Mechanistic insights into the inhibition of human placental glutathione S-transferase P1-1 by abscisic and gibberellic acids: An integrated experimental and computational study.

The interactions of the classic phytohormones gibberellic acid (gibberellin A3 , GA3 ) and abscisic acid (dormin, ABA), which antagonistically regulate several developmental processes and stress responses in higher plants, with human placental glutathione S-transferase P1-1 (hpGSTP1-1), an enzyme that plays a role in endo- or xenobiotic detoxification and regulation of cell survival and apoptosis, were investigated. The inhibitory potencies of ABA and GA3 against hpGSTP1, as well as the types of inhibition and the kinetic parameters, were determined by making use of both enzyme kinetic graphs and SPSS nonlinear regression models. The structural basis for the interaction between hpGSTP1-1 and phytohormones was predicted with the aid of molecular docking simulations. The IC50 values of ABA and GA3 were 5.3 and 5.0 mM, respectively. Both phytohormones inhibited hpGSTP1-1 in competitive manner with respect to the cosubstrates GSH and CDNB. When ABA was the inhibitor at [CDNB]f -[GSH]v and at [GSH]f -[CDNB]v , Vm , Km , and Ki values were statistically estimated to be 205 ± 16 µmol/min-mg protein, 1.32 ± 0.18 mM, 1.95 ± 0.25 mM and 175 ± 6 µmol/min-mg protein, 0.85 ± 0.06 mM, 1.85 ± 0.16 mM, respectively. On the other hand, the kinetic parameters Vm , Km , and Ki obtained with GA3 at [CDNB]f -[GSH]v and at [GSH]f -[CDNB]v were found to be 303 ± 14 µmol/min-mg protein, 1.77 ± 0.13 mM, 3.38 ± 0.26 mM and 249 ± 7 µmol/min-mg protein, 1.43 ± 0.07 mM, 2.89 ± 0.19 mM, respectively. Both phytohormones had the potential to engage in hydrogen-bonding and electrostatic interactions with the key residues that line the G- and H-sites of the enzyme's catalytic center. Inhibitory actions of ABA/GA3 on hpGSTP1-1 may guide medicinal chemists through the structure-based design of novel antineoplastic agents. It should be noted, however, that the same interactions may also render fetuses vulnerable to the potentially toxic effects of xenobiotics and noxious endobiotics.

Phase II trial of brentuximab vedotin in relapsed/refractory germ cell tumors.

Background CD-30 is highly expressed in some patients with non-seminomatous germ-cell tumors. Brentuximab vedotin is an antibody-drug conjugate directed to CD-30. We report a phase 2 trial of brentuximab vedotin in patients with chemo-refractory GCT. Patients and methods This is a single arm, two cohort phase 2 trial investigating brentuximab vedotin 1.8 mg/kg IV every 3 weeks until disease progression or intolerable toxicities in patients with relapsed GCT who have no curative options. Patients with mGCT who progressed after first line cisplatin-based chemotherapy and after at least 1 salvage regimen (high-dose or standard-dose chemotherapy) were eligible. CD30 expression was assessed and two cohorts defined: CD30 positive and CD30 negative/unknown. Results 18 patients were enrolled. Median age 34.7 (range, 23-56). All patients had non-seminoma. Median AFP 4.9 (range, 1-219,345) and hCG 282 (range, 0.6-172,064). Five patients had late relapse (> 2 years). Median number of previous chemotherapy regimens was 3 (range, 2-7). Ten patients received prior high-dose chemotherapy. Seven patients had positive CD30 staining. There were two grade 3 treatment-related adverse events. No partial or complete responses were observed. 6 patients achieved radiographic stable disease (range, 9-14.9 weeks), 5 had elevated AFP or hCG at trial entry and all 5 had transient > 50% decline in baseline AFP/hCG: 4 had CD30 -ve and 2 had CD30 + ve staining; 10 patients had progression of disease as their best response; 2 were not evaluable for response. Conclusion Brentuximab vedotin does not appear to have clinically meaningful single-agent activity in patients with refractory GCT.

Clinical and Genetic Risk Factors for Adverse Metabolic Outcomes in North American Testicular Cancer Survivors.

Background: Testicular cancer survivors (TCS) are at significantly increased risk for cardiovascular disease (CVD), with metabolic syndrome (MetS) an established risk factor. No study has addressed clinical and genetic MetS risk factors in North American TCS. Patients and Methods: TCS were aged <55 years at diagnosis and received first-line chemotherapy. Patients underwent physical examination, and had lipid panels, testosterone, and soluble cell adhesion molecule-1 (sICAM-1) evaluated. A single nucleotide polymorphism in rs523349 (5-α-reductase gene, SRD5A2), recently implicated in MetS risk, was genotyped. Using standard criteria, MetS was defined as ≥3 of the following: hypertension, abdominal obesity, hypertriglyceridemia, decreased high-density lipoprotein (HDL) cholesterol level, and diabetes. Matched controls were derived from the National Health and Nutrition Examination Survey. Results: We evaluated 486 TCS (median age, 38.1 years). TCS had a higher prevalence of hypertension versus controls (43.2% vs 30.7%; P<.001) but were less likely to have decreased HDL levels (23.7% vs 34.8%; P<.001) or abdominal obesity (28.2% vs 40.1%; P<.001). Overall MetS frequency was similar in TCS and controls (21.0% vs 22.4%; P=.59), did not differ by treatment (P=.20), and was not related to rs523349 (P=.61). For other CVD risk factors, TCS were significantly more likely to have elevated low-density lipoprotein (LDL) cholesterol levels (17.7% vs 9.3%; P<.001), total cholesterol levels (26.3% vs 11.1%; P<.001), and body mass index ≥25 kg/m2 (75.1% vs 69.1%; P=.04). On multivariate analysis, age at evaluation (P<.001), testosterone level ≤3.0 ng/mL (odds ratio [OR], 2.06; P=.005), and elevated sICAM-1 level (ORhighest vs lowest quartile, 3.58; P=.001) were significantly associated with MetS. Conclusions and Recommendations: Metabolic abnormalities in TCS are characterized by hypertension and increased LDL and total cholesterol levels but lower rates of decreased HDL levels and abdominal obesity, signifying possible shifts in fat distribution and fat metabolism. These changes are accompanied by hypogonadism and inflammation. TCS have a high prevalence of CVD risk factors that may not be entirely captured by standard MetS criteria. Cancer treatment-associated MetS requires further characterization.

Altered metabolite levels and correlations in patients with colorectal cancer and polyps detected using seemingly unrelated regression analysis.

Introduction: Metabolomics technologies enable the identification of putative biomarkers for numerous diseases; however, the influence of confounding factors on metabolite levels poses a major challenge in moving forward with such metabolites for pre-clinical or clinical applications. Objectives: To address this challenge, we analyzed metabolomics data from a colorectal cancer (CRC) study, and used seemingly unrelated regression (SUR) to account for the effects of confounding factors including gender, BMI, age, alcohol use, and smoking. Methods: A SUR model based on 113 serum metabolites quantified using targeted mass spectrometry, identified 20 metabolites that differentiated CRC patients (n = 36), patients with polyp (n = 39), and healthy subjects (n = 83). Models built using different groups of biologically related metabolites achieved improved differentiation and were significant for 26 out of 29 groups. Furthermore, the networks of correlated metabolites constructed for all groups of metabolites using the ParCorA algorithm, before or after application of the SUR model, showed significant alterations for CRC and polyp patients relative to healthy controls. Results: The results showed that demographic covariates, such as gender, BMI, BMI2, and smoking status, exhibit significant confounding effects on metabolite levels, which can be modeled effectively. Conclusion: These results not only provide new insights into addressing the major issue of confounding effects in metabolomics analysis, but also shed light on issues related to establishing reliable biomarkers and the biological connections between them in a complex disease.

1q amplification and PHF19 expressing high-risk cells are associated with relapsed/refractory multiple myeloma.

Multiple Myeloma is an incurable plasma cell malignancy with a poor survival rate that is usually treated with immunomodulatory drugs (iMiDs) and proteosome inhibitors (PIs). The malignant plasma cells quickly become resistant to these agents causing relapse and uncontrolled growth of resistant clones. From whole genome sequencing (WGS) and RNA sequencing (RNA-seq) studies, different high-risk translocation, copy number, mutational, and transcriptional markers can be identified. One of these markers, PHF19, epigenetically regulates cell cycle and other processes and is already studied using RNA-seq. In this study, we generate a large (325,025 cells and 49 patients) single cell multi-omic dataset and jointly quantify ATAC- and RNA-seq for each cell and matched genomic profiles for each patient. We identify an association between one plasma cell subtype with myeloma progression that we call relapsed/refractory plasma cells (RRPCs). These cells are associated with chromosome 1q alterations, TP53 mutations, and higher expression of PHF19. We also identify downstream regulation of cell cycle inhibitors in these cells, possible regulation by the transcription factor (TF) PBX1 on chromosome 1q, and determine that PHF19 may be acting primarily through this subset of cells.

Maintenance Oral Etoposide After High-Dose Chemotherapy (HDCT) for Patients With Relapsed Metastatic Germ-Cell Tumors (mGCT).

BACKGROUND: HDCT and peripheral-blood stem-cell transplant (PBSCT) can cure up to 60% of pts with relapsed mGCT. Maintenance daily oral etoposide after salvage therapy has demonstrated potential clinical benefit. We now evaluate the potential role of maintenance etoposide versus observation post HDCT+PBSCT in this nonrandomized retrospective analysis. METHODS: The prospectively maintained Indiana University testicular cancer database was interrogated. Patients with relapsed non-seminoma who completed HDCT+PBSCT and achieved complete serologic remission and hematologic recovery were evaluated. Outcomes of pts who received maintenance etoposide (N = 141) were compared to pts who were observed (N = 242). In this retrospective study, Kaplan-Meier method was used to analyze progression-free survival (PFS) and overall survival (OS). Univariable and multivariable cox regression models were used to determine variables associated with PFS. We also performed an additional analysis to compare the survival outcomes in the platinum-refractory patients' subgroup based on maintenance etoposide treatment. RESULTS: Two-year PFS in the maintenance etoposide vs observation group was 55% vs. 46% (P = .028). Two-year OS was 61% vs 54% (P = .04). A multivariable analysis was performed, including the factors: primary tumor site (testis vs. mediastinum), IGCCCG risk, platinum refractory, HDCT line of therapy (2nd vs ≥3rd), tumor marker amplitude at HDCT initiation, and receipt of maintenance etoposide post HDCT vs. observation. Maintenance etoposide was confirmed as an independent predictor of improved PFS with HR 0.51 [95% CI, 0.37-0.70] (P < .001). Two-year OS and PFS for platinum-refractory patients who received maintenance etoposide vs. observation group were 50.2% vs. 26.1% (P < .0001) and 44.2% vs.. 23.1% (P = .0003), respectively. There was no statistically significant difference in 2-year OS and PFS between the platinum-sensitive patients who received maintenance etoposide and those who were observed. CONCLUSION: Daily oral etoposide therapy produced encouraging efficacy results in patients with relapsed non-seminoma GCT (NSGCT) who completed HDCT and PBSCT and achieved complete serologic remission and hematologic recovery. Patients with platinum-refractory disease and poor prognostic features are potential candidates for daily maintenance oral etoposide post HDCT. These data have led to an ongoing randomized phase II clinical trial (NCT04804007).

Identifying 1q amplification and PHF19 expressing high-risk cells associated with relapsed/refractory multiple myeloma.

Multiple Myeloma is an incurable plasma cell malignancy with a poor survival rate that is usually treated with immunomodulatory drugs (iMiDs) and proteosome inhibitors (PIs). The malignant plasma cells quickly become resistant to these agents causing relapse and uncontrolled growth of resistant clones. From whole genome sequencing (WGS) and RNA sequencing (RNA-seq) studies, different high-risk translocation, copy number, mutational, and transcriptional markers have been identified. One of these markers, PHF19, epigenetically regulates cell cycle and other processes and has already been studied using RNA-seq. In this study a massive (325,025 cells and 49 patients) single cell multiomic dataset was generated with jointly quantified ATAC- and RNA-seq for each cell and matched genomic profiles for each patient. We identified an association between one plasma cell subtype with myeloma progression that we have called relapsed/refractory plasma cells (RRPCs). These cells are associated with 1q alterations, TP53 mutations, and higher expression of PHF19. We also identified downstream regulation of cell cycle inhibitors in these cells, possible regulation of the transcription factor (TF) PBX1 on 1q, and determined that PHF19 may be acting primarily through this subset of cells.

Axatilimab for Chronic Graft-Versus-Host Disease After Failure of at Least Two Prior Systemic Therapies: Results of a Phase I/II Study.

PURPOSE: Chronic graft-versus-host disease (cGVHD) remains the major cause of late morbidity after allogeneic hematopoietic cell transplantation. Colony-stimulating factor 1 receptor (CSF-1R)-dependent macrophages promote cGVHD fibrosis, and their elimination in preclinical studies ameliorated cGVHD. Axatilimab is a humanized monoclonal antibody that inhibits CSF-1R signaling and restrains macrophage development. PATIENTS AND METHODS: This phase I (phI)/phase II (phII) open-label study (ClinicalTrials.gov identifier: NCT03604692) evaluated safety, tolerability, and efficacy of axatilimab in patients age ≥ 6 years with active cGVHD after ≥ 2 prior systemic therapy lines. Primary objectives in phI were to identify the optimal biologic and recommended phII dose and in phII to evaluate the overall (complete and partial) response rate (ORR) at the start of treatment cycle 7. RESULTS: Forty enrolled patients (17 phI; 23 phII) received at least one axatilimab dose. In phI, a dose of 3 mg/kg given once every 4 weeks met the optimal biologic dose definition. Two dose-limiting toxicities occurred at the 3 mg/kg dose given once every 2 weeks. At least one treatment-related adverse event (TRAE) was observed in 30 patients with grade ≥ 3 TRAEs in eight patients, the majority known on-target effects of CSF-1R inhibition. No cytomegalovirus reactivations occurred. With the 50% ORR at cycle 7 day 1, the phII cohort met the primary efficacy end point. Furthermore, the ORR in the first six cycles, an end point supporting regulatory approvals, was 82%. Responses were seen in all affected organs regardless of prior therapy. Fifty-eight percent of patients reported significant improvement in cGVHD-related symptoms using the Lee Symptom Scale. On-target activity of axatilimab was suggested by the decrease in skin CSF-1R-expressing macrophages. CONCLUSION: Targeting profibrotic macrophages with axatilimab is a therapeutically promising novel strategy with a favorable safety profile for refractory cGVHD.

Diagnostic Evidence GAuge of Single cells (DEGAS): a flexible deep transfer learning framework for prioritizing cells in relation to disease.

We propose DEGAS (Diagnostic Evidence GAuge of Single cells), a novel deep transfer learning framework, to transfer disease information from patients to cells. We call such transferrable information "impressions," which allow individual cells to be associated with disease attributes like diagnosis, prognosis, and response to therapy. Using simulated data and ten diverse single-cell and patient bulk tissue transcriptomic datasets from glioblastoma multiforme (GBM), Alzheimer's disease (AD), and multiple myeloma (MM), we demonstrate the feasibility, flexibility, and broad applications of the DEGAS framework. DEGAS analysis on myeloma single-cell transcriptomics identified PHF19high myeloma cells associated with progression. Availability: https://github.com/tsteelejohnson91/DEGAS .

Intron retention-induced neoantigen load correlates with unfavorable prognosis in multiple myeloma.

Neoantigen peptides arising from genetic alterations may serve as targets for personalized cancer vaccines and as positive predictors of response to immune checkpoint therapy. Mutations in genes regulating RNA splicing are common in hematological malignancies leading to dysregulated splicing and intron retention (IR). In this study, we investigated IR as a potential source of tumor neoantigens in multiple myeloma (MM) patients and the relationship of IR-induced neoantigens (IR-neoAg) with clinical outcomes. MM-specific IR events were identified in RNA-sequencing data from the Multiple Myeloma Research Foundation CoMMpass study after removing IR events that also occurred in normal plasma cells. We quantified the IR-neoAg load by assessing IR-induced novel peptides that were predicted to bind to major histocompatibility complex (MHC) molecules. We found that high IR-neoAg load was associated with poor overall survival in both newly diagnosed and relapsed MM patients. Further analyses revealed that poor outcome in MM patients with high IR-neoAg load was associated with high expression levels of T-cell co-inhibitory molecules and elevated interferon signaling activity. We also found that MM cells exhibiting high IR levels had lower MHC-II protein abundance and treatment of MM cells with a spliceosome inhibitor resulted in increased MHC-I protein abundance. Our findings suggest that IR-neoAg may represent a novel biomarker of MM patient clinical outcome and further that targeting RNA splicing may serve as a potential therapeutic strategy to prevent MM immune escape and promote response to checkpoint blockade.

Effect of Sirolimus levels between days 11 and 20 after allogeneic stem cell transplantation on the risk of hepatic sinusoidal obstruction syndrome.

Sinusoidal obstruction syndrome (SOS) is a serious complication of hematopoietic stem cell transplantation (HSCT). Sirolimus plus tacrolimus is an accepted regimen for graft-versus-host disease (GVHD) prophylaxis, with both agents implicated as risk factors for SOS. We analyzed 260 consecutive patients who underwent allogeneic HSCT following myeloablative conditioning using total body irradiation (TBI)-based (n = 151) or chemotherapy only (n = 109) regimens, with sirolimus plus tacrolimus for GVHD prophylaxis. SOS occurred in 28 patients at a median of 22 (range, 12-58) days. Mean sirolimus trough levels were higher between days 11 and 20 following transplant in patients who developed SOS (10.3 vs. 8.5 ng/ml, P = 0.008), with no significant difference in mean trough levels between days 0 and 10 (P = 0.67) and days 21-30 (P = 0.37). No differences in mean tacrolimus trough levels during the same time intervals were observed between those developing SOS and others. On multivariable analysis, a mean sirolimus trough level ≥ 9 ng/ml between days 11 and 20 increased the risk of SOS (hazard ratio 3.68, 95% CI: 1.57-8.67, P = 0.003), together with a longer time from diagnosis to transplant (P = 0.004) and use of TBI (P = 0.006). Our results suggest that mean trough sirolimus levels ≥ 9 ng/mL between days 11 and 20 post transplant may increase the risk of SOS and should be avoided.

Vaccine and Cell-based Therapeutic Approaches in Acute Myeloid Leukemia.

Over the past decade, our increased understanding of the interactions between the immune system and cancer cells has led to paradigm shifts in the clinical management of solid and hematologic malignancies. The incorporation of immune-targeted strategies into the treatment landscape of acute myeloid leukemia (AML), however, has been challenging. While this is in part due to the inability of the immune system to mount an effective tumor-specific immunogenic response against the heterogeneous nature of AML, the decreased immunogenicity of AML cells also represents a major obstacle in the effort to design effective immunotherapeutic strategies. In fact, AML cells have been shown to employ sophisticated escape mechanisms to evade elimination, such as direct immunosuppression of natural killer cells and decreased surface receptor expression leading to impaired recognition by the immune system. Yet, cellular and humoral immune reactions against tumor-associated antigens (TAA) of acute leukemia cells have been reported and the success of allogeneic stem cell transplantation and monoclonal antibodies in the treatment of AML clearly provides proof that an immunotherapeutic approach is feasible in the management of this disease. This review discusses the recent progress and persisting challenges in cellular immunotherapy for patients with AML.

Risk Factors for Acute Kidney Injury During High-dose Chemotherapy and Outcomes for Patients With Relapsed Germ Cell Tumors.

BACKGROUND: Patients with relapsed germ cell tumors (GCTs) can be cured with salvage chemotherapy. We evaluated the risk factors and outcomes of patients who had developed acute kidney injury (AKI) during high-dose chemotherapy (HDCT) for relapsed GCT. PATIENTS AND METHODS: All patients were scheduled to receive 2 consecutive courses of HDCT per our standard protocol. The characteristics and outcomes of the patients with stage ≥ 3 AKI were analyzed and compared with those without stage ≥ 3 AKI. RESULTS: Of 462 patients, 21 (4.5%) developed stage ≥ 3 AKI. Of these 21 patients, 18 had required hemodialysis during HDCT and 6 had died during HDCT. Of the 15 patients who had survived HDCT, 10 experienced recovery of renal function to baseline. AKI had occurred in the first cycle of HDCT in 18 patients. These patients were also more likely to have received HDCT in a third-line setting or further, to have Eastern Cooperative Oncology Group performance status of 1 or 2, and to have experienced gastrointestinal, hepatic, pulmonary, and infectious grade ≥ 3 toxicities. At a median follow-up of 10 months after HDCT, 5 patients had no evidence of disease, 3 were alive with disease, 6 had died of disease, 6 had died of complications from HDCT, and 1 had been lost to follow-up. CONCLUSIONS: Irreversible AKI during HDCT for relapsed GCT is uncommon but is associated with greater rates of infectious, gastrointestinal, hepatic, and pulmonary complications and treatment-related death. These patients were also more heavily pretreated and had a lower baseline performance status. However, most surviving patients had recovered their renal function and 5 of 21 were alive with no evidence of disease.

Solid and Hematologic Neoplasms After Testicular Cancer: A US Population-Based Study of 24 900 Survivors.

BACKGROUND: No large US population-based study focusing on recent decades, to our knowledge, has comprehensively examined risks of second malignant solid and hematological neoplasms (solid-SMN and heme-SMN) after testicular cancer (TC), taking into account initial therapy and histological type. METHODS: Standardized incidence ratios (SIR) vs the general population and 95% confidence intervals (CI) for solid-SMN and heme-SMN were calculated for 24 900 TC survivors (TCS) reported to the National Cancer Institute's Surveillance, Epidemiology, and End Results registries (1973-2014). All statistical tests were two-sided. RESULTS: The median age at TC diagnosis was 33 years. Initial management comprised chemotherapy (n = 6340), radiotherapy (n = 9058), or surgery alone (n = 8995). During 372 709 person-years of follow-up (mean = 15 years), 1625 TCS developed solid-SMN and 228 (107 lymphomas, 92 leukemias, 29 plasma cell dyscrasias) developed heme-SMN. Solid-SMN risk was increased 1.06-fold (95% CI = 1.01 to 1.12), with elevated risks following radiotherapy (SIR = 1.13, 95% CI = 1.06 to 1.21) and chemotherapy (SIR = 1.36, 95% CI = 1.12 to 1.41) but not surgery alone (SIR = 0.83, 95% CI = 0.75 to 0.92). Corresponding risks for seminoma were 1.13 (95% CI = 1.06 to 1.21), 1.28 (95% CI = 1.02 to 1.58), and 0.87 (95% CI = 0.74 to 1.01) and for nonseminoma were 1.05 (95% CI = 0.67 to 1.56), 1.25 (95% CI = 1.08 to 1.43), and 0.80 (95% CI = 0.70 to 0.92), respectively. Thirty-year cumulative incidences of solid-SMN after radiotherapy, chemotherapy, and surgery alone were 16.9% (95% CI = 15.7% to 18.1%), 10.1% (95% CI = 8.8% to 11.5%), and 8.8% (95% CI = 7.8% to 9.9%), respectively (P < .0001). Increased leukemia risks after chemotherapy (SIR = 2.68, 95% CI = 1.70 to 4.01) were driven by statistically significant sevenfold excesses of acute myeloid leukemia 1 to 10 years after TC diagnosis. Risks for lymphoma and plasma cell dyscrasias were not elevated. CONCLUSIONS: We report statistically significant excesses of solid-SMN affecting 1 in 6 TCS 30 years after radiotherapy, and 2.7-fold risks of leukemias after chemotherapy, mostly acute myeloid leukemia. Efforts to minimize chemotherapy and radiotherapy exposures for TC should continue. TCS should be counseled about cancer prevention and screening.

Protective effect of pomegranate-derived products on UVB-mediated damage in human reconstituted skin.

Solar ultraviolet (UV) radiation, particularly its UVB (290-320 nm) component, is the primary cause of many adverse biological effects including photoageing and skin cancer. UVB radiation causes DNA damage, protein oxidation and induces matrix metalloproteinases (MMPs). Photochemoprevention via the use of botanical antioxidants in affording protection to human skin against UVB damage is receiving increasing attention. Pomegranate, from the tree Punica granatum, contains anthocyanins and hydrolysable tannins and possesses strong antioxidant and anti-tumor-promoting properties. In this study, we determined the effect of pomegranate-derived products--POMx juice, POMx extract and pomegranate oil (POMo)--against UVB-mediated damage using reconstituted human skin (EpiDerm(TM) FT-200). EpiDerm was treated with POMx juice (1-2 microl/0.1 ml/well), POMx extract (5-10 microg/0.1 ml/well) and POMo (1-2 microl/0.1 ml/well) for 1 h prior to UVB (60 mJ/cm(2)) irradiation and was harvested 12 h post-UVB to assess protein oxidation, markers of DNA damage and photoageing by Western blot analysis and immunohistochemistry. Pretreatment of Epiderm with pomegranate-derived products resulted in inhibition of UVB-induced (i) cyclobutane pyrimidine dimers (CPD), (ii) 8-dihydro-2'-deoxyguanosine (8-OHdG), (iii) protein oxidation and (iv) proliferating cell nuclear antigen (PCNA) protein expression. We also found that pretreatment of Epiderm with pomegranate-derived products resulted in inhibition of UVB-induced (i) collagenase (MMP-1), (ii) gelatinase (MMP-2, MMP-9), (iii) stromelysin (MMP-3), (iv) marilysin (MMP-7), (v) elastase (MMP-12) and (vi) tropoelastin. Gelatin zymography revealed that pomegranate-derived products inhibited UVB-induced MMP-2 and MMP-9 activities. Pomegranate-derived products also caused a decrease in UVB-induced protein expression of c-Fos and phosphorylation of c-Jun. Collectively, these results suggest that all three pomegranate-derived products may be useful against UVB-induced damage to human skin.

Inhibition of epidermal growth factor receptor signaling pathway by delphinidin, an anthocyanidin in pigmented fruits and vegetables.

Inhibitors of the epidermal growth factor receptor (EGFR) have generated considerable hope for cancer treatment, specifically for lung and breast cancers. Therefore, identification of a natural, nontoxic agent(s) as an inhibitor of EGFR is of considerable importance. Delphinidin, an anthocyanidin present in pigmented fruits and vegetables, possesses potent antioxidant and antiproliferative properties. In our study, employing EGFR positive breast cancer AU-565 cells and immortalized MCF-10A cells, we evaluated the effect of delphinidin on EGFR and its downstream signaling pathways. Delphinidin (5-40 microM; 3 hr) treatment of both AU-565 cells and MCF-10A cells inhibited the (i) phosphorylation of EGFR, (ii) activation of PI3K, (iii) phosphorylation of AKT and MAPK. Further, delphinidin treatment of AU-565 cells inhibited EGF-induced autophosphorylation of EGFR, AKT and MAPK, activation of PI3K and cell invasion. We then compared the growth inhibitory effects of delphinidin (5-40 microM; 48 hr), and found that it resulted in a decrease in cell growth of AU-565 and MCF-10A cells but had only minimal effects on normal mammary epithelial 184A1 cells. Treatment of AU-565 cells with delphinidin resulted in (i) induction of apoptosis, (ii) cleavage of PARP protein, (iii) activation of caspase-3 and (iv) downregulation of Bcl-2 with an increase in the expression of Bax. In summary, our study identifies a naturally occurring dietary agent delphinidin as an effective inhibitor of EGFR signaling in breast cancer cells. We suggest that delphinidin could be developed as an agent for the management of EGFR positive human cancers.

Consecutive epigenetically-active agent combinations act in ID1-RUNX3-TET2 and HOXA pathways for Flt3ITD+ve AML.

Co-occurrence of Flt3ITD and TET2 mutations provoke an animal model of AML by epigenetic repression of Wnt pathway antagonists, including RUNX3, and by hyperexpression of ID1, encoding Wnt agonist. These affect HOXA over-expression and treatment resistance. A comparable epigenetic phenotype was identified among adult AML patients needing novel intervention. We chose combinations of targeted agents acting on distinct effectors, at the levels of both signal transduction and chromatin remodeling, in relapsed/refractory AML's, including Flt3ITD+ve, described with a signature of repressed tumor suppressor genes, involving Wnt antagonist RUNX3, occurring along with ID1 and HOXA over-expressions. We tracked patient response to combination of Flt3/Raf inhibitor, Sorafenib, and Vorinostat, pan-histone deacetylase inhibitor, without or with added Bortezomib, in consecutive phase I trials. A striking association of rapid objective remissions (near-complete, complete responses) was noted to accompany induced early pharmacodynamic changes within patient blasts in situ, involving these effectors, significantly linking RUNX3/Wnt antagonist de-repression (80%) and ID1 downregulation (85%), to a response, also preceded by profound HOXA9 repression. Response occurred in context of concurrent TET2 mutation/hypomorphy and Flt3ITD+ve mutation (83% of complete responses). Addition of Bortezomib to the combination was vital to attainment of complete response in Flt3ITD+ve cases exhibiting such Wnt pathway dysregulation.

Predicting Cardiovascular Disease Among Testicular Cancer Survivors After Modern Cisplatin-based Chemotherapy: Application of the Framingham Risk Score.

BACKGROUND: Testicular cancer survivors (TCSs) are at increased risk of cardiovascular disease (CVD) after cisplatin-based chemotherapy (CBCT). Identifying at-risk survivors would allow early intervention, but risk prediction tools such as the Framingham Risk Score (FRS) have not been applied to TCSs given modern chemotherapy. METHODS: TCSs > 1 year post-CBCT were evaluated. Associations between FRS and clinical, socioeconomic, and lifestyle measures and treatment regimen (4 cycles, etoposide and cisplatin [EP × 4]); 3 or 4 cycles, bleomycin plus EP (BEP × 3, BEP × 4) were analyzed with general linear multivariable models. Controls from the National Health and Nutrition Examination Survey were matched 1:1 to TCSs by age, race, and education with differences in mean FRS evaluated with 2-sided t tests. RESULTS: Of 787 TCSs (median age, 37.3 years; median follow-up, 4.2 years), 284, 342, and 161 received EP × 4, BEP × 3, or BEP × 4, respectively. TCSs had higher median systolic blood pressure (126 vs. 119 mm Hg; P < .001), but fewer were smokers (8.4% vs. 28.2%; P < .001) than controls. In multivariable analysis, no significant differences in FRS between EP × 4, BEP × 3, and BEP × 4 were observed, but less than college education (P < .001) and lack of vigorous exercise (P = .006) were associated with higher FRS. Mean FRS did not differ between TCSs and controls (6.8% vs. 7.3%; P = .67). CONCLUSION: This is the first study to apply the office-based FRS to TCSs. Chemotherapy regimen (BEP × 3 vs. EP × 4) was not associated with FRS, but less educated and less vigorously active patients had higher FRS, and present a high-risk subgroup for intense follow-up and counseling.

Inhibition of UVB-mediated oxidative stress and markers of photoaging in immortalized HaCaT keratinocytes by pomegranate polyphenol extract POMx.

In recent years there has been an increase in use of botanicals with antioxidant properties as skin photoprotective agents. Pomegranate (Punica granatum L.) fruit possesses strong antioxidant and antiinflammatory properties. Recently, we have shown that pomegranate-derived products rich in anthocyanidins and ellagitannins inhibit UVB-mediated activation of nuclear factor kappa B and modulate UVA-mediated cell proliferation pathways in normal human epidermal keratinocytes. In this study, we evaluated the effect of polyphenol-rich pomegranate fruit extract (POMx) on UVB-induced oxidative stress and photoaging in human immortalized HaCaT keratinocytes. Our data show that pretreatment of HaCaT cells with POMx (10-40 microg mL(-1)) inhibited UVB (15-30 mJ cm(-2))-mediated (1) decrease in cell viability, (2) decrease in intracellular glutathione content and (3) increase in lipid peroxidation. Employing immunoblot analysis we found that pretreatment of HaCaT cells with POMx inhibited UVB-induced (1) upregulation of MMP-1, -2, -7 and -9, (2) decrease in TIMP-1, (3) phosphorylation of MAPKs and (iv) phosphorylation of c-jun, whereas no effect was observed on UVB-induced c-fos protein levels. These results suggest that POMx protects HaCaT cells against UVB-induced oxidative stress and markers of photoaging and could be a useful supplement in skin care products.

High-dose sitagliptin for systemic inhibition of dipeptidylpeptidase-4 to enhance engraftment of single cord umbilical cord blood transplantation.

Delayed engraftment remains a limitation of umbilical cord blood (UCB) transplantation. We previously showed that inhibition of dipeptidylpeptidase (DPP)-4 using sitagliptin 600 mg daily was safe with encouraging results on engraftment, but inhibition was not sustained. We evaluated the efficacy and feasibility of higher doses of sitagliptin to enhance engraftment of UCB in patients with hematological cancers. Fifteen patients, median age 41 (range, 18-59) years, received single UCB grafts matched at 4 (n=11) or 5 (n=4) of 6 HLA loci with median nucleated cell dose of 3.5 (range, 2.57-4.57) x107/kg. Sitagliptin 600 mg every 12 hours was administered days -1 to +2. All patients engrafted by day 30, with 12 (80%) engrafting by day 21. The median time to neutrophil engraftment was 19 (range, 12-30) days. Plasma DPP-4 activity was better inhibited with a mean residual trough DPP-4 activity of 70%±19%. Compared to patients previously treated with 600 mg/day, sitagliptin 600 mg every 12 hours appeared to improve engraftment, supporting the hypothesis that more sustained DPP-4 inhibition is required. In-vivo inhibition of DPP-4 using high-dose sitagliptin compares favorably with other approaches to enhance UCB engraftment with greater simplicity, and may show synergy in combination with other strategies.