Increase In Cleaved Caspase-3 Expression Research Articles

In vitro administration of sodium arsenite in mouse prepubertal testis induces germ cell loss and apoptosis

High levels of arsenic contamination in drinking water pose serious health risks in numerous countries. The documentation reporting arsenic toxicity on reproduction and development is increasing, with evidence of arsenic inducing fertility and developmental issues. Nonetheless, the impact of arsenic exposure on the development of the male reproductive system is not fully elucidated. In the present study, we have investigated the direct effects of arsenic on prepubertal mouse testis using an in vitro testicular organ culture system. Culture medium was supplemented with a range of concentrations of sodium arsenite, examining effects of low (0.5 and 1 μM) and high (10, 50, 100 μM) concentrations, in cultures of post-natal day 5 CD1 mouse testis. In vitro exposure of low arsenic concentrations (0.5 or 1 μM) for 6 days did not cause any change in the testicular morphology, germ cells density, or apoptotic marker cleaved caspase 3 (CC3) expression. In contrast, exposure of prepubertal testis to high arsenic concentrations (10, 50 or 100 μM) induced drastic changes: severe destruction of testicular morphology, with loss of seminiferous tubule integrity; a dose-dependent decrease in germ cell density, and a hundred-fold increase in CC3 expression after 50 μM arsenic exposure. In conclusion, high arsenic treatment induced a dose-dependent induction of apoptosis and germ cell loss in prepubertal mouse testis.

Changes in ki-67 and cleaved caspase-3 (CC3) with short term androgen deprivation therapy (ADT) prior to prostatectomy as part of a trial in intermediate and high-risk prostate cancer (PC).

e17502 Background: Ki-67 and CC3 are markers of cellular proliferation and apoptosis and have been associated with prognosis in localized PC. Testosterone suppression promotes a time-dependent modulation of these indices. We explored changes in Ki-67 and CC3 at different timepoints in a pilot trial using short-term ADT prior to radical prostatectomy (RP). Methods: Pts with intermediate and high-risk localized PC were randomized to receive a single dose of neoadjuvant degarelix (240 mg SC) either 4 ± 1 days (Cohort A) or 7 ± 1 days (Cohort B) prior to RP. Anatomically matched tumor foci from the pre-treatment diagnostic biopsy and the prostatectomy specimens were assessed by immunohistochemistry (IHC) for Ki-67, CC3, and PTEN expression. Results: 32 pts were randomized: 15 to Cohort A and 17 to Cohort B. Ki-67 and CC3 expression in the pre- and post-treatment samples for both cohorts are summarized in the table. A significant reduction in Ki-67 and increase in CC3 expression occurred in both cohorts (all p-values < 0.05). No difference was found when the absolute difference of Ki-67 expression was compared between Cohort A and Cohort B (p = 0.7). A significant reduction of Ki-67 and increase in CC3 expression occurred in pts independent of PTEN status. Conclusions: A reduction of Ki-67 and an increase in CC3 expression was achieved in both cohorts following a single dose of degarelix within 1 week of RP. Although loss of PTEN has been associated with androgen-independent progression, reduction of proliferation and increase in apoptosis levels were achieved independent of PTEN status in this small cohort. Further analyses are ongoing to correlate the pathologic findings with clinical outcomes. Clinical trial information: NCT01542021 . [Table: see text]

Abstract 3878: Drug-loaded microparticles as a treatment approach for pancreatic cancer

Abstract Pancreatic cancer is the fourth leading cause of cancer death in the United States with only 7% of diagnosed patients surviving 5 years. Most pancreatic cancer patients are not surgical candidates due to advanced stage at diagnosis. Also, current systemic chemotherapies have not been very effective at decreasing tumor burden. Poly(lactic-co-glycolic acid)-based (PLGA) microparticles (MPs) are a promising tool for localized drug delivery within the tumor due to their biocompatibility, flexibility in drug encapsulation and extended drug release. The present study investigated whether gemcitabine-loaded microparticles (GMPs), paclitaxel-loaded microparticles (PMPs) or sequential treatment of both, in comparison with blank (no drug) MPs, systemic treatments and no treatment control, are able to promote cancer cell killing effects and modulate drug resistance in vitro and in vivo. We previously showed in vitro studies with PANC-1 cells and we were able to complete the MPs studies with a second human pancreatic cancer cell line, MIAPaCa-2. In both cases, we tested the effect of single and combination treatments on two well-known resistance markers for gemcitabine, ribonucleotide reductase catalytic subunit M1 and cytidine deaminase, as well as on the promotion of cell death measuring cleaved caspase-3 (CC3). When treated with GMPs alone, both markers went up suggesting an increase in resistance against gemcitabine. Interestingly, both cell lines treated with PMPs alone showed an increase in CC3 expression and a significant decrease in the expression of both markers. Subsequently, we tested the in vivo efficacy of MPs by direct injection into established subcutaneous MIAPaCa-2 tumors in nude mice. Following four weeks of treatment, the tumors were excised, biopsied for protein analysis and frozen in OCT to allow visualization of fluorescent MPs and detect apoptosis by immunofluorescence. We also want to evaluate reactive oxygen species production; where we expect to see an increase in the MPs treated groups. This analysis is in progress. In conclusion, we observed a decrease in cell viability and drug resistance proteins in vitro using the drug-loaded microparticles in two grade 3 (or high grade) pancreatic cancer cell lines. The described drug delivery method has the potential to be a more efficient treatment modality than systemic gemcitabine and paclitaxel against early stage and locally advanced pancreatic cancer. Citation Format: Maria Munoz-Sagastibelza, Oluwafeyikemi Okome, Jenny E. Paredes Sanchez, Albert Palileo, Catherine Burkhart, Laura Martello-Rooney. Drug-loaded microparticles as a treatment approach for pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3878.

Abstract 1091: Paclitaxel-loaded microparticles in combination with gemcitabine-loaded microparticles decreases gemcitabine resistance and promotes cell death in pancreatic cancer cell lines

Abstract Pancreatic cancer is the fourth leading cause of cancer death in the United States with only 7% of diagnosed patients surviving 5 years. Current systemic chemotherapies have not been very effective at decreasing tumor burden, but nonetheless expose patients to the adverse side effects of treatment. Poly(lactic-co-glycolic acid)-based microparticles (MPs) are a promising tool for localized drug delivery within the tumor due to their high biocompatibility, flexibility in drug encapsulation and extended drug release. A previous in vivo experiment from our laboratory demonstrated minimal penetration of systemic gemcitabine into the tumor resulting in low levels of apoptosis compared to locally injected gemcitabine-loaded microparticles (GMPs), which demonstrated much higher levels of intratumoral apoptosis. The present study investigated whether paclitaxel-loaded microparticles (PMPs), alone or in combination with GMPs, decreased gemcitabine resistance, colony formation and promoted cell death in two human pancreatic cancer cell lines, PANC-1 and MIAPaCa-2. In vitro studies with PANC-1 cells treated with PMPs or GMPs showed enhanced cell killing and a significant decrease in colony formation in the higher MPs volume groups. Moreover, the combination treatment of PMPs followed by GMPs, showed a greater decrease in colony formation in comparison with the single treatments. Subsequently, we tested the effect of single and combination treatments on two well-known resistance markers for gemcitabine, ribonucleotide reductase catalytic subunit M1 (RRM1) and cytidine deaminase (CDA), as well as on the promotion of cell death measuring cleaved caspase-3 (CC3). Interestingly, PANC-1 cells treated with PMPs alone showed an increase in CC3 expression and a significant decrease in the expression of both markers when treated with different volumes of PMPs. MIAPaCa-2 cells exhibited similar results. The combination treatment also showed a modulation in RRM1 and CDA proteins and an increase in CC3 expression. However, when PANC-1 cells were treated with GMPs alone, both markers went up suggesting an increase in resistance against gemcitabine. These experiments are in progress for the MIAPaCa-2 cells. In addition, preliminary drug release studies with GMPs demonstrated detection of gemcitabine up to 14 days. In conclusion, our data demonstrated that PMPs and GMPs promote an increase in cancer cell death. Furthermore, the combination of the drug-loaded MPs indicated a reduction in drug resistance and clear impairment in colony formation. Further studies are in progress to investigate combination MPs injections in a mouse model of pancreatic cancer to confirm in vivo efficacy. The described drug delivery method has the potential to be a more efficient local treatment modality than systemic drug against pancreatic cancer. Citation Format: Maria Munoz-Sagastibelza, Ariella Moshchinsky, Oluwafeyikemi Okome, Jenny E. Paredes Sanchez, Raavi Gupta, Laura Martello-Rooney. Paclitaxel-loaded microparticles in combination with gemcitabine-loaded microparticles decreases gemcitabine resistance and promotes cell death in pancreatic cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1091. doi:10.1158/1538-7445.AM2017-1091