Mitotic Rate Research Articles

Gastric gastrointestinal stromal tumors and overall survival in elderly patients.

e23516 Background: The purpose of this study is to analyze the overall survival (OS) of various age groups of patients diagnosed with gastric gastrointestinal stromal tumors (GIST). Methods: We compared 4 different age groups ( < 50 years old, 50-64 years old, 65-79 years old, and 80 or greater) comprising 230 patients in terms of various clinicopathological parameters and OS from January 2010 to December 2020 within Kaiser Permanente Northern California (KPNC), using Kaplan-Meier curves, and Cox proportional hazard regression model. Results: Univariate analysis showed that mitosis greater 10 HPF was a statistically significant predictor of decreased OS from gastric GIST (p = .032). Although necrosis alone was not significant (p = .271), the combination with higher mitotic rate was significant (p = .025). Octogenarians with gastric GIST were more likely to present with bleeding, with a 5.3% ninety-day surgical mortality rate. Multivariate Cox regression showed that patients 80 years of age or greater had decreased OS when compared to the other age groups ( p= .002). Conclusions: Gastric GIST mitotic rate is significant in term of overall survival across all age groups and the combination with necrosis portends decreased overall survival. Patients over 80 have a decreased overall survival and require additional vigilance in the early peri-operative period.

Population-based validation of the melanoma institute australia (MIA) and the memorial sloan-kettering cancer center (MSKCC) predictive tool for sentinel node status in patients with melanoma.

9571 Background: Patients with primary cutaneous melanoma are selected for sentinel lymph node biopsy (SLNB) based upon their risk of a positive SLN being identified. To improve upon stage-based categorization, the Memorial Sloan Kettering Cancer Center (MSKCC) and Melanoma Institute Australia (MIA) developed predictive models. Which model is more clinically useful is currently unclear. Methods: Consecutive patients undergoing SLNB from 2007-2021 were identified from the Swedish Melanoma Registry (SweMR), which covers 99% of patients with invasive cutaneous melanoma in Sweden. The predicted probability of SLN positivity was calculated using the MSKCC and limited MIA models (using mitotic rate as absent/present instead of count and excluding the optional variable lymphovascular invasion) for each patient. The operating characteristics of the models were assessed and compared. The “clinical utility” of each model was assessed using decision curve analysis (DCA) and compared with a strategy of performing SLNB on all patients. Results: In total 10,089 patients were included; the median Breslow thickness was 1.8 mm, 33.7% were ulcerated, and 1,802 patients had a positive SLN (17.9%). Both the MSKCC and limited MIA models were well calibrated across the full range of predicted probabilities, and had similar external area under curve (AUC), MSKCC 70.8% (95% CI 69.5-72.1%) and MIA 70.3% (95% CI 68.9-71.7%). When new models were constructed to compare the incremental value of the additional parameters of either nomogram over Breslow thickness alone, the AUCs were 70.1%, 71.6% and 72.0% for Breslow alone, MIA parameters and MSKCC parameters, respectively. At a risk threshold of 5%, DCA indicated no added net benefit for either model compared to performing SLNB for all patients. At risk thresholds of 10% or higher, both models added net benefit compared to SLNB for all. The greatest benefit was observed from use of the models in patients with T2 melanomas when a threshold of 10% was used to select patients for SLNB. In that setting, use of the nomograms led to a net reduction in seven avoidable SLNBs per 100 patients for the limited MIA nomogram and eight for the MSKCC nomogram, when compared to a strategy of SLNB for all. Conclusions: This study confirms the statistical performance of both the MSKCC and limited MIA models in a large, nationally-representative dataset. However, clinical utility as assessed by DCA demonstrated that using the models only improved selection for SLNB when a risk threshold of at least 10% was employed, and then mainly for T2 melanomas. Either further development of the nomograms, or other strategies, are needed to improve selection at lower risk thresholds.

Efficacy of adjuvant therapy in patients (pts) with AJCC v8 stage IIIA cutaneous melanoma.

9518 Background: Pts with resected AJCC v8 stage IIIA melanoma have been under-represented in clinical trials of adjuvant drug therapy. The benefit of adjuvant targeted and immunotherapy in this population is unclear. Methods: In this multicenter, retrospective study, pts with stage IIIA melanoma (AJCC v8) who received adjuvant pembrolizumab or nivolumab (PD1), BRAF/MEK-targeted therapy dabrafenib + trametinib (TT), or no adjuvant treatment (OBS) were included. Recurrence free survival (RFS), distant metastasis free survival (DMFS), and toxicity rates were examined. Results: 613 pts from 34 centers across Australia, Europe and USA were included. The median follow-up was 2.6 yrs (IQR, 1.6-3.4 yrs). Pt characteristics and follow-up were similar across the cohorts (Table). Completion rates of 12-month PD1 and TT therapy were 57.0% and 69.2% respectively. 1-yr RFS was 93.3% (95% CI, 90.3-96.4) in PD1, 100% in TT and 91.3% (95% CI, 88.1-94.7) in OBS cohorts. 2-yr RFS was 79.3% (95% CI, 74.1-84.8) for PD1, 100% for TT and 84.3% (95% CI, 79.9-89.0) in OBS cohorts. 2-yr DMFS was 88.4% (95% CI, 84.3-92.8) in PD1, 100% in TT and 91.1% (95% CI, 87.7-94.7) in OBS cohorts. Risk of recurrence was associated with higher breslow thickness (HR 1.73, 95% CI 1.36-2.20), higher mitotic rate (HR 1.07, 95% CI 1.02-1.12) and neck as the site of nodal metastases (HR 2.71, 95% CI 1.45-5.06) in the overall cohort (p<0.05). Neck nodal metastases were associated with higher risk of recurrence in OBS cohort (HR 3.82, 95% CI 1.91-7.66; p<0.05) but not in the PD1 cohort. Rates of ≥ Grade 3 toxicities were 10.9% with PD1 and 20.0% with TT; discontinuation due to toxicity occurred in 25.0% and 30.0%, respectively. No new safety signals were observed. Rates of unresolved toxicity at last follow-up were 26.9% in PD1 and 7.7% in TT cohorts. Conclusions: In this large international study, adjuvant PD1 or TT did not significantly improve RFS or DMFS compared to OBS in pts with resected stage IIIA melanoma. Prognosis in stage IIIA melanoma is favourable and outcomes after adjuvant therapy in this population needs further study in prospective randomised trials. [Table: see text]

Unusual presentation of a large GIST: a challenging diagnostic dilemma

Introduction: Gastrointestinal stromal tumors (GISTs) are rare neoplasms of the gastrointestinal tract associated withhigh rates of malignant transformation. Most GISTs present asymptomatically. They are best identified by computedtomography (CT) scan and most stain positive for CD 117 (C-Kit), CD34, and/ or DOG-1. There have been many riskstratification classifications systems which are calculated based on tumor size, mitotic rate, location and perforation. Case presentation: A 42 years old man presented with dysphagia and weight loss for 2.5 months. On examination, hewas severely anaemic having a huge intra-abdominal lump. He underwent laparotomy followed by removal of tumormass with partial left lobectomy of the Liver. Conclusion:We present a case of GIST of unusual location and presentation pattern. In general, only completeresection of tumor can lead to cure, although recurrence is common after surgery. Bangladesh Crit Care J March 2023; 11 (1): 54-60

High Grade Differentiated Follicular Cell-Derived Thyroid Carcinoma Versus Poorly Differentiated Thyroid Carcinoma: A Clinicopathologic Analysis of 41 Cases.

Criteria overlap for separating between malignant follicular epithelial cell-derived thyroid gland neoplasms with high grade features of increased mitoses and tumor necrosis but lacking anaplastic histology. Patterns of growth, nuclear features, tumor necrosis, and various mitotic index cutoffs are suggested, but a reproducible Ki-67-based labeling index has not been established. Forty-one cases diagnosed as poorly differentiated thyroid carcinoma (PDTC) or high grade differentiated follicular cell-derived thyroid carcinoma (HGDFCDTC) were reviewed, with histologic features, mitotic figure counts, and Ki-67 labeling index reviewed on cases within Southern California Permanente Medical Group from 2010 to 2021 to establish any potential outcome differences. There were 17 HGDFCDTC (nine papillary thyroid carcinoma; eight oncocytic follicular thyroid carcinoma), median age 64years, affecting nine females and eight males. Tumors were large (median, 6.0cm), usually unifocal (n = 13), with only one tumor lacking invasion. Tumor necrosis was present in all; median mitotic count was 5/2 mm2 (median Ki-67 labeling index 8.3%). Three patients had metastatic disease at presentation, with additional metastases in four patients (41.2% developed metastases); 11 were without evidence of disease (median 21.2months); with the remaining six patients alive (n = 4) or dead (n = 2) with metastatic disease (median 25.8months). Criteria associated with an increased risk of developing metastatic disease: widely invasive tumors; age ≥ 55years; male; advanced tumor size and stage; extrathyroidal extension; but not increased mitotic rate or higher labeling index. There were 24 PDTC, median age 57.5years, affecting 13 females and 11 males. Tumors were large (median, 6.9cm), with 50% part of multifocal disease, with three tumors lacking invasion. Insular/trabecular/solid architecture was seen in all tumors; tumor necrosis was present in 23; and median mitotic count was 6/2 mm2 (median Ki-67 labeling index 6.9%). Five patients had metastatic disease at presentation, with additional metastases in 3 patients (29.2% developed metastases); 16 were without evidence of disease (median, 48.1months); with the remaining 8 patients alive (n = 3) or dead (n = 5) with metastatic disease (median, 22.4months). Criteria associated with an increased risk of developing metastatic disease: widely invasive tumors; male; advanced tumor size and stage; extrathyroidal extension; but not increased mitotic rate or higher labeling index. HGDFCDTC shows tumor necrosis, a median Ki-67 labeling index of 8.3%, with a high percentage (41%) of patients developing metastatic disease. Extent of invasion (non-invasive, minimally invasive, angioinvasive, widely invasive) correlates strongly with developing metastatic disease. PDTC presents at a slightly younger age, with large tumors, often in a background of multifocal tumors, with tumor necrosis nearly always seen, a median Ki-67 labeling index of 6.9%, with 29% of patients developing metastatic disease. Separation between groups is meaningful as early metastatic disease is relatively common, but mitotic counts/labeling indices are not different between the groups nor able to potentially risk stratify development of metastatic disease.

Mitotic rate as a predictive factor for positive sentinel lymph nodes in pT1 and pT2 melanomas

Sentinel lymph node biopsy is a crucial step in the management of patients affected by melanoma. The decision whether to perform it or not is based on different histological parameters, but the mitotic rate is no longer considered a prognostic variable after the release of the 8th edition of the American Joint Committee on Cancer (AJCC) guidelines. Our objective was to investigate the risk factors that increase the chance for sentinel lymph node positivity in melanomas with a Breslow thickness of less than 2.00mm, including the mitotic count. A retrospective single-center study was performed on a homogenous cohort of 408 patients treated for cutaneous melanoma. Histological and clinical features were gathered and correlated with the increased risk for sentinel lymph node positivity by means of univariate and multivariate analyses. A statistically significant correlation between a high mitotic index and a positive sentinel lymph node was found in pT1 and pT2 patients, suggesting that in the case of pT1a melanoma with a high number of mitoses, a discussion about whether a sentinel lymph node biopsy is required should be done.

Repeat-Induced Point Mutation and Gene Conversion Coinciding with Heterochromatin Shape the Genome of a Plant-Pathogenic Fungus.

Meiosis is associated with genetic changes in the genome-via recombination, gene conversion, and mutations. The occurrence of gene conversion and mutations during meiosis may further be influenced by the chromatin conformation, similar to the effect of the chromatin conformation on the mitotic mutation rate. To date, however, the exact distribution and type of meiosis-associated changes and the role of the chromatin conformation in this context are largely unexplored. Here, we determine recombination, gene conversion, and de novo mutations using whole-genome sequencing of all meiotic products of 23 individual meioses in Zymoseptoria tritici, an important pathogen of wheat. We confirm a high genome-wide recombination rate of 65 centimorgan (cM)/Mb and see higher recombination rates on the accessory compared to core chromosomes. A substantial fraction of 0.16% of all polymorphic markers was affected by gene conversions, showing a weak GC-bias and occurring at higher frequency in regions of constitutive heterochromatin, indicated by the histone modification H3K9me3. The de novo mutation rate associated with meiosis was approximately three orders of magnitude higher than the corresponding mitotic mutation rate. Importantly, repeat-induced point mutation (RIP), a fungal defense mechanism against duplicated sequences, is active in Z. tritici and responsible for the majority of these de novo meiotic mutations. Our results indicate that the genetic changes associated with meiosis are a major source of variability in the genome of an important plant pathogen and shape its evolutionary trajectory. IMPORTANCE The impact of meiosis on the genome composition via gene conversion and mutations is mostly poorly understood, in particular, for non-model species. Here, we sequenced all four meiotic products for 23 individual meioses and determined the genetic changes caused by meiosis for the important fungal wheat pathogen Zymoseptoria tritici. We found a high rate of gene conversions and an effect of the chromatin conformation on gene conversion rates. Higher conversion rates were found in regions enriched with the H3K9me3-a mark for constitutive heterochromatin. Most importantly, meiosis was associated with a much higher frequency of de novo mutations than mitosis; 78% of the meiotic mutations were caused by repeat-induced point mutations-a fungal defense mechanism against duplicated sequences. In conclusion, the genetic changes associated with meiosis are therefore a major factor shaping the genome of this fungal pathogen.

Typical Pulmonary Carcinoid Tumours and the Relevance of Ki-67 Index: A Tertiary Centre Experience in South Australia

Pulmonary carcinoid tumours are a rare type of neuroendocrine primary lung malignancy. The WHO classifies carcinoids as typical (TC) and Atypical Carcinoid (AC) based on mitotic rate and necrosis on histology [1,2]. It is widely accepted that TC has a better overall prognosis than AC. A higher stage (the presence of nodal or metastatic disease) at presentation is correlated with poorer outcomes [3].

Improving fine needle aspiration to predict the tumor biological aggressiveness in pancreatic neuroendocrine tumors using Ki-67 proliferation index, phosphorylated histone H3 (PHH3), and BCL-2

IntroductionSurgery is the only known cure for sporadic pancreatic neuroendocrine tumors (PNETs). Therefore, the prediction of the PNETs biological aggressiveness evaluated on endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) has a significant impact on clinical management. The proliferation rate of Ki-67 in PNETs can help to predict the biological aggressiveness of the tumor. In addition, there is a relatively new proliferation marker called phosphorylated histone H3 (PHH3) that can identify and quantify dividing cells in tissue samples, which is a marker highly specific to mitotic figures. Other markers such as BCL-2 also contribute to tumorigenesis and may be involved in the differentiation of neuroendocrine cells. Materials and methodsA retrospective observational study was performed on patients undergoing surveillance for PNETs from January 2010 to May 2021. Data collection included the patients' age, sex, tumor location, tumor size in the surgical specimen, and tumor grade in FNA. The 2019 World Health Organization (WHO) classification guideline was followed to diagnose PNETs, including grade and stage. Immunohistochemical stainings for Ki-67, PHH3 and BCL-2 in PNETs were performed. ResultsAfter excluding cell blocks containing fewer than 100 tumor cells, 44 patients with EUS-FNA and surgical resection specimens were included in this study. There were 19 cases of G1 PNETs, 20 cases of G2 PNETs, and 5 cases of G3 PNETs. The grade assigned based on the Ki-67 index was higher and more sensitive than that based on the mitotic count using H&E slides in some cases of G2 and G3 PNETs. However, there was no significant difference between the mitotic count using PHH3-positive tumor cells and the Ki-67 index to grade PNETs.All grade 1 tumors (19 cases) on surgical resection specimens were correctly graded on FNA (100 % concordance rate). Within the 20 G2 PNETs, 15 cases of grade 2 on surgical resection specimens were graded correctly on FNA based on the Ki-67 index only. Five cases of grade 2 PNETs on surgical resection specimens were graded as grade 1 on FNA when using only the Ki-67 index. Three of five grade 3 tumors on surgical resection specimens were graded as grade 2 on FNA based on the Ki-67 index only. Using only FNA Ki-67 to predict PNET tumor grade, the concordance (accuracy) rate was 81.8 % in total. However, all these eight cases (5 cases of G2 PNETs and 3 cases of G3 PNETs) were graded correctly by using the Ki-67 index plus mitotic rate (using PHH3 IHC stains).Four of 18 (22.2 %) patients with PNETs were positive for BCL-2 stain. In these 4 cases positive for BCL-2 stains, 3 cases were G2 PNETs and one case was G3 PNETs. ConclusionGrade and the proliferative rate in EUS-FNA can be used to predict the tumor grade in surgical resection specimens. However, when using only FNA Ki-67 to predict PNET tumor grade, about 18 % of cases were downgraded by one level. To solve the problem, immunohistochemical staining for BCL-2 and especially PHH3 would be helpful. Our results demonstrated that the mitotic count using PHH3 IHC stains not only improved the accuracy and precision of PNET grading in the surgical resection specimens, but also could reliably be used in routine scoring of mitotic figures of FNA specimens.

Patient-reported oral mucositis in solid tumour patients undergoing chemotherapy: a Ugandan experience.

The mitotic rate of the gastrointestinal tract (GIT) mucosa predisposes the entire system to chemotherapeutic-induced mucositis but the oral cavity due to its accessibility provides an opening to evaluate the extent of the problem much more easily. In addition, the oral cavity being the gateway to the GIT affects the feeding ability of the patient when the ulcers set in.It is therefore from this perspective that we embarked on a study to evaluate the extent of mucositis among patients being treated for solid tumours at our centre. Using the mouth and throat soreness (OMDQ MTS) questionnaire, we prospectively evaluated mucositis among 100 patients undergoing chemotherapy for solid tumours at the Uganda Cancer Institute. In addition to patient reported outcomes, we also had clinician assessed mucositis measurements. Approximately, 50% of the participants were breast cancer patients. The results demonstrated that patient assessment of mucositis is possible in our setting at a 76% full compliance rate. Up to 30% of our patients reported moderate-to-severe mucositis, though the figure was lower as assessed by the clinicians. The self-reported OMDQ MTS can be useful in our setting for daily mucositis evaluation, hence leading to timely hospital visits before the manifestation of severe complications.

The Enhanced Expression of ZWILCH Predicts Poor Survival of Adrenocortical Carcinoma Patients.

Zwilch kinetochore protein (ZWILCH) plays a key role in proper cell proliferation. The upregulation of the ZWILCH gene was observed in many types of cancers, but the association of ZWILCH with adrenocortical carcinoma (ACC) was not investigated so far. The main aim of the presented study was to verify if the enhanced level of the ZWILCH gene can be used as a diagnostic marker for ACC development and progression, as well as a predictor of survival time for ACC patients. The performed analyses included investigation of the ZWILCH expression profile in tumors with publicly available TCGA (The Cancer Genome Atlas) datasets and transcriptomic data from the Gene Expression Omnibus (GEO) database, as well as, in human biological samples of normal adrenal, adrenocortical carcinoma and in commercially available tissue microarrays. The findings demonstrate statistically significant higher ZWILCH gene expression in ACC tissue in comparison with normal adrenal glands. Furthermore, there is a strong correlation between ZWILCH upregulation and tumor mitotic rate and the probability of patient survival. The enhanced ZWILCH level is also connected with the activation of genes involved in cell proliferation and the inhibition of genes related to the immune system. This work contributes to a better understanding of the role of ZWILCH as an ACC biomarker and diagnostic tool.

Does "Low-Grade" Dedifferentiated Liposarcoma Exist? The Role of Mitotic Index in Separating Dedifferentiated Liposarcoma From Cellular Well-differentiated Liposarcoma.

Subjective, varying criteria identify "low-grade" dedifferentiation in well-differentiated/dedifferentiated liposarcoma (WD/DDLPS). The value of mitotic rate (MR) in defining DDLPS is not confirmed. We studied all patients with the resection of their primary or first recurrence retroperitoneal WD/DDLPS at our institution to determine the value of MR in diagnosing DDLPS and if MR associates with patient survival. Ninety-eight patients with retroperitoneal WD/DDLPS operated at our institution from January 1, 1989 to December 31, 2013 were included. Cases were defined as acellular (AC) WDLPS, LS0-4 (tumors with non-lipogenic areas and MR 0-4/10HPFs) or LS5+(non-lipogenic areas, MR≥5/10 HPFs) and graded using the French system. Kaplan-Meier survival estimates with log-rank test and multivariate Cox (mCox) analyses were performed. Follow-up was available on all patients (median 9.3y, range 0.02-23.16y). Kaplan-Meier demonstrated a significant ( P =0.004) difference in disease-specific survival (DSS) among the 3 groups. mCox demonstrated no difference in DSS between the AC and LS0-4 groups (HR 1.51; 95% CI 0.57-3.99, P =0.412) but significantly lower DSS in the LS5+group compared with the AC group (HR 2.68; 95% CI 1.07-6.71, P =0.035). The difference in DSS was not significant between grade 2 and 3 tumors ( P =0.094). DSS between MR 5-19/10 HPFs and MR20+/10 HPFs subgroups was significant ( P =0.007) but by mCox did not reach significance (HR 2.47; 95% CI 0.96-6.35, P =0.060). This study confirms that MR distinguishes DDLPS from WDLPS with non-lipogenic areas, also known as cellular WDLPS. For consistency in diagnosis and research, only WD/DDLPS with≥5 mitoses/10 HPFs should be considered DDLPS.

Abstract LB237: Functional characterisation of TRACERx reveals mechanisms of NSCLC evolution

Abstract Chromosomal instability (CIN), the increase in the rate of whole/partial chromosome gains and losses, is a driving feature of cancer identified in ~90% of solid tumours. CIN generates intratumor heterogeneity (ITH), drives evolutionary adaptation, and is associated with highly aggressive, drug-resistant tumours and poor prognosis. Aberrations in numerous pathways have been suggested to increase CIN. However, how CIN is initiated and maintained in non-small cell lung cancer (NSCLC) is largely unknown. Here we have analysed multi-region WES sequencing data from patients of the TRACERx study, and identified novel genes involved in the DNA damage response (DDR) that may contribute to chromosomal instability. Using orthogonal methods, we have identified that genetic alteration in 6 genes, particularly FAT1, culminates in deficiencies in homologous recombination repair (HRR). This is manifested by a reduction of RAD51 and BRCA1 foci formation, reduced end-resection rate and increased 53BP1 bodies. We find that these deficiencies in the HRR pathway can lead to an increase in structural CIN and deviation from the modal cjromosome number. Furthermore, we observed an early selection of FAT1 mutations in the TRACERx421 LUSC cohort, which resulted in more genome-doubling. We reprot mirrored sub-clonal allelic imbalance (MSAI) events at the FAT1 gene locus, indicating parallel evolution. We have proceeded to validate these observations using several cell lines, showing that FAT1 loss promotes whole genome doubling (WGD) through the generation of actively replicating binucleated cells through an elevated mitotic error rate. Importantly, the elevated CIN induced by FAT1 loss could be partially ameliorated by co-depletion of the oncogene YAP1. We hypothesize that elevated CIN could synergize with WGD to generate increased intratumor heterogeneity. To investigate this, we modelled the relationship between FAT1 loss and genome doubling in an isogenic cell culture system using PC9 cells which are sensitive to the EGFR inhibitor, Osimertinib. Indeed, we observed an elevated rate of acquired resistance in FAT1 KO genome-doubled PC9 cells, which we attribute to the high CIN level generated by FAT1 loss. In line with our hypothesis, we also observed a significant increase in genome content in FAT1 KO cells, suggesting FAT1 loss induces a second WGD event to escape targeted therapy. In conclusion, FAT1 is one of the most frequently mutated genes both in the TRACERx421 cohort and in somatic tissues (Martincorena et al, Science 2015). We postulate that FAT1 loss attenuates DDR and exacerbates CIN to enhance tumour heterogeneity, early in lung cancer evolution. Our observation that FAT1 loss leads to elevated EGFRi resistance also provides a unique opportunity to understand how EGFRi resistance arises through elevated CIN. Citation Format: Wei-Ting Lu, Lykourgos-Panagiotis Zalmas, Chris Bailey, Oriol Pich, Carlos M. Ruiz, James Black, Georgia Stavrou, Dhruva Biswas, Francisco Gimeno-Valiente, Kevin Litchfield, Jiri Bartek, Nicholas McGranahan, Nnennaya Kanu, Charles Swanton. Functional characterisation of TRACERx reveals mechanisms of NSCLC evolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB237.

Quantitative Nuclear Grading: An Objective, Artificial Intelligence–Facilitated Foundation for Grading Noninvasive Papillary Urothelial Carcinoma

In nonmuscle invasive bladder cancer, grade drives important treatment and management decisions. However, grading is complex and qualitative, and it has considerable interobserver and intraobserver variability. Previous literature showed that nuclear features quantitatively differ between bladder cancer grades, but these studies were limited in size and scope. In this study, we aimed to measure morphometric features relevant to grading criteria and build simplified classification models that objectively distinguish between the grades of noninvasive papillary urothelial carcinoma (NPUC). We analyzed 516 low-grade and 125 high-grade 1.0-mm diameter image samples from a cohort of 371 NPUC cases. All images underwent World Health Organization/International Society of Urological Pathology 2004 consensus pathologist grading at our institution that was subsequently validated by expert genitourinary pathologists from 2 additional institutions. Automated software segmented the tissue regions and measured the nuclear features of size, shape, and mitotic rate for millions of nuclei. Then, we analyzed differences between grades and constructed classification models, which had accuracies up to 88% and areas under the curve as high as 0.94. Variation in the nuclear area was the best univariate discriminator and was prioritized, along with the mitotic index, in the top-performing classifiers. Adding shape-related variables improved accuracy further. These findings indicate that nuclear morphometry and automated mitotic figure counts can be used to objectively differentiate between grades of NPUC. Future efforts will adapt the workflow to whole slides and tune grading thresholds to best reflect time to recurrence and progression. Defining these essential quantitative elements of grading has the potential to revolutionize pathologic assessment and provide a starting point from which to improve the prognostic utility of grade.

Association of Excision Margin Size With Local Recurrence and Survival in Patients With T1a Melanoma at Critical Structures

Melanoma guidelines recommend surgical excision with 10-mm margins for T1 melanoma. However, this procedure may be problematic at sites close to critical structures such as the scalp, face, external genitalia, acral, periumbilical, and perineal areas. To compare outcomes of wide (10-mm margins) vs narrow (5-mm margins) excision in patients with T1a melanoma near critical structures. This cohort study was a retrospective comparison of 1341 consecutive patients aged 18 years or older from the National Cancer Institute of Milan, Italy, diagnosed between 2001 and 2020 with T1a cutaneous melanoma close to critical structures who accepted wide excision vs narrow excision. Local recurrence and melanoma-specific mortality (MSM) rates with 5-mm vs 10-mm excision margins. The primary aim of the study was to ascertain whether a narrower (5-mm) vs wider (10-mm) excision margin was associated with local recurrence and MSM. The secondary aim was to compare the need for reconstructive surgery in the groups defined by excision margin width. Between April 28 and August 7, 2022, associations were assessed by weighted Cox and Fine-Gray univariable and multivariable models. A total of 1179 patients met the inclusion criteria (median [IQR] age, 50.0 [39.5-63.0] years; female, 610 [51.7%]; male, 569 [49.3%]). Six hundred twenty-six patients (53.1%) received a wide excision (434 [69.3%] with linear repair and 192 [30.7%] with flap or graft reconstruction) and 553 (46.9%) received a narrow excision (491 [88.8%] with linear repair and 62 [11.2%] with flap or graft reconstruction). The weighted 10-year MSM was 1.8% (95% CI, 0.8%-4.2%) in the wide group and 4.2% (95% CI, 2.2%-7.9%) in the narrow group; the weighted 10-year local recurrence rate was 5.7% (95% CI, 3.9%-8.3%) in the wide group and 6.7% (95% CI, 4.7%-9.5%) in the narrow group. Breslow thickness greater than 0.4 mm (subdistribution hazard ratio [sHR] for 0.6 vs 0.4 mm, 2.42; 95% CI, 1.59-3.68; P < .001) and mitotic rate greater than 1/mm2 (sHR for a single increment, 3.35; 95% CI, 2.59-4.32; P < .001) were associated with worse MSM. Multivariable analysis showed that acral lentiginous melanoma, lentigo maligna melanoma, and increasing Breslow thickness were associated with a higher incidence of local recurrence. The study's findings suggest that local excision with 5-mm margins for T1a melanoma may not be associated with an increased risk of local recurrence. Breslow thickness greater than 0.4 mm, mitotic rate greater than 1/mm2, and acral lentiginous melanoma and lentigo maligna melanoma subtypes were associated with a higher risk of recurrence. These findings may be useful for future melanoma treatment guidelines.

Abstract 5423: AI powered quantification of mitotic rate in H&amp;E stained tissue detects significant differences between treatment groups of preclinical pancreas cancer xenografts

Abstract Background: Mitotic rate is a readout routinely used for characterization of tumor samples. Standard methods to quantify cell division include manual pathologist counts based on hematoxylin and eosin (H&amp;E) staining and either manual or automated assessment of immunohistochemistry (IHC) staining using phospho-histone H3 (pHH3). These suffer the drawbacks of high inter-pathologist variability in the case of H&amp;E assessment and time inefficiency and false positive calls in case of pHH3 staining. Aiming to overcome these issues, we developed a mitosis detection model based on H&amp;E-stained tissue sections alone. Methods: To develop and evaluate the model, we used 1032 H&amp;E-stained tissue sections (156 used for model training) of pre-clinical pancreas cancer xenografts originating from mice that have undergone a series of experiments conducted to examine the pharmacodynamic effect of several anti-cancer protocols. We trained (i) a tissue segmentation model (segmenting tissue regions into ‘carcinoma’, ‘stroma’, ‘necrosis’, and ‘other’) and (ii) a segmentation model for pixel-level mitosis detection (segmenting ‘mitosis’ vs. ‘non-mitosis’). Regions predicted as mitosis were post-processed to represent individual dividing cells. The tissue segmentation model served as a filter to predict mitotic rate for carcinoma tissue areas only, which has not been accounted for in previous AI-based methods for mitotic rate prediction on H&amp;E tissue. To evaluate the model on detecting mitotic events, the model was compared against a 5-pathologist consensus of mitotic count. The mitotic rates predicted by the model were used to infer differences between treatment groups (various treatments and dosages vs. control). Results: The mitosis detection model for quantifying rates of cell division in carcinoma regions of H&amp;E-stained tissue sections showed a notable agreement with the 5-pathologist mitotic count consensus (Pearson correlation 0.92). Furthermore, the model correlated well with mitosis counts based on pHH3 IHC staining (Pearson correlation 0.78), which were available for 63 cases. Finally, when used to characterize the entire CDX cohort, the case level mitotic rate predicted by the model showed significant differences between treatment groups in line with or better than using a pHH3 IHC stain. Conclusion: This study demonstrates the efficacy and scalability of AI-based models for the quantification of mitotic rate based on H&amp;E-stained tissue alone, presenting a time- and cost-efficient approach that also mitigates inter-annotator variability. Citation Format: Sharon Ruane, Lukas Ruff, Brian Reichholf, Christina Aigner, Emil Barbuta, Stephan Tietz, Olivér Atanaszov, Rosemarie Krupar, Simon Schallenberg, Maximilian Alber, Francesca Trapani, Frederick Klauschen. AI powered quantification of mitotic rate in H&amp;E stained tissue detects significant differences between treatment groups of preclinical pancreas cancer xenografts. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5423.

Novel treatment strategy of targeting epigenetic dysregulation in pancreatic neuroendocrine tumors.

Epigenetic dysregulation is an integral step in the progression of pancreatic neuroendocrine tumors. We hypothesized that tumor suppressor repression by DNA methyltransferase 1 in pancreatic neuroendocrine tumors could be targeted with epigenetic treatment. Resected pancreatic neuroendocrine tumors from 32 patients were stained for DNA methyltransferase 1 and scored. Human (BON1) and murine (STC) pancreatic neuroendocrine tumor cells were treated with DNA methyltransferase 1 inhibitor 5-azacytidine and chemotherapeutic agents 5-fluorouracil and temozolomide. Cell proliferation assay and tumor suppressor gene analysis were performed with qRT-PCR and Clarion S microarray. Tumor measurements were compared in a murine treatment model. High DNA methyltransferase scores were associated with high Ki-67 (6.7% vs 70.6% P < .01), mitotic rate (0.0% vs 31.3%), and grade (20.0% vs 80.4%, P < .01). Treatment with 5-azacytidine and chemotherapy resulted in a reduction of cell proliferation compared to chemotherapy alone in BON1 (77.3% vs 53.1%, P < .001) and STC (73.4% vs 34.2%, P < .001). Treatment with 5-azacytidine and chemotherapy resulted in upregulation of tumor suppressors CDKN1A (7.6 rel. fold, P < .001), BRCA2 (4.3 rel. fold, P < .001), and CDH1 (6.0 rel. fold, P= .026) in BON1 and CDKN1a (14.5 rel. fold, P < .001) and CDH (17.5 rel. fold, P < .001) in STC. In microarray, 5-azacytidine drove global genetic changes in combination treatment. Invivo tumors treated with chemotherapy measured 88.6 ± 19.54 mm3 vs 52.89 ± 10.51 mm3 in those treated with combination therapy (P= .009). Epigenetic dysregulation with DNA methyltransferase 1 is associated with pancreatic neuroendocrine tumors and is a potential targetable strategy. 5-azacytidine and chemotherapy in combination can reduce cell proliferation, upregulate silenced tumor suppressor genes, and decrease invivo tumors in pancreatic neuroendocrine tumors.

327 Radiopathomics: Integration of Advanced Neuroimaging and Molecular Pathology Features in Meningiomas

OBJECTIVES/GOALS: Our overall objective is to investigate the relationship between radiologic features of meningioma with recently identified histopathological and molecular biomarkers, and to apply a machine learning (ML) approach to further demonstrate their utility in predicting clinical outcomes. METHODS/STUDY POPULATION: We have enrolled a cohort of 84 patients with meningioma diagnosed on the basis of conventional gadolinium-enhanced MRI imaging features since September 2019. Each patient has demographic and clinical data, Ga-68-DOTATATE MRI/PET SUV and dynamic metrics, DCE-MRI perfusion parameters, and histopathologic data. Various tumor subregions will be segmented semi-automatically and later confirmed by experienced neuroradiologist. Histopathologic data will include histologic grade, mitotic rate, Ki67 proliferative index, and presence of WHO established atypical histologic features, immunohistochemical parameters, and established high-grade molecular features. We will use supervised learning techniques to develop algorithms for predicting molecular features from imaging phenotypes. RESULTS/ANTICIPATED RESULTS: Anticipated results - advancements in understanding the molecular biomarkers of meningiomas has uncovered genetic alterations and epigenetic changes that more accurately determine tumor behavior. Currently, the imaging correlates of these molecular biomarkers are unknown, and utilizing radiographic data to predict prognosis and imaging-based classifications of meningiomas have not yet been investigated. Validated imaging correlates of molecular biomarkers not only provide an in-vivo assessment of tumor biology, but can also be integrated with histopathologic features ( radiopathomics models’) for more accurate disease prognostication. We anticipate that our results will identify surrogate imaging features for some of the recently emerged molecular biomarkers of meningioma. DISCUSSION/SIGNIFICANCE: There is a paucity of data on the importance of imaging phenotypes in determining tumor biology. This work has the potential of significant clinical impact by enabling a priori molecular characterization of meningiomas at the time of new diagnosis or recurrence, thereby allowing a personalized medicine approach to treatment planning.

A STUDY ON RETROSPECTIVE ANALYSIS OF GASTROINTESTINAL STROMA TUMOURS.

Introduction:.Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal neoplasms of the gastrointestinal (GI) tract. They are believed to originate from the interstitial cells of Cajal (ICCs) or from the precursors of ICCs. Most GISTs show an activating mutation in either the c-kit or platelet-derived growth factor receptor alpha (PDGFRA) gene. Tumor size, mitotic rate, and anatomic location correlate with potential malignancy and recurrence rate. The aim of our study is to Aim &amp; Objectivesobserve the demographic and clinical characteristics of GIST in our clinical environment.Method- 30 patients who are diagnosed with GIST managed during the period from Jun 2017 to July 2022 were reviewed in terms of demographics, clinical presentation, location of tumor, characteristics, management, histopathology, IHC analysis, post op follow up &amp; recurrence.The risk stratication in terms of size, mitotic index, site of tumour was observed &amp; classied using modied NIH classication. Results- Out of 30 cases male patient's- 73% (n- 22); female patients27%(n-8). Mean age was 60 years with a range of 44-81 years. Most common clinical presentation was abdominal mass, Abdominal pain. Commonly involved sites are stomach and small bowel.4 cases who underwent multiple organ resection 2 cases developed recurrence in which 1 underwent palliative Whipple's procedure and one was medically managed. GIST Conclusion- s are rare entities with a variety of clinical features, Common sites for GIST are stomach &amp; small intestine. Surgery is the main stay of management in GIST. Neo adjuvant therapy will help in down staging the tumour. Wedge resection &amp; resection &amp;anastomosis will be adequate for stomach in small bowel GIST. En-bloc resection should be considered if adjacent structures are involved. Overall GIST has favourable prognosis if diagnosed early

MicroRNA-155 and Disease-Related Immunohistochemical Parameters in Cutaneous Melanoma

Cutaneous melanoma is a severe and life-threatening form of skin cancer with growing incidences. While novel interventions have improved prognoses for these patients, early diagnosis of targeted treatment remains the most effective approach. MicroRNAs have grown to good use as potential biomarkers for early detection and as targets for treatment. miR-155 is well-studied for its role in tumor cell survival and proliferation in various tissues, although its role in melanoma remains controversial. In silico data analysis was performed in the dbDEMC v.3 to identify differentially expressed miRNA. We validated gene targets in melanoma using TarBase v8.0 and miRPath v3.0 and determined protein-protein interactions of the target genes. One hundred forty patients (age range 21-90 years) with cutaneous melanoma who underwent resection were included. Molecular assessment using Real-Time RT-qPCR, clinicopathological associations, and a literature review for the different roles of miR-155 in melanoma were performed. Analysis of the dbDEMC reveals controversial findings. While there is evidence of upregulation of miR-155 in primary and metastatic melanoma samples, others suggest decreased expression in later-stage melanoma and cases with brain metastasis. miR-155 has been overexpressed in prior cases of melanoma and precancerous lesions, and it was found to be dysregulated when compared to benign nevi. While miR-155 expression was associated with favorable outcomes in some studies, others showed an association with metastasis. Patients with high levels of miR-155 also noted reduction after receiving anti-PD-1 treatment, correlated with more prolonged overall survival. In our patient's cohort, 22.9% relapsed during treatment, and 45% developed recurrence, associated with factors such as lymph node infiltration, high mitotic index, and positive staining for CD117. Although overall analysis revealed miR-155 downregulation in melanoma specimens compared to non-cancer tissues, increased expression of miR-155 was associated with cases of superficial spreading melanoma subtype (p = 0.005) and any melanoma with a high mitotic rate (p = 0.010). The analysis did not identify optimum cutoff values to predict relapse, recurrence, or mortality. In conclusion, miR-155 could have, in part, a potential prognostic utility in cutaneous melanoma. Further mechanistic studies are required to unravel the multifunctional role of miR-155 in melanoma.