Digital odyssey: lessons learnt from a reverse transition from a digital to a manual pathology workflow.

In the fully digital Caltagirone pathology laboratory, a reverse shift from a digital to a manual workflow occurred due to a server outage in September 2023. Here, insights gained from this unplanned transition are explored. Surveying the affected pathologists and technicians revealed unanimous preferences for the time-saving and error-reducing capabilities of the digital methodology. Conversely, the return to manual methods highlighted increased dissatisfaction and reduced efficiency, emphasising the superiority of digital workflows. This case study underscores that transition challenges are not inherent to digital workflows but to transitioning itself, advocating for the adoption of digital technologies in all pathology practices.

Standardized and simplified reporting of next-generation sequencing results in advanced non-small-cell lung cancer: Practical indications from an Italian multidisciplinary group.

Molecular biomarker testing is increasingly becoming standard of care for advanced non-small cell lung cancer (NSCLC). Tissue and liquid biopsy-based next-generation sequencing (NGS) is now highly recommended and has become an integral part of the routine management of advanced NSCLC patients. This highly sensitive approach can simultaneously and efficiently detect multiple biomarkers even in scant samples. However full optimization of NGS in clinical practice requires accurate reporting and interpretation of NGS findings. Indeed, as the number of NSCLC biomarkers continues to grow, clinical reporting of NGS data is becoming increasingly complex. In this scenario, achieving standardization, simplification, and improved readability of NGS reports is key to ensuring timely and appropriate treatment decisions. In an effort to address the complexity and lengthy reporting of NGS mutation results, an Italian group of 14 healthcare professionals involved in NSCLC management convened in 2023 to address the content, structure, and ease-of-use of NGS reporting practices and proposed a standard report template for clinical use This article presents the key discussion points addressed by the Italian working group and describes the essential elements of the report template.

Validation of full-remote reporting for cervicovaginal cytology: the Caltagirone-Acireale distributed lab.

INTRODUCTION: Cervical cancer is the fourth most common cancer in women, and its prevention is based on vaccination and screening. Screening consists of molecular human papillomavirus (HPV) testing and cytologic analysis of cervical smears, which require expensive equipment and the interaction of numerous professionals such as biologists, cytologists, laboratory technicians, and pathologists. MATERIALS AND METHODS: We centralize the cervical samples from more than 51 clinics in 1 main laboratory, where automated HPV testing is performed. HPV-positive cases are collected and used to prepare a liquid-based cytology slide, which is stained and immediately scanned. The resulting whole-slide images (WSIs) are immediately available in a remote laboratory where they are examined by experienced cytologists using virtual microscopy. This setup was validated by making each of the 3 readers independently diagnose 506 specimens in random order, using both conventional light microscopy (CLM) and WSIs, with a minimum wash-out period of 3 weeks and with a final discussion for all cases. RESULTS: Intraobserver agreement among CLM and WSI ranged from 0.71 to 0.79, and interobserver agreement for the 3 readers compared with the consensus diagnosis was similar for the 2 modes of assessment. Readers subjectively felt confident in their WSI diagnosis for inadequate and negative cases, but less so in other cases. The perceived difficulty was slightly higher in WSI readings. CONCLUSIONS: Interobserver agreement in cervicovaginal cytology is moderate and does not vary if the slides are examined conventionally or digitally. Despite higher reported subjective difficulty and lower confidence in the WSI diagnosis, we did not observe a deterioration in diagnostic performance using WSI compared with CLM.

Deep learning for multi-class semantic segmentation enables colorectal cancer detection and classification in digital pathology images.

In colorectal cancer (CRC), artificial intelligence (AI) can alleviate the laborious task of characterization and reporting on resected biopsies, including polyps, the numbers of which are increasing as a result of CRC population screening programs ongoing in many countries all around the globe. Here, we present an approach to address two major challenges in the automated assessment of CRC histopathology whole-slide images. We present an AI-based method to segment multiple ([Formula: see text]) tissue compartments in the H &E-stained whole-slide image, which provides a different, more perceptible picture of tissue morphology and composition. We test and compare a panel of state-of-the-art loss functions available for segmentation models, and provide indications about their use in histopathology image segmentation, based on the analysis of (a) a multi-centric cohort of CRC cases from five medical centers in the Netherlands and Germany, and (b) two publicly available datasets on segmentation in CRC. We used the best performing AI model as the basis for a computer-aided diagnosis system that classifies colon biopsies into four main categories that are relevant pathologically. We report the performance of this system on an independent cohort of more than 1000 patients. The results show that with a good segmentation network as a base, a tool can be developed which can support pathologists in the risk stratification of colorectal cancer patients, among other possible uses. We have made the segmentation model available for research use on https://grand-challenge.org/algorithms/colon-tissue-segmentation/ .

Spatial resolution of renal amyloid deposits through MALDI-MSI: a combined digital and molecular approach to monoclonal gammopathies.

AIMS: Identification and characterisation of monoclonal gammopathies of renal significance (MGRS) is critical for therapeutic purposes. Amyloidosis represents one of the most common forms of MGRS, and renal biopsy remains the gold standard for their classification, although mass spectrometry has shown greater sensitivity in this area. METHODS: In the present study, a new in situ proteomic technique, matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI), is investigated as an alternative to conventional laser capture microdissection MS for the characterisation of amyloids. MALDI-MSI was performed on 16 cases (3 lambda light chain amyloidosis (AL), 3 AL kappa, 3 serum amyloid A amyloidosis (SAA), 2 lambda light chain deposition disease (LCDD), 2 challenging amyloid cases and 3 controls). Analysis began with regions of interest labelled by the pathologist, and then automatic segmentation was performed. RESULTS: MALDI-MSI correctly identified and typed cases with known amyloid type (AL kappa, AL lambda and SAA). A 'restricted fingerprint' for amyloid detection composed of apolipoprotein E, serum amyloid protein and apolipoprotein A1 showed the best automatic segmentation performance (area under the curve >0.7). CONCLUSIONS: MALDI-MSI correctly assigned minimal/challenging cases of amyloidosis to the correct type (AL lambda) and identified lambda light chains in LCDD cases, highlighting the promising role of MALDI-MSI for amyloid typing.

Data-driven color augmentation for H&E stained images in computational pathology.

Computational pathology targets the automatic analysis of Whole Slide Images (WSI). WSIs are high-resolution digitized histopathology images, stained with chemical reagents to highlight specific tissue structures and scanned via whole slide scanners. The application of different parameters during WSI acquisition may lead to stain color heterogeneity, especially considering samples collected from several medical centers. Dealing with stain color heterogeneity often limits the robustness of methods developed to analyze WSIs, in particular Convolutional Neural Networks (CNN), the state-of-the-art algorithm for most computational pathology tasks. Stain color heterogeneity is still an unsolved problem, although several methods have been developed to alleviate it, such as Hue-Saturation-Contrast (HSC) color augmentation and stain augmentation methods. The goal of this paper is to present Data-Driven Color Augmentation (DDCA), a method to improve the efficiency of color augmentation methods by increasing the reliability of the samples used for training computational pathology models. During CNN training, a database including over 2 million H&E color variations collected from private and public datasets is used as a reference to discard augmented data with color distributions that do not correspond to realistic data. DDCA is applied to HSC color augmentation, stain augmentation and H&E-adversarial networks in colon and prostate cancer classification tasks. DDCA is then compared with 11 state-of-the-art baseline methods to handle color heterogeneity, showing that it can substantially improve classification performance on unseen data including heterogeneous color variations.

Empowering digital pathology applications through explainable knowledge extraction tools.

Exa-scale volumes of medical data have been produced for decades. In most cases, the diagnosis is reported in free text, encoding medical knowledge that is still largely unexploited. In order to allow decoding medical knowledge included in reports, we propose an unsupervised knowledge extraction system combining a rule-based expert system with pre-trained Machine Learning (ML) models, namely the Semantic Knowledge Extractor Tool (SKET). Combining rule-based techniques and pre-trained ML models provides high accuracy results for knowledge extraction. This work demonstrates the viability of unsupervised Natural Language Processing (NLP) techniques to extract critical information from cancer reports, opening opportunities such as data mining for knowledge extraction purposes, precision medicine applications, structured report creation, and multimodal learning. SKET is a practical and unsupervised approach to extracting knowledge from pathology reports, which opens up unprecedented opportunities to exploit textual and multimodal medical information in clinical practice. We also propose SKET eXplained (SKET X), a web-based system providing visual explanations about the algorithmic decisions taken by SKET. SKET X is designed/developed to support pathologists and domain experts in understanding SKET predictions, possibly driving further improvements to the system.

Unleashing the potential of digital pathology data by training computer-aided diagnosis models without human annotations.

The digitalization of clinical workflows and the increasing performance of deep learning algorithms are paving the way towards new methods for tackling cancer diagnosis. However, the availability of medical specialists to annotate digitized images and free-text diagnostic reports does not scale with the need for large datasets required to train robust computer-aided diagnosis methods that can target the high variability of clinical cases and data produced. This work proposes and evaluates an approach to eliminate the need for manual annotations to train computer-aided diagnosis tools in digital pathology. The approach includes two components, to automatically extract semantically meaningful concepts from diagnostic reports and use them as weak labels to train convolutional neural networks (CNNs) for histopathology diagnosis. The approach is trained (through 10-fold cross-validation) on 3'769 clinical images and reports, provided by two hospitals and tested on over 11'000 images from private and publicly available datasets. The CNN, trained with automatically generated labels, is compared with the same architecture trained with manual labels. Results show that combining text analysis and end-to-end deep neural networks allows building computer-aided diagnosis tools that reach solid performance (micro-accuracy = 0.908 at image-level) based only on existing clinical data without the need for manual annotations.

The challenge of the Molecular Tumor Board empowerment in clinical oncology practice: A Position Paper on behalf of the AIOM- SIAPEC/IAP-SIBioC-SIC-SIF-SIGU-SIRM Italian Scientific Societies.

The development of innovative technologies and the advances in the genetics and genomics, have offered new opportunities for personalized treatment in oncology. Although the selection of the patient based on the molecular characteristics of the neoplasm has the potential to revolutionize the therapeutic scenario of oncology, this approach is extremely challenging. The access, homogeneity, and economic sustainability of the required genomic tests should be warranted in the clinical practice, as well as the specific scientific and clinical expertise for the choice of medical therapies. All these elements make essential the collaboration of different specialists within the Molecular Tumor Boards (MTBs). In this position paper, based on experts' opinion, the AIOM-SIAPEC/IAP-SIBioC-SIC-SIF-SIGU-SIRM Italian Scientific Societies critically discuss the available molecular profiling technologies, the proposed criteria for the selection of patients candidate for evaluation by the MTB, the criteria for the selection and analysis of biological samples, and the regulatory and pharmaco-economic issues.

A study of gene expression by RNA-seq in patients with prostate cancer and in patients with Parkinson disease: an example of inverse comorbidity.

BACKGROUND: Prostate cancer (PCa) is one of the leading causes of death in Western countries. Environmental and genetic factors play a pivotal role in PCa etiology. Timely identification of the genetic causes is useful for an early diagnosis. Parkinson's disease (PD) is the most frequent neurodegenerative movement disorder; it is associated with the presence of Lewy bodies and genetic factors are involved in its pathogenesis. Several studies have indicated that the expression of target genes in patients with PD is inversely related to cancer development; this phenomenon has been named "inverse comorbidity". The present study was undertaken to evaluate whether a genetic dysregulation occurs in opposite directions in patients with PD or PCa. METHODS AND RESULTS: In the present study, next-generation sequencing transcriptome analysis was used to assess whether a genetic dysregulation in opposite directions occurs in patients with PD or PCa. The genes SLC30A1, ADO, SRGAP2C, and TBC1D12 resulted up-regulated in patients with PD compared to healthy donors as controls and down-regulated in patients with PCa compared with the same control group. CONCLUSIONS: These results support the hypothesis of the presence of inverse comorbidity between PD and PCa.

Primary lung adenocarcinoma in three adolescent patients affected by bone sarcomas.

Pediatric primary lung carcinomas are extremely rare. Apart from known associations with congenital adenomatoid malformations, cases of primary lung adenocarcinomas after prolonged treatments of pediatric malignancy have been reported. We describe the morphological and molecular features of three cases of lung adenocarcinoma developed in adolescents aged 8 to 17 years during progression of their bone osteosarcoma or Ewing sarcomas. The morphological features overlapped those of adult lung adenocarcinoma including in situ, minimally invasive, and invasive forms. EGFR gene mutations were found in all three cases by targeted next-generation sequencing. The two patients with Ewing sarcoma had no progression of their lung cancer and no further progression of the metastatic bone tumor after additional chemo- and radio-therapy. Conversely, the osteosarcoma patient refused further treatments after thoracic surgery for metastatic osteosarcoma and locally advanced adenocarcinoma and died 2 years later of widespread distant metastases. Our results indicate that primary lung cancer might originate in pediatric patients during prolonged adjuvant therapies for primary bone neoplasm, and this possibility should be considered in the presence of suspected lung disease progression to correctly monitor the primary tumor evolution and define the appropriate therapeutic strategy at each time point. If appropriately treated, second primary lung cancer may not affect the patients' prognosis. The pathogenetic mechanisms of these rare lung adenocarcinomas are not clear, but the presence of EGFR mutations in all three cases indicates an oncogene addiction of the lung tumor, rather than a direct cancerogenic effect of the sarcoma-related treatment.

Diagnostic test assessment. Validation study of an alternative system to detect microsatellite instability in colorectal carcinoma.

The American Society for Clinical Pathology (ASCP), College of American Pathologists (CAP), Association for Molecular Pathology (AMP), and the American Society of Clinical Oncology (ASCO) have been recently strongly recommended the evaluation of mismatch repair status (MMS) as molecular biomarkers in colorectal cancer for a better prognostic stratification of patients. This recommendation is emphasized by the recent evidence of Microsatellite Instability (MSI) as a predictive marker for chemotherapy and immunotherapy.In this scenario, the validation of molecular biomarker testing methods seems to be essential to design the most appropriate tailored therapy and the most suitable care strategy, respectively.In this study, we validated an alternative method based on capillary electrophoresis system label-free PCR (Qiaxcel system) to evaluate the MSI Bethesda Panel. We also parallel the results with a standard approach.Our data showed total concordance with the standard approach, with a highly time-efficient and easy procedure combined with high sensitivity for MSI detection.Alternative capillary electrophoresis based on label-free PCR such as the Qiaxel system is a very sensitive and specific method to detect MSI for the management of patients with colorectal cancer. This procedure is adequate and suitable in diagnostic routine for the evaluation of microsatellite repeats compared to standard procedures.

Multiple Assays to Determine Methylguanine-Methyltransferase Status in Lung Carcinoids and Correlation with Clinical and Pathological Features.

BACKGROUND: O6-methylguanine-methyltransferase (MGMT) is a key enzyme for the DNA repair machinery strongly associated with response to alkylating agents in different tumors. Data on its expression and related clinical impact in neuroendocrine tumors are limited to the gastro-entero-pancreatic system, with controversial results in terms of prognostic or predictive value. In lung carcinoids, although clinical efficacy of alkylating agents has been shown in small studies, very few data to date are available on MGMT status. OBJECTIVE: To assess MGMT status in lung carcinoids using multiple assays and to compare data with major clinical and pathological features. METHODS: A retrospective series of 95 lung carcinoids and 51 control cases of high-grade neuroendocrine lung carcinomas was analyzed for MGMT promoter methylation, MGMT gene expression, and MGMT protein expression using pyrosequencing, quantitative real-time PCR, and immunohistochemistry, respectively. RESULTS: MGMT protein expression was inversely correlated with MGMT promoter methylation and positively with MGMT gene expression. MGMT promoter methylation progressively increased from carcinoids to high-grade carcinomas. In the carcinoid group, decreased MGMT gene expression was significantly associated with aggressive features (atypical histotype, grade G2, larger tumor size, higher T stage, and positive nodal status) but not with survival. MGMT promoter methylation was associated with lower stage and negative nodal status. CONCLUSIONS: Our study investigated MGMT status in a large series of lung carcinoids in the attempt to move forward a rational use of alkylating agents in these tumors. Interestingly, low MGMT gene expression defines a subgroup of lung carcinoids with aggressive features.

Cerebellar Metastasis From Ovarian Carcinoma Harboring PIK3CA-Activating Mutation: A "Clear" Explanation for Unexpected "Vertigo".

Brain metastasis is a rare and generally late manifestation of an advanced-stage, high-grade serous ovarian carcinoma. Nowadays, the improved control of intra-abdominal disease by surgery and platinum-based chemotherapy results in a longer survival, allowing distant metastasis to implant and grow in the brain parenchyma. Herein, we describe a unique case of a cerebellar metastasis from clear cell ovarian carcinoma that initially presented as a FIGO Stage IC cancer. Surprisingly, 6 mo after surgery, the patient was in good condition with complete disappearance of symptoms and no evidence of recurrence. This relatively good biologic behavior may be explained by the presence of a PIK3CA-activating mutation in exon 9 which as previously reported in the literature, may be associated with better prognosis. To the best of our knowledge, this is the first reported case of cerebellar metastasis from ovarian clear cell carcinoma. In the presence of neurological symptoms, both clinicians and pathologists must be aware of this rare possibility, to assure correct patient management and effective therapeutic options. Generally, the prognosis of epithelial ovarian cancer patients with brain metastases is poor. PIK3CA mutations could be a good prognostic indicator in clear cell carcinomas.

Clear cell carcinoma of the endometrium: an immunohistochemical and molecular analysis of 45 cases.

The aim of the present paper is to study a cohort of pure selected endometrial clear cell carcinomas (ECCCs) from an immunohistochemical and molecular perspective to provide new data about the molecular profile of this disease. In detail, 45 consecutive patients with a proven diagnosis of pure ECCC, according to World Health Organization criteria, were included into the study. We determined the incidence of KRAS, BRAF, and PIK3CA mutations as well as the immunohistochemical expression of mismatch repair (MMR) proteins (MLH1, MSH2, MSH6, PMS2), estrogen, progesterone receptors, p16, and p53. Immunohistochemical analyses for α-methylacyl-coenzyme-A racemase and Napsin A were performed to support the diagnosis of ECC. All cases were positive for at least 1 of the 2 markers. In detail, 34 of 45 (75.5%) cases were positive for α-methylacyl-coenzyme-A racemase, and 40 of 45 (88.8%) cases showed positive staining for Napsin A. All selected cases exhibited negative immunostain for estrogen receptor and progesterone receptor, a "patchy" immunostain for p16, and a "wild-type" staining pattern for p53. Fifteen patients (15/45; 33.3%) showed loss of 1 or more MMR proteins by immunohistochemistry. Seven patients showed dual loss of MSH2 and MSH6, 4 patients (8.8%) showed isolated loss of MSH6, and the remaining 4 patients showed isolated loss of PMS2, respectively. Pyrosequencing analysis revealed the presence of 5 of 45 mutations (11%) at codon 12 in exon 2 of KRAS (3/5 p.G12D, 60%; 2/5 p. G12V, 40%) and 5 of 45 (11%) mutations in PIK3CA gene, of which 3 of 5 (60%) were in exon 9 of PIK3CA (2 p.E542K and 1 p.Q546K) and 2 of 5 (40%) were in exon 20 (p.H1047R). Two synchronous mutations affecting exon 9 of PIK3CA (p.Q546K) and exon 2, codon 12 of KRAS (p.G12D) were found. No mutations were detected in the hot spot region of BRAF. In conclusion, we provided detailed immunohistochemical and molecular data in a series of ECC, demonstrating a high incidence (33%) of MMR deficiencies detected by immunohistochemistry as well as a synchronous mutation affecting PIK3CA and KRAS genes. A more extensive interrogation of the genomic features of a much larger series of clear cell carcinomas will be required to define the genomic landscape of this subtype and to determine whether there are molecular alterations that are unique to, or significantly enriched in, clear cell tumors compared to other subtypes.

Ovarian Clear Cell Carcinoma: From Morphology to Molecular Biology.

Ovarian clear cell carcinoma (oCCC) is a distinctive subtype of ovarian carcinoma, with peculiar genetic and environmental risk factors, precursor lesions, molecular events during oncogenesis, patterns of spread, and response to treatment. Because of low response to chemotherapy and poor prognosis in advanced stages, there is growing interest in investigating the molecular pathways involved in oCCC development, in order to individualize novel/molecular targeted therapies. Until now, the main molecular genetic changes associated with oCCC remain to be identified, and, although several molecular changes have been reported in clear cell tumors, most studies have analyzed a limited number of cases; therefore, the true prevalence of those changes is not known. The present review will present the clinicopathologic features of oCCC, from morphology to molecular biology, discussing the diagnostic and treatment challenges of this intriguing ovarian carcinoma.

High miR-100 expression is associated with aggressive features and modulates TORC1 complex activation in lung carcinoids.

PURPOSE: Mammalian target of rapamycin (mTOR) is a promising therapeutic target in advanced lung carcinoid patients. However, the mechanisms of mTOR modulation and of responsiveness to mTOR inhibitors are largely unclear. Our aim was to analyze the expression and functional role of specific miRNAs in lung carcinoids as an alternative mechanism targeting mTOR pathway. EXPERIMENTAL DESIGN: Seven miRNAs, selected by bioinformatic tools and literature search, were analyzed in 142 lung neuroendocrine neoplasms (92 carcinoids and a control group of 50 high grade neuroendocrine carcinomas), and compared with mTOR mRNA expression and clinical/pathological parameters. Tissue results were validated in vitro in two lung carcinoid cell lines by specific RNA interference and biological/pharmacological tests. RESULTS: Tissutal expression of five miRNAs (miR-99b, miR-100, miR-155, miR-193a-3p, miR-193a-5p) was inversely correlated with mTOR mRNA expression, supporting their role in the negative regulation of mTOR transcription. High expression of miR-100, miR-193a-3p and miR-193a-5p was associated with aggressive features and, for the former two, with shorter time to progression. In H727 and UMC11 lung carcinoid cells, miR-100 modulated mTOR RNA and TORC1 complex protein expression, positively promoted cell migration and negatively influenced cell proliferation. Moreover, miR-100 directly influenced responsiveness of H727 and UMC11 cells to rapamycin. CONCLUSIONS: MiR-100 actively participates to the regulation of mTOR expression in lung carcinoids and represents a novel candidate prognostic biomarker for this tumor type; moreover, inhibition of its expression is associated to increased responsiveness to mTOR inhibitors and might represent a novel strategy to sensitize lung carcinoids to these target agents.

Detailed genomic characterization identifies high heterogeneity and histotype-specific genomic profiles in adrenocortical carcinomas.

Molecular characterization of adrenocortical carcinoma has been recently established, but the correlation between molecular profiles and clinical and pathological characteristics is still poorly defined with no data available about genetic heterogeneity along disease progression. In this scenario, a detailed molecular profile was correlated with clinical and pathological characteristics in adrenocortical carcinoma patients to identify potentially novel biomarkers. Targeted next-generation sequencing and copy number variation analyses for 18 most frequently altered genes in adrenocortical carcinoma were assessed on 62 adult cases (including 10 with matched primary and metastatic/recurrence samples) and results correlated with major clinical and pathological characteristics of tumors. A total of 433 somatic deleterious genetic alterations (328 gene mutations and 105 copy number variations) were identified in 57/62 cases, five resulted wild type for all genes tested. TERT, CDK4, ZNRF3,and RB1 were altered in more than 30% of cases. Among histological variants genotypes were significantly different. Lowest mutation burden was found in the oncocytic type (p = 0.006), whereas the highest with a prevalence of RB1 (p = 0.001) and CDK4 (p = 0.002) was found in the conventional and myxoid ones, respectively. None of the 10 cases with matched samples showed a stable genotype along tumor progression, although allelic frequencies or percentages of altered nuclei at fluorescence in situ hybridization were in most cases similar among different tumor samples for genes that were stable along tumor progression. Among individual genes, an altered p53/Rb1 pathway was the strongest adverse molecular signature, being associated with high Ki-67 index, high tumor stage, aggressive disease status, and shorter disease-free survival. The genomic signature in adrenocortical carcinoma is changing along tumor progression and is associated with specific clinical and pathological features, including histological variant and prognosis.

Mitochondrial DNA "common deletion" in post-fine needle aspiration infarcted oncocytic thyroid tumors.

Thyroid fine needle aspiration (FNA) can rarely induce morphological changes potentially hindering the histopathological diagnosis, especially in Hurthle cell tumors (HCTs), which may easily undergo post-FNA infarction or necrosis. HCTs contain mitochondrion (mt)-rich cells that may bear mtDNA mutations, the most frequent being the so-called common deletion (CD). The aim of this study was to determine the presence and extent of the mtDNA CD in a series of thyroid HCTs that underwent extensive infarction following FNA procedure in comparison with a control series of HCTs lacking post-FNA ischemic/hemorrhagic alterations. Of 257 HCTs with available matched FNA and surgical specimens, 8 cases showed extensive (≥80%) infarction or necrosis in the resected nodule (4 adenomas, 1 carcinoma, 3 HCTs undefined for malignancy). Noninfarcted tumors in the control series included 9 adenomas, 1 carcinoma, and 1 follicular tumor of uncertain malignant potential. These lesions were significantly (P = .03) larger than infarcted nodules. The mtDNA CD was identified using semiquantitative real-time polymerase chain reaction in 2 of 8 (25%) infarcted tumors. In HCTs lacking infarction/necrosis of the control series, the CD was significantly (P = .05) more common (8/11 cases, 72.7%). In 7 of the 10 deleted cases, the CD was present also in the adjacent nonneoplastic parenchyma. In conclusion, the rare oncocytic tumors undergoing extensive infarction are smaller than those lacking ischemic changes and bear the mtDNA CD in a significantly lower proportion compared with control noninfarcted HCTs. This may suggest that mtDNA deletion confers a survival advantage to oncocytic cells in stress conditions, including FNA procedures.