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Effects of pegylated G-CSF on immune cell number and function in patients with gynecological malignancies.

Bonanno G, Procoli A, Mariotti A, Corallo M, Perillo A, Danese S, De Cristofaro R, Scambia G, Rutella S - J Transl Med (2010)

Bottom Line: In contrast, CD4+FoxP3+ bona fide Treg cells were unchanged compared with baseline.Interestingly, pegfilgrastim fostered in vitro monocytic secretion of IL-12p40 and IL-12p70 when compared with unconjugated G-CSF.Pegfilgrastim induced significant changes in immune cell number and function.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Hematology, Catholic University Med, School, Rome, Italy.

ABSTRACT

Background: Pegylated granulocyte colony-stimulating factor (G-CSF; pegfilgrastim) is a longer-acting form of G-CSF, whose effects on dendritic cell (DC) and regulatory T cell (Treg) mobilization, and on the in vivo and ex vivo release of immune modulating cytokines remain unexplored.

Methods: Twelve patients with gynecological cancers received carboplatin/paclitaxel chemotherapy and single-dose pegfilgrastim as prophylaxis of febrile neutropenia. Peripheral blood was collected prior to pegfilgrastim administration (day 0) and on days +7, +11 and +21, to quantify immunoregulatory cytokines and to assess type 1 DC (DC1), type 2 DC (DC2) and Treg cell mobilization. In vitro-differentiated, monocyte-derived DC were used to investigate endocytic activity, expression of DC maturation antigens and ability to activate allogeneic T-cell proliferation.

Results: Pegfilgrastim increased the frequency of circulating DC1 and DC2 precursors. In contrast, CD4+FoxP3+ bona fide Treg cells were unchanged compared with baseline. Serum levels of hepatocyte growth factor and interleukin (IL)-12p40, but not transforming growth factor-β1 or immune suppressive kynurenines, significantly increased after pegfilgrastim administration. Interestingly, pegfilgrastim fostered in vitro monocytic secretion of IL-12p40 and IL-12p70 when compared with unconjugated G-CSF. Finally, DC populations differentiated in vitro after clinical provision of pegfilgrastim were phenotypically mature, possessed low endocytic activity, and incited a robust T-cell proliferative response.

Conclusions: Pegfilgrastim induced significant changes in immune cell number and function. The enhancement of monocytic IL-12 secretion portends favorable implications for pegfilgrastim administration to patients with cancer, a clinical context where the induction of immune deviation would be highly undesirable.

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In vitro monocytic release of bioactive IL-12p70. Monocytes (1 × 106) purified from the peripheral blood of patients given pegfilgrastim (n = 3) or filgrastim (n = 3) were stimulated with LPS as detailed in the legend to Figure 3B. Supernatants were harvested daily and used to measure IL-12p70 by ELISA. Each point is representative of the mean value of triplicate IL-12p70 measurements.
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Figure 4: In vitro monocytic release of bioactive IL-12p70. Monocytes (1 × 106) purified from the peripheral blood of patients given pegfilgrastim (n = 3) or filgrastim (n = 3) were stimulated with LPS as detailed in the legend to Figure 3B. Supernatants were harvested daily and used to measure IL-12p70 by ELISA. Each point is representative of the mean value of triplicate IL-12p70 measurements.

Mentions: In order to more accurately substantiate the assumption that pegfilgrastim alters the balance between IL-12 and IL-10, monocytes, a prominent cellular source of both IL-12 and IL-10, were magnetically purified on day +11 from the peripheral blood of patients treated with pegfilgrastim (24 hours before the anticipated decline of serum pegfilgrastim concentration [16] and coincident with maximal monocyte mobilization) and from cancer patients treated with daily filgrastim (24 hours after the last G-CSF administration). Monocytes were routinely > 95% pure, as evaluated by flow cytometry measurements of CD14 expression (data not shown). Equal numbers of monocytes from pre-G-CSF and post-G-CSF samples were cultured for up to 96 hours in the presence of LPS as a stimulus. The LPS-induced monocytic release of IL-10 increased after pegfilgrastim administration (Figure 3B). Notably, post-pegfilgrastim monocytes secreted considerable amounts of IL-12p40 at any time-point in culture (Figure 3B). In line with previous reports [25], monocytes from filgrastim-treated patients secreted low amounts of IL-12p40. Intriguingly, IL-12p40 production by post-filgrastim monocytes was significantly lower than that measured in post-pegfilgrastim monocyte cultures at any time-point. To further reinforce the assumption that pegfilgrastim, but not unconjugated G-CSF, enhances the monocytic release of IL-12 on a per cell basis, IL-12p70 levels were measured in supernatants of monocytes purified from 3 patients given pegfilgrastim and 3 patients receiving unconjugated G-CSF. As shown in Figure 4, post-pegfilgrastim monocytes released significantly higher levels of IL-12p70 compared with monocytes isolated from cancer patients treated with unconjugated G-CSF.


Effects of pegylated G-CSF on immune cell number and function in patients with gynecological malignancies.

Bonanno G, Procoli A, Mariotti A, Corallo M, Perillo A, Danese S, De Cristofaro R, Scambia G, Rutella S - J Transl Med (2010)

In vitro monocytic release of bioactive IL-12p70. Monocytes (1 × 106) purified from the peripheral blood of patients given pegfilgrastim (n = 3) or filgrastim (n = 3) were stimulated with LPS as detailed in the legend to Figure 3B. Supernatants were harvested daily and used to measure IL-12p70 by ELISA. Each point is representative of the mean value of triplicate IL-12p70 measurements.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2992497&req=5

Figure 4: In vitro monocytic release of bioactive IL-12p70. Monocytes (1 × 106) purified from the peripheral blood of patients given pegfilgrastim (n = 3) or filgrastim (n = 3) were stimulated with LPS as detailed in the legend to Figure 3B. Supernatants were harvested daily and used to measure IL-12p70 by ELISA. Each point is representative of the mean value of triplicate IL-12p70 measurements.
Mentions: In order to more accurately substantiate the assumption that pegfilgrastim alters the balance between IL-12 and IL-10, monocytes, a prominent cellular source of both IL-12 and IL-10, were magnetically purified on day +11 from the peripheral blood of patients treated with pegfilgrastim (24 hours before the anticipated decline of serum pegfilgrastim concentration [16] and coincident with maximal monocyte mobilization) and from cancer patients treated with daily filgrastim (24 hours after the last G-CSF administration). Monocytes were routinely > 95% pure, as evaluated by flow cytometry measurements of CD14 expression (data not shown). Equal numbers of monocytes from pre-G-CSF and post-G-CSF samples were cultured for up to 96 hours in the presence of LPS as a stimulus. The LPS-induced monocytic release of IL-10 increased after pegfilgrastim administration (Figure 3B). Notably, post-pegfilgrastim monocytes secreted considerable amounts of IL-12p40 at any time-point in culture (Figure 3B). In line with previous reports [25], monocytes from filgrastim-treated patients secreted low amounts of IL-12p40. Intriguingly, IL-12p40 production by post-filgrastim monocytes was significantly lower than that measured in post-pegfilgrastim monocyte cultures at any time-point. To further reinforce the assumption that pegfilgrastim, but not unconjugated G-CSF, enhances the monocytic release of IL-12 on a per cell basis, IL-12p70 levels were measured in supernatants of monocytes purified from 3 patients given pegfilgrastim and 3 patients receiving unconjugated G-CSF. As shown in Figure 4, post-pegfilgrastim monocytes released significantly higher levels of IL-12p70 compared with monocytes isolated from cancer patients treated with unconjugated G-CSF.

Bottom Line: In contrast, CD4+FoxP3+ bona fide Treg cells were unchanged compared with baseline.Interestingly, pegfilgrastim fostered in vitro monocytic secretion of IL-12p40 and IL-12p70 when compared with unconjugated G-CSF.Pegfilgrastim induced significant changes in immune cell number and function.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Hematology, Catholic University Med, School, Rome, Italy.

ABSTRACT

Background: Pegylated granulocyte colony-stimulating factor (G-CSF; pegfilgrastim) is a longer-acting form of G-CSF, whose effects on dendritic cell (DC) and regulatory T cell (Treg) mobilization, and on the in vivo and ex vivo release of immune modulating cytokines remain unexplored.

Methods: Twelve patients with gynecological cancers received carboplatin/paclitaxel chemotherapy and single-dose pegfilgrastim as prophylaxis of febrile neutropenia. Peripheral blood was collected prior to pegfilgrastim administration (day 0) and on days +7, +11 and +21, to quantify immunoregulatory cytokines and to assess type 1 DC (DC1), type 2 DC (DC2) and Treg cell mobilization. In vitro-differentiated, monocyte-derived DC were used to investigate endocytic activity, expression of DC maturation antigens and ability to activate allogeneic T-cell proliferation.

Results: Pegfilgrastim increased the frequency of circulating DC1 and DC2 precursors. In contrast, CD4+FoxP3+ bona fide Treg cells were unchanged compared with baseline. Serum levels of hepatocyte growth factor and interleukin (IL)-12p40, but not transforming growth factor-β1 or immune suppressive kynurenines, significantly increased after pegfilgrastim administration. Interestingly, pegfilgrastim fostered in vitro monocytic secretion of IL-12p40 and IL-12p70 when compared with unconjugated G-CSF. Finally, DC populations differentiated in vitro after clinical provision of pegfilgrastim were phenotypically mature, possessed low endocytic activity, and incited a robust T-cell proliferative response.

Conclusions: Pegfilgrastim induced significant changes in immune cell number and function. The enhancement of monocytic IL-12 secretion portends favorable implications for pegfilgrastim administration to patients with cancer, a clinical context where the induction of immune deviation would be highly undesirable.

Show MeSH
Related in: MedlinePlus