The Nottingham Grading System (NGS) developed by Elston and Ellis is used to grade invasive breast cancer (IBC). Glandular (acinar)/tubule formation is a component of NGS.
To investigate the ability of pathologists to identify individual structures that should be classified as glandular (acinar)/tubule formation.
A total of 58 hematoxylin-eosin photographic images of IBC with 1 structure circled were classified as tubules (41 cases) or nontubules (17 cases) by Professor Ellis. Images were sent as a PowerPoint (Microsoft) file to breast pathologists, who were provided with the World Health Organization definition of a tubule and asked to determine if a circled structure represented a tubule.
Among 35 pathologists, the κ statistic for assessing agreement in evaluating the 58 images was 0.324 (95% CI, 0.314–0.335). The median concordance rate between a participating pathologist and Professor Ellis was 94.1% for evaluating 17 nontubule cases and 53.7% for 41 tubule cases. A total of 41% of the tubule cases were classified correctly by less than 50% of pathologists. Structures classified as tubules by Professor Ellis but often not recognized as tubules by pathologists included glands with complex architecture, mucinous carcinoma, and the “inverted tubule” pattern of micropapillary carcinoma. A total of 80% of participants reported that they did not have clarity on what represented a tubule.
We identified structures that should be included as tubules but that were not readily identified by pathologists. Greater concordance for identification of tubules might be obtained by providing more detailed images and descriptions of the types of structures included as tubules.
Histologic grading of invasive breast carcinoma (IBC) represents a morphologic assessment of the degree of tumor differentiation and is routinely reported in the pathologic assessment of IBC. Histologic grade is an important prognostic factor that is independent of lymph node involvement or tumor size and is also used to guide clinical decisions regarding the need for systemic therapies.1,2 Furthermore, the histologic grade is prognostic across all subtypes of IBC, and the impact on survival has been demonstrated for more than 10 years.3 Grading is based on microscopic evaluation of hematoxylin-eosin (H&E)–stained tumor sections; hence, it is a simple and inexpensive test for the evaluation of prognosis. It is globally included in breast cancer reporting guidelines, and in 2017 tumor grade was incorporated into the 8th edition of the American Joint Committee on Cancer (AJCC)4 Tumor Staging System for breast cancer as a component of the prognostic stage grouping for breast cancer.5–7
The Nottingham modification of the Scarff-Bloom-Richardson8 grading system, which is also known as the Nottingham Grading System (NGS), was first proposed in 1991 by Elston and Ellis.9 The NGS is the most widely recommended and used methodology for histologic grading.4–7 The NGS uses both subjective and semiquantitative criteria to evaluate 3 components of breast cancer morphology, including tubule formation, nuclear pleomorphism, and mitotic count. A score of 1 to 3 is assigned to each component, with 1 representing a favorable or better differentiated appearance and 3 representing an unfavorable, less differentiated appearance. The 3 scores are added together to give a combined score of 3 to 5, which is well differentiated; 6 to 7, which is moderately differentiated; and 8 to 9, which is poorly differentiated. Multiple studies have consistently demonstrated a highly significant independent association between higher histologic grade and poorer prognosis.9–12
It is well documented that the NGS assessment suffers from reproducibility issues, which may be related to the subjective nature of the assessment. A large number of studies have evaluated interobserver agreement in breast carcinoma grading, and it ranges from fair to excellent, with κ values ranging from 0.43 to 0.85.13–16 Methodologies demonstrated to improve reproducibility in NGS include improvement in tissue quality and fixation, as well as employment of educational and quality assurance practices.17 However, because histologic grade plays a critical role in prognosis and guiding therapeutic management, any variation in grading can translate into inaccurate prognostic staging and subsequent suboptimal patient management.
Among the 3 components comprising NGS, interobserver variability is noted in all 3. Studies generally report the lowest levels of agreement for nuclear pleomorphism and mitotic count, with a reported κ of 0.40 and 0.52, respectively.18 Although the κ value for agreement is the highest for tubule formation, it still ranges from moderate to good, with a κ of 0.53 and 0.64 in recent studies.18,19 Hence improvement in reproducibility is needed in all 3 components of NGS.
Evaluation of glandular (acinar)/tubular differentiation in NGS includes both identification of a structure as a tubule and assessment of the percentage of the entire tumor area composed of tubules. A tumor in which more than 75% has tubules is assigned a score of 1, a tumor where 10% to 75% of the tumor has tubules is assigned a score of 2, and if less than 10% of the tumor has tubules, the assigned score is 3. To assign a tubule score for Nottingham Grading a pathologist must be able to clearly identify what represents glandular (acinar)/tubule formation. As per World Health Organization (WHO) guidelines, glandular (acinar)/tubule formation is defined as “structures exhibiting a clear central lumina surrounded by polarized neoplastic cells.”7 In this multi-institutional study we sent a series of 58 unique PowerPoint (Microsoft) images of H&E-stained slides of IBC to breast pathologists and asked them to determine if a single circled structure represented a tubule as defined by the NGS.7,9 We try to identify areas of difficulty in the identification of glandular (acinar)/tubular differentiation and provide suggestions for improvement based on input from the participants.
MATERIALS AND METHODS
A survey titled “Study to evaluate concordance in assessment of tubules in Nottingham Grading in Invasive breast cancer” was sent to 39 pathologists who identified as practicing breast pathology across the United States in 2021 and 2022. The survey included a set of 65 H&E-stained microscopic slide images of IBC with a single epithelial structure circled and sent as a PowerPoint image. Images were photographed at ×10, ×20, or ×40 objectives and included representations of invasive ductal carcinoma no special type, mucinous carcinoma, invasive micropapillary carcinoma, tubular and cribriform carcinoma, and solid papillary carcinoma with invasion. Seven images were repeated to evaluate intraobserver variability; hence, there was a total of 58 unique images. The survey included a brief explanation of the study and directions on how to complete the tubule evaluation. Pathologists were asked to identify whether the circled structure on the PowerPoint image of the slide represented a tubule, and if not, they were then given 7 choices as to why not. Additionally, at the end of the survey questions were presented about the practice and experience of the participant as well as specific questions related to difficulty in tubule evaluation. The actual survey is presented as supplemental digital content (see supplemental digital content file containing 3 sections at https://meridian.allenpress.com/aplm in the October 2024 table of contents). All images were reviewed and classified by Ian Ellis, MBBS FRCPath, who, along with Elston, originally proposed the NGS in 1991.9 His opinion was used as the gold standard for tubule evaluation.9 Concordance in this study was thus defined as the agreement between a participating pathologist and Professor Ellis (gold standard) on tubule evaluation.
Statistical Analysis
Interobserver agreement in participating pathologists on evaluating H&E-stained microscopic slide images of invasive breast carcinoma with a single epithelial structure circled were examined using the κ statistic. Concordance was defined as the agreement between a participating pathologist and Ellis (the gold standard) on tubule evaluation. Concordance rate in evaluating tubule images was calculated as the number of concordant tubule images divided by the total number of tubule images designated by Ellis. Concordance rate in evaluating nontubule images was calculated as the number of concordant nontubule images divided by the total number of nontubule images designated by Ellis. Differences in concordance rate in evaluating tubule and nontubule cases between breast pathology exclusively pathologists and non–breast pathology exclusively pathologists were compared using Wilcoxon rank sum tests. All statistical analyses were performed using SAS version 9.4 (SAS Institute Inc, Cary, North Carolina). P values of .05 or less were considered statistically significant.
RESULTS
Pathologist Participant Information
We received a response from 35 of the 39 breast pathologists from 16 academic-affiliated pathology departments across the United States. These results were compared with the gold standard (Professor Ellis) classification of all images.9
A total of 18 of the 35 pathologists (51.4%) reported completing a fellowship in breast pathology. Two participants were in fellowship at the time of this study survey; 9 were in practice less than 5 years; 8 were in practice 5 to less than 10 years; 6 for 10 to less than 20 years; and 10 reported 20 or more years of practice. Although all pathologists practiced at an academic-affiliated medical center, 3 of the pathologists described their practice setting as nonacademic. A total of 17 of 35 pathologists (48.6%) reported that 100% of their surgical pathology practice was breast pathology, and among them, 9 had completed a breast fellowship. A total of 18 of 35 pathologists (51.4%) reported a surgical pathology practice that included multiple subspecialties, and among them, 9 had completed a breast fellowship. All participants in this study considered breast pathology an area of expertise.
Tubule Assessment Agreement
The 65 images presented to the pathologists in the survey are provided in the supplemental digital content (section 2) along with the gold standard classification (yes = tubule and no = nontubule) for each image (provided by Professor Ellis). Among the 65 images, there were 58 unique images, 7 of which were presented in duplicate. Among 58 images, 41 were considered to represent a tubule, and 17 were nontubule cases as determined by the gold standard. The 7 repeat images consisted of 5 tubule images and 2 nontubule images. Overall, 32 of 35 pathologists (91.4%) had the same reading for all 7 pairs, and all 35 pathologists had the same reading in at least 6 of the 7 pairs. For the 3 pathologists who each had 1 pair with a different reading, 2 were tubule cases and 1 was a nontubule case. The estimated κ statistic for assessing agreement in 35 participating pathologists in tubule evaluation was 0.326 (95% CI, 0.315–0.336) when repeat image Nos. 10, 27, 48, 42, 54, 60, and 65 were excluded, and 0.324 (95% CI, 0.314–0.335) when repeat image Nos. 1, 16, 30, 31, 47, 57, and 12 were excluded. This was considered poor overall agreement in 35 pathologists on tubule evaluation. Concordance for the 58 unique images will be presented to exclude 7 repeat images (images 10, 27, 48, 42, 54, 60, and 65 were excluded).
Concordance in the Evaluation of 17 Nontubule Cases
The median (interquartile range) concordance rate between the 35 pathologists and the gold standard for cases designated as nontubule was excellent (94.1% [88.2%–100]), with a range of 17.6% to 100%. However, a 100% concordance with the gold standard was observed in only 2 of the nontubule cases (Figure 1). The median concordance rate in evaluating all 17 nontubule cases was 100% (94.7%–100%) for the 17 pathologists who practiced only breast pathology and 94.1% (88.2%–100%) for the 18 non–breast pathology exclusive pathologists (P = .02). In all nontubule images, the explanation provided by Professor Ellis for the determination that the circled structure was not a tubule was no central lumina visible (choice A), which was also the explanation provided by most of the participating pathologists.
Two images of nontubule cases with 100% concordance between 35 pathologists and Ian Ellis (MBBS FRCPath), where the circled structure does NOT represent a tubule (hematoxylin-eosin, original magnifications ×400 [A] and ×100 [B]).
Figure 2. Two tubule images with 100% concordance between 35 pathologists and Ian Ellis (MBS FRCPath), where the circled structure represents a tubule (hematoxylin-eosin, original magnifications ×400 [A] and ×200 [B]).
Two images of nontubule cases with 100% concordance between 35 pathologists and Ian Ellis (MBBS FRCPath), where the circled structure does NOT represent a tubule (hematoxylin-eosin, original magnifications ×400 [A] and ×100 [B]).
Figure 2. Two tubule images with 100% concordance between 35 pathologists and Ian Ellis (MBS FRCPath), where the circled structure represents a tubule (hematoxylin-eosin, original magnifications ×400 [A] and ×200 [B]).
Concordance in Evaluation of 41 Tubule Cases
The median concordance rate between the 35 pathologists and the gold standard for cases classified as a tubule was 53.7% (34.2%–85.4%), with a range of 14.6% to 100%. A complete agreement of the 35 pathologists with the gold standard was observed in only 3 of 41 cases (7.3%; Figure 2). The median concordance rate was 58.5% (31.7%–75.6%) for the 17 pathologists who practiced only breast pathology and 51.3% (39%–92.7%) for the 18 non–breast pathology exclusive pathologists (P = .45).
Number of pathologists with concordant/correct classification for each of the 58 cases, including information on histologic appearance. Shown are nontubule cases (black); single tubules (white); complex/cribriform tubule patterns (hatch pattern/blue); micropapillary carcinoma with inverted tubules (diagonal lines/green); and mucinous carcinoma with tubules (checkerboard orange).
Number of pathologists with concordant/correct classification for each of the 58 cases, including information on histologic appearance. Shown are nontubule cases (black); single tubules (white); complex/cribriform tubule patterns (hatch pattern/blue); micropapillary carcinoma with inverted tubules (diagonal lines/green); and mucinous carcinoma with tubules (checkerboard orange).
In 17 of the 41 tubule cases (41.4%), most pathologists were not in concordance with the gold standard (Figure 3). The major reason why some of the gold standard tubule images were considered as nontubule by participants was architectural complexity (“such as cribriform or shape”; Figures 3 and 4). The concordance between the participants and the gold standard for the 8 tubule images from mucinous carcinoma was low (12.5% [0–75%]), with a range of 0 to 100%; Figure 3 and Figure 5, A and B). The major reason for disagreement was “a special type of carcinoma.” Interestingly, in 3 of the 4 images of micropapillary carcinoma, most of the pathologists were in agreement with the gold standard designation of tubules (Figure 5, C). The lack of polarization and high nuclear grade were the causes of misclassification in 3 cases each (Figure 6).
Three cases designated as tubule by Ian Ellis (MBBS FRCPath) where participants chose NOT a tubule because of architectural complexity, including choice B) lumen visible but contains epithelial structure or choice E) contains lumina but not a tubule due to shape. A, A total of 13 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. B, A total of 13 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. C, A total of 19 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule (hematoxylin-eosin, original magnifications ×400 [A and B] and ×200 [C]).
Figure 5. Cases designated as tubule by Ian Ellis (MBBS FRCPath) where participants chose NOT a tubule because of a special type of carcinoma hence not considered a tubule. A, Mucinous carcinoma: 21 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. B, Mucinous carcinoma: 18 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. C, Micropapillary carcinoma: 13 of 25 pathologists did not consider the outlined epithelial structure to represent a tubule (hematoxylin-eosin, original magnifications ×200 [A] and ×400 [B and C]).
Three cases designated as tubule by Ian Ellis (MBBS FRCPath) where participants chose NOT a tubule because of architectural complexity, including choice B) lumen visible but contains epithelial structure or choice E) contains lumina but not a tubule due to shape. A, A total of 13 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. B, A total of 13 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. C, A total of 19 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule (hematoxylin-eosin, original magnifications ×400 [A and B] and ×200 [C]).
Figure 5. Cases designated as tubule by Ian Ellis (MBBS FRCPath) where participants chose NOT a tubule because of a special type of carcinoma hence not considered a tubule. A, Mucinous carcinoma: 21 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. B, Mucinous carcinoma: 18 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule. C, Micropapillary carcinoma: 13 of 25 pathologists did not consider the outlined epithelial structure to represent a tubule (hematoxylin-eosin, original magnifications ×200 [A] and ×400 [B and C]).
Cases chosen by Ian Ellis (MBBS FRCPath) to represent tubules where participants chose NOT a tubule for the following reasons. A, A total of 15 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule, and the majority choice was F) contains a lumina but cells not polarized towards luminal space. B, A total of 20 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule, and the majority choice was choice C) nuclei too high grade (hematoxylin-eosin, original magnification ×400).
Cases chosen by Ian Ellis (MBBS FRCPath) to represent tubules where participants chose NOT a tubule for the following reasons. A, A total of 15 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule, and the majority choice was F) contains a lumina but cells not polarized towards luminal space. B, A total of 20 of 35 pathologists did not consider the outlined epithelial structure to represent a tubule, and the majority choice was choice C) nuclei too high grade (hematoxylin-eosin, original magnification ×400).
Overall Concordance
The median concordance rate between 35 pathologists and the gold standard in evaluating 58 unique images was 63.8% (51,7%, 84.5%), with a range of 39.7%–96.6%. Concordance with Professor Ellis for each of the individual 58 cases ranged from 100% (all pathologists agreed) to 11% (only 4 pathologists agreed). Figure 3 illustrates the number of pathologists who correctly identified each of the 58 unique images along with image type. The lowest concordance was seen for mucinous carcinoma with tubules, tubules with complex cribriform patterns, and micropapillary carcinoma with inverted tubules . The highest concordance was seen for non-tubule cases and for some cases of single tubules.
Responses to Survey Questions
Responses to the survey questions regarding tubule assessment are summarized below.
1. Do you ever use other pathologic features of the tumor to guide your decision on whether there are tubules (ie, subtype, overall grade)? If so, please describe?
A total of 19 of the 35 pathologists (54.3%) used additional features to define tubules; 11 of these did not designate tubules in a recognized subtype (ie, mucinous, micropapillary, and/or cribriform), whereas 7 considered other elements of NGS in determination of tubules.
2. Do you find it difficult to estimate the percentage of tubules present? If so, please describe?
A total of 12 of the 35 pathologists (34.3%) reported difficulty in estimating the percentage of tubules, and the most frequent explanation provided was for cases near the cut-points.
3. Do you evaluate tubule percentage based on one slide or the entire tumor?
All of the participants who responded (33 of 35; 94%) evaluated tubule percentage based on the entire tumor.
4. Do you have clarity on what represents a tubule, or do you wish for more specific guidance and if so, what might that be and where should it be available? (ie, improved text, illustrations, College of American Pathologists [CAP] website/AJCC staging manual?)
A total of 28 participants (80%) expressed the need for clarity/specific guidance, with a further specification by many (8 of 28) for assessment of tubules in cases with special histology. Two participants thought it would be helpful to better define polarized cells. Specific sites suggested included WHO, the CAP website, the AJCC staging manual, and textbooks, with a call for both improved text and illustrations.
DISCUSSION
The histologic grade of IBC represents a morphologic assessment of the degree of tumor differentiation and is one of the strongest prognostic determinants.1,2,9–12,20 Yet now, 3 decades after the NGS was proposed, there is still substantial confusion in the pathology community over the designation of tubules. We found poor overall agreement (κ = 0.326) between pathologists in tubule evaluation. The use of a gold standard (Professor Ellis) and 35 participant pathologists and 58 unique H&E PowerPoint images identified poor overall concordance largely due to the misclassification of tubules as nontubular structures (53.7% [34.2%–85.4%]), with greater concordance for nontubule structures (94.1% [88.2%–100%]). Glands with complex architecture and carcinomas that were classified as a specific subtype, in particular mucinous carcinoma, represented the greatest areas of discordance.
The predominant reasons for discordance were architectural complexity (ie, cribriform glands) or the carcinoma being of a specific subtype. For mucinous carcinoma, the median concordance rate for tubules was only 12.5% (0–75%). In their original publication on tumor grading in 1957, Bloom and Richardson8 had noted that colloid carcinoma posed difficulties in grading. They noted that “since most tumors showed attempt at tubule formation along with other favorable histologic features, they were mostly placed in grade 1.” In the 1991 publication by Ellis and Elston,9 in which NGS was first proposed, they noted that special types of IBC, such as cribriform and mucinous, have an excellent prognosis and questioned if histologic grade should be assessed in these subtypes. They reasoned that when grade is assessed on tumors of a particular histologic type it is usually found to be appropriate such that tubular or invasive cribriform carcinoma would have a grade 1 status. Some of the participants made comments suggesting subtypes such as mucinous and micropapillary carcinoma contain only pseudotubule structures which are not true tubules, and other participants suggested there is no agreement on whether cribriforming structures represent true tubules. It is unclear where this opinion comes from because a review of the original publication by Bloom and Richardson8 in 1957 clearly illustrates a structure designated as tubule which has a cribriform architecture. Notably, this work was published more than 65 years ago, hence many pathologists may not have ready access to this important work.
There were 4 images of invasive micropapillary carcinoma in our study where cell clusters contained a central space, hence an inverted tubule pattern, and all were designated as tubules by Ellis. Although most of the pathologists agreed with the tubule assignment in 3 of these 4 images, others considered the inverted tubule pattern and/or the association with “poor prognosis” to preclude a designation of any of these clusters as tubules. Although invasive micropapillary carcinoma demonstrates increased rates of lymphovascular invasion and axillary lymph node metastasis, the prognosis when adjusted for stage is similar to that for patients with invasive carcinoma no special type.21,22 Furthermore, they are luminal A or B by gene expression profiling, with more than 85% being positive for estrogen receptor expression.23,24 In the subset of cases that are human epidermal growth fact or receptor 2 (HER2) positive or the small number that are triple negative, the newer AJCC tumor staging system will place them within the appropriate prognostic stage grouping. In general, invasive micropapillary and other subtypes of invasive carcinoma associated with gland formation but with a more aggressive clinical behavior, such as invasive tubulopapillary, will likely continue to pose challenges in grading,. and this should be addressed specifically in updated directions for the identification of glandular (acinar)/tubular differentiation.25
Although the definition of tubule has not changed since 1957, the addition of the presence of cell polarization to the definition was noted to be confusing and difficult to apply by participants in our study.7–9,26 Based on our gold standard classification of tubule images, it appears that polarization need not be well developed; however, it is useful to distinguish spaces induced by shrinkage artifact due to suboptimal fixation, tissue processing or freezing, or clefts from dishesive cells or focal necrosis.27 Most of the participating pathologists (80%) stated that they do not have clarity on what represents a tubule. They suggested that updated guidelines be made available by WHO, the CAP website, the AJCC staging manual, and textbooks, with a call for both improved text and illustrations. A compilation of current definitions from recognized national and international organizations and texts is provided in the supplemental digital content (section 3). Furthermore, we hope that the information gained from this study will assist in creating an expanded description of glandular (acinar)/tubule formation as originally intended by Elston and Ellis.
As highlighted by Ellis and confirmed by our study, the term tubule formation, although used in the original NGS, needs to be clarified. Refined terminology of glandular (acinar) and tubular differentiation is preferred and already used as part of the CAP cancer protocols for invasive breast cancer.26 An excellent current description/definition/how-to guide for assignment of glandular (acinar)/tubular differentiation score is in the current International Collaboration on Cancer Reporting (ICCR) guideline for invasive carcinoma, which is provided in the supplemental digital content (section 3).28 The ICCR guideline addresses subtypes of carcinoma that include mucinous, micropapillary, and cribriform carcinoma, specifically stating they can be considered as gland/tubule forming when lumens are present. It includes the presence of such complex glandular structures as tubule/gland formation if lumens are present. As our study has demonstrated, these are areas requiring clarity for pathologists. Although artificial intelligence shows promise in assisting pathologists in the grading of IBC, defining ground truth for tubule recognition for training of these algorithms requires correct classification of structures such as tubules, which can be facilitated by improved definitions, such as those provided by the ICCR guideline.28,29
The strengths of our study include participation by 35 breast pathologists representing 15 different academic institutions, the participation by Professor Ellis to create the gold standard responses, and the design, which ensured that all pathologists evaluated the same structure as tubule or nontubule. Limitations include the fact that we focused primarily on academic pathologists, not all participants answered all the questions pertaining to practice and/or difficulties in tubule evaluation, and 4 pathologists did not respond. The addition of international participants would have strengthened our study.
The grade of IBC, as determined by the NGS, is critically important in determining prognosis and guiding therapy. In our study we tasked 35 breast pathologists with applying criteria outlined in the NGS to identify glandular (acinar)/tubular differentiation in IBC. We found poor overall agreement, with a κ statistic of 0.326 for tubule identification in 58 images of IBC. Overall concordance was lowest for identification of the presence of glandular (acinar)/tubule differentiation in tumor subtypes (ie, mucinous carcinoma) and in cases with complex architecture (ie, cribriform glands). Along with 80% of participants we support updated guidelines for recognition of glandular (acinar)/tubular differentiation in IBC, which should include specific instructions to address these issues. We hope both the information gained from this study along with the photomicrographs provided with this publication, each with tubule assignment classification as determined by Professor Ellis (yes/no), will provide improved guidance for glandular (acinar)/tubular identification.
We would like to acknowledge the editorial assistance provided by Matthew Shear in the preparation of this manuscript.
References
Author notes
Supplemental digital content is available for this article at https://meridian.allenpress.com/aplm in the October 2024 table of contents.
Competing Interests
The authors have no relevant financial interest in the products or companies described in this article.
Presented as an abstract at the United States and Canadian Academy of Pathology Annual Meeting; March 14, 2023; New Orleans, Louisiana.