Helen Ingoldsby

Prediction of Oncotype DX and TAILORx risk categories using histopathological and immunohistochemical markers by classification and regression tree (CART) analysis.

Oncotype DX is an RT-PCR assay used to predict which patients with ER-positive node-negative (NN) disease will benefit from chemotherapy. Each patient is stratified into a risk category based on a recurrence score (RS) and the TAILORx trial is determining the benefit of chemotherapy for patients with mid-range RSs. We tested if Oncotype DX and TAILORx risk categories could be predicted by standard pathological features and protein markers corresponding to 10 genes in the assay (ER, PR, Ki67, HER2, BCL2, CD68, Aurora A kinase, survivin, cyclin B1 and BAG1) on 52 patients who enrolled on TAILORx. Immunohistochemistry for the protein markers was performed on whole tissue sections. Classification and regression tree (CART) analysis correctly classified 69% of cases into Oncotype DX risk categories based on the expression of PR, survivin and nuclear pleomorphism. All tumours with PR staining (Allred score ≥ 2) and marked nuclear pleomorphism were in the high-risk category. No case with PR <2, low survivin (≤ 15.5%) and nuclear pleomorphism <3 was high-risk. Similarly, 77% of cases were correctly classified into TAILORx categories based on nuclear pleomorphism, survivin, BAG1 and cyclin B1. Ki67 was the only variable that predicted the absolute RS with a cut-off for positivity of 15% (p = 0.003). In conclusion, CART revealed key predictors including proliferation markers, PR and nuclear pleomorphism that correctly classified over two thirds of ER-positive NN cancers into Oncotype DX and TAILORx risk categories. These variables could be used as an alternative to the RT-PCR assay to reduce the number of patients requiring Oncotype DX testing.

Alcohol and unnatural deaths in the West of Ireland: a 5-year review

Aim To investigate the prevalence of alcohol in unnatural deaths in the West of Ireland between 2003 and 2007. Methods The reports of 1669 postmortem examinations carried out at Galway University Hospitals were reviewed; 379 non-homicidal unnatural deaths were eligible for the study. Alcohol levels were measured in blood and/or urine in 311 cases. For each case, gender, age, cause of death and toxicology results were recorded. Results Alcohol was detected in 162 out of 311 cases (52%); 133 (82%) cases were men and 29 (18%) were women. Alcohol levels >150 mg/100 ml were found in 99 cases (61%), most commonly in 18–49-year-olds (n=74; 75%). Road traffic crashes (RTCs) (n=38; 23%), drownings (n=38; 23%) and hangings (n=25; 15%) were common unnatural deaths associated with alcohol. The majority of RTC deaths involved the driver (n=27; 71%). The alcohol level was higher than the legal driving limit of 80 mg/100 ml in 82% (n=22) and >150 mg/100 ml in 59% (n=16) of these. Mortality of passengers (n=6; 16%) and pedestrians (n=5; 13%) was less common. Conclusions Alcohol remains a major contributor to unnatural deaths in the West of Ireland, particularly with respect to mortality in young people. Young men are especially vulnerable. Deaths in RTCs and by drowning and hanging are commonly associated with alcohol. Many driver fatalities involve alcohol levels far above legal limits. Alcohol measurement in all unnatural deaths would facilitate more accurate determination of its role.

Prognostic Significance of Deregulated Dicer Expression in Breast Cancer

Background Dicer, an RNase III-type endonuclease, is the key enzyme involved in RNA interference and microRNA pathways. Aberrant expression of Dicer is reported in several human cancers. Our aim was to assess the prognostic role of Dicer in breast cancer. Methods The entire series comprised 666 invasive breast cancers (IBCs), 480 DCIS cases (397 associated with IBC and 83 pure DCIS) and 305 lymph node metastases. Cytoplasmic Dicer expression by immunohistochemistry was scored as negative (no staining) and positive (weak, moderate or strong staining). Results Dicer staining was assessable in 446 IBC, 128 DCIS and 101 lymph node metastases. Expression of Dicer was observed in 33% (145/446) of IBCs, 34% (44/128) of DCIS and 57% (58/101) of lymph node metastases. Dicer expression was increased in nodal metastases compared to primary tumours (p<0.001); and was associated with ER negativity (p<0.001), HER2 positivity (p<0.001), high Ki67 labeling index (p<0.001) and expression of basal-like biomarkers (p = 0.002). Dicer positivity was more frequent in the HER2 overexpressing (p<0.001) and basal-like (p = 0.002) subtypes compared to luminal A subtype. Dicer expression was associated with reduced overall survival (OS) on univariate analysis (p = 0.058) and remained an independent predictor of OS on multivariate analysis (HR 2.84, 95% CI 1.43–5.62, p = 0.003), with nodal status (HR 2.61, 95% CI 1.18–5.80, p = 0.018) and PR (HR 0.28, 95% CI 0.13–0.59, p = 0.001). Further, moderate or strong expression of Dicer was associated with improved disease-free survival in the HER2-overexpressing subtype compared to negative or weak expression (p = 0.038). Conclusion Deregulated Dicer expression is associated with aggressive tumour characteristics and is an independent prognostic factor for OS. Our findings suggest that Dicer is an important prognostic marker in breast cancer and that its prognostic role may be subtype specific.

RET protein expression in papillary renal cell carcinoma

OBJECTIVE To examine the role of RET in renal malignancy, in particular papillary renal cell carcinoma (RCC). MATERIALS AND METHODS A cohort of 111 archival renal samples was used consisting of 94 renal cancers (66 papillary RCC, 18 conventional clear cell carcinoma, 10 chromophobe RCC), 4 benign oncocytomas, and 13 normal kidney tissues. RET protein expression was examined by immunohistochemistry and expression levels were correlated with clinicopathologic and patient survival data. RESULTS Positive RET staining was seen in 34/66 (52%) papillary RCCs, 4/10 (40%) chromophobe carcinomas, 4/4 (100%) oncocytomas, and 11/13 (85%) normal kidney samples. All 18 cases of conventional clear cell carcinoma had negative RET staining. RET expression was associated with low Fuhrman nuclear grade. CONCLUSIONS RET protein may be contributing in part to an adaptation of a papillary growth pattern in certain renal malignancies. Given the possible therapeutic benefit of small molecule inhibitors of RET activation, further work needs to be done to highlight the functional relevance of RET protein expression in papillary RCC.

Abstract P6-04-01: Engineering mesenchymal stem cells(MSCs) to secrete tumour-suppressing exosomes for breast cancer therapy

Introduction: Mesenchymal Stem Cells(MSCs) are multipotent stromal cells that are known to engraft into tumours, raising their potential as tumour-targeted delivery vehicles. MSCs secrete tiny vesicles known as exosomes, which contain genetic material including microRNAs, and are effectively taken up by recipient cells. This study aimed to characterise a tumour-suppressing microRNA, miR-379, and engineer MSCs to secrete exosomes enriched with the microRNA. Methods: The mechanism of action of miR-379 In Vivo was determined through lentivirus-mediated upregulation of miR-379 in breast tumours, and analysis of changes in tumour angiogenesis, proliferation and progression. Subsequently, MSCs were engineered with lenti-379 and any impact on MSC migration, proliferation and morphology was assessed. MSC-secreted exosomes were isolated and characterised using Transmission Electron Microscopy(TEM) and Western Blot. The exosomal microRNA content was analysed by RQ-PCR, and transfer between cell populations visualised using confocal microscopy. Results: While elevated miR-379 expression did not impact tumour size In Vivo, an increase in tumour necrosis and decrease in invasion was observed. MSCs were successfully transduced with miR-379, resulting in a distinct change in cell morphology. Despite this, MSC-379 cells maintained inherent tumour-targeted migratory capacity, with no impact on proliferation observed. MSC-379 derived exosomes were 30-120nm in size and expressed the exosome-associated protein CD63. A 5-fold increase in miR-379 was observed in engineered exosomes. Successful transfer of RFP-labelled MSC-derived exosomes to breast cancer cells was visualised using confocal microscopy. Conclusion: Engineering tumour-targeted MSCs to secrete exosomes enriched with miR-379 holds exciting potential as a novel therapy for breast cancer. Citation Format: O9Brien KP, Khan S, Thompson K, Joyce D, Lalor P, Dockery P, Ingoldsby H, Kerin MJ, Dwyer RM. Engineering mesenchymal stem cells(MSCs) to secrete tumour-suppressing exosomes for breast cancer therapy [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-04-01.

Abstract 1708: Identification of triple negative breast cancer (TNBC) subtypes by an immunohistochemistry (IHC) panel with impact on clinical outcomes

Background TNBC comprises 10-20% of breast cancers. Lehmann et al. identified 6 TNBC subtypes by gene expression profiling: BL1, BL2, IM, M, MSL & LAR. Despite the diversity of TNBC, standard of care is combination chemotherapy as in non TNBCs. Identification of chemosensitive TNBC subtypes is necessary. Chemoresistant TNBCs need alternative targets to improve treatment strategies. Aims 1. To validate an IHC biomarker panel to define molecular subtypes of TNBC 2. To correlate molecular subtypes with prognosis to identify appropriate therapy 3. To improve diagnostic tools to individualize therapy based on TNBC subtypes Methods A TMA was constructed of 197 TNBCs diagnosed from 1999 - 2014. An 8-protein IHC panel was developed to identify TNBC subtypes on FFPE tissue. The panel includes markers for key pathways to discriminate between 6 subtypes: AR, Bcl2, c-myc, TIE1, PDGFC, MMP2, Il2R, MSH2. To date, AR & Bcl2 have been stained and assessed, together with p53 & Ki67. 10% was used as the cut off for positivity. Clinical data were obtained from hospital records and incorporated in the database. Results On initial observation, AR+ tumors had an older age at diagnosis than AR- (60 v 56), lower rates of family history (36 v 87%) and longer DFS (31 v 21 months). Bcl2+ tumors had a younger age at diagnosis than Bcl2- (55 v 59), lower recurrence rates (25 v 31%) and longer DFS (33 v 15 months). High Ki67 tumors had a younger age at diagnosis than low Ki67 (56 v 65), higher rates of family history (30 v 14%), lower recurrence rates (16 v 25%) and longer DFS (30 v 2 months). By the time of presentation, the entire panel of 8 proteins will be analyzed. The clinicopathological association of specific TNBC subtypes and the impact of TNBC subtype on chemotherapy response will be statistically assessed. Analysis will include response, duration, DFS and OS. This study will ultimately correlate TNBC molecular subtypes with prognosis to aid clinical decision making, individualize therapies and improve patient outcomes. Citation Format: Elaine M. Walsh, Aliaa Shalaby, Laura Murillo, Mark Webber, Michael Kerin, Sharon Glynn, Grace Callaghy, Helen Ingoldsby, Maccon Keane. Identification of triple negative breast cancer (TNBC) subtypes by an immunohistochemistry (IHC) panel with impact on clinical outcomes. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1708. doi:10.1158/1538-7445.AM2015-1708

Through the looking glass: primary epithelioid angiosarcoma in the left atrium, a unique site for a rare malignancy.

Malignant cardiac tumours occurring on the left side are vanishingly rare entities. We describe a case of a 73‐year‐old male who underwent surgery for a left‐sided cardiac tumour following initial presentation with transient ischaemic attacks. In addition to the unusual presentation and subsequent metastatic pattern to the femur, the tumours pathological diagnosis was that of an epithelioid variant of an angiosarcoma which has not been previously described in this anatomical location.