Extracellular vesicles (EVs) are microvesicles secreted from various cell types. We aimed to discover a new biomarker for high Gleason score (GS) prostate cancer (PCa) in urinary EVs via quantitative ...proteomics. EVs were isolated from urine after massage from 18 men (negative biopsy n = 6, GS 6 PCa n = 6, or GS 8-9 PCa n = 6). EV proteins were labeled with iTRAQ and analyzed by LC-MS/MS. We identified 4710 proteins and quantified 3528 proteins in the urinary EVs. Eleven proteins increased in patients with PCa compared to those with negative biopsy (ratio >1.5, p-value < 0.05). Eleven proteins were chosen for further analysis and verified in 29 independent urine samples (negative n = 11, PCa n = 18) using selected reaction monitoring/multiple reaction monitoring. Among these candidate markers, fatty acid binding protein 5 (FABP5) was higher in the cancer group than in the negative group (p-value = 0.009) and was significantly associated with GS (p-value for trend = 0.011). Granulin, AMBP, CHMP4A, and CHMP4C were also higher in men with high GS prostate cancer (p-value < 0.05). FABP5 in urinary EVs could be a potential biomarker of high GS PCa.
Proteomic analysis of urinary extracellular vesicles (EVs) is a powerful approach to discover potential bladder cancer (BCa) biomarkers, however urine contains numerous EVs derived from the kidney ...and normal urothelial epithelium, which can obfuscate information related to BCa cell‐derived EVs. In this study, we combined proteomic analysis of urinary EVs and tissue‐exudative EVs (Te‐EVs), which were isolated from culture medium of freshly resected viable BCa tissues. Urinary EVs were isolated from urine samples of 11 individuals (7 BCa patients and 4 healthy individuals), and Te‐EVs were isolated from 7 BCa tissues. We performed tandem mass tag (TMT)‐labeling liquid chromatography (LC‐MS/MS) analysis for both urinary EVs and Te‐EVs and identified 1960 proteins in urinary EVs and 1538 proteins in Te‐EVs. Most of the proteins identified in Te‐EVs were also present in urinary EVs (82.4%), with 55 of these proteins showing upregulated levels in the urine of BCa patients (fold change > 2.0; P < .1). Among them, we selected 22 membrane proteins as BCa biomarker candidates for validation using selected reaction monitoring/multiple reaction monitoring (SRM/MRM) analysis on urine samples from 70 individuals (40 BCa patients and 30 healthy individuals). Six urinary EV proteins (heat‐shock protein 90, syndecan‐1, myristoylated alanine‐rich C‐kinase substrate (MARCKS), MARCKS‐related protein, tight junction protein ZO‐2, and complement decay‐accelerating factor) were quantified using SRM/MRM analysis and validated as significantly upregulated in BCa patients (P < .05). In conclusion, the novel strategy that combined proteomic analysis of urinary EVs and Te‐EVs enabled selective detection of urinary BCa biomarkers.
Proteomic analysis of urinary extracellular vesicles (EVs) is a powerful approach to discovering potential BCa biomarkers, however urine contains numerous EVs derived from kidney and normal urothelial epithelium that could dilute the information of cancer BCa cell‐derived EVs. In this study, we performed combined proteomic analysis of both urinary EVs and tissue‐extracted EVs (Te‐EVs) to identify reliable BCa biomarkers. This novel strategy presented here identified reliable urinary EV biomarker proteins exhibiting high levels of specificity and sensitivity for non‐invasive BCa detection.
Objective
18
F-labeled prostate-specific membrane antigen (PSMA) ligand,
18
FPSMA-1007, has the benefit of a higher synthetic yield and minimal excretion in the urine. High detection efficacy was ...reported in biochemical recurrence (BCR) of prostate cancer after radical prostatectomy. Thus, we evaluated the preliminary diagnostic utility of
18
FPSMA-1007 PET in patients with prostate cancer, focusing on the BCR which is not detected on conventional imaging.
Methods
We enrolled a total of 28 patients (age 51–79 years) with BCR of prostate cancer. BCR was defined as a continuous increase in PSA after radical prostatectomy or radiation therapy without any apparent recurrent lesions on conventional diagnostic imaging (CT and bone scintigraphy). PSMA-PET scanning was performed approximately 60 min after intravenous injection of
18
FPSMA-1007 (259 ± 37 MBq). PSMA-PET images were evaluated for lesion detection as well as its relation to PSA values and location.
Results
Abnormal uptake, which was suspected to be recurrence or metastasis, was detected in 92.9% (26/28) of patients with BCR. The SUVmax was 8.4 ± 6.4 in local recurrence, 11.5 ± 11.8 in pelvic lymph nodes (LN), and 4.1 ± 1.6 in bone metastasis. The detection rates were 66.7% in the PSA group-1 (0.1–0.5 ng/mL), 85.7% in the PSA group-2 (0.5–1.0 ng/mL), and 100% in the PSA group-3 (above 1.0 ng/mL). Among the PET-positive BCR patients (
n
= 26), local recurrence was detected in 57.7% (15/26), pelvic LN in 42.3% (11/26), and bone metastasis in 15.4% (4/26). In 53% (8/15) of BCR patients who were suspected of local recurrence, focal uptake was detected adjacent to the bladder on
18
FPSMA-1007 PET. This suggested the significant advantage of having minimal physiological urine excretion.
Conclusions
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FPSMA-1007 PET showed a high detection rate in recurrent and metastatic lesions. In patients with BCR, its high detection led to suitable treatment strategies, such as salvage radiation therapy or surgical removal of recurrent lymph nodes.
Trial registration
(UMIN Clinical Trials Registry) UMIN000037697.
Most upper tract urothelial carcinomas (UTUC) are muscle invasive at the time of diagnosis. Current standard methods for the diagnosis of UTUC are invasive. Urine cytology is the only non‐invasive ...test for detecting UTUC, but its sensitivity is low. A novel non‐invasive assay for UTUC detection would improve patient outcome. This study aimed to investigate the mutation of cell‐free DNA (cfDNA) in urine supernatant to develop a reliable diagnostic biomarker for UTUC patients. We studied urinary cfDNA from 153 individuals, including 56 patients with localized UTUC, and carried out droplet digital PCR assay for TERT promoter and FGFR3 hotspot mutations. We could detect mutations of TERT C228T in 22/56 (39.3%), TERT C250T in 4/56 (7.1%), and FGFR3 S249C in 9/56 (16.1%) patients. FGFR3 mutation was detected only in ≤pT1 tumors (positive predictive value: 100.0%). In combination with cytology results, the sensitivity was 78.6%, and the specificity was 96.0%. Although these data need to be validated in a larger‐scale cohort, mutation analysis of TERT promoter and FGFR3 in urinary cfDNA has the potential to be a non‐invasive diagnostic marker and reliable factor for tumor staging.
TERT promoter and FGFR3 hotspot mutations in urinary cell‐free DNA were analyzed. In combination with urine cytology, the sensitivity was 78.6%, and specificity was 96.0% for upper tract urothelial carcinoma diagnosis.
Reliable biomarkers for renal cell carcinoma (RCC) have yet to be determined. Circulating tumor DNA (ctDNA) is an emerging resource to detect and monitor molecular characteristics of various tumors. ...The present study aims to clarify the clinical utility of ctDNA for RCC. Fifty‐three patients histologically diagnosed with clear cell RCC were enrolled. Targeted sequencing was carried out using plasma cell‐free DNA (cfDNA) and tumor DNA. We applied droplet digital PCR (ddPCR) to validate detected mutations. cfDNA fragment size was also evaluated using a microfluidics‐based platform and sequencing. Proportion of cfDNA fragments was defined as the ratio of small (50‐166 bp) to large (167‐250 bp) cfDNA fragments. Association of mutant allele frequency of ctDNA with clinical course was analyzed. Prognostic potential was evaluated using log‐rank test. A total of 38 mutations across 16 (30%) patients were identified from cfDNA, including mutations in TP53 (n = 6) and VHL (n = 5), and median mutant allele frequency of ctDNA was 10%. We designed specific ddPCR probes for 11 mutations and detected the same mutations in both cfDNA and tumor DNA. Positive ctDNA was significantly associated with a higher proportion of cfDNA fragments (P = .033), indicating RCC patients with ctDNA had shorter fragment sizes of cfDNA. Interestingly, the changes of mutant allele frequency in ctDNA concurrently correlated with clinical course. Positive ctDNA and fragmentation of cfDNA were significantly associated with poor cancer‐specific survival (P < .001, P = .011). In conclusion, our study shows the clinical utility of ctDNA status and cfDNA fragment size as biomarkers for prognosis and disease monitoring in RCC.
We evaluated somatic mutations and fragmentation of circulating tumor DNA (ctDNA) using next‐generation sequencing and droplet digital PCR in renal cell carcinoma (RCC). ctDNA can be promising tools for monitoring and predicting prognosis of RCC.
Excessive intake of animal fat and resultant obesity are major risk factors for prostate cancer. Because the composition of the gut microbiota is known to change with dietary composition and body ...type, we used prostate-specific
knockout mice as a prostate cancer model to investigate whether there is a gut microbiota-mediated connection between animal fat intake and prostate cancer. Oral administration of an antibiotic mixture (Abx) in prostate cancer-bearing mice fed a high-fat diet containing a large proportion of lard drastically altered the composition of the gut microbiota including
and
, inhibited prostate cancer cell proliferation, and reduced prostate
expression and circulating insulin-like growth factor-1 (IGF1) levels. In prostate cancer tissue, MAPK and PI3K activities, both downstream of the IGF1 receptor, were suppressed by Abx administration. IGF1 directly promoted the proliferation of prostate cancer cell lines DU145 and 22Rv1
. Abx administration also reduced fecal levels of short-chain fatty acids (SCFA) produced by intestinal bacteria. Supplementation with SCFAs promoted tumor growth by increasing IGF1 levels. In humans, IGF1 was found to be highly expressed in prostate cancer tissue from obese patients. In conclusion, IGF1 production stimulated by SCFAs from gut microbes influences the growth of prostate cancer via activating local prostate MAPK and PI3K signaling, indicating the existence of a gut microbiota-IGF1-prostate axis. Disrupting this axis by modulating the gut microbiota may aid in prostate cancer prevention and treatment. SIGNIFICANCE: These results suggest that intestinal bacteria, acting through short-chain fatty acids, regulate systemic and local prostate IGF1 in the host, which can promote proliferation of prostate cancer cells.
High-fat diet (HFD) could induce prostate cancer progression. The aim of this study is to identify mechanisms of HFD-induced prostate cancer progression, focusing on inflammation.
We administered HFD ...and celecoxib to autochthonous immunocompetent Pb-Cre
;
(fl/fl) model mice for prostate cancer. Tumor growth was evaluated by tumor weight and Ki67 stain, and local immune cells were assessed by flow cytometry at 22 weeks of age. Cytokines which correlated with tumor growth were identified, and the changes of tumor growth and local immune cells after inhibition of the cytokine signals were evaluated in the mice. IHC analyses using prostatectomy specimens of obese patients were performed.
HFD accelerated tumor growth and increased the myeloid-derived suppressor cells (MDSCs) fraction and M2/M1 macrophage ratio in the model mice. Celecoxib-suppressed tumor growth, and decreased both local MDSCs and M2/M1 macrophage ratio in HFD-fed mice. HFD-induced tumor growth was associated with IL6 secreted by prostatic macrophages, as were phosphorylated STAT3 (pSTAT3)-positive tumor cells. Anti-IL6 receptor antibody administration suppressed tumor growth, and decreased local MDSCs and pSTAT3-positive cell fractions in HFD-fed mice. The tumor-infiltrating CD11b-positive cell count was significantly higher in prostatectomy specimens of obese than those of nonobese patients with prostate cancer.
HFD increased MDSCs and accelerated prostate cancer tumor growth via IL6/pSTAT3 signaling in the mice. This mechanism could exist in obese patients with prostate cancer. IL6-mediated inflammation could be a therapeutic target for prostate cancer.
.
To present mature results of high-dose-rate brachytherapy (HDR-BT) as monotherapy for intermediate- and high-risk prostate cancer.
From 1995 through 2012, 190 patients, 79 with intermediate-risk and ...111 with high-risk prostate cancer, were treated with HDR-BT alone using 48 Gy/8 fractions, 54 Gy/9 fractions, or 45.5 Gy/7 fractions over 4 to 5 days. Neoadjuvant with or without adjuvant androgen deprivation therapy was administered to 139 patients, 35 intermediate- and 104 high-risk.
Median follow-up time was 92 months (range, 10-227 months), with a minimum of 2 years for surviving patients. Respective rates of cause-specific survival, overall survival, metastasis-free survival, and biochemical no evidence of disease for the intermediate-risk patients were 100%, 100%, 96%, and 93% at 5 years, and 100%, 96%, 91%, and 91% at 8 years. Corresponding rates for the high-risk patients were 97%, 93%, 84%, and 81% at 5 years, and 93%, 81%, 74%, and 77% at 8 years. The cumulative incidence of late grade 2 to 3 genitourinary toxicity was 5% at 5 years and 10% at 8 years, and that of late grade 3 was 0 at 5 years and 1% at 8 years. The cumulative incidence of late grade 2-3 gastrointestinal toxicity was 4% at 5 years and 6% at 8 years, and that of late grade 3 was 0 at 5 years and 2% at 8 years. No grade 4 or 5 toxicity was detected.
Our single-institution study with a median 8-year follow-up showed that HDR-BT as monotherapy was safe and effective for patients with intermediate- and high-risk prostate cancer.
Prostate cancer is the second leading cause of cancer death in men in the United States. Several novel therapeutic agents have been developed for castration-resistant prostate cancer (CRPC), but the ...prognosis for patients with CRPC remains poor. The identification of novel therapeutic targets for CRPC is an urgent issue. Exosomes are small vesicles secreted by a variety of cells, and exosomes derived from cancer cells have been reported to circulate in the patient’s bodily fluids, promoting metastasis and invasion. We aimed to identify novel therapeutic targets for CRPC by proteomic analysis of serum exosomes. Exosomes were isolated by ultracentrifugation of sera from 36 men with metastatic prostate cancer: untreated (n = 8), well-controlled with primary androgen deprivation therapy (ADT) (n = 8), and CRPC (n = 20). We identified 823 proteins in the serum exosomes. Six proteins were increased in CRPC patients compared with untreated patients. In contrast, only ACTN4 was increased in the CRPC patients compared to the ADT patients. We focused on ACTN4 as a candidate for targeted therapeutics. ACTN4 was highly expressed in the prostate cancer cell line DU145 as well as exosomes from this line. RNA interference-mediated downregulation of ACTN4 significantly attenuated cell proliferation and invasion in DU145 cells. ACTN4 could be a potential therapeutic target for CRPC.
•Proteomic analysis of serum exosomes from patients with metastatic prostate cancer was performed.•ACTN4 was highly expressed in serum exosomes of castration-resistant prostate cancer patients.•Knockdown of ACTN4 attenuated proliferation and invasion of prostate cancer cells.
Inactivated hemagglutinating virus of Japan envelope (HVJ‐E) has an antitumor effect and tumor immunity. We undertook an open‐label, phase I, dose‐escalation study in patients with ...castration‐resistant prostate cancer (CRPC) to determine the safety and efficacy of intratumoral and s.c. injection of HVJ‐E (GEN0101). Patients with CRPC, who were resistant to or unable to receive standard of care, were included. GEN0101 was injected directly into the prostate and s.c. in two 28‐day treatment cycles. The primary end‐points were to evaluate the safety and tolerability of GEN0101 and determine its recommended dose. The secondary end‐points were to analyze the antitumor effect and tumor immunity. Three patients received 30 000 mNAU GEN0101 and 6 received 60 000 mNAU. There was no dose‐limiting toxicity, and the recommended dose of GEN0101 was defined as 60 000 mNAU. Radiographically, 1 patient had stable disease and 2 had progressive disease in the low‐dose group, whereas 5 patients had stable disease and 1 had progressive disease in the high‐dose group. Three patients in the high‐dose group showed reduction in lymph node metastasis. Prostate‐specific antigen increase rates in the high‐dose group were suppressed more than those in the low‐dose group. Natural killer cell activity was enhanced in 2 patients of the low‐dose group and in 5 patients in the high‐dose group. In conclusion, intratumoral and s.c. injections of GEN0101 were well‐tolerated and feasible to use. The study is registered with the UMIN Clinical Trials Registry (no. UMIN000017092).
Inactivated hemagglutinating virus of Japan envelope (HVJ‐E) has an antitumor effect and tumor immunity. We undertook an open‐label, phase I, dose‐escalation study in patients with castration‐resistant prostate cancer (CRPC). Intratumoral and s.c. injections of GEN0101 were well‐tolerated and feasible to use; antitumor effects were observed in patients with CRPC.
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