Ki-67 Score Research Articles

Abstract PD13-06: Neoadjuvant giredestrant (GDC-9545) + palbociclib versus anastrozole + palbociclib in postmenopausal women with estrogen receptor-positive, HER2-negative, untreated early breast cancer: Primary analysis of the randomized, open-label, phase II coopERA breast cancer study

Abstract Background Endocrine therapy, the therapeutic mainstay for estrogen receptor-positive breast cancer, targets estrogen receptor activity and/or estrogen synthesis. CDK4/6 inhibitors cause cell cycle arrest and significantly decrease expression of the proliferation biomarker Ki67 when used in conjunction with aromatase inhibitors such as anastrozole. Giredestrant, a highly potent, nonsteroidal, oral, selective estrogen receptor antagonist and degrader, achieves robust estrogen receptor occupancy, is well tolerated, and has encouraging antitumor activity as a monotherapy and in combination with the CDK4/6 inhibitor palbociclib in metastatic breast cancer. coopERA Breast Cancer (NCT04436744) is a phase II study investigating 2 weeks of giredestrant versus anastrozole in a window-of-opportunity phase, followed by 4 months of giredestrant plus palbociclib versus anastrozole plus palbociclib in a neoadjuvant phase in postmenopausal women with estrogen receptor-positive, HER2-negative, untreated early breast cancer. We will report the results of the primary analysis. Methods Eligible patients who had measurable cT1c (≥1.5 cm)-cT4a-c estrogen receptor-positive, HER2-negative, untreated early breast cancer and baseline Ki67 score ≥5% were randomized 1:1 to 1 mg oral, daily anastrozole or 30 mg oral, daily giredestrant on Days 1-14 (window-of-opportunity phase lasting 14 days) followed by daily dosing for four 28-day cycles in combination with 125 mg oral palbociclib on Days 1-21 (neoadjuvant phase lasting 16 weeks) before surgery. Patients were stratified according to T status, Ki67 score, and progesterone receptor status. The primary efficacy endpoint was centrally assessed geometric mean relative Ki67 score change from baseline to Week 2 during the window-of-opportunity phase, which is reflective of the ability of endocrine therapies to suppress tumor-cell proliferation, and is a surrogate marker for clinical outcomes. The secondary efficacy endpoint is complete cell cycle arrest rate (CCCA), defined as Ki67 score ≤2.7%, at Week 2. Safety was also assessed. Results Results of a previous interim analysis (including 83 of the planned 202 patients) demonstrated a greater relative reduction of Ki67 at 2 weeks with giredestrant (reduction from baseline to Week 2 geometric mean = 80%; 95% CI = -85%, -72%) compared with anastrozole (reduction from baseline to Week 2 geometric mean = 67%; 95% CI = -75%, -56%; P = 0.0222). Similarly, consistent Ki67 suppression was observed in patients with baseline Ki67 ≥20% (83% reduction with giredestrant versus 71% reduction with anastrozole) or <20% (65% versus 24% reductions). At Week 2, 25% of tumors exhibited CCCA with giredestrant versus 5.1% with anastrozole (a 20% difference; 95% CI = -37%, -3%). Safety results were consistent with the known safety profile for giredestrant. Fewer patients experienced adverse events (AEs) related to giredestrant (28%) than to anastrozole (38%), and no grade ≥3 AEs or serious adverse events were assessed as related to giredestrant. We will present the results of the primary analysis, which will include data from all enrolled patients, and will report the primary and secondary efficacy endpoints (including patients with Ki67 >20%), and updated safety. Conclusions The study will proceed to the primary analysis. We expect to see encouraging results based on the favorable interim analysis data that demonstrated the superior activity of giredestrant, an oral selective estrogen receptor antagonist and degrader, compared with anastrozole. Citation Format: Sara A Hurvitz, Vanesa Quiroga, Yeon Hee Park, Aditya Bardia, Vanesa López-Valverde, Jutta Steinseifer, Tharu M Fernando, Gonzalo Spera, Cloris Xue, Peter A Fasching. Neoadjuvant giredestrant (GDC-9545) + palbociclib versus anastrozole + palbociclib in postmenopausal women with estrogen receptor-positive, HER2-negative, untreated early breast cancer: Primary analysis of the randomized, open-label, phase II coopERA breast cancer study [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD13-06.

Abstract P1-02-17: Artificial intelligence-based whole slide scoring of nuclear breast cancer IHC markers Ki67, ER, and PR matches performance of manual clinical scoring

Abstract Background: 2.1 million breast cancers are newly diagnosed each year. Current guidelines endorse routine testing for estrogen receptor (ER) and progesterone receptor (PR), while use of the Ki67 biomarker can provide additional prognostic value. All three biomarkers currently require quantitative evaluation using manual review of a glass slide, resulting in reproducibility issues across labs due to interpretative and scoring variabilities. Current on-market image analysis algorithms only offer limited field-of-view (FOV) that analyze a tiny fraction of the entire tissue. Whole slide image (WSI) analysis, in comparison, analyzes the entire tissue and, therefore, more closely mimics how pathologists assess these slides in clinical practice. In this work, we developed three deep learning artificial intelligence (AI) based algorithms for WSI analysis (IA) of digitized images from Ki67, ER, and PR stained slides that address these variabilities and allow pathologists across labs to consistently score at the same accuracy as selected expert labs. The complete software solution delivers high throughput analyzing whole-slide images in less than 2 minutes during pre-computation on conventional computer hardware and returning results on user-provided annotations in milliseconds. Methods: We assembled a benchmark validation data set of 312 breast cancer cases (100 Ki67, 102 ER, and 110 PR slides, stained at multiple sites) representative of breast cancer subtypes (i.e. ductal, lobular, mucinous, medullary, tubular), score (i.e. 0%-100% positivity), tumor grade (i.e. well, moderately, and poorly differentiated), and specimen type (i.e. biopsy and resection). Three pre-clinical validation studies were performed using the Roche uPath enterprise software and each of the ER, PR and Ki67 image analysis algorithms. A total of 6 pathologists participated in the study split into expert (n=3) and study (n=3) readers. A non-inferiority Ground-Truth (GT) study design was implemented in which the study and expert readers performed manual read (MR) followed by AI-assisted scoring. The expert manual scores were used as GT to which the readers’ manual and AI scores were compared for each marker and case. Results: The overall concordance rates between AI scores and expert GT was as follows: For Ki67, OPA=97.2% (95% CI: 94.0, 99.7), NPA=97.8% (95% CI: 93.4,100), and PPA=96.7% (95% CI: 91.3, 100), for ER, OPA=95.4% (CI:91.4,98.4), NPA=96.4% (CI:92.5,99.4), and PPA=94.4% (CI:87.4,100), and for PR, OPA=96.1% (95% CI:92.7,99.1), NPA=96.7% (95% CI:92.5,100), and PPA=95.6% (95% CI:89.9,100). The differences between AI and MR overall concordance rates (AI-MR) when compared to the expert GT were: for Ki67: OPA-diff=1.4% (2-sided 95% CI:-0.7,3.7), NPA-diff=3.8% (CI:0.6,7.8), PPA-diff=-1.0% (CI:-3.5,0.0), for ER: OPA-diff=-0.9% (CI:-3.3,1.0), NPA-diff=-0.1% (CI:-3.1,3.0), PPA-diff=-1.8% (CI:-6.2,0.0), and for PR: OPA-diff=-1.5% (CI:-3.9,0.6), NPA-diff=-2.4% (CI:-6.8,1.2), PPA-diff= -0.7%(CI:-2.8,1.1) using the cutoffs 20% (Ki67), 1% (ER), and 1% (PR) respectively. Conclusion: Our preliminary feasibility data shows that pathologists using WSI analysis assisted scoring was equivalent to manual scoring and an expert panel GT using a truly representative benchmark data set. Additionally, image analysis algorithms are known to provide high reproducibility and precision. We will provide those numbers at a later stage as they were not fully available at time of submission. Our results show the value and potential of deep learning technologies to improve the diagnosis and care of patients with breast cancer. Citation Format: Monesh Kapadia, Mehrnoush Khojasteh, Margarita Kouzova, Carol Jones, Xiao-Meng Xu, Matthew T. Olson, Sarah Gladden, Nancy Sapanara, Shalini Singh, Chen Chun Chen, Isaac Bai, Jim Ranger-Moore, Landon J. Inge, Uday Kurkure, Ipshita Bhattacharya, Margaret Zhao, Karel Zuiderveld, Chandana Chintakindi, Bryan Lopez, Christoph Guetter. Artificial intelligence-based whole slide scoring of nuclear breast cancer IHC markers Ki67, ER, and PR matches performance of manual clinical scoring [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P1-02-17.

Abstract P4-04-08: Determination of PD-L1 expression on tumorspheres cultured from the peripheral blood in patients with breast cancer

Abstract Background Cancer stem cells (CSCs) are the critical subpopulation of cancer cells involved in tumor formation, progression, metastasis and drug resistance. CSCs also have been considered as the main cause of cancer recurrence. The exact role of CSCs in escape of cancer from immunosurveillance is still unknown, but recent studies revealed that enhanced PDL-1 expression in cancer stem cells is linked to immune evasion and could crucially contribute to the maintenance of CSC self-renewal. Understanding the mechanisms behind this PDL-1 overexpression in cancer stem cells is critical for developing more effective anti-PD-1/PD-L1 therapy. Therefore the aim of the study was to determine the number of tumorspheres and expression of PD-L1 on tumorspheres cultured from the peripheral blood in patients with breast cancer.Methods:110 patients with breast cancer in different stages of disease were included in this study. The determination of circulating tumor cells and circulating cancer stem cells (cCSC) was performed using maintrac and tumorsphere forming assay, respectively. Additionally anti-PDL-1 antibody staining was applied to examine PDL-1 expression on breast tumorspheres.Results:We developed an innovative in vitro platform for cultivation of CSCs from peripheral blood of solid cancer patients and confirmed for the first time that tumorspheres are positive for PDL-1 in breast cancer patients. Our data also show a considerable heterogeneity in the PDL-1 expression of tumorspheres. We confirmed once again that the number of tumorspheres increased significantly with tumor progression, especially with presence of metastasis. Patients with metastatic disease had statistically more tumorspheres as compared to patients without metastasis (30 vs 10/100µl blood, p<0.05). Patients with multiple metastasis had more tumorspheres compared to patients with single metastases (60 vs 30/100µl blood, p<0.05). The number of tumorspheres was positively correlated with proliferating index Ki-67 and grade score in primary breast tumor. Moreover, patients with HER2 positive tumors had more tumorspheres compared to patients with HER2 negative primary tumors (35 vs 10/100 µl blood, p<0.005). Conclusion:The number of tumorspheres cultured from peripheral blood directly reflects aggressiveness and proliferation capacity of primary tumor. The presence of tumorspheres with expression of PDL-1 might suggest their immunregulatory potential, but further studies are needed to validate the impact of PDL-1 expression on tumorspheres in patients with breast cancer. Citation Format: Monika Pizon, Dorothea Schott, Ulrich Pachmann, Katharina Pachmann. Determination of PD-L1 expression on tumorspheres cultured from the peripheral blood in patients with breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P4-04-08.

Abstract P4-05-01: Results from the first UK NEQAS ICC & ISH external quality assessment for Ki-67 demonstration in breast cancer

Abstract AIMS: We previously described use of digital image analysis (DIA) to improve objectivity and reproducibility in external quality assessment (EQA) for immunocytochemical (ICC) staining in pathology [1]. Here we extend the work and report on: (1) use in EQA of the proliferation marker, Ki-67 in breast cancer, (2) use of a commercial prototype cell-line microarray (CLMA). MATERIALS AND METHODS: 34 laboratories stained slides provided by UK NEQAS using their routine ICC methods and returned them for central review and analysis. Slides were provided in duplicate; 29 laboratories returned both and 5 one slide (N = 63). Centrally performed DIA used the Cognition Master Professional Suite (CogM, VMscope GmbH) to quantify Ki67 labelling index (LI) [2]. Qualitative assessment aimed to separate submissions that were not useful from those which could be reported. This followed UK NEQAS standard methods. Materials assessed comprised two different breast cancer samples and a Ki-67 CLMA (Karpas Prototype, Array Science). RESULTS: Descriptive statistics on Ki-67 scores for each sample are presented in Table 1. Analysis of individual sample types for difference in Ki-67 expression associated with primary antibody showed the following: mean Ki-67 LI on breast cancer A had a range of 13.6% (K2, Leica Biosystems) to 17.6% (30-9, Ventana Medical Systems), but no significant difference was found for any primary antibody clone. For breast cancer sample B, the range was 2.0% (MIB-1, Dako Agilent) to 3.2% (30-9), and the difference was significant at P<0.05. In contrast, the CLMA showed no significant difference in expression for either MIB-1 or 30-9 (K2 could not be reported).The Ki-67-positive Karpas cell line in the CLMA had a small nuclear size compared to that of ‘standard’ breast cancer, which led to undercalling by CogM and potentially other DIA software; it also demonstrated a restricted dynamic range of high staining intensity, which reduced its ability to demonstrate differential staining in response to methodological variations.When assessed qualitatively 11.1% of submissions were not reportable (fail), 12.7% were sub-optimal but were scoreable (borderline), 23.8% had minor defects (adequate) and 52.4% were optimally stained (good). For both breast cancer samples, but not for the CLMA these qualitative categories were associated with increasing Ki-67 LI scores. For breast cancer A the mean Ki-67 LI score for the non-reportable group was significantly different from all other categories, which were not significantly different from each other; for breast cancer B, significant differences were seen between the non-reportable group and the adequate and good groups only (all at P<0.05). CONCLUSIONS: Centrally performed DIA of Ki-67 LI on breast cancer tissue samples produces additional information that allows the identification of statistically significant differences between primary antibodies (MIB-1 and 30-9). It also validates qualitative assessment by demonstrating significant differences between qualitative categories and is likely to be useful to identify lab-specific differences in staining performance in future rounds of EQA.The prototype CLMA produced using the Karpas cell-line did not demonstrate statistically significant differences between primary antibody clones or qualitative categories and is not a useful tool for use in EQA.REFERENCES: 1. Warrick N, et al. UK IBCS 2020. Breast Cancer Res Treat 180, 527-596 (2020)2. http://vmscope.com/index BC_ABC_BCLMA_0%CLMA_5%CLMA_10%CLMA_20%CLMA_30%Number of values124125164165110165165Minimum0.30.00.00.01.93.30.025% Percentile11.31.00.01.74.07.613.5Median15.22.20.02.35.09.416.375% Percentile20.03.50.23.26.111.519.8Maximum32.212.112.09.111.721.335.995% CI of medianActual confidence level96.2%95.1%96.5%95.7%95.5%95.7%95.7%Lower confidence limit14.01.80.02.04.99.115.5Upper confidence limit16.62.60.02.65.59.817.1Mean15.72.50.42.65.510.017.1Std. Deviation6.52.11.71.42.23.35.7Std. Error of Mean0.60.20.10.10.20.30.4Lower 95% CI of mean14.52.20.22.45.09.516.2Upper 95% CI of mean16.82.90.72.85.910.518.0 Citation Format: Andrew Dodson, Dawn Wilkinson, Mitch Dowsett, Suzanne Parry. Results from the first UK NEQAS ICC & ISH external quality assessment for Ki-67 demonstration in breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P4-05-01.

MCM6 versus Ki-67 in diagnosis of luminal molecular subtypes of breast cancers

BackgroundCurrently, breast cancers are divided into four major molecular subtypes. The distinction between the luminal A and luminal B subtypes is mainly based on the cellular proliferation indices and is assessed by the Ki-67 scoring. Due to the limitations in the assessment and expression of Ki-67, we hypothesized that minichromosome maintenance protein 6 (MCM6) might be taken as a surrogate marker to differentiate molecular subtypes and aid in more precise grading of tumors.MethodsWe performed a retrospective, cross-sectional study on 124 samples of breast cancer and 40 samples of normal breast tissue. Relevant clinical information was retrieved from the Cancer Institute database.ResultsMCM6 could discriminate between various categories of histologic grades, tubule formation, mitotic indices, and nuclear pleomorphism (P = 0.002 for tubule formation and P < 0.001 for other). Moreover, the MCM6 score exhibited a significant correlation with the mitotic count (P < 0.001). However, the Ki-67 score could not discriminate subgroups of the mitotic index and nuclear pleomorphism. Compared to the luminal A subtype, luminal B exhibited a higher MCM6 score (P = 0.01). Besides, MCM6 scores were higher for certain subtypes with more aggressive behaviors, such as hormone receptor (HR)-negative disease, and human epidermal growth factor receptor 2 (HER2)-enriched and triple-negative breast cancers, as there was a significantly higher MCM6 mean score in the HR-negative in comparison to the luminal breast cancers (P < 0.001). Similarly, higher MCM6 scores were observed among samples with more advanced nuclear grades, tubule formation, and overall grades.ConclusionMCM6 can differentiate luminal A and luminal B subtypes and is correlated with mitotic counts. However, this study was unable to prove the superiority of MCM6 in differentiating between molecular subtypes compared to the Ki-67 score. Nevertheless, in our study, MCM6 was superior to Ki-67 in exhibiting correlations with the mitotic grade, tubule formation, and nuclear grades. More studies are needed to standardize its assessment methods, determine more robust cut-off values, and evaluate its associations with prognostic features of breast cancer.

FDA Approval Summary: Abemaciclib With Endocrine Therapy for High-Risk Early Breast Cancer.

The US Food and Drug Administration approved abemaciclib in combination with endocrine therapy (ET) for the adjuvant treatment of adult patients with hormone receptor-positive, human epidermal growth factor receptor 2-negative, node-positive, early breast cancer (EBC) at high risk of recurrence and a Ki-67 score ≥ 20%. The approval was based on monarchE, a phase III, open-label, 2-cohort, multicenter trial of patients with EBC randomly assigned to receive abemaciclib plus ET (n = 2,808) or ET alone (n = 2,829). Abemaciclib was given at 150 mg orally twice daily for 2 years. Invasive disease-free survival (IDFS) in the intent-to-treat population was statistically significant at the second IDFS interim analysis (IA; March 2020; hazard ratio [HR; 95% CI], 0.747 [0.598 to 0.932]; P = .0096); however, only 12.5% of patients had completed adjuvant therapy, and the HR for overall survival (OS) was > 1. A prespecified, controlled analysis of IDFS in patients with Ki-67 ≥ 20% in cohort 1 was statistically significant at the final IDFS analysis (July 2020; HR [95% CI], 0.643 [0.475 to 0.872]; P = .0042). At the first OS IA (April 2021), the majority of patients had completed adjuvant therapy, IDFS remained consistent, and potential detriment in OS was not observed for this subgroup (HR [95% CI], 0.767 [0.511 to 1.152]). The HR for OS in the intent-to-treat population at OS IA remained > 1 (HR [95% CI], 1.091 [0.818 to 1.455]). More patients in the abemaciclib plus ET arm experienced treatment emergent adverse events (all grades 98.4% v 88.8%, grade 3 ≥ 49.7% v 16.3%). The approval of abemaciclib in adjuvant EBC was limited to patients with high risk of recurrence and Ki-67 ≥ 20%, given their favorable benefit:risk with a statistically significant IDFS advantage and no observed detriment on survival.

Visual Counting and Automated Image-analytic Assessment of Ki-67 and their Prognostic Value in Synovial Sarcoma.

Ki-67 is a widely used proliferation marker reflecting prognosis in various tumors. However, visual assessment and scoring of Ki-67 suffers from marked inter-observer and intra-observer variability. We aimed to assess the concordance of manual counting and automated image-analytic scoring methods for Ki-67 in synovial sarcoma. Tissue microarrays from 34 patients with synovial sarcoma were immunostained for Ki-67 and scored both visually and with 3DHistech QuantCenter. The automated assessment of Ki-67 expression was in good agreement with the visually counted Ki-67 (r Pearson =0.96, p<0.001). In a Cox regression model automated [hazard ratio (HR)=1.047, p=0.024], but not visual (HR=1.063, p=0.053) assessment method associated high Ki-67 scores with worse overall survival. The automated Ki-67 assessment method appears to be comparable to the visual method in synovial sarcoma and had a significant association to overall survival.

Relationship between 18F-FDG PET/CT Semi-Quantitative Parameters and International Association for the Study of Lung Cancer, American Thoracic Society/European Respiratory Society Classification in Lung Adenocarcinomas

ObjectiveTo investigate the relationship between 18F-FDG PET/CT semi-quantitative parameters and the International Association for the Study of Lung Cancer, American Thoracic Society/European Respiratory Society (IASLC/ATS/ERS) histopathologic classification, including histological subtypes, proliferation activity, and somatic mutations.Materials and MethodsThis retrospective study included 419 patients (150 males, 269 females; median age, 59.0 years; age range, 23.0–84.0 years) who had undergone surgical removal of stage IA–IIIA lung adenocarcinoma and had preoperative PET/CT data of lung tumors. The maximum standardized uptake values (SUVmax), background-subtracted volume (BSV), and background-subtracted lesion activity (BSL) derived from PET/CT were measured. The IASLC/ATS/ERS subtypes, Ki67 score, and epidermal growth factor/anaplastic lymphoma kinase (EGFR/ALK) mutation status were evaluated. The PET/CT semi-quantitative parameters were compared between the tumor subtypes using the Mann–Whitney U test or the Kruskal–Wallis test. The optimum cutoff values of the PET/CT semi-quantitative parameters for distinguishing the IASLC/ATS/ERS subtypes were calculated using receiver operating characteristic curve analysis. The correlation between the PET/CT semi-quantitative parameters and pathological parameters was analyzed using Spearman’s correlation. Statistical significance was set at p < 0.05.ResultsSUVmax, BSV, and BSL values were significantly higher in invasive adenocarcinoma (IA) than in minimally IA (MIA), and the values were higher in MIA than in adenocarcinoma in situ (AIS) (all p < 0.05). Remarkably, an SUVmax of 0.90 and a BSL of 3.62 were shown to be the optimal cutoff values for differentiating MIA from AIS, manifesting as pure ground-glass nodules with 100% sensitivity and specificity. Metabolic-volumetric parameters (BSV and BSL) were better potential independent factors than metabolic parameters (SUVmax) in differentiating growth patterns. SUVmax and BSL, rather than BSV, were strongly or moderately correlated with Ki67 in most subtypes, except for the micropapillary and solid predominant groups. PET/CT parameters were not correlated with EGFR/ALK mutation status.ConclusionAs noninvasive surrogates, preoperative PET/CT semi-quantitative parameters could imply IASLC/ATS/ERS subtypes and Ki67 index and thus may contribute to improved management of precise surgery and postoperative adjuvant therapy.

A narrative review of the clinical development of CDK4/6 inhibitor abemaciclib in breast cancer

Background and Objective: Advanced or metastatic breast cancer (MBC) is associated with poor prognosis and presents many challenges in medical management and treatment decisions. Anticancer drugs that act on cell cycle mechanisms have shown great potential in preclinical studies. In clinical trials, abemaciclib, a reversible ATP-competitive cyclin-dependent kinase 4/6 (CDK4/6) inhibitor developed by Eli Lilly and Company, combined with endocrine therapy (ET) were associated with superior outcomes compared with ET alone in patients with advanced or metastatic hormone receptor positive (HR+)/human epidermal growth factor receptor 2 negative (HER2−) breast cancer, representing a new standard-of-care in this population. Abemaciclib has been approved by the U.S. Food and Drug Administration (FDA) for use in HR+/HER2− MBC. In China, abemaciclib was also approved by the National Medical Products Administration (NMPA) based on findings from the MONARCH plus trial. Recently, abemaciclib have been approved as the first and only CDK4/6 inhibitor by FDA and NMPA for use in HR+/HER2−, node-positive, early breast cancer (EBC) at high risk of recurrence and Ki-67 score ≥20%. Further trials of abemaciclib are ongoing. This is an overview of the clinical development of abemaciclib in breast cancer.

Prognostic impact of PDGFRA gain/amplification and MGMT promoter methylation status in patients with IDH wild-type glioblastoma.

BackgroundPlatelet-derived growth factor receptor alpha (PDGFRA) is the second most frequently mutated tyrosine kinase receptor in glioblastoma (GBM). However, the prognostic impact of PDGFRA amplification on GBM patients remains unclear. Herein, we evaluated this impact by retrospectively analyzing outcomes of patients with IDH wild-type GBM.MethodsUsing a custom-made oncopanel, we evaluated PDGFRA gain/amplification in 107 GBM samples harboring wild-type IDH, along with MGMT promoter (MGMTp) methylation status.ResultsWe detected PDGFRA gain/amplification in 31 samples (29.0%). PDGFRA gain/amplification predicted poor prognosis (P = .003). Compared to unamplified PDGFRA, PDGFRA gain/amplification in GBM was associated with higher patient age (P = .031), higher Ki-67 score (P = .019), and lower extent of surgical resection (P = .033). Unmethylated MGMTp also predicted poor prognosis (P = .005). As PDGFRA gain/amplification and unmethylated MGMTp were independent factors for poor prognosis in multivariate analyses, we grouped GBM cases based on PDGFRA and MGMTp status: poor (PDGFRA gain/amplification and unmethylated MGMTp), intermediate (PDGFRA gain/amplification or unmethylated MGMTp), and good (PDGFRA intact and methylated MGMTp) prognosis. The Kaplan-Meier survival analysis indicated that these groups significantly correlated with the OS of GBM patients (P < .001).ConclusionsHere we report that PDGFRA gain/amplification is a predictor of poor prognosis in IDH wild-type GBM. Combining PDGFRA gain/amplification with MGMTp methylation status improves individual prognosis prediction in patients with IDH wild-type GBM.

Disparity Analysis of Clinicopathologic Hallmarks between Transformed DLBCL and Primary DLBCL.

A variety of indolent lymphomas, particularly marginal zone lymphoma (MZL) and follicular lymphoma (FL) can be histologically transformed to diffuse large B-cell lymphoma (DLBCL). Little is known about the disparity of clinicopathologic characteristics between transformed DLBCL (tDLBCL) and primary DLBCL (pDLBCL). This retrospective study analyzed the clinicopathological hallmarks of 10 tDLBCL (7 MZL and 3 FL) and 40 pDLBCL from the Affiliated Hospital of Xuzhou Medical University. Patients of tDLBCL had a higher ECOG score, more B-symptoms, and lower serum albumin level than those in pDLBCL (60.0% vs. 7.50%, 40.0% vs. 10.0%, and 90.0% vs. 10.0%, respectively, p < 0.01). Pathologically, tDLBCL had more c-Myc and BCL-2 dual-expression than that in pDLBCL (60.0% vs. 25.0%, p < 0.01). The positive rate of CD5 expression and the proportion of high Ki-67 score in tDLBCL were higher than those in pDLBCL (50.0% vs. 7.5%, 50.0% vs. 32.5%, respectively, p < 0.01). The median overall survival and progression-free survival were 14 months and 11 months in tDLBCL, 35 months and 28 months in pDLBCL (p < 0.05 and p < 0.001). Our results demonstrate that tDLBCL manifested aggressive clinical course and pathological features of Myc/BCL-2 expression, CD5 expression, and high Ki-67 score.

The role of 18F-FDG PET/CT in diagnosis and treatment evaluation for ocular adnexal mucosa-associated lymphoid tissue lymphoma.

The purpose of this study is to evaluate the value of fluorine-18-fludeoxyglucose positron emission tomography (18F-FDG PET)/CT in the diagnosis and treatment evaluation of ocular adnexal mucosa-associated lymphoid tissue (MALT) lymphoma. 70 patients with OAML who received radiotherapy were recruited in our study. All the patients had the 18F-FDG PET/CT examination before the treatment. We retrospectively reviewed the medical records, pathological reports, laboratory results, and imaging features of all patients. The associations between 18F-FDG PET/CT parameters and Epstein-Barr virus antibodies, treatment response, MRI data, and Ki-67 expression were investigated. The PET/CT scan indicated that 80% (56/70) of the patients showed orbital FDG avidity. The median level of maximum standardized uptake value (SUVmax) of the lesions was 4.65 ± 3.00 (range:1.2-13.5). 92.0% (46/50) of the mass-forming lesions showed 18F-FDG avidity, while only 50.0% (10/20) of the non-massive lesions had 18F-FDG avidity (χ2 = 13.23, p=0.01). The SUVmax in orbit, conjunctiva, and lacrimal gland lymphoma were 5.6, 2.9, and 3.7, respectively. A significant difference was identified of SUVmax among the three locations' lymphoma using one-way ANOVA analysis (F = 5.039, p = 0.01). After completion of radiotherapy, the complete remission rate was achieved in 30.8% (4/13) of the patients without 18F-FDG avidity, and 70.4% (38/54) in cases with 18F-FDG avidity (χ2 = 5.43, p = 0.02). The correlation between high Ki-67 score and 18F-FDG avidity was confirmed (χ2 = 3.916, p = 0.048); however, no significant correlation was found between the SUVmax and Ki-67 score of the lesions (p = 0.971). Three patients (3/70, 4.3%) were upregulated the stage via PET/CT. 18F-FDG PET/CT had some potential values in the diagnosis and assessment of treatment response in patients with OAML. The value of 18F-FDG PET/CT for patients with OAML.

Abstract 14414: Embryonic Stem Cell Derived Exosomes Enhance Anti-Tumor Efficacy of Doxorubicin and Improve Cardiac Function in Triple-Negative Breast Cancer Xenograft Mice

Background: Doxorubicin (Dox) is a potent chemotherapeutic drug widely used for the treatment of triple negative breast cancer (TNBC). However, Dox treatment results in inadvertent cardiotoxic effects which often require supplemental therapy. We previously showed that treatment with embryonic stem cells derived exosomes (ES-Exos) enhanced chemosensitivity of Dox in breast cancer cells. Here we tested the hypothesis whether adjuvant therapy with ES-Exos in mice with orthotopic TNBC xenograft can enhance the anti-tumor effect of Dox and improve cardiac function. Methods and Results: Athymic female nude mice (6-wk old) were injected with human MDA-MB-231-Luc cell line in mammary intraductal T4 region to generate orthotopic xenografts of TNBC. After 2 weeks of tumor growth, mice (n=10/group) were treated for 4 weeks with: Saline (Con); Dox (5 mg/kg/week); ES-Exos (50 μg, twice a week); Dox+ES-Exos; and MEF (50 μg/twice a week; non-stem cell exosomes derived from mouse fibroblasts). ES-Exos were labelled with ExoGlow-Vivo near-IR dye to track the localization of exosomes in the tumor. Treatment with Dox+ES-Exos and Es-Exos showed improved survival as compared to Dox (2 deaths) and Con (3 deaths) (Fig.A) . Tumor size was significantly reduced with Dox+ES-Exos and ES-Exos treatments as compared to Con and Dox treatment (Fig.B) . Cardiac function measured by echocardiography showed improved ejection fraction in Dox+ES-Exos and ES-Exos as compared to Con group (Fig.C) . Cell proliferation rate assessed by Ki-67 score with immunofluorescence showed significantly higher rate in Con as compared to Dox and Dox+ES-Exos groups. RNA seq analysis identified upregulation of unique antitumorigenic regulators such as Semaphorin 5 (Sema5) and FOXO15 in ES-Exos compared to MEF, which may contribute to the therapeutic effect. Conclusion: Treatment with ES-Exos may be a promising novel strategy to improve chemosensitivity with attenuation of its cardiotoxic effect in TNBC.

Application of diffusion kurtosis imaging to the study of edema in solid and peritumoral areas of glioma

ObjectiveWhen gliomas grow in an infiltrative form, high-grade malignant glioma tissue extends beyond the contrast-enhancing tumor boundary, and this diffuse non-enhancing tumor infiltration is not visible on conventional MRI. The purpose of this study was to evaluate the of diffusion kurtosis imaging (DKI)-derived parameters in a group of patients with pre-operative gliomas, evaluating changes in the solid tumor and peritumoral edema area, and investigating their use for evaluating the recurrence and prognosis of gliomas. MethodsIn this retrospective study, 51 patients with gliomas who underwent biopsy or surgery underwent DKI scans before surgery. DKI scans were performed to generate DKI parameter maps of the solid tumor and peritumoral edema areas. In the solid tumor area, the kurtosis parameters showed the highest area under the curve (AUC), sensitivity, and specificity for distinguishing high- and low-grade gliomas (all P < 0.01). ResultsIn the peritumoral edema area, significant differences were found between groups with grade III and IV gliomas (P < 0.05). DKI parameters were found to correlate with clinical Ki-67 scores within the solid tumor area (MK: R2 = 0.288, P < 0.001; Kr: R2 = 0.270, P < 0.001; Ka: R2 = 0.274, P < 0.001; MD: R2 = 0.223, P < 0.001; FA: R2 = 0.098, P < 0.01). No significant correlations were found between Ki-67 and kurtosis parameters of peritumoral edema. ConclusionsIn this study, DKI showed potential utility for studying solid tumor and peritumoral edema of high grade gliomas.

Ki67 and P53 expression in breast cancer and their correlation with clinicopathological parameters

Breast carcinoma is the most common malignancy in females and is a leading cause of death. Treatment depends upon various pathological and prognostic markers like lymph node status, size, type and grade of the tumour which influences the outcome of breast cancer. Markers like Ki-67 and p53 have been studied extensively and their roles in breast cancer are yet to be established.We evaluated the expression of Ki67 and P53 in breast cancer and their association with other clinico-pathological factors was studied. Ki67 and P53 expression was assessed in 60 breast cancer cases admitted to our hospital over a period of one year. Association with other prognostic parameters was evaluated. Statistical analysis was done by Chi square test and a p value of &amp;#60;0.05 was taken as significant. 43.33% cases had low proliferative Ki-67 score whereas 56.67% of the cases were highly proliferative. p53 expression was seen in 41.66% cases. Ki-67 and p53 expression were not significantly related to age, menopausal status, and tumour size whereas a significant correlation was seen with positive axillary lymph node status, high histological grade, negative hormone receptor status (ER, PR) and positive HER2/neu expression. Significant association was seen between Ki-67 and p53 expression.Ki67 and P53 may be considered as a valuable biomarker in breast cancer patients which can help in planning treatment strategies.

Combined Use of cyclinD1 and Ki67 for Prognosis of Luminal-Like Breast Cancer Patients.

BackgroundKi67 is a biomarker of proliferation to be used in immunohistochemistry (IHC)-based surrogate assay to determine the necessity of cytotoxic therapy for Luminal-like breast cancer patients. cyclinD1 is another frequently used biomarker of proliferation. A retrospective study was performed here to investigate if these two biomarkers may be combined to improve the prognosis of Luminal-like patients.MethodsBoth Ki67 and cyclinD1 protein levels were measured absolutely and quantitatively using Quantitative Dot Blot method in 143 Luminal-like specimens. Optimized cutoffs for these two biomarkers were developed to evaluate their prognostic roles using Kaplan–Meier overall survival (OS) analysis.ResultscyclinD1 was found as an independent prognostic factor from Ki67 in univariate and multivariate OS analyses. At optimized cutoffs (cyclinD1 at 0.44 μmol/g and Ki67 at 2.31 nmol/g), the subgroup with both biomarkers below the cutoffs (n = 65) had 10-year survival probability at 90% in comparison to those with both biomarkers above the cutoffs (n = 18) with 8-year survival probability at 26% (log-rank test, p <0.0001). This finding was used to modify the surrogate assay using IHC-based cyclinD1 scores, with p-value decreased from 0.031 to 0.00061 or from 0.1 to 0.02, when the Ki67 score of 14 or 20% was used as cutoff, respectively, in the surrogate assay.ConclusionThe current study supports the prospective investigation of cyclinD1 relevance in the clinic.

Clinical Outcome of Refractory Lymphoma Patients with Non Response to Chimeric Antigen Receptor T Cell Therapy

BackgroundChimeric antigen receptor T cell (CART) therapy has remarkably improved the outcome of patients (pts) with relapsed refractory B cell non-Hodgkin lymphoma. Anti-CD19 directed CART therapy have shown to yield durable remission in 40%-50% of patients with relapsed refractory (r/r) diffuse large B cell lymphoma (Neelapu et al. NEJM 2017; NCT 02348216). However, pts who had disease progression post CART therapy (non-responders) have poor outcomes. There is paucity of information regarding negative predictors for disease progression and outcome of non-responders post CART therapy.MethodsWe conducted a single center retrospective study of 11 patients with relapsed refractory large B cell lymphoma who underwent treatment with CART in our institution from December 2019 to June 2021 and further studied the baseline characteristics, and clinical outcome of 8 pts who were non responders to CART therapy.ResultsAll patients received low dose fludarabine cyclophosphamide based lymphodepleting conditioning regimen followed by anti-CD19 directed CART cells using CD28 as costimulatory domain with a target dose of 2 x 10 6 cells /kilogram of body weight.Baseline characteristics as shown in [Table 1]. Median age at CART therapy was 61 years (range 49-71). Median ECOG score at CART therapy was 1. All patients had chemo refractory disease at time of CART therapy with 9 patients having refractory diffuse large B cell lymphoma and 2 patients had transformed large B cell lymphoma. Genomic studies showed high risk disease in 4 patients with double hit lymphoma, 3 patients with complex cytogenetics with one having 17 p deletion and 2 patients with double expressors of c-myc and bcl2. Only 5 patients had available Ki-67 score > 80%. All patients had stage III-IV disease at the time of CART therapy. Seven pts had extra nodal disease and one patient had CNS involvement. Median R-IPI score at time of CART therapy was 3 (range 2-4). Median LDH, serum ferritin and serum CRP at the time of CART therapy was 388 (range 136-1489); 831 (range 116.9-1503); and 28.1 (range 7.4-226) respectively. Median number of prior therapies were 3 (range 2-9). One patient had prior autologous stem cell transplant. Median absolute lymphocyte count (ALC) and platelet count at CART therapy was 0.01/ul (range 0.01-2.5) and 89 x 10 3/ul (range 10-385) respectively. Median time from last salvage chemotherapy to CART therapy was 62 days (range 28-492).Total six pts had cytokine release syndrome (CRS) with median time for CRS onset was 3.5 days (range 1-8) with all of them having grade 1-2 CRS. Total seven pts had neurotoxicity with median time for onset was 5 days (range 1-41) post CART with 4 pts having > grade 3 immune effector cell associated neurotoxicity syndrome (ICANS). Overall, six and seven pts received tociluzumab and steroids respectively for treatment of CART related CRS and ICANS. Day 30 PET/CT scan showed three responders (2CR,1PR) and eight non responder patients with disease progression.Non responder pts to CART, when compared with responders had higher value of median LDH (438 vs 300; p = 0.92); median CRP (29 vs 13; p = 0.497) ; higher extra nodal involvement ( 6 pts vs one), median R-IPI score at the time of CART ( 4 vs 3), median number of prior therapies (4 vs 3), and median time from last salvage therapy to CART ( 73 days vs 50 days ; p = 0.91). No difference in baseline ALC count and platelet count was noted between the two groups. Compared to responders, non-responder pts had lower incidence of CRS (100% vs 37.5%) and ICANS (100% vs 50%). Post CART therapy, non-responder pts were treated with Nivolumab based immunotherapy in 5 pts, Ipilimumab based therapy in 1 pt, Selinexor in 1 patient and Polatuzumab based therapy in 2 pts.Median progression free survival and overall survival of non-responders was only 30.5 days and 94 days respectively. 5 out of 8 non responder pts died (4 due to disease progression and 1 due to sepsis). All three responder pts were alive and disease free at the end of study. Median follow up of our study was 108 days (range 32- 561).ConclusionsDespite smaller sample size, our study showed dismal outcome of pts who don't respond to CART therapy, showing an unmet need for better salvage therapies in such pts. Refractory lymphoma pts with extra nodal disease involvement, higher LDH and CRP at the time of CART therapy could be negative predictors for response. Further prospective studies with larger sample size required to further validate these findings. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Matrix metalloproteinase‑1 expression is regulated by HIF‑1‑dependent and epigenetic mechanisms and serves a tumor‑suppressive role in gastric cancer progression.

The matrix metalloproteinase (MMP) family is associated with degradation of the extracellular matrix and is known to promote cancer invasion. The present study aimed to investigate the biological role of MMP-1 in gastric cancer cells and analyze the association between MMP-1 expression and the clinical outcomes of gastric cancer patients. In the present study, hypoxia accelerated invasion, accompanied by elevated MMP-1 expression in the gastric cancer cell line 58As9. Additionally, hypoxia-inducible factor-1α (HIF-1α) knock- down in 58As9 cells reduced MMP-1 expression under hypoxic conditions. Treatment with 5-aza-2-deoxycytidine and trichostatin A restored MMP-1 expression in the MMP-1-deficient cell lines MKN45 and MKN74. These results indicated that MMP-1 expression was controlled by both HIF-1α-dependent and epigenetic mechanisms in gastric cancer cell lines. In addition, MMP-1 knockdown impaired the hypoxia-induced invasiveness of 58As9 cells, implicating MMP-1 in the elevated invasion. By contrast, knockdown enhanced the proliferative ability of 58As9 cells, whereby expression of cell cycle-related genes was subsequently altered. In nude mouse models, the knockdown accelerated the growth of xenograft tumor and the development of peritoneal dissemination. In an immunohistochemical study using 161 surgically resected cancer tissues, the Ki67 score was significantly higher in the group with low MMP-1 expression (P<0.001). Disease-free survival (DFS) and disease-specific survival (DSS) were both significantly reduced in patients with low MMP-1 expression (log-rank test; DFS: P=0.005; DSS: P=0.022). Multivariate analysis demonstrated that MMP-1 expression was an independent prognostic factor for DFS and DSS [DFS: HR=2.11 (1.22–3.92) P=0.005, DSS: HR=2.90 (1.23–8.50) P=0.012]. In conclusion, the present study indicated that MMP-1 may serve as a tumor-suppressive factor that inhibits gastric cancer progression, although it promoted invasion in vitro.

Comparison of Different Molecular Subtypes with 14% Ki-67 Cut-off Threshold in Breast Cancer Patients of Pakistan- An Indication of Poor Prognosis

Ki-67 is a proliferation marker that is used not only to categorize patients in luminal A and B subtypes of breast cancers, but also to determine the aggressiveness of the disease in triple negative and human epidermal growth factor 2 (HER2) over expressed molecular subtypes. The present study was designed to evaluate the role of Ki-67 with cut off value of 14% in molecular subgroups and its association with patient prognosis. Immunostaining was performed on histopathologically confirmed sections (n = 278) to assess expression of Ki-67, estrogen receptor (ER), progesterone receptor (PR) and HER2. Immunoreactivity of molecules was recorded as percentage scoring. Adopting a cut off value of 14%, Ki-67 was high in 88%of the cases included in the study. High Ki-67 was significantly associated with pathological parameters including histological grade, advanced stage and nodal/distant metastasis. Immunoexpression of ER, PR and HER2 also showed strong correlation with high expression of Ki-67. Based on the St. Gallen classification, the cases were categorized into luminal A (10%) and luminal B (51%), triple negative (20%) and HER2 enriched (18%). Ki-67 index was also significantly high in 98% of HER2 enriched and 95% of TNBC patients. Interestingly, Ki-67 score with cut off value of 14% proved to be significant in deciphering prognosis in luminal patients. Moreover, high expression of Ki-67 also proved to be a marker of poor prognosis, especially in triple negative patients. We suggest that utilization of IHC4 status i.e. ER, PR, HER2 and Ki-67 along with pathological findings and molecular subtyping can considerably affect clinical as well as therapeutic decisions.

Independent Clinical Validation of the Automated Ki67 Scoring Guideline from the International Ki67 in Breast Cancer Working Group.

Ki67 is an important biomarker with prognostic and potential predictive value in breast cancer. However, the lack of standardization hinders its clinical applicability. In this study, we aimed to investigate the reproducibility among pathologists following the guidelines of the International Ki67 in Breast Cancer Working Group (IKWG) for Ki67 scoring and to evaluate the prognostic potential of this platform in an independent cohort. Four algorithms were independently built by four pathologists based on our study cohort using an open-source digital image analysis (DIA) platform (QuPath) following the detailed guideline of the IKWG. The algorithms were applied on an ER+ breast cancer study cohort of 157 patients with 15 years of follow-up. The reference Ki67 score was obtained by a DIA algorithm trained on a subset of the study cohort. Intraclass correlation coefficient (ICC) was used to measure reproducibility. High interobserver reliability was reached with an ICC of 0.938 (CI: 0.920–0.952) among the algorithms and the reference standard. Comparing each machine-read score against relapse-free survival, the hazard ratios were similar (2.593–4.165) and showed independent prognostic potential (p ≤ 0.018, for all comparisons). In conclusion, we demonstrate high reproducibility and independent prognostic potential using the IKWG DIA instructions to score Ki67 in breast cancer. A prospective study is needed to assess the clinical utility of the IKWG DIA Ki67 instructions.