A Novel Liver Cancer-Selective Histone Deacetylase Inhibitor Is Effective Against Hepatocellular Carcinoma and Induces Durable Responses with Immunotherapy.

Hepatocellular cancer (HCC) progression is facilitated by gene-silencing chromatin histone hypoacetylation due to histone deacetylases (HDACs) activation. However, inhibiting HDACs, an effective treatment for lymphomas, has shown limited success in solid tumors. We report the discovery of a class of HDAC inhibitors (HDACi) that demonstrates exquisite selective cytotoxicity against human HCC cells. The lead compound STR-V-53 (3) showed favorable safety profile in mice and robustly suppressed tumor growth in orthotopic xenograft models of HCC. When combined with the anti-HCC drug sorafenib, STR-V-53 showed greater in vivo efficacy. Moreover, STR-V-53 combined with anti-PD1 therapy increased the CD8+ to regulatory T-cell (Treg) ratio and survival in an orthotopic HCC model in immunocompetent mice. This combination therapy resulted in durable responses in 40% of the mice. Collectively, our data demonstrate that the novel HDACi STR-V-53 is an effective anti-HCC agent that can induce profound responses when combined with standard immunotherapy.

A missense SNP in the tumor suppressor SETD2 reduces H3K36me3 and mitotic spindle integrity in Drosophila.

Mutations in SETD2 are among the most prevalent drivers of renal cell carcinoma (RCC). We identified a novel single nucleotide polymorphism (SNP) in SETD2, E902Q, within a subset of RCC patients, which manifests as both an inherited or tumor-associated somatic mutation. To determine if the SNP is biologically functional, we used CRISPR-based genome editing to generate the orthologous mutation within the Drosophila melanogaster Set2 gene. In Drosophila, the homologous amino acid substitution, E741Q, reduces H3K36me3 levels comparable to Set2 knockdown, and this loss is rescued by reintroduction of a wild-type Set2 transgene. We similarly uncovered significant defects in spindle morphogenesis, consistent with the established role of SETD2 in methylating α-Tubulin during mitosis to regulate microtubule dynamics and maintain genome stability. These data indicate the Set2 E741Q SNP affects both histone methylation and spindle integrity. Moreover, this work further suggests the SETD2 E902Q SNP may hold clinical relevance.

Discovery of 5'-Substituted 5-Fluoro-2'-deoxyuridine Monophosphate Analogs: A Novel Class of Thymidylate Synthase Inhibitors.

5-Fluorouracil and 5-fluorouracil-based prodrugs have been used clinically for decades to treat cancer. Their anticancer effects are most prominently ascribed to inhibition of thymidylate synthase (TS) by metabolite 5-fluoro-2'-deoxyuridine 5'-monophosphate (FdUMP). However, 5-fluorouracil and FdUMP are subject to numerous unfavorable metabolic events that can drive undesired systemic toxicity. Our previous research on antiviral nucleotides suggested that substitution at the nucleoside 5'-carbon imposes conformational restrictions on the corresponding nucleoside monophosphates, rendering them poor substrates for productive intracellular conversion to viral polymerase-inhibiting triphosphate metabolites. Accordingly, we hypothesized that 5'-substituted analogs of FdUMP, which is uniquely active at the monophosphate stage, would inhibit TS while preventing undesirable metabolism. Free energy perturbation-derived relative binding energy calculations suggested that 5'(R)-CH3 and 5'(S)-CF3 FdUMP analogs would maintain TS potency. Herein, we report our computational design strategy, synthesis of 5'-substituted FdUMP analogs, and pharmacological assessment of TS inhibitory activity.

Mutation of the Drosophila melanogaster serotonin transporter dSERT impacts sleep, courtship, and feeding behaviors.

The Serotonin Transporter (SERT) regulates extracellular serotonin levels and is the target of most current drugs used to treat depression. The mechanisms by which inhibition of SERT activity influences behavior are poorly understood. To address this question in the model organism Drosophila melanogaster, we developed new loss of function mutations in Drosophila SERT (dSERT). Previous studies in both flies and mammals have implicated serotonin as an important neuromodulator of sleep, and our newly generated dSERT mutants show an increase in total sleep and altered sleep architecture that is mimicked by feeding the SSRI citalopram. Differences in daytime versus nighttime sleep architecture as well as genetic rescue experiments unexpectedly suggest that distinct serotonergic circuits may modulate daytime versus nighttime sleep. dSERT mutants also show defects in copulation and food intake, akin to the clinical side effects of SSRIs and consistent with the pleomorphic influence of serotonin on the behavior of D. melanogaster. Starvation did not overcome the sleep drive in the mutants and in male dSERT mutants, the drive to mate also failed to overcome sleep drive. dSERT may be used to further explore the mechanisms by which serotonin regulates sleep and its interplay with other complex behaviors.

Digital haptics improve speed of visual search performance in a dual-task setting.

Dashboard-mounted touchscreen tablets are now common in vehicles. Screen/phone use in cars likely shifts drivers' attention away from the road and contributes to risk of accidents. Nevertheless, vision is subject to multisensory influences from other senses. Haptics may help maintain or even increase visual attention to the road, while still allowing for reliable dashboard control. Here, we provide a proof-of-concept for the effectiveness of digital haptic technologies (hereafter digital haptics), which use ultrasonic vibrations on a tablet screen to render haptic perceptions. Healthy human participants (N = 25) completed a divided-attention paradigm. The primary task was a centrally-presented visual conjunction search task, and the secondary task entailed control of laterally-presented sliders on the tablet. Sliders were presented visually, haptically, or visuo-haptically and were vertical, horizontal or circular. We reasoned that the primary task would be performed best when the secondary task was haptic-only. Reaction times (RTs) on the visual search task were fastest when the tablet task was haptic-only. This was not due to a speed-accuracy trade-off; there was no evidence for modulation of VST accuracy according to modality of the tablet task. These results provide the first quantitative support for introducing digital haptics into vehicle and similar contexts.

Urine-Based Metabolomics and Machine Learning Reveals Metabolites Associated with Renal Cell Carcinoma Stage.

Urine metabolomics profiling has potential for non-invasive RCC staging, in addition to providing metabolic insights into disease progression. In this study, we utilized liquid chromatography-mass spectrometry (LC-MS), nuclear magnetic resonance (NMR), and machine learning (ML) for the discovery of urine metabolites associated with RCC progression. Two machine learning questions were posed in the study: Binary classification into early RCC (stage I and II) and advanced RCC stages (stage III and IV), and RCC tumor size estimation through regression analysis. A total of 82 RCC patients with known tumor size and metabolomic measurements were used for the regression task, and 70 RCC patients with complete tumor-nodes-metastasis (TNM) staging information were used for the classification tasks under ten-fold cross-validation conditions. A voting ensemble regression model consisting of elastic net, ridge, and support vector regressor predicted RCC tumor size with a R2 value of 0.58. A voting classifier model consisting of random forest, support vector machines, logistic regression, and adaptive boosting yielded an AUC of 0.96 and an accuracy of 87%. Some identified metabolites associated with renal cell carcinoma progression included 4-guanidinobutanoic acid, 7-aminomethyl-7-carbaguanine, 3-hydroxyanthranilic acid, lysyl-glycine, glycine, citrate, and pyruvate. Overall, we identified a urine metabolic phenotype associated with renal cell carcinoma stage, exploring the promise of a urine-based metabolomic assay for staging this disease.

Machine Learning-Enabled Renal Cell Carcinoma Status Prediction Using Multiplatform Urine-Based Metabolomics.

Renal cell carcinoma (RCC) is diagnosed through expensive cross-sectional imaging, frequently followed by renal mass biopsy, which is not only invasive but also prone to sampling errors. Hence, there is a critical need for a noninvasive diagnostic assay. RCC exhibits altered cellular metabolism combined with the close proximity of the tumor(s) to the urine in the kidney, suggesting that urine metabolomic profiling is an excellent choice for assay development. Here, we acquired liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR) data followed by the use of machine learning (ML) to discover candidate metabolomic panels for RCC. The study cohort consisted of 105 RCC patients and 179 controls separated into two subcohorts: the model cohort and the test cohort. Univariate, wrapper, and embedded methods were used to select discriminatory features using the model cohort. Three ML techniques, each with different induction biases, were used for training and hyperparameter tuning. Assessment of RCC status prediction was evaluated using the test cohort with the selected biomarkers and the optimally tuned ML algorithms. A seven-metabolite panel predicted RCC in the test cohort with 88% accuracy, 94% sensitivity, 85% specificity, and 0.98 AUC. Metabolomics Workbench Study IDs are ST001705 and ST001706.

A panel of selected serum protein biomarkers for the detection of aggressive prostate cancer.

Background: Current PSA-based tests used to detect prostate cancer (PCa) lack sufficient specificity, leading to significant overdetection and overtreatment. Our previous studies showed that serum fucosylated PSA (Fuc-PSA) and soluble TEK receptor tyrosine kinase (Tie-2) had the ability to predict aggressive (AG) PCa. Additional biomarkers are needed to address this significant clinical problem. Methods: A comprehensive Pubmed search followed by multiplex immunoassays identified candidate biomarkers associated with AG PCa. Subsequently, multiplex and lectin-based immunoassays were applied to a case-control set of sera from subjects with AG PCa, low risk PCa, and non-PCa (biopsy negative). These candidate biomarkers were further evaluated for their ability as panels to complement the prostate health index (phi) in detecting AG PCa. Results: When combined through logistic regression, two panel of biomarkers achieved the best performance: 1) phi, Fuc-PSA, SDC1, and GDF-15 for the detection of AG from low risk PCa and 2) phi, Fuc-PSA, SDC1, and Tie-2 for the detection of AG from low risk PCa and non-PCa, with noticeable improvements in ROC analysis over phi alone (AUCs: 0.942 vs 0.872, and 0.934 vs 0.898, respectively). At a fixed sensitivity of 95%, the panels improved specificity with statistical significance in detecting AG from low risk PCa (76.0% vs 56%, p=0.029), and from low risk PCa and non-PCa (78.2% vs 65.5%, p=0.010). Conclusions: Multivariate panels of serum biomarkers identified in this study demonstrated clinically meaningful improvement over the performance of phi, and warrant further clinical validation, which may contribute to the management of PCa.

Liver-Targeting Class I Selective Histone Deacetylase Inhibitors Potently Suppress Hepatocellular Tumor Growth as Standalone Agents.

Dysfunctions in epigenetic regulation play critical roles in tumor development and progression. Histone deacetylases (HDACs) and histone acetyl transferase (HAT) are functionally opposing epigenetic regulators, which control the expression status of tumor suppressor genes. Upregulation of HDAC activities, which results in silencing of tumor suppressor genes and uncontrolled proliferation, predominates in malignant tumors. Inhibition of the deacetylase activity of HDACs is a clinically validated cancer therapy strategy. However, current HDAC inhibitors (HDACi) have elicited limited therapeutic benefit against solid tumors. Here, we disclosed a class of HDACi that are selective for sub-class I HDACs and preferentially accumulate within the normal liver tissue and orthotopically implanted liver tumors. We observed that these compounds possess exquisite on-target effects evidenced by their induction of dose-dependent histone H4 hyperacetylation without perturbation of tubulin acetylation status and G0/G1 cell cycle arrest. Representative compounds 2 and 3a are relatively non-toxic to mice and robustly suppressed tumor growths in an orthotopic model of HCC as standalone agents. Collectively, our results suggest that these compounds may have therapeutic advantage against HCC relative to the current systemic HDACi. This prospect merits further comprehensive preclinical investigations.

Serotonergic modulation of visual neurons in Drosophila melanogaster.

Sensory systems rely on neuromodulators, such as serotonin, to provide flexibility for information processing as stimuli vary, such as light intensity throughout the day. Serotonergic neurons broadly innervate the optic ganglia of Drosophila melanogaster, a widely used model for studying vision. It remains unclear whether serotonin modulates the physiology of interneurons in the optic ganglia. To address this question, we first mapped the expression patterns of serotonin receptors in the visual system, focusing on a subset of cells with processes in the first optic ganglion, the lamina. Serotonin receptor expression was found in several types of columnar cells in the lamina including 5-HT2B in lamina monopolar cell L2, required for spatiotemporal luminance contrast, and both 5-HT1A and 5-HT1B in T1 cells, whose function is unknown. Subcellular mapping with GFP-tagged 5-HT2B and 5-HT1A constructs indicated that these receptors localize to layer M2 of the medulla, proximal to serotonergic boutons, suggesting that the medulla neuropil is the primary site of serotonergic regulation for these neurons. Exogenous serotonin increased basal intracellular calcium in L2 terminals in layer M2 and modestly decreased the duration of visually induced calcium transients in L2 neurons following repeated dark flashes, but otherwise did not alter the calcium transients. Flies without functional 5-HT2B failed to show an increase in basal calcium in response to serotonin. 5-HT2B mutants also failed to show a change in amplitude in their response to repeated light flashes but other calcium transient parameters were relatively unaffected. While we did not detect serotonin receptor expression in L1 neurons, they, like L2, underwent serotonin-induced changes in basal calcium, presumably via interactions with other cells. These data demonstrate that serotonin modulates the physiology of interneurons involved in early visual processing in Drosophila.

Molecular characteristics and markers of advanced clear cell renal cell carcinoma: Pitfalls due to intratumoral heterogeneity and identification of genetic alterations associated with metastasis.

OBJECTIVES: To identify clear cell renal cell carcinoma-related gene mutations potentially associated with aggressive disease, sarcomatoid differentiation or poor prognosis. METHODS: We carried out genomic analysis of 217 tumor foci from 25 patients with conventional clear cell renal cell carcinoma (14 patients), clear cell renal cell carcinoma with sarcomatoid differentiation (six patients) and non-clear cell renal cell carcinoma (five patients). Each tumor nodule on the tissue block that corresponded to the same focus on the slide was separated from the normal parenchyma and other histologically distinct areas of tumor. The isolated tumor foci were used for subsequent analyses and sequencing. Deoxyribonucleic acid from the formalin-fixed paraffin-embedded tissues was extracted. Multiplex bar-coded polymerase chain reaction amplification was carried out using next-generation sequencing libraries. RESULTS: Overall, 67 protein alterations, including amino acid alterations, frame shifts and splice site mutations in seven genes were identified in the cohort of renal cell carcinoma tumors included in this study. Fewer patients with clear cell renal cell carcinoma with sarcomatoid differentiation had clear cell renal cell carcinoma-related mutations in comparison with patients with conventional clear cell renal cell carcinoma. Additionally, the average number of unique clear cell renal cell carcinoma-related protein alterations per patient was significantly lower in clear cell renal cell carcinoma with sarcomatoid differentiation than in conventional clear cell renal cell carcinoma. Mutations in PBRM1 were identified in a higher proportion of patients with high-grade tumors (World Health Organization/International Society of Urological Pathology grade 4) and in the primary tumors of six of 10 (60%) patients with metastatic disease. CONCLUSIONS: Although there are pitfalls due to intratumoral heterogeneity and sampling bias, mutations in PBRM1 may be associated with metastasis and aggressive disease in clear cell renal cell carcinoma.

The JNK inhibitor AS602801 Synergizes with Enzalutamide to Kill Prostate Cancer Cells In Vitro and In Vivo and Inhibit Androgen Receptor Expression.

In our previous study, we observed that androgen deprivation therapy (ADT) may induce a compensatory increase in MAPK or JNK signaling. Here, we tested the effects of the MEK inhibitors PD0325901 and GSK1120212, ERK1/2 inhibitor GDC-0994, and the JNK inhibitor AS602801 alone and in combination with the AR inhibitor enzalutamide (ENZ) in androgen-sensitive LNCaP cells and androgen-resistant C4-2 and 22Rv1 cells. Enzalutamide combined with AS602801 synergistically killed LNCaP, C4-2, and 22Rv1 cells, and decreased migration and invasion of LNCaP and C4-2 cells. We studied the combination of enzalutamide with AS602801 in vivo using luciferase labeled LNCaP xenografts, and observed that combination of ENZ with AS602801 significantly suppressed tumor growth compared with either drug alone. Importantly, combination therapy resulted in dramatic loss of AR mRNA and protein. Surprisingly, mechanistic studies and Nanostring data suggest that AS602801 likely activates JNK signaling to induce apoptosis. Since AS602801 had sufficient safety and toxicity profile to advance from Phase I to Phase II in clinical trials, repurposing of this compound may represent an opportunity for rapid translation for clinical therapy of CRPC patients.

Discovery and mechanisms of host defense to oncogenesis: targeting the ß-defensin-1 peptide as a natural tumor inhibitor.

Human beta-defensin-1 (hBD-1) is one of a number of small cationic host-defense peptides. Besides its well-known broad-spectrum antimicrobial function, hBD-1 has recently been identified as a chromosome 8p tumor-suppressor gene. The role of hBD-1 in modulating the host immune response to oncogenesis, associated with cell signaling and potential therapeutic applications, has become increasingly appreciated over time. In this study, multiple approaches were used to illustrate hBD-1 anti-tumor activities. Results demonstrate that hBD-1 peptide alters human epidermal growth factor receptor 2 (HER2) signal transduction and represses retroviral-mediated transgene expression in cancer cells. Loss of orthologous murine defense-1 (mBD1) in mice enhances nickel sulfate-induced leiomyosarcoma and causes mouse kidney cells to exhibit increased susceptibility to HPV-16 E6/7-induced neoplastic transformation. Furthermore, for the first time, a novel function of the urine-derived hBD-1 peptide was discovered to suppress bladder cancer growth and this may lead to future applications in the treatment of malignancy.

Preoperative Metabolic Signatures of Prostate Cancer Recurrence Following Radical Prostatectomy.

Technological advances in mass spectrometry (MS), liquid chromatography (LC) separations, nuclear magnetic resonance (NMR) spectroscopy, and big data analytics have made possible studying metabolism at an "omics" or systems level. Here, we applied a multiplatform (NMR + LC-MS) metabolomics approach to the study of preoperative metabolic alterations associated with prostate cancer recurrence. Thus far, predicting which patients will recur even after radical prostatectomy has not been possible. Correlation analysis on metabolite abundances detected on serum samples collected prior to surgery from prostate cancer patients ( n = 40 remission vs n = 40 recurrence) showed significant alterations in a number of pathways, including amino acid metabolism, purine and pyrimidine synthesis, tricarboxylic acid cycle, tryptophan catabolism, glucose, and lactate. Lipidomics experiments indicated higher lipid abundances on recurrent patients for a number of classes that included triglycerides, lysophosphatidylcholines, phosphatidylethanolamines, phosphatidylinositols, diglycerides, acyl carnitines, and ceramides. Machine learning approaches led to the selection of a 20-metabolite panel from a single preoperative blood sample that enabled prediction of recurrence with 92.6% accuracy, 94.4% sensitivity, and 91.9% specificity under cross-validation conditions.

Preoperative botulinum neurotoxin A for children with bilateral cerebral palsy undergoing major hip surgery: a randomized double-blind placebo-controlled trial.

AIM: To assess whether preoperative botulinum neurotoxin A (BoNT-A) affects pain after major hip surgery for children with bilateral cerebral palsy (CP). METHOD: This was a randomized, parallel arms, placebo-contolled trial. Children with hypertonic CP aged 2 to 15 years awaiting bony hip surgery at a tertiary hospital were randomized to receive either BoNT-A or placebo injections into the muscles of the hip on a single occasion immediately before surgery. The primary outcome was the paediatric pain profile (PPP), which was assessed at baseline and weekly for 6 weeks. Treatment allocation was by minimization. Participants, clinicians, and outcome assessors were masked to group assignment. RESULTS: Twenty-seven participants (17 males, 10 females; mean 8y 8mo [SD 3y 9mo], range 3y 4mo-15y 10mo) were allocated to BoNT-A and 27 participants (14 males, 13 females; mean 8y 11mo [SD 3y 5mo], range 4y 1mo-15y 2mo) to placebo. Mean (SD) PPP at 6 weeks for the BoNT-A group (n=24 followed up) was 10.96 (7.22) and for the placebo group (n=26) was 10.04 (8.54) (p=0.69; 95% confidence interval [CI] -4.82, 3.18). There were 16 serious adverse events in total during 6 months of follow-up (n=6 in BoNT-A group). INTERPRETATION: Use of BoNT-A immediately before bony hip surgery for reducing postoperative pain for children with CP was not supported. WHAT THIS PAPER ADDS: Botulinum neurotoxin A (BoNT-A) does not reduce postoperative pain following bony hip surgery. BoNT-A also does not affect postoperative quality of life.

NEUROTÓXINA A BOTULÍNICA PREOPERATORIA PARA NIÑOS CON PARÁLISIS CEREBRAL BILATERAL QUE VAN A SER SOMETIDOS A UNA CIRUGÍA MAYOR DE CADERA: UN ENSAYO ALEATORIO, DOBLE CIEGO, CONTROLADO CON PLACEBO: OBJETIVE: Evaluar si la neurotóxina A botulínica preoperatoria (BoNT-A) afecta el dolor después de una cirugía mayor de cadera en niños con parálisis cerebral bilateral (PC). MÉTODO: Este fue un ensayo aleatorio, con brazos paralelos, controlado con placebo. Los niños con PC hipertónica de 2 a 15 años de edad que esperaban una cirugía de cadera en un hospital terciario se escogieron al azar para recibir inyecciones de BoNT-A o de placebo en los músculos de la cadera una sola administración previa a la cirugía. El resultado primario fue el perfil de dolor pediátrico (PPP, siglas en ingles), que se evaluó al inicio del estudio y semanalmente durante 6 semanas. La asignación del tratamiento fue por minimización. Tanto los participantes, como los clínicos y evaluadores de resultados, eran desconocidos para la asignación de grupo. RESULTADOS: Veintisiete participantes (17 varones y 10 mujeres; medios 8 años 8 meses) [Desviación Estandar SD 3 años 9 meses], rango 3 años 4 meses-15 años 10 meses) se les administro BoNT-A y 27 participantes (14 varones y 13 mujeres; media 8 años 11 meses [SD 3 años 5 meses], rango 4 años 1 mes - 15 años 2 meses) a placebo. La PPP media (SD) a las 6 semanas para el grupo de BoNT-A (n = 24 seguidas) fue de 10,96 (7,22) y para el grupo de placebo (n = 26) fue de 10,04 (8,54) (p = 0,69; intervalo de confianza del 95% [CI, siglas en ingles] -4,82, 3,18). Hubo 16 eventos adversos graves en total durante 6 meses de seguimiento (n = 6 en el grupo BoNT-A). INTERPRETACIÓN: El uso de BoNT-A inmediatamente antes de la cirugía de cadera con el fin de reducir el dolor postoperatorio en niños con PC no fue consistente.

NEUROTOXINA BOTULÍNICA A PRÉ-OPERATÓRIA PARA CRIANÇAS COM PARALISIA CEREBRAL BILATERAL SUBMETIDAS A GRANDE CIRURGIA DE QUADRIL: UM ESTUDO RANDOMIZADO, DUPLO-CEGO, CONTROLADO POR PLACEBO: OBJETIVO: Avaliar se a neurotoxina botulínica tipo A (BTA) pré-operatória A afeta a dor após grande cirurgia de quadril em crianças com paralisia cerebral bilateral (PC). MÉTODO: Este foi um estudo randomizado, com braços paralelos e controlado por placebo. Crianças com PC espástica com idade entre 2 a 15 anos aguardando cirurgia óssea de quadril em um hospital terciário foram randomizadas para receber ou BTA ou injeções de placebo nos músculos do quadril em uma única ocasião imediatamente antes da cirurgia. O desfecho primário foi o perfil de dor pediátrica (PDP), que foi avaliado na linha de base e semanalmente por 6 semanas. A alocação de tratamento foi por minimização. Os participantes, clínicos e avaliadores de resultados foram cegados quanto a atribuição de grupo. RESULTADOS: Vinte e sete participantes (17 homens, 10 mulheres; média de 8 anos e 8 meses [DP 3 anos e 9 meses], com idade entre 3 anos e 4 meses à 15 anos e 10 meses) foram alocados para o grupo BTA e 27 participantes (14 homens, 13 mulheres; média de 8 anos e 11 meses [DP 3 anos e 5 meses], com idade entre 4anos e 1mês à 15anos e 2 meses) foram alocados no grupo placebo. A média (DP) do PDP às 6 semanas para o grupo BTA (n = 24) foi de 10,96 (7,22) e para o grupo placebo (n = 26) foi de 10,04 (8,54) (p = 0,69; intervalo de confiança de 95% [IC] -4,82, 3,18). Houve 16 eventos adversos sérios no total durante 6 meses de acompanhamento (n = 6 no grupo BTA). INTERPRETAÇÃO: O uso da BTA imediatamente antes da cirurgia óssea do quadril para reduzir a dor pós-operatória em crianças com PC não foi apoiado.

Perlecan/HSPG2 and matrilysin/MMP-7 as indices of tissue invasion: tissue localization and circulating perlecan fragments in a cohort of 288 radical prostatectomy patients.

Prostate cancer (PCa) cells use matrix metalloproteinases (MMPs) to degrade tissue during invasion. Perlecan/HSPG2 is degraded at basement membranes, in reactive stroma and in bone marrow during metastasis. We previously showed MMP-7 efficiently degrades perlecan. We now analyzed PCa tissue and serum from 288 prostatectomy patients of various Gleason grades to decipher the relationship between perlecan and MMP-7 in invasive PCa. In 157 prostatectomy specimens examined by tissue microarray, perlecan levels were 18% higher than their normal counterparts. In Gleason grade 4 tissues, MMP-7 and perlecan immunostaining levels were highly correlated with each other (average correlation coefficient of 0.52) in PCa tissue, regardless of grade. Serial sections showed intense, but non-overlapping, immunostaining for MMP-7 and perlecan at adjacent borders, reflecting the protease-substrate relationship. Using a capture assay, analysis of 288 PCa sera collected at prostatectomy showed elevated levels of perlecan fragments, with most derived from domain IV. Perlecan fragments in PCa sera were associated with overall MMP-7 staining levels in PCa tissues. Domain IV perlecan fragments were present in stage IV, but absent in normal, sera, suggesting perlecan degradation during metastasis. Together, perlecan fragments in sera and MMP-7 in tissues of PCa patients are measures of invasive PCa.

Honokiol bis-dichloroacetate (Honokiol DCA) demonstrates activity in vemurafenib-resistant melanoma in vivo.

The majority of human melanomas bears BRAF mutations and thus is treated with inhibitors of BRAF, such as vemurafenib. While patients with BRAF mutations often demonstrate an initial dramatic response to vemurafenib, relapse is extremely common. Thus, novel agents are needed for the treatment of these aggressive melanomas. Honokiol is a small molecule compound derived from Magnolia grandiflora that has activity against solid tumors and hematopoietic neoplasms. In order to increase the lipophilicity of honokiol, we have synthesized honokiol DCA, the dichloroacetate ester of honokiol. In addition, we synthesized a novel fluorinated honokiol analog, bis-trifluoromethyl-bis-(4-hydroxy-3-allylphenyl) methane (hexafluoro). Both compounds exhibited activity against A375 melanoma in vivo, but honokiol DCA was more active. Gene arrays comparing treated with vehicle control tumors demonstrated induction of the respiratory enzyme succinate dehydrogenase B (SDHB) by treatment, suggesting that our honokiol analogs induce respiration in vivo. We then examined its effect against a pair of melanomas, LM36 and LM36R, in which LM36R differs from LM36 in that LM36R has acquired vemurafenib resistance. Honokiol DCA demonstrated in vivo activity against LM36R (vemurafenib resistant) but not against parental LM36. Honokiol DCA and hexafluoro inhibited the phosphorylation of DRP1, thus stimulating a phenotype suggestive of respiration through mitochondrial normalization. Honokiol DCA may act in vemurafenib resistant melanomas to increase both respiration and reactive oxygen generation, leading to activity against aggressive melanoma in vivo.