Catriona McKenzie

A Phase 3 Randomized Trial of Nicotinamide for Skin-Cancer Chemoprevention

BACKGROUND Nonmelanoma skin cancers, such as basal-cell carcinoma and squamous-cell carcinoma, are common cancers that are caused principally by ultraviolet (UV) radiation. Nicotinamide (vitamin B3) has been shown to have protective effects against damage caused by UV radiation and to reduce the rate of new premalignant actinic keratoses. METHODS In this phase 3, double-blind, randomized, controlled trial, we randomly assigned, in a 1:1 ratio, 386 participants who had had at least two nonmelanoma skin cancers in the previous 5 years to receive 500 mg of nicotinamide twice daily or placebo for 12 months. Participants were evaluated by dermatologists at 3-month intervals for 18 months. The primary end point was the number of new nonmelanoma skin cancers (i.e., basal-cell carcinomas plus squamous-cell carcinomas) during the 12-month intervention period. Secondary end points included the number of new squamous-cell carcinomas and basal-cell carcinomas and the number of actinic keratoses during the 12-month intervention period, the number of nonmelanoma skin cancers in the 6-month postintervention period, and the safety of nicotinamide. RESULTS At 12 months, the rate of new nonmelanoma skin cancers was lower by 23% (95% confidence interval [CI], 4 to 38) in the nicotinamide group than in the placebo group (P=0.02). Similar differences were found between the nicotinamide group and the placebo group with respect to new basal-cell carcinomas (20% [95% CI, -6 to 39] lower rate with nicotinamide, P=0.12) and new squamous-cell carcinomas (30% [95% CI, 0 to 51] lower rate, P=0.05). The number of actinic keratoses was 11% lower in the nicotinamide group than in the placebo group at 3 months (P=0.01), 14% lower at 6 months (P<0.001), 20% lower at 9 months (P<0.001), and 13% lower at 12 months (P=0.001). No noteworthy between-group differences were found with respect to the number or types of adverse events during the 12-month intervention period, and there was no evidence of benefit after nicotinamide was discontinued. CONCLUSIONS Oral nicotinamide was safe and effective in reducing the rates of new nonmelanoma skin cancers and actinic keratoses in high-risk patients. (Funded by the National Health and Medical Research Council; ONTRAC Australian New Zealand Clinical Trials Registry number, ACTRN12612000625875.).

A phase II randomized controlled trial of nicotinamide for skin cancer chemoprevention in renal transplant recipients

DEAR EDITOR, Immunosuppressed organ transplant recipients have an 80-fold increased risk of squamous cell carcinoma (SCC) and a 16-fold increased risk of basal cell carcinoma (BCC); SCCs are more aggressive and more likely to metastasize in these patients. While mammalian target of rapamycin (mTOR) inhibitors may help reduce the incidence of SCC, the current mainstay of nonmelanoma skin cancer (NMSC) chemoprevention post-transplant is oral retinoids, which have side-effects, including liver and lipid abnormalities, mucocutaneous dryness and teratogenicity. Nicotinamide (vitamin B3) enhances repair of photodamaged DNA and prevents the inhibitory effects of ultraviolet radiation on the immune system without altering baseline immune reponses. In 386 immunocompetent participants at high risk of having skin cancer, nicotinamide 500 mg twice daily reduced new NMSCs by 23% (P = 0 02), with 20% fewer BCCs and 30% fewer SCCs compared with placebo. Actinic keratoses (AKs) were also significantly reduced by nicotinamide in the same phase III randomized controlled trial. However, it is unknown whether nicotinamide prevents NMSCs and is safe in immunosuppressed renal transplant recipients. Eligible adult immunosuppressed participants included those who had received a renal transplant ≥ 12 months ago, had stable renal function and a history of ≥ 2 NMSCs in the previous 12 months. Participants were ineligible if they commenced retinoids or mTOR inhibitors within 6 months, or used nicotinamide supplements or AK field treatments within the 4 weeks prior to enrolment. This double-blind study was conducted at the Royal Prince Alfred Hospital (Sydney, Australia; Australian New Zealand Clinical Trials Registry ACTRN12612000628842) after ethical approval was received. All participants provided written informed consent. Participants were enrolled (by A.C.C. and R.A.D.) and randomized 1:1 to receive either nicotinamide 500 mg twice daily or matched placebo (Insolar; Blackmores, Warriewood, Australia) by centralized randomization (permuted blocks; National Health and Medical Research Council Clinical Trials Centre) stratified by lifetime NMSC history (< 6 or ≥ 6 NMSCs), oral retinoid use and mTOR inhibitor use. Compliance was monitored by tablet counts at each visit (A.C.C). Skin cancer checks were performed at baseline and twice monthly up to 6 months by dermatologists blinded to treatment allocation (D.L.D., P.M.L.). AKs on the face, scalp, forearms and hands were counted at baseline and twice monthly up to 6 months by a dermatology Fellow blinded to allocation (A.C.C). All squamous lesions were reviewed by a single (blinded) histopathologist (C.A.M.) to ensure consistent reporting of SCC differentiation. Blood and urine samples were taken at baseline, 2 and 4 weeks, and 2, 4 and 6 months for full blood count, electrolytes, renal and liver function tests, drug levels and urine microalbumin/creatinine ratio. Blood pressure and weight were measured at baseline and twice monthly for 6 months. The primary end point was the number of new NMSCs (BCC + SCC, including invasive and in situ SCC) up to 6 months. Secondary end points included new BCCs, new SCCs, AK counts up to 6 months and safety. The planned sample size was 80 participants, in order to provide 80% power to detect a 50% reduction in the 6-month NMSC rate at the 5% level of significance, assuming that NMSC counts followed a Poisson distribution with a mean of 1 5 for the placebo group, allowing for ≤ 10% noncompliance. However, owing to slow recruitment because of an overestimation of the eligible participant population at our site, the study was stopped early; 22 participants were recruited over 1 5 years. Analyses were by intention to treat. As per the provision specified in the statistical analysis plan, a negative binomial model was used to analyse NMSC, SCC and BCC data, owing to overdispersion that rendered the Poisson model inappropriate. Models included an offset term to account for variation in follow-up duration. The 2-monthly postbaseline AK data were analysed using a mixed model for repeated measures, including treatment group, baseline value, time point and a time-by-treatment group interaction as covariates. From August 2012 to March 2014, 25 participants were assessed for eligibility and 22 were randomized [11 in each group; Figure S1 (see Supporting Information)]. The groups had similar baseline characteristics (Table 1). Follow-up was from August 2012 to August 2014. Median compliance was 93% and 98% for the placebo and nicotinamide groups, respectively. The 6-month NMSC rate was not significantly lower for the nicotinamide group [mean 2 7, 95% confidence interval (CI) 1 4–5 3; total 30 cancers] compared with placebo (mean 4 2, 95% CI 2 2–7 8; total 45 cancers); however, the numeric trend was dominated by one patient in the placebo group

The Effect of Aging on Acetaminophen Pharmacokinetics, Toxicity and Nrf2 in Fischer 344 Rats

We investigated the effect of aging on hepatic pharmacokinetics and the degree of hepatotoxicity following a toxic dose of acetaminophen. Young and old male Fischer 344 rats were treated with 800 mg/kg acetaminophen (young n = 8, old n = 5) or saline (young n = 9, old n = 9). Serum measurements showed old rats treated with acetaminophen had significantly lower serum alanine aminotransferase and higher acetaminophen and acetaminophen glucuronide levels and creatinine, compared with acetaminophen treated young rats (p < .05). Immunoblotting and activity assays showed old saline-treated rats had twofold lower cytochrome P450 2E1 activity and threefold higher NAD(P)H quinone oxireductase 1 protein expression and activity than young saline-treated rats (p < .05), although Nrf2, glutathione cysteine ligase-modulatory subunit, glutathione cysteine ligase-catalytic subunit, and cytochrome P450 2E1 protein expressions were unchanged. Primary hepatocytes isolated from young rats treated with 10 mM acetaminophen had lower survival than those from old rats (52.4% ± 5.8%, young; 83.6% ± 1.7%, old, p < .05). The pharmacokinetic changes described may decrease susceptibility to acetaminophen-induced hepatotoxicity but may increase risk of nephrotoxicity in old age.

Acetaminophen hepatotoxicity in mice: Effect of age, frailty and exposure type.

Acetaminophen is a commonly used analgesic that can cause severe hepatotoxicity in overdose. Despite old age and frailty being associated with extensive and long-term utilization of acetaminophen and a high prevalence of adverse drug reactions, there is limited information on the risks of toxicity from acetaminophen in old age and frailty. This study aimed to assess changes in the risk and mechanisms of hepatotoxicity from acute, chronic and sub-acute acetaminophen exposure with old age and frailty in mice. Young and old male C57BL/6 mice were exposed to either acute (300 mg/kg via oral gavage), chronic (100 mg/kg/day in diet for six weeks) or sub-acute (250 mg/kg, t.i.d., for three days) acetaminophen, or saline control. Pre-dosing mice were scored for the mouse clinical frailty index, and after dosing serum and liver tissue were collected for assessment of toxicity and mechanisms. There were no differences with old age or frailty in the degree of hepatotoxicity induced by acute, chronic or subacute acetaminophen exposure as assessed by serum liver enzymes and histology. Age-related changes in the acetaminophen toxicity pathways included increased liver GSH concentrations, increased NQO1 activity and an increased pro- and anti-inflammatory response to acetaminophen in old age. Frailty-related changes included a negative correlation between frailty index and serum protein, albumin and ALP concentrations for some mouse groups. In conclusion, although there were changes in some pathways that would be expected to influence susceptibility to acetaminophen toxicity, there was no overall increase in acetaminophen hepatotoxicity with old age or frailty in mice.

The effect of ageing on isoniazid pharmacokinetics and hepatotoxicity in Fischer 344 rats.

Isoniazid is the first‐line treatment for tuberculosis; however, its use is limited by hepatotoxicity. Age‐related differences in isoniazid pharmacokinetics and hepatotoxicity are uncertain. We aimed to investigate these in young (3 ± 0 months, n = 26) and old (23.0 ± 0.2 months, n = 27) male Fischer 344 rats following a low‐ or high‐dose toxic regimen of isoniazid or vehicle (4 doses/day over 2 days; low: 100, 75, 75, 75 mg/kg; high: 150, 105, 105, 105 mg/kg i.p. every 3 h). Fifteen hours after the last dose, animals were euthanized and sera and livers were prepared for analysis. Isoniazid treatment increased serum hepatotoxicity markers (alanine and aspartate transaminase) in young animals but not in old animals, and only reached significance with the high dose in young animals. Isoniazid treatment caused a trend towards an increase in necrosis in young animals with both doses. In contrast, microvesicular steatosis was increased in old isoniazid‐treated animals, reaching significance only with the low dose (steatosis prevalence in old: vehicle 1/9, isoniazid 4/5; P < 0.05). Among isoniazid‐treated animals, concentrations of toxic intermediates acetylhydrazine and hydrazine were higher in old than young animals (P < 0.05). With both doses, hepatic cytochrome P450 2E1 activity was higher in young animals compared with old (P < 0.05). There were no other age effects seen on any of the other measured enzymes involved in isoniazid metabolism (N‐acetyl transferase, amidase, glutathione‐S‐transferase). These results show age‐related changes in isoniazid pharmacokinetics may contribute towards differential patterns of toxicity and confirm that standard hepatotoxicity markers do not detect isoniazid‐induced microvesicular steatosis.

N-Acetyl cysteine does not prevent liver toxicity from chronic low-dose plus subacute high-dose paracetamol exposure in young or old mice.

Paracetamol is an analgesic commonly used by people of all ages, which is well documented to cause severe hepatotoxicity with acute overexposures. The risk of hepatotoxicity from nonacute paracetamol exposures is less extensively studied, and this is the exposure most common in older adults. Evidence on the effectiveness of N‐acetyl cysteine (NAC) for nonacute paracetamol exposures, in any age group, is lacking. This study aimed to examine the effect of long‐term exposure to therapeutic doses of paracetamol and subacute paracetamol overexposure, in young and old mice, and to investigate whether NAC was effective at preventing paracetamol hepatotoxicity induced by these exposures. Young and old male C57BL/6 mice were fed a paracetamol‐containing (1.33 g/kg food) or control diet for 6 weeks. Mice were then dosed orally eight times over 3 days with additional paracetamol (250 mg/kg) or saline, followed by either one or two doses of oral NAC (1200 mg/kg) or saline. Chronic low‐dose paracetamol exposure did not cause hepatotoxicity in young or old mice, measured by serum alanine aminotransferase (ALT) elevation, and confirmed by histology and a DNA fragmentation assay. Subacute paracetamol exposure caused significant hepatotoxicity in young and old mice, measured by biochemistry (ALT) and histology. Neither a single nor double dose of NAC protected against this toxicity from subacute paracetamol in young or old mice. This finding has important clinical implications for treating toxicity due to different paracetamol exposure types in patients of all ages, and implies a need to develop new treatments for subacute paracetamol toxicity.

The effect of aging on mitochondrial and cytosolic hepatic intrinsic death pathway and apoptosis associated proteins in Fischer 344 rats

Apoptosis is increased in the liver in old age and is a common pathological feature of liver disease. The mitochondria play a key role in regulating apoptosis via the intrinsic death pathway. As the effect of aging on this pathway is unclear, we aimed to characterize the impact of aging on the hepatic intrinsic death pathway and apoptosis. Livers from young adult (6.6 ± 0.3 months, n = 9) and old (25.4 ± 0.7 months, n = 9) male Fischer 344 rats were extracted for cellular fractionation and immunobloting. In old age there were lower mitochondrial protein levels of pro-apoptotic BAK, BID, tBID and VDAC1 (p < 0.05) and of anti-apoptotic Bcl-2. Compared to young, old rats had lower cytosolic protein levels of pro-apoptotic BAX, BAK, BID, tBID and anti-apoptotic Bcl-xL (p < 0.05). BAK, Bcl-2 and Bcl-xL were found in the cytosol. Furthermore with old age, cytosolic protein levels of cytochrome C, AIF and cleaved caspase-9 did not change but activation of caspase-3, -6 and -7 increased (p < 0.05) and DNA fragmentation trended to increase. Our results suggest an age-related decline in the levels of a number of proteins involved in the intrinsic death pathway, an uncoupling of intermediate apoptosis signaling and increased cellular apoptosis in the liver in old age.

Classification of high risk basal cell carcinoma subtypes: experience of the ONTRAC study with proposed definitions and guidelines for pathological reporting

Classification of high risk basal cell carcinoma subtypes: experience of the ONTRAC study with proposed definitions and guidelines for pathological reporting CATRIONA A. MCKENZIE, ANDREW C. CHEN, BONITA CHOY, PABLO FERNANDEZ-PENAS, DIONA L. DAMIAN AND RICHARD A. SCOLYER 1Tissue Pathology and Diagnostic Oncology, Royal Prince Alfred Hospital, 2Dermatology and Bosch Institute, The University of Sydney at Royal Prince Alfred Hospital, 3Dermatology, The University of Sydney at Westmead Hospital, 4Melanoma Institute Australia, North Sydney, Sydney, and 5Discipline of Pathology, Sydney Medical School, The University of Sydney, NSW, Australia

A case report of invasive ductal carcinoma of the breast with prominent sebaceous differentiation.

Background Sebaceous carcinoma of the breast is a rare entity, with only nine examples reported previously (WHO, 2012). Sebaceous carcinoma is defined as a breast carcinoma with prominent sebaceous differentiation in no less than 50% of cells (WHO, 2012). Limited data is available regarding the significance of sebaceous differentiation. Aim To present a case of diagnostic interest. Method and results We present a case of a 77-year-old woman with a self-detected left breast mass. After an initial core biopsy she underwent a mastectomy with sentinel lymph node biopsy. His-topathology confirmed an ER-positive/PR-positive/Her2-equiv-ocal invasive ductal carcinoma, with prominent sebaceous differentiation, and negative sentinel nodes. Discussion The amount of sebaceous differentiation falls just short of the greater than 50% required for diagnosis of sebaceous carcinoma. The differential diagnosis includes carcinoma with apocrine differentiation, lipid-rich carcinoma, and liposarcoma. A review of the available literature highlights the scarcity of knowledge of this rare entity. Very little is understood about the clinical course and prognosis of sebaceous carcinoma, and follow up data is limited. There is an association in the cases reported, with high grade tumours and high mitotic counts, and the predominant type of breast cancer associated is ductal. The two cases that have reported metastasis have been late in onset, and have occurred in sites which are not common for breast malignancies (scalp and skin). The significance of sebaceous differentiation is difficult to establish, and further research is warranted.

Metastatic adenoid cystic carcinoma of the liver confirmed with MYB rearrangement

S S135 Background: CD5 is an antigen normally expressed on T-cells but can also be expressed in a small subset of B-cells and some non-Hodgkin B-cell lymphoproliferative disorders such as small lymphocytic lymphoma, marginal zone lymphoma and mantle cell lymphoma. When using a single immunohistochemical stain for each antigen on separate slides, it can be challenging to compare and correlate whether CD5 positivity is being seen in neoplastic B-cells or normal T-cells. This problem is amplified when the CD5 expressing B-cells are present in small numbers, such as in a bone marrow trephine. Aim: In addition to the usual panel of immunohistochemical stains, we describe the application of two T-cell markers in a combined stain, CD5 (red, cytoplasmic) / CD3 (brown, cytoplasmic), to aid in the identification of CD5 co-expressing Bcells in small B-cell lymphoproliferative disorders. Method: In a suspected small B-cell lymphoproliferative disorder, the red CD5 immunohistochemical stain is applied first and is positive in all normal T-cells and CD5-expressing B-cells. The brown CD3 stain is applied second, and also stains all the normal T-cells that express both CD3 and CD5, but does not stain any of the B-cells. As the darker brown stain is applied over the red, all CD3 and CD5 expressing T-cells that have been stained twice only appear brown, whereas the CD5 expressing Bcells appear red. Conclusion: Following the application of these two T-cell markers to a single slide, it can be deduced that all brown staining cells are normal CD3 / CD5 expressing T-cells whereas the red staining cells are CD5-expressing B-cells. 55. NEOPLASIA IN GASTRIC ADENOCARCINOMA AND PROXIMAL POLYPOSIS SYNDROME (GAPPS): GASTRIC RATHER THAN INTESTINAL PHENOTYPE Priyanthi Kumarasinghe, W. Bastiaan de Boer, M. Hooi Ee