An in vitro investigation of metabolically sensitive biomarkers in breast cancer progression.

Epigenetic biomarkers are emerging as determinants of breast cancer prognosis. Breast cancer cells display unique alterations in major cellular metabolic pathways and it is becoming widely recognized that enzymes that regulate epigenetic alterations are metabolically sensitive. In this study, we used microarray data from the GEO database to compare gene expression for regulators of metabolism and epigenetic alterations among non-invasive epithelial (MCF-7, MDA-MB-361, and T-47D) and invasive mesenchymal (MDA-MB-231, Hs-578T, and BT-549) breast cancer cell lines. The expression of genes, including GLS1, GFPT2, LDHA, HDAC9, MYST2, and SUV420H2, was assessed using RT-PCR. There was differential expression between epithelial and mesenchymal cell lines. MYST2 and SUV420H2 regulate the levels of the epigenetic biomarkers histone H4 lysine 16 acetylation (H4K16ac) and histone H4 lysine 20 trimethylation (H4K20me3), respectively. Reduced amounts of H4K16ac and H4K20me3 correlated with lower levels of MYST2 and SUV420H2 in mesenchymal cells and, along with reduced amounts of histone H3 lysine 9 acetylation (H3K9ac), were found to distinguish epithelial from mesenchymal cells. In addition, both GLS1 and GFPT2 play roles in glutamine metabolism and were observed to be more highly expressed in mesenchymal cell lines, and when glutamine and glutamate levels reported in the NCI-60 metabolomics dataset were compared, the ratio of glutamate/glutamine was found to be higher in mesenchymal cells. Blocking the conversion of glutamine to glutamate using an allosteric inhibitor, Compound 968, against GLS1, increased H4K16ac in T-47D and MDA-MB-231 cells, linking glutamine metabolism to a particular histone modification in breast cancer. These findings support the concept that metabolically sensitive histone modifications and corresponding histone modifying enzymes can be used as diagnostic and prognostic biomarkers for breast cancer. It also further emphasizes the importance of glutamine metabolism in tumor progression and that inhibitors of cellular metabolic pathways may join histone deacetylase inhibitors as a form of epigenetic therapy.

FDA Approval Summary: Belantamab Mafodotin for Patients with Relapsed or Refractory Multiple Myeloma.

On August 5, 2020, the FDA granted accelerated approval to belantamab mafodotin-blmf (BLENREP; GlaxoSmithKline) for the treatment of adult patients with relapsed or refractory multiple myeloma who have received at least four prior therapies including an anti-CD38 monoclonal antibody, a proteasome inhibitor, and an immunomodulatory agent. Substantial evidence of effectiveness was obtained from the phase II, multicenter DREAMM-2 trial. Patients received belantamab mafodotin 2.5 or 3.4 mg/kg intravenously once every 3 weeks until disease progression or unacceptable toxicity. The trial demonstrated an overall response rate of 31% in the 2.5 mg/kg cohort and 34% in the 3.4 mg/kg cohort. Keratopathy was the most frequent adverse event, occurring in 71% and 77% of patients, respectively. Other ocular toxicities included changes in visual acuity, blurred vision, and dry eye. The U.S. prescribing information for belantamab mafodotin includes a boxed warning for ocular toxicity, and belantamab mafodotin is available only through a restricted program under a Risk Evaluation and Mitigation Strategy. This article summarizes the data and the FDA review process supporting accelerated approval of belantamab mafodotin 2.5 mg/kg intravenously once every 3 weeks. This approval may be contingent upon verification and description of clinical benefit in confirmatory trial(s).

FDA Approval Summary: Tagraxofusp-erzs For Treatment of Blastic Plasmacytoid Dendritic Cell Neoplasm.

Tagraxofusp-erzs (Elzonris, Stemline) is a cytotoxin that targets CD123-expressing cells. On December 21, 2018, FDA approved tagraxofusp-erzs for the treatment of blastic plasmacytoid dendritic cell neoplasms (BPDCN) in adult and pediatric patients 2 years and older. Approval was based on the response rate in a single-arm trial, Study STML-401-0114; the pivotal cohort included 13 patients with treatment-naïve BPDCN who received tagraxofusp-erzs monotherapy. The complete response or clinical complete response (CR/CRc) rate in the pivotal cohort was 54% (95% CI: 25%-81%), and the median duration of CR/CRc was not reached with a median follow-up of 11.5 months (range: 0.2-12.7). In a separate exploratory cohort, a CR/CRc was achieved by 2 (13%) patients with R/R BPDCN. Safety was assessed in 94 patients with myeloid neoplasms treated with tagraxofusp-erzs at the approved dose and schedule. The major toxicity was capillary leak syndrome (CLS), which occurred in 55% of patients and was fatal in 2%. Hepatotoxicity and hypersensitivity reactions were reported in 88% and 46% of patients, respectively. Other common (≥30%) adverse reactions were nausea, fatigue, peripheral edema, pyrexia, and weight increase. A high proportion of patients (85%) developed neutralizing antidrug antibodies. Tagraxofusp-erzs is the first FDA-approved treatment for BPDCN.

Nonclinical Evaluations of Small-Molecule Oncology Drugs: Integration into Clinical Dose Optimization and Toxicity Management.

Multidisciplinary approaches that incorporate nonclinical pharmacologic and toxicologic characterization of small-molecule oncology drugs into clinical development programs may facilitate improved benefit-risk profiles and clinical toxicity management in patients. The performance of the current nonclinical safety-testing scheme was discussed, highlighting current strengths and areas for improvement. While current nonclinical testing appears to predict the clinical outcome where the prevalence of specific adverse effects are high, nonclinical testing becomes less reliable for predicting clinical adverse effects that occur infrequently, as with some kinase inhibitors. Although adverse effects associated with kinase inhibitors can often be predicted on the basis of target biology, drugs can be promiscuous and inhibit targets with poorly defined function and associated risks. Improvements in adverse effect databases and better characterization of the biologic activities of drug targets may enable better use of computational modeling approaches in predicting adverse effects with kinase inhibitors. Assessing safety of a lead candidate in parallel with other drug properties enables incorporation of a molecule's best features during chemical design, eliminates the worst molecules early, and permits timely investigation/characterization of toxicity mechanisms for identified liabilities. A safety lead optimization and candidate identification strategy that reduces intrinsic toxicity and metabolic risk and enhances selectivity can deliver selective kinase inhibitors that demonstrate on-target adverse effects identified nonclinically. Integrating clinical and nonclinical data during drug development can facilitate better identification and management of oncology drugs. Follow-up nonclinical studies may be used to better understand the risks in a given patient population and minimize or manage these risks more appropriately. Clin Cancer Res; 22(11); 2618-22. ©2016 AACR SEE ALL ARTICLES IN THIS CCR FOCUS SECTION, "NEW APPROACHES FOR OPTIMIZING DOSING OF ANTICANCER AGENTS".

Modifying metabolically sensitive histone marks by inhibiting glutamine metabolism affects gene expression and alters cancer cell phenotype.

The interplay of metabolism and epigenetic regulatory mechanisms has become a focal point for a better understanding of cancer development and progression. In this study, we have acquired data supporting previous observations that demonstrate glutamine metabolism affects histone modifications in human breast cancer cell lines. Treatment of non-invasive epithelial (T-47D and MDA-MB-361) and invasive mesenchymal (MDA-MB-231 and Hs-578T) breast cancer cell lines with the glutaminase inhibitor, Compound 968, resulted in cytotoxicity in all cell lines, with the greatest effect being observed in MDA-MB-231 breast cancer cells. Compound 968-treatment induced significant downregulation of 20 critical cancer-related genes, the majority of which are anti-apoptotic and/or promote metastasis, including AKT, BCL2, BCL2L1, CCND1, CDKN3, ERBB2, ETS1, E2F1, JUN, KITLG, MYB, and MYC. Histone H3K4me3, a mark of transcriptional activation, was reduced at the promoters of all but one of these critical cancer genes. The decrease in histone H3K4me3 at global and gene-specific levels correlated with reduced expression of SETD1 and ASH2L, genes encoding the histone H3K4 methyltransferase complex. Further, the expression of other epigenetic regulatory genes, known to be downregulated during apoptosis (e.g., DNMT1, DNMT3B, SETD1 and SIRT1), was also downregulated by Compound 968. These changes in gene expression and histone modifications were accompanied by the activation of apoptosis, and decreased invasiveness and resistance of MDA-MB-231 cells to chemotherapeutic drug doxorubicin. The results of this study provide evidence to a link between cytotoxicity caused by inhibiting glutamine metabolism with alterations of the epigenome of breast cancer cells and suggest that modification of intracellular metabolism may enhance the efficiency of epigenetic therapy.

Research resource: enhanced genome-wide occupancy of estrogen receptor α by the cochaperone p23 in breast cancer cells.

p23 is a chaperone with multiple heat shock protein 90 dependent and independent cellular functions, including stabilizing unliganded steroid receptors and modulating receptor-DNA dynamics. p23 protein is also up-regulated in several cancers, notably breast cancer. We previously demonstrated that higher expression of p23 in the estrogen-dependent breast cancer line MCF-7 (MCF-7+p23) selectively increased estrogen receptor (ER) target gene transcription and ER recruitment to regulatory elements, promoted cell invasion, and predicted a poor prognosis in breast cancer patients. To probe the impact of p23 on ER binding throughout the human genome, we compared ER occupancy in MCF-7+p23 cells relative to MCF-7-control cells by using chromatin immunoprecipitation followed by ultrahigh-throughput DNA sequencing in the absence and presence of 17ß-estradiol (E2) treatment. We found that increased expression of p23 resulted in a 230% increase in the number of E2-induced ER-binding sites throughout the genome compared with control cells and also increased ER binding under basal conditions. Motif analysis indicated that ER binds to a similar DNA sequence regardless of p23 status. We also observed that ER tends to bind closer to genes that were induced, rather than repressed by either E2 treatment or p23 overexpression. Interestingly, we also found that the increased invasion of MCF-7+p23 cells was not only p23 dependent but also ER dependent. Thus, a small increase in the expression of p23 amplifies ER-binding genome wide and, in combination with ER, elicits an invasive phenotype. This makes p23 an attractive target for combating tumor cell metastasis in breast cancer patients.

Alterations in histone H4 lysine 20 methylation: implications for cancer detection and prevention.

SIGNIFICANCE: Cancer development and progression are associated with numerous genetic, epigenetic, and metabolic changes. RECENT ADVANCES: A number of epigenetic aberrations have been characterized in cancer, including DNA methylation and various histone modification changes. One of the most unique and enigmatic epigenetic marks that is noticeably altered in several major human cancers is methylation of histone H4 lysine 20; however, there is insufficient knowledge of the underlying molecular mechanisms associated with this abberation. CRITICAL ISSUES: This review presents current evidence of the role of histone H4 lysine 20 methylation in normal and cancer cells and during tumorigenesis induced by genotoxic and nongenotoxic carcinogens. Additionally, it describes molecular mechanisms that may cause this alteration and highlights the significance of this epigenetic mark as an early indicator of carcinogenesis. FUTURE DIRECTIONS: Accumulating evidence suggests that dietary components may be significant regulators of the cellular epigenome, including histone methylation, by providing and maintaining the adequate levels of S-adenosyl-L-methionine, flavin adenine dinucleotide, α-ketoglutarate, and iron. Future research should elucidate the potential for modifying cellular metabolism through dietary intervention for timely regulation of the epigenome as means for the prevention of cancer development.

High levels of Hsp90 cochaperone p23 promote tumor progression and poor prognosis in breast cancer by increasing lymph node metastases and drug resistance.

p23 is a heat shock protein 90 (Hsp90) cochaperone located in both the cytoplasm and nucleus that stabilizes unliganded steroid receptors, controls the catalytic activity of certain kinases, regulates protein-DNA dynamics, and is upregulated in several cancers. We had previously shown that p23-overexpressing MCF-7 cells (MCF-7+p23) exhibit increased invasion without affecting the estrogen-dependent proliferative response, which suggests that p23 differentially regulates genes controlling processes linked to breast tumor metastasis. To gain a comprehensive view of the effects of p23 on estrogen receptor (ER)-dependent and -independent gene expression, we profiled mRNA expression from control versus MCF-7+p23 cells in the absence and presence of estrogen. A number of p23-sensitive target genes involved in metastasis and drug resistance were identified. Most striking is that many of these genes are also misregulated in invasive breast cancers, including PMP22, ABCC3, AGR2, Sox3, TM4SF1, and p8 (NUPR1). Upregulation of the ATP-dependent transporter ABCC3 by p23 conferred resistance to the chemotherapeutic agents etoposide and doxorubicin in MCF-7+p23 cells. MCF-7+p23 cells also displayed higher levels of activated Akt and an expanded phosphoproteome relative to control cells, suggesting that elevated p23 also enhances cytoplasmic signaling pathways. For breast cancer patients, tumor stage together with high cytoplasmic p23 expression more accurately predicted disease recurrence and mortality than did stage alone. High nuclear p23 was found to be associated with high cytoplasmic p23, therefore both may promote tumor progression and poor prognosis by increasing metastatic potential and drug resistance in breast cancer patients.