Emmett Pinney

Abstract 2265: Sub-fractions of conditioned medium, from hypoxia-induced cells with multipotent potential, exhibit a significant anti-oncogenic activity

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA We have previously shown that a human soluble cell conditioned media (CCM) from hypoxia-induced, potentially multipotent cells, grown in a perfusion bioreactor system inhibits tumor cell growth in vitro and in two animal models. Here we present data that indicate that a low molecular weight sub-fraction from the CCM has an increased anti-oncogenic activity. This activity was demonstrated in orthotopic pancreatic models, nRAS melanoma assays and over 10 additional human cancer cell lines in vitro. In an orthotopic BxPC3 pancreatic cancer model, there was a statistically significant improvement in survival following weekly intravenous injections of 0.1ml of sub fractioned CCM, as well as a significant improvement with 0.5ml intraperitoneal injections post-resection, as measured by Kaplan Meier analysis (p<0.05) (figures below). Uveal and nRAS melanoma cancer cell growth was inhibited by greater than 90% in in-vitro proliferation assays. The mechanism of action of the CCM is the induction of apoptosis through the upregulation of Caspase 3 and 9, as demonstrated by immunostaining of Annexin V and immunoblot ayalysis. In a Miapaca nude mouse daily dose study, tumor mass and metastatic extent was substantially reduced in treated mouse versus the control. Further, in a BXPC3 nude mouse daily dose study, 60% (p<0.05) of treated mice were tumor free after four weeks of treatment, while only 10% were tumor free in the control group . Little or no apparent drug toxicity was observed. These results indicate that the CCM sub-fraction could be a useful raw material for a treatment for a large range of neoplastic diseases. Citation Format: Emmett Pinney, Mayra Montes-Camacho, Kayler Brintle, Christian Posch, Rhiana Menen, David Easter, Michael Bouvet, Robert Hoffman, Gail Naughton. Sub-fractions of conditioned medium, from hypoxia-induced cells with multipotent potential, exhibit a significant anti-oncogenic activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2265. doi:10.1158/1538-7445.AM2014-2265

Abstract 2689: Inhibition of metastasis of circulating tumor cells by extracellular matrix secreted by human embryonic cells

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL We have previously demonstrated the increased metastatic potential of human prostate cancer PC-3 CTCs compared to their parental counterparts in both chick embryo and mouse models. In the present study, we tested the effects of human embryonic extracellular matrix (ECM), secreted by human embryonic cells including conditioned medium (CM) and a semi-solid form (hECM), on metastasis of PC-3 CTC in the chick embryo model. The chorioallentoic membrane (CAM) of 18 chicken embryos were inoculated with either 106 PC-3 human prostate cancer cells or PC-3 CTCs, both stably expressing GFP. Twelve hours later, embryos were divided into 6 groups each containing three embryos: PC-3 parental control; PC-3 + CM; PC-3 + hECM; CTC control; CTC + CM; and CTC+ hECM. Twelve hours following inoculation of the cells, a single dose of 100 μL of either treatment was given to the appropriate group. Embryo brains were removed on day 8 post-inoculations, and processed for cryo-sectioning, generating 3 slides of brain tissue per embryo taken at various depths. Imaging was performed using the IV-100 scanning laser microscope (Olympus Corp, Tokyo, Japan) in order to count metastatic foci. PC-3 controls had an average of 11.1 metastatic foci compared to 2.55 in the PC-3 + hECM group (p< 0.0001) and 2.76 in the PC-3 + CM group (p< 0.0001) The treatment showed even greater response on the CTC cells which an average of 30.9 metastatic foci in the CTC controls compared to 4.38 in the CTC + hECM group (p< 0.0001) and 4.18 in the CTC + CM group (p< 0.0001). Thus, human embryonic secreted ECM compound drastically decreased the metastatic potential of human prostate cancer CTCs in the chick embryo model. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2689. doi:1538-7445.AM2012-2689

Abstract 2676: A secreted tissue engineered extracellular matrix (shECM) prevents recurrence in post-resection pancreatic cancer models

Pancreatic cancer is a highly treatment-resistant cancer. Gemcitabine, which is first-line therapy, results in toxicity without meaningful survival benefit. We report here that a extracellular matrix (ECM) secreted by human embryone fibroblasts in culture inhibit human pancreatic cancer growth and post-surgical recurrence in an orthotopic nude mouse models. Post-resection, all untreated control mice had recurrent disease. In contrast, only 2 of 8 mice (p=0.004) in the treatment group had palpable masses after 6 weekly intravenous treatments. In a second study, targeting the one surgical site with ECM treatment followed by 8 weekly i.p. treatments of ECM, 7 of 8 treated mice survived beyond 10 weeks while none of the untreated control mice survival 8 weeks. No measurable toxicity was observed in the treatment group. These results demonstrate the potential of ECM treatment of pancreatic cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2676. doi:1538-7445.AM2012-2676

Abstract 432: A tissue engineered human extra cellular matrix (hECM) inhibits tumor growth

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL A human extracellular matrix (hECM) was extracted from deposits made by neonatal dermal fibroblasts that were seeded on dextran microspheres and cultivated in a stirred bioreactor. The resulting hECM is high in collagen content with reduced hydroxylation as well as proteins found primarily in fetal ECM. These include high concentrations of SPARC, fibulin-2, tenascin, decorin, lumican, and fibromodulin. This hECM was used to treat tumor lines that were introduced onto the CAM of the fertilized chicken egg or growing subcutaneously in the nude mouse. In the CAM model significant tumor inhibition by hECM was seen with the following tumor lines: B16 melanoma was reduced 67%, C6 glioma 58% and MDA-MB-435 breast cancer 78%. Histology indicated that where the cells are in contact with hECM the cells appeared apoptotic. In the nude mouse models similar inhibition by hECM was observed: B16 melanoma was reduced 61%, C6 glioma 60% and MDA-MB-435 breast cancer reduced 58%. Apoptotic pathway activation was confirmed with caspase 9 upregulation in a dose-dependent response to hECM. These data indicate that an ECM rich in fetal proteins is capable of retarding tumor growth. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 432. doi:10.1158/1538-7445.AM2011-432

Abstract 4281: Real-time imaging of the cell-cycle in vivo

FUCCI (Fluorescent Ubiquitination-based Cell Cycle Indicator) cells are labeled orange in the G1 phase nuclei (monomeric Kusabira-Orange2: mKO2) and labeled green in the S/G2/M phases (monomeric Azami-Green1: mAG1) (Cell 132, 487-498, 2008). FUCCI-expressing MKN45 gastric cancer cells were used for imaging cell-cycle dynamics in vivo and in vitro. When a confluent monolayer of MKN45-FUCCI was scratched, the edge of the wound turned green within 24hr, indicating that healing of the wound required proliferation of MKN45 cells. FUCCI-expressing MKN45 cells were injected subcutaneously in the nude mice. The injected cells gradually grew and emitted both green and red fluorescence for 14 days, indicating cell cycle progression. In the chorioallantonic membrane (CAM) chick embryo model, green and red fluorescence FUCCI cells and blood vessels were observed for 5 days. FUCCI cells of the edge in the tumor migrated into the transparent egg membrane. Mouse and CAM FUCCI tumor models will be used to study the role of cell cycle dynamics in metastasis including dormancy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4281. doi:10.1158/1538-7445.AM2011-4281