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1.
Nanoscale ; 16(20): 9875-9886, 2024 May 23.
Article in English | MEDLINE | ID: mdl-38687003

ABSTRACT

Black metal oxides with varying concentrations of O-vacancies display enhanced optical and catalytic properties. However, direct solution syntheses of this class of materials have been limited despite being highly advantageous given the different synthetic handles that can be leveraged towards control of the targeted material. Herein, we present an alternate colloidal synthesis of black In2O3-x nanoparticles from the simple reaction between In(acac)3 and oleyl alcohol. Growth studies by PXRD, TEM, and STEM-EDS coupled to mechanistic insights from 1H, 13C NMR revealed the particles form via two paths, one of which involves In0. We also show that variations in the synthesis atmosphere, ligand environment, and indium precursor can inhibit formation of the black In2O3-x. The optical spectrum for the black nanoparticles displayed a significant redshift when compared to pristine In2O3, consistent with the presence of O-vacancies. Raman spectra and surface analysis also supported the presence of surface oxygen vacancies in the as-synthesized black In2O3-x.

2.
J Sports Sci Med ; 23(1): 1-7, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38455439

ABSTRACT

This study analysed critical movement demands of tennis match-play to better inform contemporary approaches to athlete preparation and training. HawkEye data from matches during the 2021 and 2022 Australian Open were utilised. Distance was aggregated for movement cycles, points, games, sets, and matches, alongside total shots played. Data was collated for males (best-of-five sets) and females (best-of-three sets) allowing comparisons both within and between sexes. Overall, no differences within sexes were evident for total distance, however males traversed further per match than females (MDE = 809 ± 139m, ES = 0.86). Female players travelled further in their deciding (third) sets compared to set 1 (ES = 0.28) and while this effect wasn't as discernible for males, the deciding (fifth) set showed some evidence of elevated distance requirements and variability. Between sexes, only female set 3 was different to male set 3 (ES = 0.29). Female and male tiebreak games (i.e. game 13) required players travel further distance compared to other games (ES = ~1.45). Between sex differences were observed for tiebreak games compared to games 1 to 12 (female ES = 1.36 and male ES = 1.53). Players from both sexes generally covered similar distances during points and movement cycles, with between-shot distances of 4.2m-4.5m, notably longer than previous reports. Further, total shots and total match distance (r > 0.97; p < 0.01) shared similar linear relationships. These results highlight that the between shot or movement cycle demands of professional hard court tennis are substantially higher than described in the literature (Roetert et al., 2003). The findings also reveal competitiveness as a key influence on set level distance demands during professional tennis match-play, a consideration in player preparation programs.


Subject(s)
Athletic Performance , Tennis , Humans , Male , Female , Australia , Athletes , Movement
3.
iScience ; 27(2): 108984, 2024 Feb 16.
Article in English | MEDLINE | ID: mdl-38327800

ABSTRACT

Olaparib is a pioneering PARP inhibitor (PARPi) approved for treating castration-resistant prostate cancer (CRPC) tumors harboring DNA repair defects, but clinical resistance has been documented. To study acquired resistance, we developed Olaparib-resistant (OlapR) cell lines through chronic Olaparib treatment of LNCaP and C4-2B cell lines. Here, we found that IGFBP3 is highly expressed in acquired (OlapR) and intrinsic (Rv1) models of Olaparib resistance. We show that IGFBP3 expression promotes Olaparib resistance by enhancing DNA repair capacity through activation of EGFR and DNA-PKcs. IGFBP3 depletion enhances efficacy of Olaparib by promoting DNA damage accumulation and subsequently, cell death in resistant models. Mechanistically, we show that silencing IGFBP3 or EGFR expression reduces cell viability and resensitizes OlapR cells to Olaparib treatment. Inhibition of EGFR by Gefitinib suppressed growth of OlapR cells and improved Olaparib sensitivity, thereby phenocopying IGFBP3 inhibition. Collectively, our results highlight IGFBP3 and EGFR as critical mediators of Olaparib resistance.

4.
Cancers (Basel) ; 15(21)2023 Nov 03.
Article in English | MEDLINE | ID: mdl-37958444

ABSTRACT

Current common treatments for castration-resistant prostate cancer (CRPC) typically belong to one of three major categories: next-generation anti-androgen therapies (NGAT) including enzalutamide, abiraterone acetate, apalutamide, and darolutamide; taxane therapy represented by docetaxel; and PARP inhibitors (PARPi) like olaparib. Although these treatments have shown efficacy and have improved outcomes for many patients, some do not survive due to the emergence of therapeutic resistance. The clinical landscape is further complicated by limited knowledge about how the sequence of treatments impacts the development of therapeutic cross-resistance in CRPC. We have developed multiple CRPC models of acquired therapeutic resistance cell sublines from C4-2B cells. These include C4-2B MDVR, C4-2B AbiR, C4-2B ApaR, C4-2B DaroR, TaxR, and 2B-olapR, which are resistant to enzalutamide, abiraterone, apalutamide, darolutamide, docetaxel, and olaparib, respectively. These models are instrumental for analyzing gene expression and assessing responses to various treatments. Our findings reveal distinct cross-resistance characteristics among NGAT-resistant cell sublines. Specifically, resistance to enzalutamide induces resistance to abiraterone and vice versa, while maintaining sensitivity to taxanes and olaparib. Conversely, cells with acquired resistance to docetaxel exhibit cross-resistance to both cabazitaxel and olaparib but retain sensitivity to NGATs like enzalutamide and abiraterone. OlapR cells, significantly resistant to olaparib compared to parental cells, are still responsive to NGATs and docetaxel. Moreover, OlapR models display cross-resistance to other clinically relevant PARP inhibitors, including rucaparib, niraparib, and talazoparib. RNA-sequencing analyses have revealed a complex network of altered gene expressions that influence signaling pathways, energy metabolism, and apoptotic signaling, pivotal to cancer's evolution and progression. The data indicate that resistance mechanisms are distinct among different drug classes. Notably, NGAT-resistant sublines exhibited a significant downregulation of androgen-regulated genes, contrasting to the stable expression noted in olaparib and docetaxel-resistant sublines. These results may have clinical implications by showing that treatments of one class can be sequenced with those from another class, but caution should be taken when sequencing drugs of the same class.

5.
Molecules ; 28(10)2023 May 20.
Article in English | MEDLINE | ID: mdl-37241953

ABSTRACT

In this work, a continuous system to produce multi-hundred-gram quantities of aryl sulfonyl chlorides is described. The scheme employs multiple continuous stirred-tank reactors (CSTRs) and a continuous filtration system and incorporates an automated process control scheme. The experimental process outlined is intended to safely produce the desired sulfonyl chloride at laboratory scale. Suitable reaction conditions were first determined using a batch-chemistry design of experiments (DOE) and several isolation methods. The hazards and incompatibilities of the heated chlorosulfonic acid reaction mixture were addressed by careful equipment selection, process monitoring, and automation. The approximations of the CSTR fill levels and pumping performance were measured by real-time data from gravimetric balances, ultimately leading to the incorporation of feedback controllers. The introduction of process automation demonstrated in this work resulted in significant improvements in process setpoint consistency, reliability, and spacetime yield, as demonstrated in medium- and large-scale continuous manufacturing runs.

6.
J Sports Sci Med ; 22(1): 133-141, 2023 03.
Article in English | MEDLINE | ID: mdl-36876180

ABSTRACT

Understanding on-court movement in tennis allows for enhanced preparation strategies to improve player readiness and performance. Here, we explore expert physical preparation coaches' perceptions of elite training strategies for preparation and performance in tennis, with special reference to lower limb activity. Thirteen world renowned tennis strength and conditioning coaches were interviewed in a semi-structured method that explored four key topic areas of physical preparation for tennis: i) the physical demands; ii) load monitoring practice; iii) the direction of ground reaction forces application during match-play; and iv) the application of strength and conditioning for tennis. Three higher-order themes emerged from these discussions: i) off-court training for tennis should be specific to the demands of the sport, ii) the mechanical understanding of tennis lags our physiological approach, and iii) our understanding of the lower limb's contribution to tennis performance is limited. These findings provide valuable insights into the importance of improving our knowledge relevant to the mechanical demands of tennis movement, whilst highlighting important practical considerations from leading tennis conditioning experts.


Subject(s)
Sports , Tennis , Humans , Fitness Trackers , Knowledge , Lower Extremity
7.
Int J Sports Physiol Perform ; 18(5): 512-522, 2023 May 01.
Article in English | MEDLINE | ID: mdl-36965486

ABSTRACT

PURPOSE: To quantify match load associated with padel and compare responses with both singles and doubles tennis. METHODS: On separate days, 12 participants (7 men and 5 women) played 60-minute padel (PADEL), singles tennis (SINGLES), and doubles tennis (DOUBLES) simulated games. Participants wore a 10-Hz GPS/100-Hz triaxial accelerometer unit and heart-rate monitor. Exercise-related sensations and blood lactate concentration were monitored every 20 minutes. Match-play characteristics (temporal structure) and shot selection were derived from video analysis. Vertical jump ability was assessed before and after each game. RESULTS: Heart rate, exercise-related sensations (overall perceived exertion and limb discomfort), and physical load (total distance covered, PlayerLoad, acceleration density and load) for SINGLES were higher compared with DOUBLES and PADEL (all P ≤ .05). Blood lactate concentrations remained low (1-2 mmol·L-1) and did not differ between conditions. Effective playing time (P < .001) was lower in SINGLES and DOUBLES compared with PADEL. The number of forehands (P = .002) and backhands (P < .001) was greater for SINGLES than for DOUBLES and PADEL. The number of volleys/smashes and lobs (P < .05) was greater for PADEL compared with SINGLES and DOUBLES. Performance for squat, countermovement, and multirebound jumps was similarly reduced below baseline after match play (P < .05), independent of condition. CONCLUSION: Padel imposes a unique match load on players that is different from singles tennis and more closely resembles that of doubles tennis. Cardiovascular stimulation and physical load are highest in singles tennis, while padel sees players hit a larger variety of shots with higher effective playing percentages.


Subject(s)
Tennis , Male , Humans , Female , Tennis/physiology , Physical Exertion/physiology , Lactic Acid , Exercise , Heart Rate/physiology
8.
Nanoscale ; 15(6): 2650-2658, 2023 Feb 09.
Article in English | MEDLINE | ID: mdl-36722489

ABSTRACT

Devising synthetic strategies to control crystal structure is of great importance as materials properties are governed by structure. MnS is a great model system as it has three known stable polymorphs. Herein, we show the selective synthesis of colloidal wurtzite- and rock-salt-type MnS under identical reactions conditions changing only the manganese halide precursor. Mixtures of Mn halides or halide surrogate (e.g., NH4Cl) also enabled polymorph control. Powder X-ray diffraction aliquot studies of the reactions revealed the crystal structure at the onset of nucleation and that of the final product is the same, unlike the Ostwald ripening transformation observed in other systems. The halide-driven selectivity was also observed in the synthesis of manganese selenide nanoparticles. In this system, variation of the Mn halide precursor allowed access to the wurtzite- and rock salt-type polymorphs of MnSe, as well as the pyrite-MnSe2 phase. Based on this work, the mixing of metal salts might be a simple and effective strategy towards polymorph control and access materials with new crystal structures.

9.
Mol Cancer Ther ; 21(4): 677-685, 2022 04 01.
Article in English | MEDLINE | ID: mdl-35086956

ABSTRACT

PARP inhibition represents the dawn of precision medicine for treating prostate cancer. Despite this advance, questions remain regarding the use of PARP inhibitors (PARPi) for the treatment of this disease, including (i) how specifically do PARPi-sensitive tumor cells respond to treatment, and (ii) how does PARPi resistance develop? To address these questions, we characterized response to olaparib in sensitive LNCaP and C4-2B cells and developed two olaparib-resistant derivative cell line models from each, termed LN-OlapR and 2B-OlapR, respectively. OlapR cells possess distinct morphology from parental cells and display robust resistance to olaparib and other clinically relevant PARPis, including rucaparib, niraparib, and talazoparib. In LNCaP and C4-2B cells, we found that olaparib induces massive DNA damage, leading to activation of the G2-M checkpoint, activation of p53, and cell-cycle arrest. Furthermore, our data suggest that G2-M checkpoint activation leads to both cell death and senescence associated with p21 activity. In contrast, both LN-OlapR and 2B-OlapR cells do not arrest at G2-M and display a markedly blunted response to olaparib treatment. Interestingly, both OlapR cell lines harbor increased DNA damage relative to parental cells, suggesting that OlapR cells accumulate and manage persistent DNA damage during acquisition of resistance, likely through augmenting DNA repair capacity. Further impairing DNA repair through CDK1 inhibition enhances DNA damage, induces cell death, and sensitizes OlapR cells to olaparib treatment. Our data together further our understanding of PARPi treatment and provide a cellular platform system for the study of response and resistance to PARP inhibition.


Subject(s)
Phthalazines , Prostatic Neoplasms , Cell Cycle Checkpoints , Cell Line, Tumor , Humans , Male , Phthalazines/pharmacology , Piperazines/pharmacology , Poly(ADP-ribose) Polymerase Inhibitors/pharmacology , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/genetics
10.
J Strength Cond Res ; 36(12): 3415-3421, 2022 Dec 01.
Article in English | MEDLINE | ID: mdl-32898037

ABSTRACT

ABSTRACT: Thornton, HR, Armstrong, CR, Gamble, T, Rigby, A, Johnston, RD, and Duthie, GM. Quantifying the movement characteristics of Australian Football League Women's competition. J Strength Cond Res 36(12): 3415-3421, 2022-The purpose is to provide an overview of the externally measured movement characteristics of Australian Football League Women's (AFLW) competition, and the variability in this between matches. A range of movement variables were measured from 28 athletes who wore 10-Hz global positioning system devices during games and were summarized according to positional groups. The variance in each athlete's match loads for each round were expressed using standardized scores ( z -scores), and the change compared with the first game played was calculated and expressed as a standard deviation. Furthermore, using the raw export, moving means (1-10 minutes) of speed (m·min -1 ) and acceleration (m·s -2 ) were calculated. Following log transformation of the maximal means, intercept and slopes were calculated. Linear mixed models identified differences between positional groups for match loads, and intercept and slopes. Effects were described using standardized effect sizes (ESs) and magnitude-based decisions. There were no substantial and unclear differences between positional groups for match loads (ES range; ±confidence limits = 0.10-0.80; ±0.30-4.30) and for intercept and slopes (ES range; 0.04-0.44; ±0.52-2.11). Large within-athlete variation in match demands between rounds was observed ( z -score up to -1.8 SD for distance), and the maximal means for speed and acceleration demonstrate the extensive physical demands of AFLW competition. These data describe the intense and variable physical demands of AFLW competition, and further provide novel information regarding the maximal mean intensities and intercept and slopes, which should assist practitioners in planning and prescribing training in preparation for competition.


Subject(s)
Athletic Performance , Running , Team Sports , Female , Humans , Acceleration , Australia , Geographic Information Systems , Movement
11.
BJU Int ; 129(3): 345-355, 2022 03.
Article in English | MEDLINE | ID: mdl-34185954

ABSTRACT

OBJECTIVES: To investigate the genetic alterations of patients with prostate cancer (PCa) with and without intraductal carcinoma of the prostate (IDC-P). PATIENTS AND METHODS: We performed targeted sequencing of plasma cell-free DNA on 161 patients with prostate adenocarcinoma (PAC) with IDC-P and 84 without IDC-P. Genomic alterations were compared between these two groups. The association between genetic alterations and patients' survival outcomes was also explored. RESULTS: We identified that 29.8% (48/161) and 21.4% (18/84) of patients with and without IDC-P harboured genomic alterations in DNA repair pathways, respectively (P = 0.210). Pathogenic germline DNA repair alterations were frequently detected in IDC-P carriers compared to IDC-P non-carriers (11.8% [19/161] vs 2.4% [two of 84], P = 0.024). Germline BReast CAncer type 2 susceptibility protein (BRCA2) and somatic cyclin-dependent kinase 12 (CDK12) defects were specifically identified in IDC-P carriers relative to PAC (BRCA2: 8.7% [14/161] vs 0% and CDK12: 6.8% [11/161] vs 1.2% [one of 84]). Patients with IDC-P had a distinct androgen receptor (AR) pathway alteration, characterised by an enrichment of nuclear receptor corepressor 2 (NCOR2) mutations compared with patients with pure PAC (21.1% [34/161] vs 6.0% [five of 84], P = 0.004). Increased AR alterations were detected in patients harbouring tumours with an IDC-P proportion of ≥10% vs those with an IDC-P proportion of <10% (6.4% [five of 78] vs 18.1% [15/83], P = 0.045). For IDC-P carriers, tumour protein p53 (TP53) mutation was associated with shorter castration-resistant-free survival (median 10.9 vs 28.9 months, P = 0.026), and BRCA2 alteration was related to rapid prostate-specific antigen progression for those receiving abiraterone treatment (median 9.1 vs 11.9 months, P = 0.036). CONCLUSION: Our findings provide genomic evidence explaining the aggressive phenotype of tumours with IDC-P, highlighting the potential therapeutic strategies for this patient population.


Subject(s)
Carcinoma, Intraductal, Noninfiltrating , Circulating Tumor DNA , Prostatic Neoplasms , Carcinoma, Intraductal, Noninfiltrating/genetics , Carcinoma, Intraductal, Noninfiltrating/pathology , Circulating Tumor DNA/genetics , Humans , Male , Phenotype , Prostate/pathology , Prostatic Neoplasms/pathology
12.
Nat Commun ; 12(1): 5262, 2021 09 06.
Article in English | MEDLINE | ID: mdl-34489456

ABSTRACT

TFE3-translocation renal cell carcinoma (TFE3-tRCC) is a rare and heterogeneous subtype of kidney cancer with no standard treatment for advanced disease. We describe comprehensive molecular characteristics of 63 untreated primary TFE3-tRCCs based on whole-exome and RNA sequencing. TFE3-tRCC is highly heterogeneous, both clinicopathologically and genotypically. ASPSCR1-TFE3 fusion and several somatic copy number alterations, including the loss of 22q, are associated with aggressive features and poor outcomes. Apart from tumors with MED15-TFE3 fusion, most TFE3-tRCCs exhibit low PD-L1 expression and low T-cell infiltration. Unsupervised transcriptomic analysis reveals five molecular clusters with distinct angiogenesis, stroma, proliferation and KRAS down signatures, which show association with fusion patterns and prognosis. In line with the aggressive nature, the high angiogenesis/stroma/proliferation cluster exclusively consists of tumors with ASPSCR1-TFE3 fusion. Here, we describe the genomic and transcriptomic features of TFE3-tRCC and provide insights into precision medicine for this disease.


Subject(s)
Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/genetics , Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/mortality , Kidney Neoplasms/genetics , Kidney Neoplasms/mortality , Adolescent , Adult , Aged , Carcinoma, Renal Cell/pathology , Carcinoma, Renal Cell/therapy , Child , Child, Preschool , Female , Gene Expression Regulation, Neoplastic , Gene Fusion , Humans , Intracellular Signaling Peptides and Proteins/genetics , Kidney Neoplasms/pathology , Kidney Neoplasms/therapy , Male , Middle Aged , Mutation , Prognosis , Sequence Analysis, RNA , Tumor Microenvironment/genetics , Tumor Microenvironment/immunology , Exome Sequencing , Young Adult
13.
Am J Clin Exp Urol ; 9(4): 292-300, 2021.
Article in English | MEDLINE | ID: mdl-34541028

ABSTRACT

Current therapies for treating castration resistant prostate cancer (CRPC) include abiraterone and enzalutamide which function by inhibiting androgen signaling by targeting androgen synthesis and antagonizing the androgen receptor (AR) respectively. While these therapies are initially beneficial, resistance inevitably develops. A number of pathways have been identified to contribute to CRPC progression and drug resistance. Among these is aberrant androgen signaling perpetuated by increased expression and activity of androgenic enzymes. While abiraterone inhibits the androgenic enzyme, CYP17A1, androgen synthesis inhibition by abiraterone is incomplete and sustained androgenesis persists, in part due to increased levels of AKR1C3 and steroid sulfatase (STS). Expression of both of these enzymes is increased in CRPC and is associated with resistance to anti-androgens. A number of studies have identified methods for targeting these enzymes. Indomethacin, a non-steroidal anti-inflammatory drug commonly used to treat inflammatory arthritis has been well established as an inhibitor of AKR1C3. Treatment of CRPC cells with indomethacin reduces cell growth and improves the response to enzalutamide and abiraterone. Similarly, STS inhibitors have been shown to reduce intracrine androgens and also reduce CRPC growth and enhance anti-androgen treatment. In this review, we provide an overview of androgen synthesis in CRPC and strategies aimed at inhibiting intracrine androgens.

14.
Oncogene ; 40(35): 5379-5392, 2021 09.
Article in English | MEDLINE | ID: mdl-34272475

ABSTRACT

Targeting androgen signaling with the second-generation anti-androgen drugs, such as enzalutamide (Enza), abiraterone (Abi), apalutamide (Apal), and darolutamide (Daro), is the mainstay for the treatment of castration-resistant prostate cancer (CRPC). While these treatments are effective initially, resistance occurs frequently. Continued expression of androgen receptor (AR) and its variants such as AR-V7 despite AR-targeted therapy contributes to treatment resistance and cancer progression in advanced CRPC patients. This highlights the need for new strategies blocking continued AR signaling. Here, we identify a novel AR/AR-V7 degrader (ARVib) and found that ARVib effectively degrades AR/AR-V7 protein and attenuates AR/AR-V7 downstream target gene expression in prostate cancer cells. Mechanistically, ARVib degrades AR/AR-V7 protein through the ubiquitin-proteasome pathway mediated by HSP70/STUB1 machinery modulation. ARVib suppresses HSP70 expression and promotes STUB1 nuclear translocation, where STUB1 binds to AR/AR-V7 and promotes its ubiquitination and degradation. ARVib significantly inhibits resistant prostate tumor growth and improves enzalutamide treatment in vitro and in vivo. These data suggest that ARVib has potential for development as an AR/AR-V7 degrader to treat resistant CRPC.


Subject(s)
Prostatic Neoplasms , Receptors, Androgen , Humans , Male , Signal Transduction
15.
Mol Cancer Ther ; 20(10): 2061-2070, 2021 10.
Article in English | MEDLINE | ID: mdl-34326198

ABSTRACT

Docetaxel and cabazitaxel-based taxane chemotherapy are critical components in the management of advanced prostate cancer. However, their efficacy is hindered due to de novo presentation with or the development of resistance. Characterizing models of taxane-resistant prostate cancer will lead to creation of strategies to overcome insensitivity. We have previously characterized docetaxel-resistant C4-2B and DU145 cell line derivatives, TaxR and DU145-DTXR, respectively. In the present study, we characterize cabazitaxel-resistant derivative cell lines created from chronic cabazitaxel exposure of TaxR and DU145-DTXR cells, CabR and CTXR, respectively. We show that CabR and CTXR cells are robustly resistant to both taxanes but retain sensitivity to antiandrogens. Both CabR and CTXR cells possess increased expression of ABCB1, which is shown to mediate resistance to treatment. Interestingly, we also present evidence for coordinated overexpression of additional genes present within the 7q21.12 gene locus where ABCB1 resides. This locus, known as the ABCB1 amplicon, has been demonstrated to be amplified in multidrug-resistant tumor cells, but little is known regarding its role in prostate cancer. We show that two ABCB1-amplicon genes other than ABCB1, RUNDC3B and DBF4, promote cellular viability and treatment resistance in taxane-resistant prostate cancer models. We present evidence that coordinated amplification of ABCB1-amplicon genes is common in a subset of prostate cancer patients. These data together suggest that ABCB1-amplicon activation plays a critical role in taxane resistance.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Biomarkers, Tumor/metabolism , Drug Resistance, Neoplasm , Gene Expression Regulation, Neoplastic , Prostatic Neoplasms, Castration-Resistant/drug therapy , ATP Binding Cassette Transporter, Subfamily B/genetics , ATP Binding Cassette Transporter, Subfamily B/metabolism , Apoptosis , Biomarkers, Tumor/genetics , Cell Proliferation , Docetaxel/administration & dosage , Humans , Male , Prostatic Neoplasms, Castration-Resistant/genetics , Prostatic Neoplasms, Castration-Resistant/pathology , Taxoids/administration & dosage , Tumor Cells, Cultured
16.
Eur J Pharm Sci ; 159: 105702, 2021 Apr 01.
Article in English | MEDLINE | ID: mdl-33429045

ABSTRACT

Due to the complexity in the interactions of variables and mechanisms leading to blend segregation, quantifying the segregation propensity of an Active Pharmaceutical Ingredient (API) has been challenging. A high-throughput segregation risk prediction workflow for early drug product development has been developed based on the dispensing mechanism of automated powder dispensing technology. The workflow utilized liquid handling robots and high-performance liquid chromatography (HPLC) with a well-plate autosampler for sample preparation and analysis. Blends containing three different APIs of varying concentrations and particle sizes of different constituents were evaluated through this automated workflow. The workflow enabled segregation evaluation of different API blends in very small quantities (~7g) compared to other common segregation testers that consume hundreds of grams. Segregation patterns obtained were well explained with vibration induced percolation-based segregation phenomena. Segregation risk was translated quantitatively using relative standard deviation (RSD) calculations, and the results matched well with large-scale segregation studies. The applied approach increased the throughput, introduced a simple and clean walk-up method with minimized equipment space and API exposures to conduct segregation studies. Results obtained can provide insights about optimizing particle size distributions, as well as selecting appropriate formulation constituents and secondary processing steps in early drug product development when the amount of available API is very limited.


Subject(s)
Chemistry, Pharmaceutical , Technology, Pharmaceutical , Excipients , Powders , Technology
17.
Clin Cancer Res ; 27(6): 1734-1743, 2021 03 15.
Article in English | MEDLINE | ID: mdl-33414138

ABSTRACT

PURPOSE: Fumarate hydratase-deficient renal cell carcinoma (FH-deficient RCC) is a rare but lethal subtype of RCC. Little is known about the genomic profile of FH-deficient RCC, and the therapeutic options for advanced disease are limited. To this end, we performed a comprehensive genomics study to characterize the genomic and epigenomic features of FH-deficient RCC. EXPERIMENTAL DESIGN: Integrated genomic, epigenomic, and molecular analyses were performed on 25 untreated primary FH-deficient RCCs. Complete clinicopathologic and follow-up data of these patients were recorded. RESULTS: We identified that FH-deficient RCC manifested low somatic mutation burden (median 0.58 mutations per megabase), but with frequent somatic copy-number alterations. The majority of FH-deficient RCCs were characterized by a CpG sites island methylator phenotype, displaying concerted hypermethylation at numerous CpG sites in genes of transcription factors, tumor suppressors, and tumor hallmark pathways. However, a few cases (20%) with low metastatic potential showed relatively low DNA methylation levels, indicating the heterogeneity of methylation pattern in FH-deficient RCC. Moreover, FH-deficient RCC is potentially highly immunogenic, characterized by increased tumor T-cell infiltration but high expression of immune checkpoint molecules in tumors. Clinical data further demonstrated that patients receiving immune checkpoint blockade-based treatment achieved improved progression-free survival over those treated with antiangiogenic monotherapy (median, 13.3 vs. 5.1 months; P = 0.03). CONCLUSIONS: These results reveal the genomic features and provide new insight into potential therapeutic strategies for FH-deficient RCC.


Subject(s)
Biomarkers, Tumor/metabolism , Carcinoma, Renal Cell/pathology , Epigenesis, Genetic , Fumarate Hydratase/deficiency , Gene Expression Regulation, Neoplastic , Kidney Neoplasms/pathology , Mutation , Adolescent , Adult , Aged , Biomarkers, Tumor/genetics , Carcinoma, Renal Cell/drug therapy , Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/metabolism , DNA Methylation , Female , Follow-Up Studies , Fumarate Hydratase/genetics , Humans , Immune Checkpoint Inhibitors/therapeutic use , Kidney Neoplasms/drug therapy , Kidney Neoplasms/genetics , Kidney Neoplasms/metabolism , Male , Middle Aged , Neoplasm Metastasis , Prognosis , Retrospective Studies , Survival Rate , Young Adult
18.
Clin Cancer Res ; 26(22): 6064-6074, 2020 11 15.
Article in English | MEDLINE | ID: mdl-32928794

ABSTRACT

PURPOSE: Most patients with prostate cancer receiving enzalutamide or abiraterone develop resistance. Clinical evidence indicates that serum levels of dehydroepiandrosterone sulfate (DHEAS) and biologically active DHEA remain in the high range despite antiandrogen treatment. The conversion of DHEAS into DHEA by steroid sulfatase (STS) may contribute to sustained intracrine androgen synthesis. Here, we determine the contribution of STS to treatment resistance and explore the potential of targeting STS to overcome resistance in prostate cancer. EXPERIMENTAL DESIGN: STS expression was examined in patients and cell lines. In vitro, STS activity and expression were modulated using STS-specific siRNA or novel STS inhibitors (STSi). Cell growth, colony formation, androgen production, and gene expression were examined. RNA-sequencing analysis was conducted on VCaP cells treated with STSi. Mice were treated with STSis with or without enzalutamide to determine their effects in vivo. RESULTS: STS is overexpressed in patients with castration-resistant prostate cancer (CRPC) and resistant cells. STS overexpression increases intracrine androgen synthesis, cell proliferation, and confers resistance to enzalutamide and abiraterone. Inhibition of STS using siRNA suppresses prostate cancer cell growth. Targeting STS activity using STSi inhibits STS activity, suppresses androgen receptor transcriptional activity, and reduces the growth of resistant C4-2B and VCaP prostate cancer cells. STSis significantly suppress resistant VCaP tumor growth, decrease serum PSA levels, and enhance enzalutamide treatment in vitro and in vivo. CONCLUSIONS: These studies suggest that STS drives intracrine androgen synthesis and prostate cancer proliferation. Targeting STS represents a therapeutic strategy to treat CRPC and improve second-generation antiandrogen therapy.


Subject(s)
Androgens/biosynthesis , Prostatic Neoplasms, Castration-Resistant/drug therapy , Prostatic Neoplasms/drug therapy , Steryl-Sulfatase/genetics , Androgen Antagonists/pharmacology , Androgens/genetics , Androstenes/adverse effects , Androstenes/pharmacology , Benzamides/adverse effects , Benzamides/pharmacology , Carcinogenesis/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Dehydroepiandrosterone/metabolism , Dehydroepiandrosterone Sulfate/metabolism , Drug Resistance, Neoplasm/genetics , Gene Expression Regulation, Neoplastic/drug effects , Humans , Male , Neoplasm Staging , Nitriles/adverse effects , Nitriles/pharmacology , Phenylthiohydantoin/adverse effects , Phenylthiohydantoin/pharmacology , Prostatic Neoplasms, Castration-Resistant/blood , Prostatic Neoplasms, Castration-Resistant/genetics , Prostatic Neoplasms, Castration-Resistant/pathology , RNA-Seq
19.
Eur J Pharm Sci ; 154: 105516, 2020 Nov 01.
Article in English | MEDLINE | ID: mdl-32814162

ABSTRACT

The segregation of an active pharmaceutical ingredient (API) within a powder blend is one of the major manufacturing obstacles in achieving content uniformity. Segregation can be due to differences in physicochemical properties of formulation components and/or perturbations experienced during secondary processing steps, such as granulation, fluidization, die-filling and compression. A near-infrared (NIR) spectrometer embedded segregation tester, which could mimic the external stimulations (vibration and fluidization) experienced by a blend in a manufacturing facility, was used to evaluate and predict blend segregation. Two different GlaxoSmithKline (GSK) product blends with variations in the API particle size and concentration were tested. Drug content was further measured at different locations along the powder bed by NIR to sketch the segregation profile and calculate the overall segregation intensity of each blend. The study indicated that the segregation potential was dependent on the particle sizes of API and excipients, as well as the type of stimulus applied (vibration vs fluidization). Drug concentration profiles obtained from this mode of analysis decoded the underlying segregation mechanisms (sieving, trajectory and air elutriation) easily. The employed NIR-based segregation tester proved to be a useful small-scale predictive tool to evaluate and rank the segregation risk of the studied pharmaceutical blends.


Subject(s)
Chemistry, Pharmaceutical , Pharmaceutical Preparations , Technology, Pharmaceutical , Drug Compounding , Excipients , Particle Size , Powders , Spectrum Analysis , Tablets
20.
Mol Cancer Ther ; 19(8): 1708-1718, 2020 08.
Article in English | MEDLINE | ID: mdl-32430485

ABSTRACT

The next-generation antiandrogen drugs, XTANDI (enzalutamide), ZYTIGA (abiraterone acetate), ERLEADA (apalutamide) and NUBEQA (darolutamide) extend survival times and improve quality of life in patients with advanced prostate cancer. Despite these advances, resistance occurs frequently and there is currently no definitive cure for castration-resistant prostate cancer. Our previous studies identified that similar mechanisms of resistance to enzalutamide or abiraterone occur following treatment and cross-resistance exists between these therapies in advanced prostate cancer. Here, we show that enzalutamide- and abiraterone-resistant prostate cancer cells are further cross-resistant to apalutamide and darolutamide. Mechanistically, we have determined that the AKR1C3/AR-V7 axis confers this cross-resistance. Knockdown of AR-V7 in enzalutamide-resistant cells resensitize cells to apalutamide and darolutamide treatment. Furthermore, targeting AKR1C3 resensitizes resistant cells to apalutamide and darolutamide treatment through AR-V7 inhibition. Chronic apalutamide treatment in C4-2B cells activates the steroid hormone biosynthesis pathway and increases AKR1C3 expression, which confers resistance to enzalutamide, abiraterone, and darolutamide. In conclusion, our results suggest that apalutamide and darolutamide share similar resistant mechanisms with enzalutamide and abiraterone. The AKR1C3/AR-V7 complex confers cross-resistance to second-generation androgen receptor-targeted therapies in advanced prostate cancer.


Subject(s)
Aldo-Keto Reductase Family 1 Member C3/metabolism , Alternative Splicing , Androgen Receptor Antagonists/pharmacology , Drug Resistance, Neoplasm , Gene Expression Regulation, Neoplastic/drug effects , Prostatic Neoplasms, Castration-Resistant/drug therapy , Receptors, Androgen/chemistry , Aldo-Keto Reductase Family 1 Member C3/genetics , Androgen Receptor Antagonists/classification , Apoptosis , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Cell Proliferation , Humans , Male , Prostatic Neoplasms, Castration-Resistant/genetics , Prostatic Neoplasms, Castration-Resistant/pathology , Receptors, Androgen/genetics , Tumor Cells, Cultured
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