Molecular Classification Of Tumors Research Articles

Genetic Test, Risk Prediction, and Counseling.

Advancement in technology has nurtured the new era of genetic tests for personalized medicine. In this chapter, we will introduce the current development, challenges, and the outlook of genetic test, disease risk prediction, and genetic counseling. In the first section, we will present the success cases in the areas of molecular classification of tumors, pharmacogenomics, and Mendelian disorders, and the challenges of genetic tests implementations. In the second section, common methods for genetic risk prediction models and evaluation measures will be introduced, as well as challenges in feature reliability, risk model stability, and clinical utility. In the final section, key components of genetic counseling will be introduced, covering individual communications, psychosocial concerns, risk assessments, and follow-ups. Current evidences have shown a promising future for genetic testing and risk prediction; we expect that the advancement of analytical methods, technology, integration of omics data, and the increasing clinical implementation and regulation will continue to pave the way for precision medicine in future.

Next-generation sequencing in thyroid cancer.

Next-generation sequencing (NGS) in thyroid cancer allows for simultaneous high-throughput sequencing analysis of variable genetic alterations and provides a comprehensive understanding of tumor biology. In thyroid cancer, NGS offers diagnostic improvements for fine needle aspiration (FNA) cytology of thyroid with indeterminate features. It also contributes to patient management, providing risk stratification of patients based on the risk of malignancy. Furthermore, NGS has been adopted in cancer research. It is used in molecular tumor classification, and molecular prediction of recurrence and metastasis in papillary thyroid carcinoma. This review covers previous NGS analyses in variable types of thyroid cancer, where samples including FNA cytology, fresh frozen tissue, and formalin-fixed, paraffin-embedded tissues were used. This review also focuses on the clinical and research implications of using NGS to study and treat thyroid cancer.

Expression and methylation patterns partition luminal-A breast tumors into distinct prognostic subgroups.

BackgroundBreast cancer is a heterogeneous disease comprising several biologically different types, exhibiting diverse responses to treatment. In the past years, gene expression profiling has led to definition of several “intrinsic subtypes” of breast cancer (basal-like, HER2-enriched, luminal-A, luminal-B and normal-like), and microarray based predictors such as PAM50 have been developed. Despite their advantage over traditional histopathological classification, precise identification of breast cancer subtypes, especially within the largest and highly variable luminal-A class, remains a challenge. In this study, we revisited the molecular classification of breast tumors using both expression and methylation data obtained from The Cancer Genome Atlas (TCGA).MethodsUnsupervised clustering was applied on 1148 and 679 breast cancer samples using RNA-Seq and DNA methylation data, respectively. Clusters were evaluated using clinical information and by comparison to PAM50 subtypes. Differentially expressed genes and differentially methylated CpGs were tested for enrichment using various annotation sets. Survival analysis was conducted on the identified clusters using the log-rank test and Cox proportional hazards model.ResultsThe clusters in both expression and methylation datasets had only moderate agreement with PAM50 calls, while our partitioning of the luminal samples had better five-year prognostic value than the luminal-A/luminal-B assignment as called by PAM50. Our analysis partitioned the expression profiles of the luminal-A samples into two biologically distinct subgroups exhibiting differential expression of immune-related genes, with one subgroup carrying significantly higher risk for five-year recurrence. Analysis of the luminal-A samples using methylation data identified a cluster of patients with poorer survival, characterized by distinct hyper-methylation of developmental genes. Cox multivariate survival analysis confirmed the prognostic significance of the two partitions after adjustment for commonly used factors such as age and pathological stage.ConclusionsModern genomic datasets reveal large heterogeneity among luminal breast tumors. Our analysis of these data provides two prognostic gene sets that dissect and explain tumor variability within the luminal-A subgroup, thus, contributing to the advancement of subtype-specific diagnosis and treatment.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-016-0724-2) contains supplementary material, which is available to authorized users.

Application of carbon nanoparticles followed by microscale methylthioninum chloride in sentinel lymph node biopsy for breast cancer

Objective To explore the application value of Carbon Nanoparticles followed by microscale Methylthioninum Chloride in sentinel lymph node biopsy for breast cancer. Methods Two hundred and eight breast cancer patients were selected in the First Affiliated Hospital of Zhengzhou University during recent two years who performed sentinel lymph node biopsy .They were randomly divided into five groups according to different lymphatic tracer injected: Sequential group (A group, 128 patients), Carbon Nanoparticles group (B group, 44 patients) and Methylthioninum Chloride group (C group, 36 patients). According to different Methylthioninum Chloride doses, sequential groups were randomly divided into low dose group (A1 group, 48 patients), middle dose group (A2 group, 42 patients), high dose group (A3 group, 38 patients), If sentinel lymph nodes were negative no longer performed axillary lymph nodes dissection. Receive the five groups statistics of detection rate, the condition of lymphatic tracer, the incidence of local skin irritation and so on. Results (1)The detection rate of A, B and C three groups had no obvious difference.The incidence of lymphatic tracer dyed not ideal in B group was higher than A and B two groups, the incidence of local skin stimulation in C group was higher than A and B two groups.(2)A1, A2, A3, three groups had no difference of the detection rate and lymphatic tracer, but the incidence of local skin stimulation in A3 group is higher than A1, A2 two groups. 3. 5 groups of SLN detection rate had no relationship with patients′ age, BMI, tumor size, tumor location, tumor molecular classification factors (P>0.05). Conclusions Carbon Nanoparticles followed by microscale Methylthioninum Chloride detection rate was similar with two separate application, but can reduce the lymphatic vessels not developing situation when used Carbon Nanoparticles alone and reduce the local irritation caused by large dose of Methylthioninum Chloride , so it can be a good choice of SLNB in clinical. Key words: Breast neoplasms; Sentinel lymph node biopsy; Carbon Nanoparticles suspension; Methylthioninum Chloride injection

Important molecular genetic markers of colorectal cancer.

Colorectal cancer (CRC) ranks third in the incidences of cancer morbidity and mortality worldwide. CRC is rather heterogeneous with regard to molecular genetic characteristics and pathogenic pathways. A wide spectrum of biomarkers is used for molecular subtype determination, prognosis, and estimation of sensitivity to different drugs in practice. These biomarkers can include germline and somatic mutations, chromosomal aberrations, genomic abnormalities, gene expression alterations at mRNA or protein level and changes in DNA methylation status. In the present review we discuss the most important and well-studied CRC biomarkers, and their potential clinical significance and current approaches to molecular classification of colorectal tumors.

The Influence of the Global Gene Expression Shift on Downstream Analyses.

The assumption that total abundance of RNAs in a cell is roughly the same in different cells is underlying most studies based on gene expression analyses. But experiments have shown that changes in the expression of some master regulators such as c-MYC can cause global shift in the expression of almost all genes in some cell types like cancers. Such shift will violate this assumption and can cause wrong or biased conclusions for standard data analysis practices, such as detection of differentially expressed (DE) genes and molecular classification of tumors based on gene expression. Most existing gene expression data were generated without considering this possibility, and are therefore at the risk of having produced unreliable results if such global shift effect exists in the data. To evaluate this risk, we conducted a systematic study on the possible influence of the global gene expression shift effect on differential expression analysis and on molecular classification analysis. We collected data with known global shift effect and also generated data to simulate different situations of the effect based on a wide collection of real gene expression data, and conducted comparative studies on representative existing methods. We observed that some DE analysis methods are more tolerant to the global shift while others are very sensitive to it. Classification accuracy is not sensitive to the shift and actually can benefit from it, but genes selected for the classification can be greatly affected.

ENDOCRINE TUMOURS: The genomics of adrenocortical tumors.

The last decade witnessed the emergence of genomics, a set of high-throughput molecular measurements in biological samples. These pan-genomic and agnostic approaches have revolutionized the molecular biology and genetics of malignant and benign tumors. These techniques have been applied successfully to adrenocortical tumors. Exome sequencing identified new major drivers in all tumor types, including KCNJ5, ATP1A1, ATP2B3 and CACNA1D mutations in aldosterone-producing adenomas (APA), PRKACA mutations in cortisol-producing adenomas (CPA), ARMC5 mutations in primary bilateral macronodular adrenocortical hyperplasia (PBMAH) and ZNRF3 mutations in adrenocortical carcinomas (ACC). Moreover, the various genomic approaches - including exome sequencing, transcriptome, miRNome, genome and methylome - converge into a single molecular classification of adrenocortical tumors. Especially for ACC, two main molecular groups have emerged, showing major differences in outcomes. These ACC groups differ by their gene expression profiles, but also by recurrent mutations and specific DNA hypermethylation patterns in the subgroup of poor outcome. The clinical impact of these findings is just starting. The main altered signaling pathways now become therapeutic targets. The molecular groups of diseases individualize robust subtypes within diseases such as APA, CPA, PBMAH and ACC. A revised nosology of adrenocortical tumors should impact the clinical research. Obvious consequences also include genetic counseling for the new genetic diseases such as ARMC5 mutations in PBMAH, and a better prognostication of ACC based on targeted measurements of a few discriminant molecular alterations. Identifying the main molecular groups of adrenocortical tumors by extensively gathering the molecular variations is a significant step forward towards precision medicine.

Single-cell approaches for molecular classification of endocrine tumors.

In this review, we summarize recent developments in single-cell technologies that can be employed for the functional and molecular classification of endocrine cells in normal and neoplastic tissue. The emergence of new platforms for the isolation, analysis, and dynamic assessment of individual cell identity and reactive behavior enables experimental deconstruction of intratumoral heterogeneity and other contexts where variability in cell signaling and biochemical responsiveness inform biological function and clinical presentation. These tools are particularly appropriate for examining and classifying endocrine neoplasias, as the clinical sequelae of these tumors are often driven by disrupted hormonal responsiveness secondary to compromised cell signaling. Single-cell methods allow for multidimensional experimental designs incorporating both spatial and temporal parameters with the capacity to probe dynamic cell signaling behaviors and kinetic response patterns dependent upon sequential agonist challenge. Intratumoral heterogeneity in the provenance, composition, and biological activity of different forms of endocrine neoplasia presents a significant challenge for prognostic assessment. Single-cell technologies provide an array of powerful new approaches uniquely well suited for dissecting complex endocrine tumors. Studies examining the relationship between clinical behavior and tumor compositional variations in cellular activity are now possible, providing new opportunities to deconstruct the underlying mechanisms of endocrine neoplasia.

Advancement in phase-transitional ultrasound molecular probe

With the rapid development of molecular imaging technology and materials science, it is very possible for the medical imaging techniques to enable the real-time and multi-modality visualization, characterization, and quantification of biologic processes taking place at the organ, tissue, cellular and molecular levels within intact living subjects. Via the comprehensive progress in terms of tumor early diagnosis based on molecular targets, a revolutionary tumor molecular classification, tumor boundary determination, and surgical navigation, the precision medicine will be achieved. Ultrasound (US) molecular imaging, which is a new branch of molecular imaging, has developed rapidly in the latest decades. The phase-transitional molecular probe has shown significantly more superiority than the traditional US contrast agents owing to its good penetration, stability and the ability for theranostical integration, which indicates a better potential in the basic researches and the clinical applications. In this paper, we reviewed the progress in the studies of the phase-transitional ultrasound molecular imaging probe and its applications in biomedicine.

IMP2/p62 induces genomic instability and an aggressive hepatocellular carcinoma phenotype.

Hepatocellular carcinoma (HCC) represents the third leading cause of cancer-related deaths and commonly develops in inflammatory environments. The IGF2 mRNA-binding protein IMP2-2/IGF2BP2-2/p62 was originally identified as an autoantigen in HCC. Aim of this study was to investigate a potential pathophysiological role of p62 in hepatocarcinogenesis. Human HCC tissue showed overexpression of IMP2, which strongly correlated with the fetal markers AFP and DLK1/Pref-1/FA-1 and was particularly elevated in tumors with stem-like features and hypervascularization. Molecular classification of IMP2-overexpressing tumors revealed an aggressive phenotype. Livers of mice overexpressing the IMP2 splice variant p62 highly expressed the stem cell marker DLK1 and secreted DLK1 into the blood. p62 was oncogenic: diethylnitrosamine (DEN)-treated p62 transgenic mice exhibited a higher tumor incidence and multiplicity than wild types. Tumors of transgenics showed a more aggressive and stem-like phenotype and displayed more oncogenic chromosomal aberrations determined with aCGH analysis. DEN-treated p62 transgenic mice exhibited distinct signs of inflammation, such as inflammatory cytokine expression and oxidative stress markers, that is, thiobarbituric acid-reactive substance (TBARS) levels. Reactive oxygen species (ROS) production was elevated in HepG2 cells, which either overexpressed p62 or were treated with DLK1. p62 induced this ROS production by a DLK1-dependent induction and activation of the small Rho-GTPase RAC1, activating NADPH oxidase and being overexpressed in human HCC. Our data indicate that p62/IMP2 promotes hepatocarcinogenesis by an amplification of inflammation.

Abstract B21: Intratumoral hormone receptor heterogeneity is a characteristic of high-grade breast cancer and is associated with inter-laboratory hormone receptor status discordance

Abstract Introduction: Intratumoral hormone receptor (HR) heterogeneity may have implications for molecular classification of breast tumors which, in turn, may impact clinical decision making and epidemiologic research. Prior studies show that 10-20% of patients have discordant estrogen receptor (ER) or progesterone receptor (PR) status upon repeat assay. While technical factors contribute to this discordance, intratumoral HR heterogeneity may also play a role. We examined the association between intratumoral HR heterogeneity, measured at a single research laboratory, and clinical HR status, tumor characteristics and demographic variables. Methods: We used paraffin-embedded blocks from 932 invasive breast cancer cases, of which 497 were African-American, from the population-based Carolina Breast Cancer Study Phase 3, part of the African-American Breast Cancer Epidemiology and Risk (AMBER) Consortium. Two to four 1mm tumor cores per case were assembled into tissue microarrays and stained for ER and PR at a single research laboratory. ER and PR status (positive: ≥1% positive tumor cells; negative: <1%) were assigned to each core by automated digital image analysis and ER and PR status were assigned to each case by core-to-case collapsing, using the weighted average of positive cells across all cores. Intratumoral heterogeneity status (i.e. ‘homogeneous’ vs. ‘heterogeneous’) was assigned based on ‘identical’ or ‘different’ HR status between cores from a single case. Multivariable logistic regression was used to evaluate the association between heterogeneity status and patient and tumor characteristics, overall and stratified by menopausal status. Clinical HR status (positive vs. negative) was abstracted from medical records and concordance between clinical and research ER and PR status were examined, overall and stratified by heterogeneity status. Results: Of 932 cases, 95 (10%) had ER heterogeneity while 149 (16%) had PR heterogeneity. Overall, there was high agreement between clinical and research ER status (93% concordance; 95% confidence interval (CI) 92-95%) and PR status (89%; 95%CI 87-91%), consistent with previously reported inter-laboratory concordance rates. However, agreement was lower in cases with intratumoral heterogeneity (ER: 74%; 95%CI 65-83% and PR: 61%; 95%CI 53-69%), relative to homogeneous cases (ER: 96%; 95%CI 95-97% and PR: 94%; 95%CI 92-96%). Obesity, defined as BMI≥30kg/m2, was associated with increased odds of ER heterogeneity (adjusted odds ratio (OR)obese vs. normal weight 1.76; 95%CI 0.95-3.29), with a stronger association among postmenopausal women (OR 2.52; 95%CI 1.14-5.56). High tumor grade was associated with increased odds of both ER and PR heterogeneity (ORgrade III vs. I 3.14; 95%CI 1.47-6.70 and ORgrade III vs. I 2.28; 95%CI 1.26-4.14, respectively). There were no associations between intratumoral heterogeneity and age, race or menopausal status. Conclusions: Intratumoral heterogeneity may account for a portion of the well-documented inter-laboratory discordance in ER and PR status, with implications for tumor subtype classification. Intratumoral heterogeneity was more prevalent in high grade tumors, indicating that it may be a feature of aggressive breast cancer. Furthermore, obese postmenopausal women were more likely to have intratumoral ER heterogeneity, suggesting a potential mechanism underlying the association between obesity and worse breast cancer prognosis. Although these data do not support an association between intratumoral heterogeneity and race, given that both high grade disease and obesity are more common in African Americans, future studies should explore a possible role for intratumoral heterogeneity in breast cancer racial disparities. Citation Format: Emma H. Allott, Joseph Geradts, Xuezheng Sun, Stephanie M. Cohen, Thaer Khoury, Wiam Bshara, Gary Zirpoli, Julie R. Palmer, Christine B. Ambrosone, Andrew F. Olshan, Melissa A. Troester. Intratumoral hormone receptor heterogeneity is a characteristic of high-grade breast cancer and is associated with inter-laboratory hormone receptor status discordance. [abstract]. In: Proceedings of the Seventh AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 9-12, 2014; San Antonio, TX. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2015;24(10 Suppl):Abstract nr B21.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Molecular dissection of ependymomas.

Molecular dissection of ependymomas.

Current and future molecular profiling of cancer by next-generation sequencing.

Advances in the new sequencing technologies have enabled us to explore global genetic alterations including driver genes in a wide range of cancers. Concordantly, successes of molecular target therapy promoted the validity of tumor classification based on the combination of targetable genetic abnormalities, and next-generation sequencing-based genetic profiling using target gene capturing or multiplex-polymerase chain reaction has already been tested or adapted in many cancer centers. Driver gene-based classification may be applicable beyond organs, and clinical trials incorporating this genomic information, called as a basket trial, have been executed, although it should be considered that similar therapeutic efficacies against driver mutations are not invariably maintained among different cancer types. Research efforts to identify still missing driver genes in rare cancers, to complete functional annotation of infrequent driver genes, and multiple-layered omics approaches are further expected for better classification of tumor. Emerging clinical interests in the development of immunotherapies postulate a new molecular classification of tumors. Recent studies reported that total number of mutations and the frequent appearance of neo-antigens by protein-coding mutations were associated with a better response, and genetic evaluation of both tumor and host immune system by sequencing is expected to contribute to the personalized immunotherapies in the near future. Lastly intratumoral molecular heterogeneity challenges the current 'static' molecular classification of tumor. For example, dynamic change in clonal constitution within tumor plays an important role in acquired drug resistance. It has been extensively explored whether liquid biopsy-based molecular profiling can resolve currently confronting difficulties.

Pharmacists in Australian general practice: an opportunity for expertise in precision medicine.

Pharmacists in Australian general practice: an opportunity for expertise in precision medicine.

Advances in Radiation Therapy in Pediatric Neuro-oncology.

Radiation therapy remains a highly effective therapy for many pediatric central nervous system tumors. With more children achieving long-term survival after treatment for brain tumors, late-effects of radiation have become an important concern. In response to this problem, treatment protocols for a variety of pediatric central nervous system tumors have evolved to reduce radiation fields and doses when possible. Recent advances in radiation technology such as image guidance and proton therapy have led to a new era of precision treatment with significantly less exposure to healthy tissues. These developments along with the promise of molecular classification of tumors and targeted therapies point to an optimistic future for pediatric neuro-oncology.

Abstract 3704: Demographics of colorectal cancer patients vary by aspirin use

Abstract Purpose: Regular NSAID (Non-Steroidal Anti-Inflammatory Drug) use results in an overall reduction in colorectal cancer (CRC) risk; however, this reduction may vary among molecularly defined subsets of CRC. Characterization of patients by regular NSAID use and molecular subtype will provide unique insight into the biology of NSAID-based CRC prevention. Methods: A sample of patients was selected from the Assessment of Targeted Therapies Against Colorectal Cancer (ATTACC) protocol, which is designed to molecularly profile tumors of patients with refractory metastatic CRC. Demographic data and NSAID use were collected from a risk-factor survey. Demographic characteristics were compared between 106 non-NSAID users, 75 regular aspirin users and 78 regular non-aspirin NSAID users. Chi-square analysis was used to compare categorical variables and ANOVA was used for continuous variables. Logistic regression was used to assess predictors of aspirin or non-aspirin NSAID use compared to non-use. Results: Preliminary results indicate that aspirin users tend to be older, male and more frequently engage in vigorous physical activity compared to non-users and users of non-aspirin NSAIDs (Table I). In univariate analyses, patients who are older or engage in more frequent vigorous physical activity are at increased odds of using aspirin. When controlling for other variables, only age at diagnosis predicts aspirin use. Table 1.Description and AssociationsNo NSAID useAspirin useNon-Aspirin usep valueN1067578Age at Stage IV Diagnosis, Mean (SE)55.2 (1.0)59.5 (1.1)53.1 (1.3)0.0006Gender, N (%)11884810.0251Male72 (61.0)59 (72.8)44 (52.4)Female46 (39.0)22 (27.2)40 (47.6)Vigorous Physical Activity, N (%)11780840.0214Little or none50 (42.7)18 (22.5)34 (40.5)Once/week10 (8.6)14 (17.5)13 (15.5)Twice/week or more57 (48.7)48 (60.0)37 (44.1)Logistic RegressionUnadjusted Odds of Aspirin UseAdjusted Odds of Aspirin Use*OR (95% CI)p valueOR (95% CI)p valueAge at Stage IV Diagnosis1.047 (1.013, 1.081)0.00571.049 (1.014, 1.085)0.0055Gender (Female)0.584 (0.316, 1.078)0.08550.701 (0.356, 1.379)0.3032Vigorous Physical Activity (overall effect)0.00850.0486Little or none (Ref)1.001.00Once/week3.889 (1.468, 10.300)0.03692.481 (0.892, 6.906)0.2997Twice/week or more2.339 (1.207, 4.533)0.58952.305 (1.137, 4.673)0.2662*Each variable is adjusted for the other variables in the table Conclusions: These preliminary data indicate that we will be able to distinguish aspirin users from non-NSAID users in a sample of late-stage CRC patients. Going forward, we will combine these demographic data with tumor molecular classification to determine the subtypes of CRC that are more or less prevalent in aspirin/NSAID users. Use of NSAIDs and overall survival will also be evaluated among different molecular subtypes of CRC. This data, combined with analysis of risk factors for each molecular subtype will provide unique insight into the biology of NSAID-based CRC prevention. Citation Format: Jennifer S. Davis, Shailesh Advani, Emilyn Banfield, Michael Overman, Zhi-Qin Jiang, Shanequa Manuel, Carrie Daniel, Shine Chang, Scott Kopetz. Demographics of colorectal cancer patients vary by aspirin use. [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 3704. doi:10.1158/1538-7445.AM2015-3704

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Molecular Classification of Ependymal Tumors across All CNS Compartments, Histopathological Grades, and Age Groups

Ependymal tumors across age groups are currently classified and graded solely by histopathology. It is, however, commonly accepted that this classification scheme has limited clinical utility based on its lack of reproducibility in predicting patients' outcome. We aimed at establishing a uniform molecular classification using DNA methylation profiling. Nine molecular subgroups were identified in a large cohort of 500 tumors, 3 in each anatomical compartment of the CNS, spine, posterior fossa, supratentorial. Two supratentorial subgroups are characterized by prototypic fusion genes involving RELA and YAP1, respectively. Regarding clinical associations, the molecular classification proposed herein outperforms the current histopathological classification and thus might serve as a basis for the next World Health Organization classification of CNS tumors.

Validation of Tissue Microarray for Molecular Profiling of Canine and Feline Mammary Tumours

Tissue microarray (TMA) is a high-throughput method adopted for simultaneous molecular profiling of tissue samples from large patient cohorts. The aim of this study was to validate the TMA method for the molecular classification of canine and feline mammary tumours. Twelve samples, five feline and five canine mammary tumours and two canine haemangiosarcomas, were collected. TMA construction was based on Kononen's method of extracting a cylindrical core of paraffin wax-embedded 'donor' tissue and inserting it into a 'recipient' wax block. Seven consecutive sections from each tissue array block were subjected to immunohistochemistry (IHC) using primary antibodies specific for oestrogen receptor (OR), progesterone receptor (PR), c-erbB-2, cytokeratin (CK) 5/6, CK14, CK19 and p63. The same panel of antibodies was applied to the full sections from all cases. Comparison between full sections and TMA scores revealed different results depending on the antibodies. Labelling for OR, PR, CK19 and p63 showed total concordance, c-erbB2 (score +2, +3) was concordant in nine out of ten cases, CK5/6 and CK14 in eight out of ten cases. The TMA platform preserves the molecular profile of canine and feline mammary tumour markers, representing a useful tool for rapid and cost-effective analysis for the first phenotypic screening using OR, PR and c-erbB2 antibodies. Basal cytokeratin, used for triple negative identification, shows a multifocal 'niche' expression pattern, for which IHC of the full section or multiple core array is recommended.