Núñez-Nateras, Rafael

ABSTRACT Despite the annual increase in liver transplants, a persistent disparity between suitable donors and recipients continues to challenge the transplant community. The introduction of machine perfusion emerged as a novel approach to organ preservation. This technique, which utilizes extended criteria donor grafts, is expected to reduce rejection rates and expand the donor pool. Its principle, like that of any ex vivo machine perfusion, is based on the continuous supply of nutrients and oxygen to maintain cellular metabolism while interrupting anaerobic processes. This helps prevent conditions such as ischemia-reperfusion injury, early graft dysfunction, and ischemic cholangiopathy, which can lead to graft loss and the need for retransplantation. There are two main modes of ex vivo machine perfusion: hypothermic and normothermic, differentiated by the operating temperature. Utilizing ex situ perfusion machines presents a promising opportunity to increase liver graft usage and reduce waitlist times. However, it is crucial to evaluate how much this technology would raise the overall cost of transplants. In Brazil, implementing a machine perfusion program may encounter unique challenges not seen in other countries. These challenges include limited access to the healthcare system for high-risk liver transplant candidates, long waiting lists, inadequate support for organ donors, reliance solely on deceased brain-dead donors, since donation after cardiac death is not ethically accepted, and geographical barriers.

287 Background: Our preliminary immunohistochemical studies of bladder cancer patients have shown that the inflammatory/immune responses associated with bladder cancer appear to be Th2-polarized as judged by the preponderance of tumor infiltrating GATA-3+ (Th2) vs. T-bet+ (Th1) T cells, as well as by evidence of tumor-associated eosinophil infiltration and activation. The standard-of-care treatment for Tis bladder cancer is intravesical administration of BCG, a Th1-polarizing immunomodulator. The aim of the present study was to assess the Th2/Th1 ratio correlation with response to BCG therapy in patients with Tis bladder cancer. Methods: Two groups of patients were studied. Group A included 20 patients that responded to BCG, and Group B included 20 patients that did not respond to therapy. Response to BCG was determined as presence/absence of disease at 6 months post-BCG therapy. The initial diagnostic biopsies of these patients were subjected to immunohistochemical analysis using commercially available antibodies specific for Th2 (GATA-3+) and Th1 (T-bet+) lymphocytes, and our unique monoclonal antibody specific for eosinophil peroxidase (EPX-mAb). Ten random high powered (400X) fields at the maximum focus of mononuclear infiltration were examined and the number of GATA 3+ and T-bet+ tumor infiltrating T cells were counted to define the G/T ratio for each patient sample. Eosinophils and their degranulation activity were counted in the same fields. Results: Patients with response to BCG therapy had a G/T ratio of ≥ 3 and patients with non-response to BCG therapy had a G/T ratio of ≤ 1.5. Correlation of these scores with subsequent patient outcomes following BCG immune therapy demonstrated that patient responsiveness (i.e., tumor elimination at 6 months) trended with the higher G/T ratio patients. In addition, initial biopsies of BCG responsive patients display a higher number of eosinophils and degranulation relative to BCG non-responsive patients. Conclusions: Evaluations of initial biopsies revealed a strong correlation between Th1 and Th2 immune response with BCG therapy outcomes. The development of specific and novel tools to identify/measure Th2 immune responses in Tis bladder cancer provides a unique opportunity to predict outcomes at the time of diagnosis.

214 Background: In our preliminary in vitro studies, we have demonstrated evidence of enhanced apoptosis and inhibition of cellular proliferation in both hormone sensitive and castrate resistant prostate cancer (PCa) cell lines using a combination of an antiandrogen (Bicalutamide) and a selective estrogen receptor modulator (Raloxifene). The aim of this study was to study the effect of the administration of these two drugs in in vivo models of castrate resistant PCa. Methods: In vivo model consisted on NCr Nude: Mice bearing s.c. human prostate (PC3 cell line) xenografts. Based on the treatment received, mice were divided into 5 groups as follows: Group 1: No drugs (control); Group 2: Bicalutamide 50mg; Group 3: Raloxifene 60 mg; Group 4: Combined administration of Bicalutamide 50 mg and Raloxifene 60 mg; Group 5 Combined administration of Bicalutamide 150 mg and Raloxifene 120 mg. A total of 10 mice where included in each group. All drugs dosages were converted to their equivalent in the mice. Drugs were administered by gavage technique to the mice once per day for a total of 14 days. Results: As expected, Bicalutamide administered alone causes minimal inhibition without reaching statistical significance (Group 2: 0.34 g Vs Group 1: 0.40 g; p=0.073). Although Raloxifene causes some marked growth inhibition, its effect is not statistically significant (Group 3: 0.31 Vs Group 1: 0.40 g; p=0.062). Bicalutamide and Raloxifene, when administered in combination, induced prominent growth inhibition in PC3 tumors when compared to the control group (Group 4: 0.26 g Vs Group 1: 0.40 g; p=0.038). Growth inhibition is significantly more evident when the drugs dosages are increased (Group 5: 0.17 g Vs Group 1: 0.40 g; p=0.024). Conclusions: The simultaneous administration of Bicalutamide and Raloxifene appears to have a synergistic effect on tumor growth inhibition in PC3 xenografts. The pathway(s) responsible for this observation may be independent of the androgen receptor as PC3 cells are AR negative and still affected by the combination over the drugs administered alone. Research is warranted to identify these potential pathways.

290 Background: Previous studies have shown robot assisted radical cystectomy (RARC) to have equivalent perioperative outcomes to open radical cystectomy. There are few reports that have examined the oncologic results of RARC specifically with respect to node-positive patients. We report the outcomes of node-positive patients who have undergone RARC with medium-term (at least 1 year) follow-up. Methods: A total of 275 patients underwent RARC at two institutions for invasive bladder cancer between 2005-present. We examined the 50 patients with node-positive disease that had a minimum of one year follow-up. Oncologic outcomes, recurrence free survival (RFS), and disease specific survival (DSS) were analyzed and compared to the open literature. Results: Mean clinical follow up in this case series was 29 months (range 12–64 months). The mean number of lymph nodes removed was 18 (range 5–35), and mean number of positive LNs was 3.1 (range 1–12). Overall rate of LN positivity was 26%. Mean LN density was 18%. Seventeen (34%) patients had ≤ pT2 disease and 33 (66%) pT3/T4 disease. At this follow-up, 29 patients have recurred, 21 patients died of disease, giving a RFS and DSS of 42% and 58%, respectively. Mean (median) time to recurrence was 10.2 months (9 months). A total of 60% of patients received peri-operative chemotherapy in this cohort. These findings are consistent with prior reports of such oncologic outcomes in node-positive patients in open series. Conclusions: The oncologic follow-up of patients undergoing RARC with LN positive disease appears to have acceptable outcomes during medium term (mean 29 months) follow-up. As our follow-up increases, we expect to continue to accurately define the long-term clinical suitability and oncologic success of this procedure in this high-risk population. No significant financial relationships to disclose.

288 Background: We report our experience with robot assisted radical cystectomy (RARC) with regard to medium-term (at least 2 year) oncologic outcomes. Methods: A total of 275 patients have undergone RARC and urinary diversion at two institutions for invasive bladder cancer between 2005-present. We performed a retrospective analysis of the 139 patients who underwent RARC with a minimum of 2 years follow-up. Medium term oncologic outcomes including recurrence rates, time to recurrence, recurrence free survival (RFS), disease specific survival (DSS) were analyzed. Follow-up was measured from time of surgery to time of most recent clinical follow-up. Results: This cohort of patients consisted of 108 men (78%) and 31 women (22%) at a mean age of 67.3 years (range 45-86 years). Sixty-one (44%) patients had ≤ pT2 disease, 38 (27%) pT3/T4 disease, and 40 (29%) N+ disease. The mean number of lymph nodes removed was 18 (range 3-41). The average clinical follow up in this case series was nearly 3 years with a mean of 35.9 months (range 24-64 months). At this follow-up, 39 patients have recurred, 27 patients died of disease, and 5 patients died of other causes giving an overall RFS, DSS, and OS rates of 80%, 71%, and 68%, respectively. The mean (median) time to recurrence was 12.3 months (10 months). These findings are consistent with prior reports of the oncologic outcomes for open radical cystectomy. Conclusions: The oncologic follow-up of patients undergoing RARC appears to be favorable with acceptable outcomes in the medium-term (mean – 3 years). As our follow-up increases, we should expect to truly define the long-term clinical appropriateness and oncologic success of this procedure. No significant financial relationships to disclose.

168 Background: Androgens and estrogens have been shown to play an important role in normal prostate development and function as well as carcinogenesis and development of the castration resistant phenotype of disease. The aim of this study was to evaluate the effect of a simultaneous administration of an androgen receptor antagonist (bicalutamide) and a selective estrogen receptor modulator (raloxifene) on both androgen sensitive and androgen insensitive prostate cancer cell lines. Methods: Experiments were performed on LNCaP, PC3 and DU145 cell lines. Western blot was utilized for the identification of androgen and estrogen receptors (a andb) in the cell lines. Drug concentrations required to achieve IC 50 were obtained using the MTT assay; such concentrations were identified for the drugs individually and when used in combination. The effect of the drugs on apoptosis was assessed using flow cytometry. Results: Results of the IC 50 for the drugs alone and in combination by each cell line are shown in the table. An enhanced effect was observed when the drugs were used in combination in all the cell lines. It was evident that the combination of the drugs decreased the total drug required to achieve the IC50 decreases considerably. Apoptosis rates were also affected by the simultaneous administration of bicalutamide and raloxifene. The synergistic effect of the combination was reflected in the increase of the apoptosis rate in all cell lines. Conclusions: The simultaneous administration of bicalutamide and raloxifene has a synergistic effect on cell death and apoptosis of DU145, PC3 and LNCaP cell lines. The pathway(s) responsible for this observation may be independent of the androgen receptor as both AR negative cell lines were still affected by the combination over the SERM alone. [Table: see text] No significant financial relationships to disclose.

Purpose: To determine whether on-clamp partial nephrectomy (ON-PN) has any significant impact on long-term renal function in a two kidney model. Patients and Methods: From November 1999 to July 2013, 607 patients underwent PN at our institution. After excluding patients with solitary kidneys, multiple renal masses, and follow-up less than 90 days, 331 remained. Patient demographics were assessed, as was renal function based on pre- and postoperative mercaptoacetyltriglycine (MAG-3) renal scans and change in estimated glomerular filtration rate (eGFR) using the preoperative and most recent recorded creatinine levels. Results: There were a total of 236 patients who underwent ON-PN and 95 who underwent off-clamp PN (OFF-PN) during the study period. The longest follow-up was 12.6 years with mean follow-up of 3 years. Mean ischemia time of patients undergoing ON-PN was 25 minutes (range 8–63 min). No differences were noted between the ON-PN and OFF-PN cohorts with respect to estimated change in eGFR (ON-PN: −6.07 mL/min/1.73 m 2 vs OFF-PN: −6.00 mL/min/1.73 m 2 , P =0.69). No differences were noted in the % change in the MAG-3 renal scans (ON-PN: −0.77% vs OFF-PN: −1.1%, P =0.94). A post hoc sensitivity analysis of the same two variables stratified by age revealed no differences in change in estimated GFR or % change in differential function on renal scan. Conclusions: In the two kidney model, ischemia does not appear to affect long-term renal function outcomes after PN. These data provide evidence that ON-PN is perfectly acceptable in the appropriately selected patient with two kidneys.

Objective To present the first age‐stratified assessment of outcomes after holmium laser enucleation of the prostate ( HoLEP ) for the treatment of lower urinary tract symptoms resulting from prostate enlargement. Patients and Methods We retrospectively analysed and compared the morbidity, and the peri‐operative and functional outcomes of patients aged 50–59, 60–69, 70–79 and ≥80 years. Complications at 30 days were stratified using the C lavien system. Functional outcomes were assessed using the I nternational P rostate S ymptom S core ( IPSS ), maximum urinary flow rate ( Q max ), post‐void residual urine volume ( PVR ) and urinary continence. Results A total of 311 patients underwent HoLEP for obstructive voiding symptoms from A ugust 2007 to J une 2011, of whom 22 patients were aged 50–59 years, 91 were aged 60–69 years, 153 were aged 70–79 years, and 45 were aged ≥80 years. The overall morbidity rates were similar among the age groups (20, 24.4, 21.6 and 22.1% for groups 1, 2, 3 and 4, respectively), as were the incidence of significant complications ( C lavien grade ≥ III ), change in serum haemoglobin level, and length of hospital stay. Patients ≥80 years did have a longer catheterization time (3.4 days) than patients aged 50–59 years (1.68 days). By 1 year there were no significant differences in urinary continence, IPSS , Q max , or PVR among the age groups. Conclusions Overall morbidity, hospital stay, and 1‐year functional outcomes of HoLEP were similar among all age groups. This study shows that HoLEP is a safe and effective treatment for benign prostatic hyperplasia regardless of age.