Treatment resistance in potentially malignant disorders-'Nature' or 'Nurture' ?

BACKGROUND: Contemporary potentially malignant disorder management is based upon provisional histological diagnosis followed by interventional surgery to excise or ablate 'high-risk' mucosal lesions. Although the majority of patients achieve disease-free status post-treatment, others develop further or persistent disease unresponsive to intervention. METHODS: A detailed, retrospective clinico-pathological review of treatment resistant potentially malignant lesions, from a 590 patient cohort treated by CO2 laser surgery and followed for a mean of 7.3 years, was undertaken. Clinical outcome was determined at study census date (31 December 2014). RESULTS: A total of 87 patients (15%) exhibited PMD disease resistant to treatment: 34 (6%) became disease free following further treatment, whilst 53 (9%) had persistent disease despite intervention. Disease-free patients were younger, changed lesion appearance from erythroleukoplakia to leukoplakia (P = .004), developed further lesions at new sites, demonstrated reduction in dysplasia severity with time and required multiple treatments to achieve disease-free status (P = .0005). In contrast, persistent disease patients were older, male, often presented with proliferative verrucous leukoplakia (PVL) on gingival and alveolar sites, displayed less severe dysplasia initially and underwent laser ablation rather than excision (P = .027). CONCLUSION: Despite clinico-pathological profiling of treatment resistant patients, the precise inter-relationship between the inherent nature of potentially malignant disease and the external influence of treatment intervention remains obscure.

Profiling cancer risk in oral potentially malignant disorders-A patient cohort study.

BACKGROUND: Oral potentially malignant disorders harbour variable and unpredictable risk for squamous carcinoma development. Whilst current management strategies utilise histopathological diagnoses, dysplasia grading and targeted intervention for "high-risk" lesions, clinicians are unable to predict malignant potential. METHODS: Detailed, retrospective clinico-pathological analysis of potentially malignant lesions undergoing malignant transformation, from a 590 patient cohort treated by interventional laser surgery and followed for a mean of 7.3 years, was undertaken. Clinical outcome was documented at study census date (31 December 2014). RESULTS: A total of 99 patients (16.8%) developed cancer: 71 (12%) seen "unexpectedly" upon excision and 28 (4.8%) progressing to malignancy at a median of 87.3 months post-surgery. Thirty "unexpected" excisions were micro-invasive (42.3%) arising primarily in severely dysplastic precursors (75%) at ventro-lateral tongue and floor of mouth sites (54.5%); 1 patient (1.4%) had a cancer-related death, whilst 58 (81.7%) were disease free. A total of 19 of 28 "progressive" cancers (67.9%) arose at new sites, with erythroleukoplakia a significant predictor of malignancy (P = .0019). Nine (32.1%) developed at the same precursor site, with 6 (77.7%) on the ventro-lateral tongue and floor of mouth. Three (10.7%) were micro-invasive, 9 patients (32.1%) died from metastatic disease and 12 (42.9%) were disease free (P < .001). CONCLUSION: Squamous carcinoma may arise at the site of a precursor lesion as transformation or new-site development via field cancerisation. Whilst interventional surgery facilitates early diagnosis and treatment of occult disease, thus reducing risk from same-site transformation, new-site cancer is a significant long-term risk for patients with potentially malignant disorder.

Efficacy of oral brush biopsy in potentially malignant disorder management.

BACKGROUND: Oral potentially malignant disorders (PMD) harbour unpredictable risk for squamous cell carcinoma development. Current management requires tissue biopsy for histopathology characterisation, dysplasia grading and targeted intervention to "high-risk" lesions, although evidence-based guidelines are limited and diagnoses subjective. This study investigated the use of adjunctive oral brush biopsy techniques during the management of PMD in a UK hospital population. METHODS: Retrospective review of a 310 PMD patient cohort presenting to Maxillofacial Surgery in Newcastle upon Tyne with new, single-site lesions between December 2009 and May 2014. Patients underwent Orcellex® brush biopsy and liquid-based cytology examination in addition to conventional biopsy techniques, with management proceeding along established care pathways. Patient demographics, cytology data, most significant histopathology diagnoses and clinical outcome were all documented at the study census date (31.12.15). RESULTS: A total of 170 male & 140 female patients (age range 18-91 years), exhibiting primarily leukoplakia (86.5%) at floor of mouth and ventrolateral tongue sites (44.9%), were identified. Management comprised: observation (49.7%), laser surgery (44.9%), antifungal treatment (3.5%) and Head & Neck clinic referral following cancer diagnosis (1.9%). Clinical outcomes were as follows: disease free (51.3%), persistent PMD (42.3%) and malignant transformation (6.4%). Histology and cytology diagnoses strongly correlated (r = .305). Treatment modality, lesion site, histology and cytology diagnoses were the best predictors of clinical outcome. CONCLUSIONS: Orcellex® brush cytology provides reliable diagnoses consistent with conventional histopathology and offers less invasive, adjunctive assessment appropriate for long-term monitoring of patients in specialist clinics.

Chronic stress and Rosiglitazone increase indices of vascular stiffness in male rats.

Prolonged and/or frequent exposure to psychological stress responses may lead to deterioration of organs and tissues, predisposing to disease. In agreement with this, chronic psychosocial stress is linked to greater cardiovascular risk, including increased incidence of atherosclerosis, myocardial ischemia, coronary heart disease, and death. Thus the association between stress and cardiovascular dysfunction represents an important node for therapeutic intervention in cardiovascular disease. Here we report that 2weeks of chronic variable stress (CVS) increased indices of vascular stiffness, including increased collagen deposition in the aortic adventitia and increased resting pulse pressure, in male rats. Thus CVS may represent a useful rodent model for stress-associated CVD, especially for aging populations for which widening pulse pressure is a well-known risk factor. Additionally, we report that the thiazolidinedione Rosiglitazone (RSG) blunts chronic stress-associated increases in circulating corticosterone. Despite this, RSG was not protective against adverse cardiovascular outcomes associated with chronic stress. Rather RSG itself is associated with increased pulse pressure, and this is exacerbated by chronic stress-highlighting that chronic stress may represent an additional contributor to RSG-associated cardiovascular risk.

Interventional laser surgery for oral potentially malignant disorders: a longitudinal patient cohort study.

Oral squamous cell carcinoma (OSCC) is a lethal disease, with rising incidence. There were 6767 new OSCC cases and 2056 deaths in the UK in 2011. Cancers are preceded by oral potentially malignant disorders (PMDs), recognizable mucosal diseases harbouring increased SCC risk, offering clinicians a 'therapeutic window' to intervene. Contemporary practice remains unable to predict lesion behaviour or quantify malignant transformation risk. No clear management guidelines exist and it is unclear from the literature whether early diagnosis and intervention prevents cancer. Between 1996 and 2014, 773 laser treatments were performed on 590 PMD patients in Newcastle maxillofacial surgery departments. The efficacy of the intervention was examined by review of the clinicopathological details and clinical outcomes of the patients (mean follow-up 7.3 years). Histopathology required up-grading in 36.1% on examining excision specimens. Seventy-five percent of patients were disease-free, mostly younger patients with low-grade dysplasia; 9% exhibited persistent disease and were generally older with proliferative verrucous leukoplakia. Disease-free status was less likely for erythroleukoplakia (P=0.022), 'high-grade' dysplasia (P<0.0001), and with lichenoid inflammation (P=0.028). Unexpected OSCC was identified in 12.0%, whilst 4.8% transformed to malignancy. Interventional laser surgery facilitates definitive diagnosis and treatment, allows early diagnosis of OSCC, identifies progressive disease, and defines outcome categories. Evidence is lacking that intervention halts carcinogenesis. Multicentre, prospective, randomized controlled trials are needed to confirm the efficacy of surgery.

Oral precursor lesions and malignant transformation--who, where, what, and when?

Oral potentially malignant disorders (PMD) are recognisable mucosal conditions that have an unpredictable risk of transformation to squamous cell carcinoma (SCC), a lethal and deforming disease of rising incidence. Contemporary management is based on clinical recognition of suspicious lesions and incisional biopsy to enable histopathological assessment and grading of dysplasia, together with excision of high-risk lesions and long-term surveillance. However, it is impossible to predict clinical outcome or risk of malignant transformation. Our aim was to evaluate the relevance of previously identified oral precursor lesions for the development of SCC and staging of disease. We therefore retrospectively reviewed 1248 cases of SCC diagnosed in oral and maxillofacial surgery units at Newcastle upon Tyne and Sunderland hospitals between 1996 and 2009. Of them, 58 identifiable precursor lesions became malignant but only 25 had been dysplastic on initial biopsy; 19 of 33 non-dysplastic lesions exhibited lichenoid inflammation only. SCC arose most often on the ventrolateral tongue and floor of the mouth, with a mean transformation time of 29.2 months. Transformation time was significantly shorter in men (p=0.018) and those over 70 years of age (p=0.010). Patients who consumed more than 21 units of alcohol/week and those who had had interventional laser surgery to treat precursor lesions, had higher-staged tumours (p=0.048). Although retrospective, this study shows that the results of incisional biopsy and grading of dysplasia have limited use as predictive tools, and supports the view that cancer may arise in the absence of recognisable epithelial dysplasia. Our findings confirm the importance of clinical vigilance and active surveillance in the management of all patients with clinically suspicious oral lesions, irrespective of the histological findings.

Oral and maxillofacial surgery: "publication hot spots" in the United Kingdom.

We aimed to evaluate the number of articles in the British Journal of Oral and Maxillofacial Surgery (BJOMS) written by consultants and trainees within deaneries in the UK. We reviewed the authorship of articles published between January 2010 and July 2011, and subdivided them by deanery and first author. In total, 76 full length articles were published (30 written by consultants and 46 by trainees), and half came from the London, Wessex, and Mersey deaneries. Technical notes were mostly written by trainees; 5 of the 22 published came from London. A total of 38 short communications were published; most were from the Wessex (21%), Yorkshire and Humber (29%), and Mersey (13%) deaneries, and in 32% a consultant was first author. A league table of all publications showed that most came from Wessex (n=25), Mersey (n=23), and Yorkshire and Humber (n=23). Most publications by trainees came from Wessex (n=20), followed by Yorkshire (n=18) and London (n=11). The study shows that the amount of published material varies considerably between oral and maxillofacial surgery (OMFS) units in the UK. To realise their ambitions, potential trainees with an interest in research and publications may consequently find the best support in these deaneries.

Complications following interventional laser surgery for oral cancer and precancerous lesions.

Interventional carbon dioxide laser surgery is the preferred method to treat oral precancerous lesions and early invasive squamous cell carcinomas (SCCs). Little is known, however, about the complications that patients experience after such treatment. We retrospectively reviewed the hospital records of 82 patients with new dysplastic oral lesions or early invasive oral SCCs treated by laser surgery in the maxillofacial unit at Newcastle General Hospital. The most common postoperative complications were pain for more than two weeks after operation (n=28), bleeding (n=4), difficulties with speech (n=5), paraesthesia of the lingual nerve (n=17), difficulty swallowing (n=2), obstructive swelling of the submandibular gland (n=22), and tethering of the tongue (n=10). Overall, 78% of patients had one or more complication. In the absence of randomised controlled trials, this study provides the best available evidence for complication rates following interventional surgery. In addition to aiding in the preoperative counselling of patients, the data will help to inform and advise patients particularly during the immediate postoperative period.

Management of oral carcinoma: benefits of early precancerous intervention.

Management of oral precancerous lesions remains polarised between interventional surgery and conservative treatment. We have previously shown the efficacy of carbon dioxide laser excision for both diagnosis and treatment of oral precancerous lesions. The aim of this study was to review the clinicopathological details of a group of patients in whom pre-existing but occult invasive carcinoma was diagnosed histopathologically in specimens excised by laser. We retrospectively reviewed 169 patients who attended the Maxillofacial Dysplasia Clinic at Newcastle General Hospital with single, new oral premalignant lesions over a 5-year period (2004-2008). They were all treated by laser excision of lesions that were confirmed to be dysplastic from examination of preoperative incisional biopsy specimens. There was a significant correlation between the results of diagnostic incisional, and laser excision, biopsy specimens (p < 0.01), but 15 patients had signs of occult invasive carcinoma in the excision specimens (9%). In all cases the carcinomas were completely excised by the laser. Carbon dioxide laser excision is not only an effective treatment of precancerous lesions, but also facilitates early diagnosis and management of oral carcinoma at a stage when it is otherwise clinically undetectable.

Clinicopathological behaviour of multiple oral dysplastic lesions compared with that of single lesions.

Oral precancerous lesions may be solitary or multifocal, the latter being difficult to manage because of extensive field change. The aim of this study was to characterise differences in clinicopathological features, proliferative labelling indexes for cyclin A, cyclin B1, and Ki67, and clinical outcome 5 years after laser resection in a group of patients presenting with single and multiple oral precancerous lesions. Ninety-six patients with 132 lesions (78 single and 18 multiple) were recruited, and there were no significant differences between those with single and multiple lesions with respect to age, sex, smoking, or alcohol consumption, although multiple lesions were significantly more common in smokers who ate little fruit and vegetables (p=0.02). Clinically, most lesions were leukoplakia, with ulcerated or exophytic lesions appearing singly. There were significant differences in site, single lesions being most common on the floor of the mouth and the ventrolateral tongue, and multiple lesions preferring the buccal mucosa (p=0.0002). The most severe dysplasia was seen in single lesions (p=0.001) with labelling indexes for cyclin A and Ki67 being significantly higher in these (p=0.04 and p=0.01, respectively). Oral squamous cell carcinoma developed in 3/78 single lesions and 4/18 multiple ones. There are distinct differences between single and multiple lesions that have implications for the prophylaxis and management of oral cancer.

The role of alcohol in oral precancer: observations from a North-East England population.

Alcohol is known to be a risk factor for oral precancerous lesions, but evidence has been weakened by subjective estimates of alcohol intake from patients, and confounded by their use of tobacco. Red cell macrocytosis, assessed by calculation of mean corpuscular volume (MCV), may be a useful objective indicator of chronic alcohol intake. The aim of this study was to compare subjective and objective measures of alcohol intake in patients with oral precancerous lesions and assess the use of reported alcohol intake and MCV on assessing the degree of dysplasia at presentation and their role as markers of the behaviour of such lesions by assessing clinical outcome after treatment. Fifty-four new patients were recruited. All were smokers and had histologically confirmed single dysplastic oral precancerous lesions, but had had no previous treatment. Subjective data about their alcohol consumption were recorded, and blood samples taken for the assessment of MCV. All patients had laser excision of their lesions, which were assessed histopathologically for signs of dysplasia. Patients were followed up for 2 years. The significance of differences was assessed using Fisher's exact test. Alcohol intake of more than 28 units/week and MCV of over 100 were associated with increased dysplasia at presentation (p=0.01 and p=0.03, respectively). Thirty-six patients were disease-free at 2 years, but 18 developed further disease, often at new sites. Alcohol intake of more than 28 units/week was significantly associated with an increased risk of further disease (p=0.03), particularly recurrence at the same site (p=0.02).

Predicting recurrence after oral precancer treatment: use of cell cycle analysis.

Prediction of the behaviour of oral precancerous lesions (OPLs) is unreliable in clinical practice. The aim of this study was to analyse the efficacy of cell cyclin markers A and B1, and the proliferative marker Ki67, in predicting clinical outcome for patients with OPLs. A cohort of previously-treated patients with single OPLs were retrieved from the MaxilloFacial Dysplasia database and reviewed. All had dysplastic lesions excised by laser and were followed up for 5 years post-treatment. Outcome was determined as no recurrence or further disease. Excision specimens were re-examined immunohistochemically and labelling indices (LIs) for cyclin A, B1 and Ki67 determined. Forty patients, aged between 31 and 91 years, were recruited. There were no differences in age or sex. OPLs were predominantly leukoplakias on the floor of mouth or ventro-lateral tongue (65%), most of which exhibited moderate or severe dysplasia. Cyclin A LIs ranged from 3.9% to 31.3%, B1 0 to 28.3% and Ki67 3.5% to 54.5%. Using median LIs as 'cut off points' (12% cyclins; 22% Ki67) Kaplan-Meier survival analysis showed a significant risk of further progression of disease in patients with OPL LIs exceeding median values (Cyclin A p=0.02, Cyclin B1 p=0.01, Ki67 p=0.025). By combining analysis of both Cyclin A and B LI, the significance of the difference was increased (p<0.01). Cell cycle analysis is effective in identifying patients at risk of further progression of disease following treatment of OPLs. Multi-centre, longitudinal trials are needed to assess the precise role of cell cycle markers in their management.

Cyclin A activity predicts clinical outcome in oral precancer and cancer.

Accurate, predictive assessment of the behaviour of oral cancers and precancers remains elusive. Increasing dysregulation of cell proliferation is a feature of carcinogenesis, and alterations in cyclin proteins regulating cell cycle progression are involved in enhanced cell proliferation. The authors of the present study have previously demonstrated increased proliferative activity in oral dysplastic lesions and poorly differentiated carcinomas, and hypothesize that cell proliferation can be used as a predictive agent in clinical management. In this preliminary study, immunohistochemical quantification of cyclin A expression was carried out for 33 excised oral lesions (ranging from mild dysplasia to invasive squamous cell carcinoma, SCC). Clinical outcome was determined as: no disease after 2 years follow-up, persistent disease, or further disease presentation. Labelling Indices (LIs) ranged from 5.5 to 32.1%, and whilst a trend to increased labelling in increasingly dysplastic and neoplastic tissue was seen, this was not statistically significant (P=0.06). High LIs were related to poor clinical outcome (P=0.003), suggesting a definite role for cyclin A measurement as a predictive tool in clinical management.

A study of public opinion on the use of tissue samples from living subjects for clinical research.

AIMS: To assess public opinion on the use of tissue samples from living adults and children for clinical research. METHODS: A questionnaire study of 100 healthy volunteers (100% response rate) from a Newcastle NHS dental practice. The issues investigated were the types of tissues that individuals were prepared to donate for research, the type of research donors would be prepared to consent to, and attitudes to research on children's tissues. RESULTS: Eighteen per cent of the participants said that they would not give consent for research to be carried out on their tissues, 50% would not give consent for the donation of a child's tissues. Only 26% of subjects said that they would give consent for research on genetic cloning compared with 82% for cancer research. Sex differences existed in the responses. CONCLUSIONS: Greater research attention needs to be given to public opinion on the use of tissue from living subjects for medical research to facilitate drafting of new legislation.

Regulation of heat shock transcription factor 1 by stress-induced SUMO-1 modification.

Heat shock transcription factor 1 (HSF1) mediates the induction of heat shock protein gene expression in cells exposed to elevated temperature and other stress conditions. In response to stress HSF1 acquires DNA binding ability and localizes to nuclear stress granules, but the molecular mechanisms that mediate these events are not understood. We report that HSF1 undergoes stress-induced modification at lysine 298 by the small ubiquitin-related protein called SUMO-1. Antibodies against SUMO-1 supershift the HSF1 DNA-binding complex, and modification of HSF1 in a reconstituted SUMO-1 reaction system causes conversion of HSF1 to the DNA-binding form. HSF1 colocalizes with SUMO-1 in nuclear stress granules, which is prevented by mutation of lysine 298. Mutation of lysine 298 also results in a significant decrease in stress-induced transcriptional activity of HSF1 in vivo. This work implicates SUMO-1 modification as an important modulator of HSF1 function in response to stress.

Sumo-1 modification regulates the DNA binding activity of heat shock transcription factor 2, a promyelocytic leukemia nuclear body associated transcription factor.

Heat shock transcription factor 2 (HSF2) is a transcription factor that regulates heat shock protein gene expression, but the mechanisms regulating the function of this factor are unclear. Here we report that HSF2 is a substrate for modification by the ubiquitin-related protein SUMO-1 and that HSF2 colocalizes in cells with SUMO-1 in nuclear granules. Staining with anti-promyelocytic leukemia antibodies indicates that these HSF2-containing nuclear granules are PML bodies. Our results identify lysine 82 as the major site of SUMO-1 modification in HSF2, which is located in a "wing" within the DNA-binding domain of this protein. Interestingly, SUMO-1 modification of HSF2 results in conversion of this factor to the active DNA binding form. This is the first demonstration that SUMO-1 modification can directly alter the DNA binding ability of a transcription factor and reveals a new mechanism by which SUMO-1 modification can regulate protein function.