozygosity (LOH) of the DCC gene and/or adjacent sequences was detected in all adenoma derived cells as well as the carcinoma cell lines. The level of p53 protein was also investigated as increased cellular p53 protein had previously been associated with mutation of the p53 gene. A further aim was to investigate genetic changes in our in vitro model of tumour progression, where the adenoma derived PC/AA cell line has previously been converted in vitro to two distinct tumorigenic phenotypes, producing either an adenocarcinoma or a mucinous carcinoma in athymic nude mice. Progression to the adenocarcinoma phenotype was shown to involve a specific chromosome 1 rearrangement, loss of both normal copies of chromosome 18 (although DCC gene sequences were retained), loss of the remaining wild type allele of k-ras resulting in homozygosity for the k-ras codon 12 mutation and increased cellular p53 protein as detected by SDS-PAGE Western blotting. The increase in p53 protein was shown not to be due to the acquisition of a mutation in the p53 gene. Interestingly, progression of the adenoma derived PC/AA cell line to the mucinous malignant phenotype did not involve any of these molecular rearrangements, suggesting that different genetically distinct pathways are involved in colorectal carcinogenesis. These studies show that the genetic changes in our in vitro model of human colorectal tumour progression are similar to those observed in in vivo studies.

TGF-beta isoforms fail to modulate inositol phosphates and cAMP in normal and tumour-derived human oral keratinocytes.

This study examined inositol phosphate and cAMP regulation by TGF-beta 1, -beta 2 and -beta 3 in normal and tumour-derived human oral keratinocytes. Previous findings indicated that the cell lines expressed TGF-beta cell surface receptors and had a range of response to exogenous TGF-beta 1, -beta 2 and -beta 3 from being refractory to the ligand to marked inhibition. Basal levels of inositol phosphates broadly reflected the differentiation status of the cells as demonstrated by involucrin expression, but did not correlate with responsiveness to TGF-beta 1, as measured previously by thymidine incorporation. Treatment of cells with bradykinin or serum caused up-regulation of inositol phosphate levels; by contrast, TGF-beta 1, -beta 2 and -beta 3 failed to modulate inositol phosphates. In two tumour-derived cell lines, the TGF-beta isoforms had no effect on cAMP levels, despite a significant increase in cAMP using a potent agonist of adenylate cyclase (forskolin). Furthermore, the cAMP analogue, dibutyryl cAMP, failed to mimic the inhibitory or refractory responses of TGF-beta in these cells lines. The results demonstrate that in normal and tumour-derived human oral keratinocytes, TGF-beta signal transduction is not mediated by inositol phosphates or cAMP.

Cloning and sequence analysis of an intron-containing domain from a peptide synthetase-encoding gene of the entomopathogenic fungus Metarhizium anisopliae.

A gene encoding a putative peptide synthetase has been cloned and partially sequenced from the filamentous fungus, Metarhizium anisopliae. The deduced amino acid sequence of one entire domain and the following spacer is typical of fungal peptide synthetases, showing good conservation of the six expected core sequences. There are two introns within this region, the first interrupting core 5 (RLDLTDIE) of the domain and the second in a conserved area of the spacer region.

Cloning and characterisation of a gene encoding a cuticle-degrading protease from the insect pathogenic fungus Metarhizium anisopliae.

Metarhizium anisophilae (Ma) secretes a range of proteases when grown in vitro on insect cuticle. A trypsin-like serine protease, PR2, was purified from culture filtrates by anion exchange chromatography and the N-terminal sequence determined. Using oligodeoxyribonucleotide probes based on this sequence and that of the highly conserved trypsin active site, a gene was isolated from a lambda EMBL3 genomic library of Ma isolate ME1. Sequencing of the gene and RT-PCR revealed that the gene contains two introns which are 94 and 40 bp long. The deduced protein consists of 254 amino acids, has a putative signal sequence to allow transport into the endoplasmic reticulum and probably undergoes a second proteolytic processing step at its N terminus to yield the mature enzyme. The putative mature enzyme has extensive homology with other serine proteases of the trypsin subclass and, in particular, with the trypsin characterised from Fusarium oxysporum.

Misonidazole in patients receiving radical radiotherapy: pharmacokinetic effects of phenytoin, tumor response and neurotoxicity.

In 1978, a pilot study began of 29 patients with advanced tumors of the head and neck. The study showed a initial peripheral neuropathy rate of 55%, despite a dose limitation of 12g/m2 of misonidazole. Tumor response at 9 months was most encouraging. We are now able to examine tumor response and persistence of neuropathy in these patients 2 1/2 years after radical radiotherapy. The results are comparable with those obtained with hyperbaric oxygen in a clinical trial at this center during the 1970's. Neuropathy was a serious side effect but the drug phenytoin has been shown to shorten the half-life of misonidazole. We have examined the effect of phenytoin on the pharmacokinetics of misonidazole in 13 patients who received radical radiotherapy for advanced head and neck tumors or oesophageal tumors. Misonidazole was given in multiple doses, i.e. daily or weekly as it would be used in conventional therapy. Phenytoin was given either daily throughout treatment, or it was withdrawn during treatment. There were dramatic changes in the half-life of misonidazole, but the concentration at the time of irradiation was little affected. The significant changes in the half-life of misonidazole and the increased concentration of the metabolite desmethylmisonidazole are discussed.

ad and ay subtypes of hepatitis B antigen in a case of hypogammaglobulinaemia.

We describe the finding of d and y specificities of hepatitis B surface antigen (HBsAg) in a case of hypogammaglobulinaemia of the 'common variable' type treated with fresh frozen plasma infusions. Absorption studies show that the two specificities are on separate particles, suggesting dual infection. It raises important questions regarding the relationship between HBsAg persistence and the immune status of the carrier.

Under-registration of melanoma in Wales in 1998: use of the capture-recapture method to estimate the 'true' incidence.

Several studies have suggested that melanomas may be significantly under-recorded in cancer registries and that smaller, thinner, better prognosis lesions are the ones most likely to be missed. A systematic search of three independent sources of melanoma data in Wales for 1998 revealed a total of 406 histologically confirmed cases, of which only 194 were known to the cancer registry. Eighty-one per cent of the total cases were registered on a specialist melanoma register, compared with 48% on the cancer registry database. From the cancer registry data alone, the world age-standardized incidence rates (WASRs) were 4.3 and 5.8 per 100,000 for males and females, respectively, but these increased to 8.2 and 10.2 with the addition of histologically confirmed cases discovered from other sources. The capture-recapture method estimated the number of melanomas not ascertained by either means to be 140, resulting in a 'true' incidence of 546 cases for 1998 compared with just 194 cases from the cancer registry data alone. The 'true' WASRs are 11.2 and 13.4 per 100,000 for males and females, respectively, which are some of the highest in Europe. There was evidence to support the hypothesis that smaller, thinner melanomas are more likely to be recorded on a specialist melanoma register than on the cancer registry database.

A review of inherited cancer syndromes and their relevance to oral squamous cell carcinoma.

This paper examines the genetic defects associated with inherited cancer syndromes and their relevance to oral cancer. Tumour suppressor genes are now thought of as either gatekeepers or caretakers according to whether they control cell growth directly by inhibiting cell proliferation and/or promoting cell death (gatekeepers) or whether they maintain the integrity of the genome by DNA repair mechanisms (caretakers). In disorders such as xeroderma pigmentosum, ataxia telangiectasia, Bloom syndrome and Fanconi's anaemia, where there are defective caretaker genes, there is an increased incidence of second primary malignancies, including oral cancer. By contrast, with the exception of Li Fraumeni syndrome, abnormalities of gatekeeper genes do not predispose to oral cancer. Not only do Li Fraumeni patients develop second primary malignancies, but defects of the p53 pathway (p53 mutation, MDM2 over-expression, CDKN2A deletion) appear to be a ubiquitous feature of sporadic oral cancer as it occurs in the West. The findings suggest that genetic instability is of fundamental importance in the pathogenesis of oral cancer.

Functional significance of MMP-2 and MMP-9 expression by human malignant oral keratinocyte cell lines.

This study examined the expression of MMP-2 and MMP-9 in normal and human malignant oral keratinocytes. The expression of pro-MMP-2 and pro-MMP-9 was heterogeneous in the malignant cell lines. Normal oral keratinocytes expressed less pro-MMP-2 and more pro-MMP-9 than their malignant counterparts. Cells that expressed high levels of both MMP-2 and MMP-9 showed the greatest degree of invasion through Matrigel in vitro compared to cells with either low or variable levels of these enzymes; normal keratinocytes were non-invasive in these conditions. The degree to which the cells invaded through Matrigel was similar to their motility in the absence of Matrigel and was not influenced by the activation of the pro-enzymes or the inhibition of enzyme activity using a chemical inhibitor of gelatinases. Cells were transplanted orthotopically to athymic mice and demonstrated a variable capacity not only to form tumours at the site of inoculation but, also, to metastasise; normal oral keratinocytes were non-tumorigenic. There was no correlation between the expression of either MMP-2 or MMP-9 and the tumorigenic/metastatic phenotype. The results emphasise the limitations of correlating in vitro and in vivo assays of tumour cell behaviour and suggest that invasion/motility in vitro may be a distinct phenotype from tumorigenicity/metastasis in vivo.

Homeopathy: what is it and is it of value in the care of patients with cancer?

The underlying principles of homeopathy include treating 'like with like' by remedies which are potentized by serial dilution and succussion. These distinguish homeopathy from other forms of alternative or complementary therapy. Conventional scientific wisdom dictates that homeopathy should have no effect above and beyond placebo but experiments on ultra-high dilutions of solutes together with some clinical data suggest the intriguing possibility that it might do in some circumstances. However, most clinical evidence comes from treating relatively minor self-limiting diseases and little comes from treating life-threatening disorders such as cancer. This paper explains the principles of homeopathy and reviews the data on its use in cancer care.

Consent to treatment: somebody's moved the goalposts.

It is timely to consider the issue of consent to treatment in clinical oncology in a period of changing expectations where signed consent has become obligatory. Consent has a dual purpose to protect patient and doctor; if one is achieved without the other then the task is incomplete. In terms of meaningful consent the consent form is of relatively little value and it is necessary to appreciate that, if patients are to truly exercise their right of choice, they need information in ways and in situations which may mean oncologists having to alter their work practices. Oncologists work without necessarily knowing what other oncologists do or do not tell their patients, or of what ought to be told. It is unlikely that the probable wide variations for similar patients across the country will be tolerated in the future and this should act as an incentive to arrive at agreed guidelines which are protective of all the legitimate interests. The problems and the possible solutions are discussed.

The concept of intellectual property and its implication for oncology.

Oncologists and scientists in oncology centres work in an environment that seeks and promotes innovation and are under an ethical obligation to disseminate new knowledge for the benefit of society. That knowledge (which may give rise to intellectual property rights) may have substantial commercial value, and with it comes the need to protect employers' and employees' legitimate interests through patents and copyright. Employers and employees may not fully appreciate the legal and ethical obligations surrounding ownership of such knowledge, so both should co-operate in formulating policies which balance the need for disseminating new knowledge and the need to protect institutional interests. This is particularly appropriate for the new NHS trusts which may undertake research jointly with commercial organizations. This article outlines the important issues for consideration.

A detailed baseline audit of radiotherapy for primary breast cancer: old habits, new principles.

In the UK, fundamental changes are occurring to the organization and delivery of services to breast cancer patients, prompted by the Calman/Hine initiative and by the perception that outcomes in the UK compare unfavourably with the rest of Europe and North America. Breast cancer, being a common malignancy, has often been treated by general surgeons and oncologists rather than by site-specialized multidisciplinary teams. Outcome results for many of these surgeons and oncologists may be unknown, so it becomes difficult, in any particular centre, to quantify changes that arise from the reorganization of treatment services and adherence to new guidelines without first determining baseline results. A retrospective audit of the 350 patients with primary breast cancer who received radiotherapy in Velindre NHS Trust in 1988-1989 has been performed to provide a baseline against which future results can be measured following the adoption of agreed guidelines. The endpoints for this study were: the completeness of data; actual treatment received versus current guideline recommendations; locoregional recurrence rates; and radiation related morbidity. The adherence to agreed guidelines is likely to ensure more appropriate surgery of the primary disease and will limit radiotherapy compared with 1988-1989 in that much radiation to the axilla and Internal Mammary Chain and 'booster' treatment to the primary site will be avoided in the future. The incidence of brachial plexus neuropathy was consistent with the radiation doses used and an analysis of this data suggests that the adoption of the guidelines will probably result in the complete avoidance of this complication in the future. Likewise, it is expected that the incidence of soft tissue fibrosis, telangiectasia and pneumonitis should diminish, but this and the effect of the clinical guidelines on local recurrence rates remains to be demonstrated by future comparable audits.

Thyroid cancer in Wales 1985-1996: a cancer registry-based study.

No dedicated clinicopathological database for thyroid cancer exists in Wales to provide information about the incidence, histology, treatment and outcome of this uncommon malignancy. We have used cancer registry data to examine the epidemiology of thyroid cancer in Wales between 1985 and 1996 and have searched the database for clinically relevant data about stage, treatment and survival. A total of 699 cases were identified in this 12-year period, with no significant differences in their distribution between the five health authorities. Further analysis revealed a very wide age range (8-93 years) with a predominance of females (F:M ratio 2.8:1) and survival strongly influenced by gender, age and histological type. There were insufficient data to stage these cancers and only limited data about the surgical procedures undertaken. There was no information on non-surgical treatments (radio-iodine, radiotherapy and chemotherapy) in the database. Cancer registry data is well able to sustain an analysis of the epidemiology of thyroid cancer but further work is necessary to improve the quality of clinically relevant information about stage and treatment that could be used for audit.

The treatment of squamous cell carcinoma of the paranasal sinuses.

Of 72 cases of paranasal sinus malignancy that were identified, 47 were biopsy proven squamous cell carcinomas. Actuarial analysis of these patients receiving both radical and palliative treatment revealed an overall 5-year survival of 37.4%, and a relapse-free survival of 31.6%. For patients receiving radical treatments (i.e. radiotherapy alone, or preoperative radiotherapy followed by elective surgery), the 5-year survival results were identical (46%). In this series radiotherapy alone is an effective treatment, and subsequent planned surgery did not improve the survival.

Effect of deprivation on survival of patients with head and neck cancer: a study of 20,131 cases.

There is increasing evidence that, for many cancers, the survival of socioeconomically deprived patients is worse compared with those who are more affluent. This study provides additional evidence that this is true for patients with head and neck cancers. However, the detrimental effects of deprivation were not found to be lifelong, and in this study, were confined to the first 12-18 months after diagnosis. After this there were no significant deprivation-associated effects on subsequent survival. The reasons for the initial increased mortality in the deprived are not clear but may be related to more advanced stage, more biologically aggressive cancers, greater co-morbidity or worse treatment.

The influence of breast size on late radiation reaction following excision and radiotherapy for early breast cancer.

The correlation between late radiation reaction and brassiere size (bust and cup size) in 133 patients with T1 and T2 breast cancer treated by excision and radiotherapy was examined. The radiotherapy schedule used was 4875 cGy in 15 x 3.25 Gy fractions over 39 days treating five times per fortnight. Patients were followed up to a minimum of 2 years (median 4.2 years). The incidence of unacceptable late radiation reaction rose significantly with bust size (P less than 0.001) and cup size (P less than 0.02). It is intended to treat ladies with large breasts with a reduced fraction size or on a daily basis.