Copyright © 2019 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. 474 Original article 0960-8931 Copyright © 2019 Wolters Kluwer Health, Inc. All rights reserved. DOI: 10.1097/CMR.0000000000000594 Nuclear morphometric analysis in tissue as an objective tool with potential use to improve melanoma staging Tatiana W.N. Nunes a, *, Eduardo C. Filippi-Chiela b, *, Sídia M. Callegari-Jacques c , Vinicius D. da Silva g , Tatiana Sansonowicz d , Guido Lenz e and Adriana V. Roehe f Alterations in nuclear size and shape are commonly observed in cancers, and its objective evaluation may provide valuable clinical information about the outcome of the disease. Here, we applied the nuclear morphometric analysis in tissues in hematoxylin and eosin-digitized slides of nevi and melanoma, to objectively contribute to the prognostic evaluation of these tumors. To this, we analyzed the nuclear morphometry of 34 melanomas classified according to the TNM stage. Eight cases of melanocytic nevi were used as non-neoplastic tissues to set the non-neoplastic parameters of nuclear morphology. Our samples were set as G1 (control, nevi), G2 (T1T2N0M0), G3 (T3T4N0M0), G4 (T1T2N1M1), and G5 (T3T4N1M1). Image-Pro Plus 6.0 software was used to acquire measurements related to nuclear size (variable: Area) and shape (variables: Aspect, AreaBox, Roundness, and RadiusRatio, which were used to generate the Nuclear Irregularity Index). From these primary variables, a set of secondary variables were generated. All the seven primary and secondary variables related to the nuclear area were different among groups (Pillai’s trace P<0.001), whereas Nuclear Irregularity Index, which is the variable related to nuclear shape, did not differ among groups. The secondary variable ‘Average Area of Large Nuclei’ was able to differ all pairwise comparisons, including thin nonmetastatic from thin metastatic tumors. In conclusion, the objective quantification of nuclear area in hematoxylin and eosin slides may provide objective information about the risk stratification of these tumors and has the potential to be used as an additional method in clinical decision making. Melanoma Res 29:474–482 Copyright © 2019 Wolters Kluwer Health, Inc. All rights reserved. Melanoma Research 2019, 29:474–482 Keywords: computer assisted, experimental, image analysis, melanoma, neoplasm staging, prognosis a Laboratory of Pathology of the Hospital Nossa Senhora da Conceição de Porto Alegre, b Department of Morphological Sciences, Federal University of Rio Grande do Sul, c Department of Statistics, Institute of Mathematics and Statistics of the Federal University of Rio Grande do Sul, d Program of Medical Residency and Pathology of Hospital Nossa Senhora da Conceição de Porto Alegre, e Department of Biophysics and Biotechnology Center, Federal University of Rio Grande do Sul, f Department of Pathology, Federal University of Health Sciences of Porto Alegre, Porto Alegre and g Department of Pathology, Barretos Cancer Hospital, Barretos, Brazil Correspondence to Tatiana W.N. Nunes, PhD, Hospital Conceição, Av. Francisco Trein, 596 - Cristo Redentor, Porto Alegre, Porto Alegre, Rio Grande do Sul 91350-200, Brazil Tel/fax: +55 51 3357 2000; e-mail: tatianawnn@gmail.com *Tatiana W.N. Nunes and Eduardo C. Filippi-Chiela contributed equally to the writing of this article. Received 13 September 2018 Accepted 24 January 2019 Introduction Cutaneous melanoma corresponds to 4% of all skin malig- nant neoplasms [1]. However, because of its aggressive- ness, it is responsible for ~90% of skin cancer-associated deaths [2]. Numerous advances made in recent decades have allowed the classification of melanoma on the basis of molecular alterations [3]. However, concerning the prognosis of patients, the morphological features evalu- ated by optical microscopy in clinical routine still provide the most relevant criteria [4]. The criteria to melanoma staging, defined by the American Joint Committee on Cancer, are based on the most relevant prognostic factors in clinics, consid- ering the accessibility, reproducibility, and applicability. The Melanoma Expert Panel revised and published the eighth edition of the American Joint Committee on Cancer melanoma staging system, formally implemented nationwide on 1 January 2018. Key changes have been made in the new system to improve staging, prediction, and risk stratification of patients [5]. However, primary melanoma thickness and ulceration remain to define T category [6]. Survival among patients with T1 melanoma is primarily related to tumor thickness [7], with a clin- ically important threshold in the region of 0.7–0.8 mm [8]. Furthermore, among patients with T1 melanoma, tumor thickness and ulceration were stronger predictors of survival than mitotic rate. Sentinel lymph node (SLN) with metastasis occurs in less than 5% of cases among patients with T1a melanomas (< 0.8 mm without ulcera- tion), reaching over to 50% in T4b tumors ( > 4.0 mm with ulceration) [5]. Evidence-based TNM staging is essential for pathologic staging and personalized-medicine approach. Patients with stages I and II melanoma have localized disease, whereas those with stages III and IV melanoma have regional and distant metastatic disease, respectively. Although partially defined by the absence of regional Supplemental Digital Content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal’s website, www.melanomaresearch.com.