Relationship of pathological features and a 21 gene expression assay in younger versus older women with node-negative endocrine receptor-positive breast cancer.

PURPOSE: Determining the need for adjuvant chemotherapy in estrogen receptor (ER)+ disease can be influenced by pathological characteristics and gene expression assays [i.e., Oncotype Dx recurrence scores (RSs)]. The primary objective of this study is to investigate the relationship between the RSs and pathological markers in younger (< 50) versus older (≥ 50) women with early-stage node-negative ER+ breast cancer. METHODS: This was a single academic-center retrospective cohort study. Subjects who underwent Oncotype gene expression testing were retrospectively and sequentially identified. 436 Subjects were identified of which 344 were eligible for analysis (133 younger subjects < 50 years of age, and 211 older subjects ≥ 50 years). Pathological data assessed included the progesterone receptor (PR), histological grade (grade), Ki-67, and P53. A multivariable regression analysis was performed using age, PR, and grade as predictor variables for RS. Adjusted R2 was determined. To investigate the primary objective, subjects were stratified based on age, PR, and grade status in that sequence. Within each tumor subtype as determined by PR and grade statuses, the RSs in the younger versus older age group were compared using Student's t-test and the differences in the 95% confidence interval limits in RS means calculated. Age influence on adjuvant chemotherapy recommendation was also assessed by stratifying subjects based on age (< 50 vs. ≥ 50) and then by RS risk group (≤ 10, 11-25, ≥ 26). Subsequently, the proportions of younger versus older subjects within identical RS risk groups who were explicitly advised by their oncologist to proceed with chemotherapy as documented in their electronic health records were compared using χ2 test. RESULTS: Based on the multivariable regression analysis, the adjusted R2 was 0.229232 and RS was found to be independent of age (p = 0.7169). Between younger and older subjects with tumors with similar PR and grade pathological features, the differences in the RS were insignificant (p > 0.05). Chemotherapy was recommended in younger versus older women, in 0% when the RS was ≤ 10, 39% and 40% when the RS was 11-25 (p = 0.82), and 100% and 98% when the RS was ≥ 26 (p = 0.51), respectively. CONCLUSIONS: The relationship between pathological features and RS is consistent irrespective of age; therefore, models predicting RS may be applicable irrespective of age.

Impact of tissue-based genomic profiling on clinical decision making in the management of patients with metastatic breast cancer at academic centers.

BACKGROUND: Genomic profiling can identify targetable mutations; however, the impact of tissue-based genomic profiling on clinical decision making for patients with metastatic breast cancer has not been well characterized. METHODS: Patients with stage IV breast cancer who had undergone genomic profiling between 7/2013 and 3/2015 were identified at three academic cancer centers. Genomic analysis was determined to have impacted clinical decision if (A) a patient was enrolled onto a genotype-matched clinical trial or (B) prescribed off-label an FDA-approved therapy targeting an identified mutation. The frequency of mutated genes was determined. RESULTS: A total of 117 patients with stage IV breast cancer were identified. Median age was 46 (25-75). Fifty-three patients (45%) had ER-positive/HER2-negative disease, 50 (43%) had ER-negative/HER2-negative disease, and 14 (12%) had ER-any/HER2-positive disease. Median number of previous therapies received prior to genomic profiling was 2 (range 0-15), and median follow-up after testing was obtained after 5.8 months (range 0-24.4 months). Commercial reports indicated that 85 (73%) patients had at least one mutation targetable by an FDA-approved medication, and 112 (96%) patients had at least one clinical trial available; however, clinical management was only affected in 11 patients (9%). The most frequent mutations observed were those in TP53, FGF, PI3KCA, MYC, ZNF, FGFR, CCND, ARID1A, GATA3, and MAP; frequencies of these mutations varied by clinical subtype. CONCLUSIONS: Tumor genomic profiling affected clinical management in a minority of patients with metastatic breast cancer, thus these data do not support the routine use of genomic profiling outside of a clinical trial.

Quantitative characterization of T-cell repertoire and biomarkers in kidney transplant rejection.

BACKGROUND: T-cell-mediated rejection (TCMR) remains a major cause of kidney allograft failure. The characterization of T-cell repertoire in different immunological disorders has emerged recently as a novel tool with significant implications. We herein sought to characterize T-cell repertoire using next generation sequencing to diagnose TCMR. METHODS: In this prospective study, we analyzed samples from 50 kidney transplant recipients. We collected blood and kidney transplant biopsy samples at sequential time points before and post transplant. We used next generation sequencing to characterize T-cell receptor (TCR) repertoire by using illumina miSeq on cDNA synthesized from RNA extracted from six patients' samples. We also measured RNA expression levels of FOXP3, CD8, CD4, granzyme and perforin in blood samples from all 50 patients. RESULTS: Seven patients developed TCMR during the first three months of the study. Out of six patients who had complete sets of blood and biopsy samples two had TCMR. We found an expansion of the TCR repertoire in blood at time of rejection when compared to that at pre-transplant or one-month post transplant. Patients with TCMR (n = 7) had significantly higher RNA expression levels of FOXP3, Perforin, Granzyme, CD4 and CD8 in blood samples than those with no TCMR (n = 43) (P = 0.02, P = 0.003, P = 0.002, P = 0.017, and P = 0.01, respectively). CONCLUSIONS: Our study provides a potential utilization of TCR clone kinetics analysis in the diagnosis of TCMR. This approach may allow for the identification of the expanded T-cell clones associated with the rejection and lead to potential noninvasive diagnosis and targeted therapies of TCMR.

Management of Metastatic HER2-Positive Breast Cancer: Where Are We and Where Do We Go From Here?

Human epidermal growth factor receptor 2 (HER2)/neu-positive breast cancer has changed from being an aggressive disease with a poor prognosis to a disease that is highly treatable, with prolonged survival possible even in patients with metastatic disease. A better understanding of HER2 biology has led to the development of powerful targeted therapies, and four drugs are already approved by the US Food and Drug Administration for treatment in the metastatic setting (trastuzumab, pertuzumab, lapatinib, and trastuzumab emtansine). Optimizing how these drugs are delivered and in what sequence is an important part of modern management of HER2-positive breast cancer. However, while the prognosis has improved, metastatic disease is still not curable; newer, better drugs are needed. This review will summarize the current standard of care; key issues that arise when treating patients with HER2-positive disease; and developments in novel therapeutics, including small-molecule inhibitors, nanoparticles, immunotherapy, and agents targeting resistance pathways.

Targeting Suppressor of Variegation 3-9 Homologue 2 (SUV39H2) in Acute Lymphoblastic Leukemia (ALL).

Although recent progress in understanding the biology and optimizing the treatment of acute lymphoblastic leukemia (ALL) has improved cure rates of childhood ALL to nearly 90%, the cure rate in adult ALL remains less than 50%. The poor prognosis in adult ALL has in part been attributed to larger proportion of high-risk leukemia showing drug resistance. Thus, identifying novel therapeutic targets in ALL is needed for further improvements in treatment outcomes of adult ALL. Genetic aberration of chromatin-modifying molecules has been recently reported in subtypes of ALL, and targeting components of chromatin complexes has shown promising efficacy in preclinical studies. Suppressor of variegation 3-9 homologue 2 (SUV39H2), also known as KMT1B, is a SET-domain-containing histone methyltransferase that is upregulated in solid cancers, but its expression is hardly detectable in normal tissues. Here, we show that SUV39H2 is highly expressed in ALL cells but not in blood cells from healthy donors and also that SUV39H2 mRNA is expressed at significantly higher levels in bone marrow or blood cells from patients with ALL obtained at diagnosis compared with those obtained at remission (P = .007). In four ALL cell lines (Jurkat and CEM derived from T-ALL and RS4;11 and REH derived from B-ALL), SUV39H2 knockdown resulted in a significant decrease in cell viability (~77%, P < .001), likely through induction of apoptosis. On the other hand, SUV39H2 overexpression made cells more resistant to chemotherapy. We conclude that SUV39H2 is a promising therapeutic target and further investigation of this therapeutic approach in ALL is warranted.

T-LAK cell-originated protein kinase presents a novel therapeutic target in FLT3-ITD mutated acute myeloid leukemia.

Gain-of-function mutations of FLT3 (FLT3-ITD), comprises up to 30% of normal karyotype acute myeloid leukemia (AML) and is associated with an adverse prognosis. Current FLT3 kinase inhibitors have been tested extensively, but have not yet resulted in a survival benefit and novel therapies are awaited. Here we show that T-LAK cell-originated protein kinase (TOPK), a mitotic kinase highly expressed in and correlated with more aggressive phenotype in several types of cancer, is expressed in AML but not in normal CD34+ cells and that TOPK knockdown decreased cell viability and induced apoptosis. Treatment of AML cells with TOPK inhibitor (OTS514) resulted in a dose-dependent decrease in cell viability with lower IC50 in FLT3-mutated cells, including blasts obtained from patients relapsed after FLT3-inhibitor treatment. Using a MV4-11-engrafted mouse model, we found that mice treated with 7.5 mg/kg IV daily for 3 weeks survived significantly longer than vehicle treated mice (median survival 46 vs 29 days, P < 0.001). Importantly, we identified TOPK as a FLT3-ITD and CEBPA regulated kinase, and that modulating TOPK expression or activity resulted in significant decrease of FLT3 expression and CEBPA phosphorylation. Thus, targeting TOPK in FLT3-ITD AML represents a novel therapeutic approach for this adverse risk subset of AML.

Preclinical efficacy of maternal embryonic leucine-zipper kinase (MELK) inhibition in acute myeloid leukemia.

Maternal embryonic leucine-zipper kinase (MELK), which was reported to be frequently up-regulated in various types of solid cancer, plays critical roles in formation and maintenance of cancer stem cells. However, little is known about the relevance of this kinase in hematologic malignancies. Here we report characterization of possible roles of MELK in acute myeloid leukemia (AML). MELK is expressed in AML cell lines and AML blasts with higher levels in less differentiated cells. MELK is frequently upregulated in AML with complex karyotypes and is associated with worse clinical outcome. MELK knockdown resulted in growth inhibition and apoptosis of leukemic cells. Hence, we investigated the potent anti-leukemia activity of OTS167, a small molecule MELK kinase inhibitor, in AML, and found that the compound induced cell differentiation and apoptosis as well as decreased migration of AML cells. MELK expression was positively correlated with the expression of FOXM1 as well as its downstream target genes. Furthermore, MELK inhibition resulted in downregulation of FOXM1 activity and the expression of its downstream targets. Taken together, and given that OTS167 is undergoing a phase I clinical trial in solid cancer, our study warrants clinical evaluation of this compound as a novel targeted therapy for AML patients.

Cell-specific multifunctional processing of heterogeneous cell systems in a single laser pulse treatment.

Current methods of cell processing for gene and cell therapies use several separate procedures for gene transfer and cell separation or elimination, because no current technology can offer simultaneous multifunctional processing of specific cell subsets in highly heterogeneous cell systems. Using the cell-specific generation of plasmonic nanobubbles of different sizes around cell-targeted gold nanoshells and nanospheres, we achieved simultaneous multifunctional cell-specific processing in a rapid single 70 ps laser pulse bulk treatment of heterogeneous cell suspension. This method supported the detection of cells, delivery of external molecular cargo to one type of cells and the concomitant destruction of another type of cells without damaging other cells in suspension, and real-time guidance of the above two cellular effects.