PMC

(a) Comparative pharmacokinetics of parenteral NanoCurc™ versus free curcumin in corn oil, administered in non-tumor bearing mice. Plasma concentrations of curcumin (ng/mL) were determined at 1, 2, 4, 8 and 24 hours after single intraperitoneal administration of either formulation, at an equivalent dose of 25 mg/kg curcumin. See text for analytical details.
(b) Tissue and plasma levels of curcumin assessed following necropsy in non-tumor bearing mice receiving 4 weeks of parenteral NanoCurc™. Curcumin levels were measured by LC-MS/MS, and are expressed as μg/g for tissues, and in μg/mL for plasma. Horizontal line indicates mean of levels measured in three mice. The anatomic site is indicated on the x-axis.
(c) No significant alterations in body mass (grams) are observed in cohorts of non-tumor bearing mice (N=3) receiving either NanoCurc™ or void NVA622 polymer for four weeks.
(d) Histopathological assessment of visceral tissues (clockwise from top left, kidney, pancreas, lung, liver, spleen, and intestine) obtained at necropsy from mice receiving 4 weeks of NanoCurc™ demonstrate no abnormalities.

(a) Graphical illustration of tumor volumes for orthotopic Pa03C pancreatic cancer xenografts treated with NanoCurc™, gemcitabine, or the combination, compared to void NVA622 polymer. Single agent NanoCurc™ leads to significant inhibition of the “primary” tumor volume compared to void polymer (double asterisks), and the effect is further accentuated upon addition of gemcitabine (triple asterisks). Note that the combination also has a more significant effect on tumor volume compared to single-agent gemcitabine (double asterisks). All treatments were carried out for a period of three weeks; horizontal lines represent the average of measurements in seven mice per arm.
(b) Representative excised Pa03C xenografts from the control (far left) and three treatment arms (NanoCurc™, gemcitabine, and combination, respectively) are designated.
(c) Control mice receiving void polymer (VP) demonstrate extensive micrometastases to the lungs, lymph nodes and peritoneum (also see ). Both single-agent therapy arms demonstrate considerable reduction in micrometastatic disease (albeit still present in the lymph nodes), while the combination arm demonstrates complete abrogation of micrometastatic disease in all examined viscera.

(a) Subcutaneous xenografts were established using the low-passage Pa03C (a.k.a. LZ10.7) human pancreatic cancer cell line, and mice were randomized to four arms, including (i) control (void NVA622 polymer), (ii) NanoCurc™, (iii) gemcitabine, and (iv) combination of NanoCurc™ and gemcitabine. Treatment was culminated at three weeks. Representative xenografted mice from each of the four arms are illustrated. While single-agent NanoCurc™ significantly blocked tumor growth compared to void NVA622 arm (see panel b), the results were even more striking upon administration with gemcitabine, wherein four of five xenografts demonstrated complete regression in the combination arm. Photomicrographs to the right depict a representative xenograft from the gemcitabine arm, and the single residual “nubbin” from the combination therapy group.
(b) Graphical depiction of the tumor volumes in the four arms, over the three week time course of therapy. Single-agent NanoCurc™ demonstrates significant reduction in tumor volume compared to void NVA622 (control) arm at three weeks (double asterisk, P<0.01). No significant difference is observed in the tumor volumes between single-agent gemcitabine and combination arms; however, this data underestimates the effect of combination therapy, as four of the five xenografts had undergone complete histological regression (see panel a), and only a residual “nubbin” of tumor from a single xenograft was available for measurement. The inset demonstrates the comparative tumor volume data for gemcitabine and combination arms using a magnified Y-axis, which illustrates the clear separation in growth curves between the two arms during the third week.

(a) In subcutaneous Pa03C xenografts, immunohistochemistry for nuclear p65 (active NFκB) demonstrates significant reduction in nuclear labeling in NanoCurc™-treated xenografts (hashed bar) compared to those receiving void NVA622 polymer (black bar). The Y-axis designates number of cells with nuclear p65 staining per high power filed (40X), over an average of 10 randomly selected fields. Bottom panel illustrates photomicrograph of p65 immunohistochemistry in the two arms, at the given (40X) objective. Please note that due to only a single residual “nubbin” of tumor in the combination therapy arm, immunohistochemistry could not be reliably performed in this case.
(b) Electrophoretic mobility shift assay (EMSA) was performed as a “gold standard” for NFκB activity in orthotopic Pa03C xenografts, treated with vehicle (Lanes 1 and 2), NanoCurc™ (Lanes 3 and 4), combination (Lanes 5 and 6) and gemcitabine (Lanes 7 and 8); Lane 9 is a Jurkat cell line nuclear lysate with activated NFκB (positive control). The intra-tumoral curcumin concentrations for each pair of xenografts in each arm are represented at the bottom (Y-axis equals concentration in μg/g of tissue). Reduction in DNA binding ability of NFκB (as indicated by gel shift) is observed in both xenografts in the combination arm (Lanes 5 and 6), as well as in one of two xenografts receiving single-agent NanoCurc™ (Lane 3), all three of which have robust levels of intra-tumoral curcumin (>2.5μg/g of tissue). In contrast, one of the two xenografts receiving single agent NanoCurc™ (Lane 4) with retained NFκB activity also demonstrates ∼4-fold lower intra-tumoral curcumin concentration (∼0.6μg/g). Expectedly, no reduction in DNA binding ability is seen for the xenografts treated with vehicle or single agent gemcitabine only.