Natalie B. Jones

IL-12 Enhances the Antitumor Actions of Trastuzumab via NK Cell IFN-γ Production

The antitumor effects of therapeutic mAbs may depend on immune effector cells that express FcRs for IgG. IL-12 is a cytokine that stimulates IFN-γ production from NK cells and T cells. We hypothesized that coadministration of IL-12 with a murine anti-HER2/neu mAb (4D5) would enhance the FcR-dependent immune mechanisms that contribute to its antitumor activity. Thrice-weekly therapy with IL-12 (1 μg) and 4D5 (1 mg/kg) significantly suppressed the growth of a murine colon adenocarcinoma that was engineered to express human HER2 (CT-26HER2/neu) in BALB/c mice compared with the result of therapy with IL-12, 4D5, or PBS alone. Combination therapy was associated with increased circulating levels of IFN-γ, monokine induced by IFN-γ, and RANTES. Experiments with IFN-γ–deficient mice demonstrated that this cytokine was necessary for the observed antitumor effects of therapy with IL-12 plus 4D5. Immune cell depletion experiments showed that NK cells (but not CD4+ or CD8+ T cells) mediated the antitumor effects of this treatment combination. Therapy of HER2/neu-positive tumors with trastuzumab plus IL-12 induced tumor necrosis but did not affect tumor proliferation, apoptosis, vascularity, or lymphocyte infiltration. In vitro experiments with CT-26HER2/neu tumor cells revealed that IFN-γ induced an intracellular signal but did not inhibit cellular proliferation or induce apoptosis. Taken together, these data suggest that tumor regression in response to trastuzumab plus IL-12 is mediated through NK cell IFN-γ production and provide a rationale for the coadministration of NK cell-activating cytokines with therapeutic mAbs.

VEGF Secretion is Inhibited by Interferon-Alpha in Several Melanoma Cell Lines

Interferon-alpha (IFN-alpha) is employed in the treatment of malignant melanoma; however, it mediates regression of disease in only 10-15% of patients. Currently, its mechanism of action is uncharacterized. Low-dose IFN-alpha exerts anti-angiogenic effects when used in the treatment of life-threatening hemangiomas of infancy, suggesting anti-angiogenesis as a mechanism of action. IFN-alpha may exert its anti-tumor effect in the setting of advanced malignancy by inhibiting the secretion of vascular endothelial growth factor (VEGF), a pro-angiogenic substance. We hypothesized that IFN-alpha would decrease the release of VEGF by melanoma tumors. We studied the effect of IFN-alpha on VEGF production in nine human melanoma cell lines. We also examined VEGF levels in 49 patients with advanced malignancies who received low-dose IFN-alpha and interleukin-12 (IL-12) on an NCI-sponsored phase I trial. Human melanoma cell lines produced varying amounts of VEGF in vitro (60-1500 pg/mL at 48 h). Certain melanoma cell lines such as 18105 MEL secreted low levels of VEGF (152 pg/mL) after 48 h of culture, whereas other lines secreted very high levels (FO-1 3,802 pg/mL). Treatment of melanoma cells with IFN-alpha (2000 U/mL) decreased VEGF secretion by 40-60% in VEGF-high cell lines; however, this effect was not demonstrated in VEGF-low cell lines. In cancer patients, pretreatment VEGF plasma levels varied from 471 to 4200 pg/mL. A decrease in VEGF plasma levels after treatment directly correlated with the number of treatment cycles administered (Pearson correlation, p = 0.04). In summary, IFN-alpha inhibits VEGF secretion by melanoma cell lines in vitro and may have similar actions in malignancies that respond to IFN-alpha treatment.

Use of a nanoporous biodegradable miniature device to regulate cytokine release for cancer treatment

The clinical management of locally recurrent or unresectable malignant melanoma continues to pose a significant challenge. These lesions are typically painful and currently available treatments, such as repeated intratumoral injections of interferon-alpha (IFN-α), are costly and inconvenient. Nanotechnology offers promise as a novel means of drug delivery. A capsule-like nanoporous miniature device (NMD) based on a biodegradable polymer, poly(polycaprolactone) (PCL) was developed for controlling the local delivery of immunological agents to the tumor microenvironment. The device consists of a nanoporous release gate, a fabricated drug reservoir loaded with IFN-α and a protective layer. To improve the biocompatibility of the device, a hydrophilic poly(ethylene glycol) monoacrylate was applied to the outside wall of the device via covalent bonding techniques. Microscopic visualization of the nanoporous gate from in vitro experiments exhibited good pore stability over a two-month period. In vitro experiments demonstrated a constant release rate of IFN-α from the NMD and showed that the release rate could be regulated by the gate area. The released IFN-α was biologically functional. Cytokine-containing supernatants from release experiments phosphorylated signal transducer and activator of transcription (STAT1) in peripheral blood mononuclear cells. Subcutaneous implantation of the NMDs was well tolerated and associated with an anti-tumor effect in a human xenograft model of melanoma. There was no evidence of a significant inflammatory response to the NMD or encapsulation of the NMD by fibrosis. These experiments show that the NMD can be fabricated and employed in vivo as a versatile drug delivery platform.

Clinical factors predictive of malignant and premalignant cystic neoplasms of the pancreas: A single institution experience

BACKGROUND As cystic neoplasms of the pancreas are discovered with advanced imaging techniques, pancreatic surgeons often struggle with identifying who is at risk of having or developing pancreatic cancer. We sought to review our experience with the surgical management of cystic neoplasms of the pancreas to determine pre-operative clinical indicators of malignancy or premalignant (i.e. mucinous) lesions. METHODS Between 1996 and 2007, 114 consecutive patients with cystic neoplasms of the pancreas underwent a pancreatectomy. Invasive adenocarcinoma was identified in 35 whereas 79 had benign lesions. Mucinous lesions were considered premalignant and consisted of 29 intraductal papillary mucinous neoplasms (IPMN) and 17 mucinous cystic neoplasms (MCN). The remaining 33 benign lesions were serous microcystic adenomas. Descriptive statistics were calculated and multivariate logistic regression was performed. Receiver-operating characteristic (ROC) curves were constructed for continuous variables and the area under the curves compared. Likelihood ratios were calculated from the combinations of predictors. RESULTS Patients with pancreatic cancer arising from a cystic neoplasm were older than those with benign cysts. Mucinous lesions with or without associated cancer were more likely to be symptomatic and present with elevated serum carbohydrate antigen (CA)19-9 levels. Cancers more commonly presented in the head of the pancreas and were associated with longer hospitalizations after resection. Using multivariate logistic regression, size and elevated CA19-9 were predictors of malignancy whereas male gender and size were predictors of mucinous lesions with or without malignancy. Size, however, was not an accurate test to determine premalignant or malignant lesions using area under the ROC curve analysis whereas CA19-9 performed the best regardless of gender or lesion location. CONCLUSIONS Based upon our single institution experience with resection of cystic neoplasms of the pancreas, we advocate an aggressive surgical approach to any patient with a cystic neoplasm of the pancreas and associated elevated CA19-9.

Prognostic factors and staging for soft tissue sarcomas: an update.

Soft tissue sarcoma (STS) staging is a constantly evolving process. Grading is still of utmost importance and has been adapted into a three-tier system. The STS most difficult to categorize are those with uncertain malignant potential, such as solitary fibrous tumors, gastrointestinal stromal tumors, and glomus tumors, some of which have developed completely separate staging systems and may not even be considered sarcomas. Beyond the current TNM staging system, a multitude of prognostic factors for STS will continue to be discovered and ultimately incorporated into future revisions of the staging system.

NK Cell–Mediated Antitumor Effects of a Folate-Conjugated Immunoglobulin Are Enhanced by Cytokines

Many tumors express receptors for the vitamin folate (FRs). Folate-conjugated IgG, which can bind both FR+ tumors and NK-cell receptors for IgG, induced potent NK-cell antitumor responses that were further augmented by cytokine therapy. Optimally effective antitumor therapies would not only activate immune effector cells but also engage them at the tumor. Folate conjugated to immunoglobulin (F-IgG) could direct innate immune cells with Fc receptors to folate receptor–expressing cancer cells. F-IgG bound to human KB and HeLa cells, as well as murine L1210JF, a folate receptor (FR)–overexpressing cancer cell line, as determined by flow cytometry. Recognition of F-IgG by natural killer (NK) cell Fc receptors led to phosphorylation of the ERK transcription factor and increased NK cell expression of CD69. Lysis of KB tumor cells by NK cells increased by about 5-fold after treatment with F-IgG, an effect synergistically enhanced by treatment with IL2, IL12, IL15, or IL21 (P < 0.001). F-IgG also enhanced the lysis of chronic lymphocytic leukemia cells by autologous NK cells. NK cells significantly increased production of IFNγ, MIP-1α, and RANTES in response to F-IgG–coated KB target cells in the presence of the NK cell–activating cytokine IL12, and these coculture supernatants induced significant T-cell chemotaxis (P < 0.001). F-IgG-coated targets also stimulated FcR-mediated monocyte effector functions. Studies in a murine leukemia model confirmed the intratumoral localization and antitumor activity of F-IgG, as well as enhancement of its effects by IL12 (P = 0.05). The antitumor effect of this combination was dependent on NK cells and led to decreased tumor cell proliferation in vivo. Thus, F-IgG can induce an immune response against FR-positive tumor cells that is mediated by NK cells and can be augmented by cytokine therapy. Cancer Immunol Res; 4(4); 323–36. ©2016 AACR.

Cytopathologic Evaluation of the In Situ Margin in Patients Undergoing Hepatectomy

Margin status is a predictor of outcome for patients with liver malignancies, although what constitutes a negative margin is controversial. Traditionally, the completeness of resection is estimated by surgical histopathology of the resected specimen margin, despite the in situ margin being potentially more important. The true margin is often altered by parenchymal transection techniques. The authors propose that cytologic assessment of the in situ margin is more specific for determining the true margin.

Abstract 2686: Anti-tumor effects of a folate-immunoglobulin conjugate are enhanced by cytokine treatment in vitro and in vivo

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Folate conjugation is a means of selectively targeting therapeutics to folate receptor (FR)-expressing cancer cells. A novel folate-bound immunoglobulin (F-IgG) was tested for its ability to target natural killer (NK) cells to folate-receptor expressing cancer cells in the presence or absence of NK-activating cytokines. FR expression by the KB and HeLa cell lines was confirmed by immunoblot analysis (IB) and flow cytometry. Binding of F-IgG to NK cell Fc receptors led to increased phosphorylation of the epidermal growth factor related kinase (ERK) as measured by IB. Lysis of FR+ KB tumor cells by NK cells was increased 8-fold following treatment with F-IgG as compared to C-IgG (p<0.0001 across E:T ratios from 6.25:1 to 50:1). NK cell lysis of F-IgG- coated KB target cells was significantly enhanced following treatment of NK cells with IL-2, IL-12, IL-15 and IL-21 (all at 10ng/mL). NK cell production of IFN-γ, RANTES and MIP-1α was significantly enhanced by IL-12 in response to F-IgG-coated KB target cells as compared to control-treated cells (p<0.005; IFN-γ-2100 vs. 1000pg/mL, RANTES-800 vs. 150pg/mL, MIP-1α 1800 vs. 500pg/mL, respectively). Studies using the L1210JF murine leukemia model confirmed the anti-tumor activity of F-IgG and the ability of NK-activating cytokines to significantly enhance its effects. NK cell depletion in tumor-bearing mice demonstrated that the anti-tumor effects of IL-12 and F-IgG are dependent on NK cells. These studies indicate that F-IgG induces an immune response by NK cells against FR-positive cancer cell lines and that cytokine treatment has a synergistic effect on this response both in vitro and in vivo. Thus F-IgG has a potential to be used as a therapeutic antibody for the treatment of FR-positive cancers in combination with immune modulatory cytokines. 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 2686. doi:10.1158/1538-7445.AM2011-2686