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Neoadjuvant administration of Semliki Forest virus expressing interleukin-12 combined with attenuated Salmonella eradicates breast cancer metastasis and achieves long-term survival in immunocompetent mice.

Kramer MG, Masner M, Casales E, Moreno M, Smerdou C, Chabalgoity JA - BMC Cancer (2015)

Bottom Line: Moreover, pre-treatment with LVR01 seemed to suppress SFV-IL-12 antiangiogenic effects associated to lower IL-12 expression in this group.To our knowledge, these are the most encouraging results obtained to date in a post-operatory setting using the highly aggressive 4T1 animal model.SFV-IL-12-based gene therapy combined with Salmonella LVR01 neoadjuvant administration has a synergic antitumor effect and may be a promising therapeutic option to prevent and/or eradicate pre-operatory metastasis in locally advanced breast cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Instituto de Higiene, Facultad de Medicina, Universidad de la República, (UdelaR), Av. A. Navarro 3051, 11600, Montevideo, Uruguay. mgkramer@higiene.edu.uy.

ABSTRACT

Background: Metastatic breast cancer is a major cause of death among women worldwide; therefore efficient therapeutic strategies are extremely needed. In this work we have developed a gene therapy- and bacteria-based combined neoadjuvant approach and evaluated its antitumor effect in a clinically relevant animal model of metastatic breast cancer.

Methods: 2×10(8) particles of a Semliki Forest virus vector expressing interleukin-12 (SFV-IL-12) and/or 2×10(7) units of an aroC (-) Samonella Typhimurium strain (LVR01) were injected into 4T1 tumor nodules orthotopically implanted in mice. Tumors were surgically resected and long-term survival was determined. IL-12 and interferon-γ were quantified by Enzyme-Linked ImmunoSorbent Assay, bacteria was visualized by inmunohistochemistry and the number of lung metastasis was calculated with a clonogenic assay.

Results: SFV-IL-12 and LVR01 timely inoculated and followed by surgical resection of tumors succeeded in complete inhibition of lethal lung metastasis and long-term survival in 90% of treated mice. The combined therapy was markedly synergistic compared to each treatment alone, since SFV-IL-12 monotherapy showed a potent antiangiogenic effect, being able to inhibit tumor growth and extend survival, but could not prevent establishment of distant metastasis and death of tumor-excised animals. On the other hand, LVR01 alone also showed a significant, although limited, antitumor potential, despite its ability to invade breast cancer cells and induce granulocyte recruitment. The efficacy of the combined therapy depended on the order in which both factors were administered; inasmuch the therapeutic effect was only observed when SFV-IL-12 was administered previous to LVR01, whereas administration of LVR01 before SFV-IL-12 had negligible antitumor activity. Moreover, pre-treatment with LVR01 seemed to suppress SFV-IL-12 antiangiogenic effects associated to lower IL-12 expression in this group. Re-challenged mice were unable to reject a second 4T1 tumor; however 100% of them could be totally cured by applying the same neoadjuvant combined regimen. To our knowledge, these are the most encouraging results obtained to date in a post-operatory setting using the highly aggressive 4T1 animal model.

Conclusions: SFV-IL-12-based gene therapy combined with Salmonella LVR01 neoadjuvant administration has a synergic antitumor effect and may be a promising therapeutic option to prevent and/or eradicate pre-operatory metastasis in locally advanced breast cancer.

No MeSH data available.


Related in: MedlinePlus

Neoadjuvant effect of SFV-IL-12. a Schematic representation of the therapeutic protocol performed in this study. Two doses of SFV-LacZ, SFV-IL-12 or PBS were i.t. injected as in Fig. 2. Three days later (i.e. day 16 after 4T1 cells inoculation), treated tumors were surgically removed (S-16). b Kaplan–Meier plot shows the survival rate of treated and control mice (n = 5-10). Non-significant (n.s), p< 0.001 (***)
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Fig3: Neoadjuvant effect of SFV-IL-12. a Schematic representation of the therapeutic protocol performed in this study. Two doses of SFV-LacZ, SFV-IL-12 or PBS were i.t. injected as in Fig. 2. Three days later (i.e. day 16 after 4T1 cells inoculation), treated tumors were surgically removed (S-16). b Kaplan–Meier plot shows the survival rate of treated and control mice (n = 5-10). Non-significant (n.s), p< 0.001 (***)

Mentions: In order to mimic frequent management of patients with LABC, we have implemented a therapeutic protocol that included surgical removal of primary tumors after a preliminary treatment. Unlike conventional neadjuvant systemic approaches, here we performed local administration of the therapeutic agent (Fig. 3a). Two doses of SFV-IL-12 were injected inside the tumor mass at days 10 and 13 after 4T1 cells inoculation. Tumors were surgically removed 3 days later (day 16) and survival outcome was compared to SFV-LacZ or PBS treated mice. As shown in Fig. 3b, the overall survival of SFV-IL-12 treated animals was significantly higher than that of controls. In addition, 20 % of long-term survivors were observed in the IL-12-based neoadjuvant post-operatory setting. All of the non-survivor animals experienced breathing difficulties before death and showed numerous macroscopic lung metastases as was previously observed in untreated mice (see Fig. 1c). In contrast, mice that survived at the end of this study did not present any visible metastasis. However, the fact that long-term survival was only achieved in a small percentage of treated mice indicated that our strategy to treat this type of cancer needed to be improved. Based on several studies that report the benefit obtained when combining IL-12-based gene therapy with complementary strategies or drugs [36–40], we next decided to combine SFV-IL-12 with another biological agent that could collaborate in inducing a more efficient response against the metastatic disease.Fig. 3


Neoadjuvant administration of Semliki Forest virus expressing interleukin-12 combined with attenuated Salmonella eradicates breast cancer metastasis and achieves long-term survival in immunocompetent mice.

Kramer MG, Masner M, Casales E, Moreno M, Smerdou C, Chabalgoity JA - BMC Cancer (2015)

Neoadjuvant effect of SFV-IL-12. a Schematic representation of the therapeutic protocol performed in this study. Two doses of SFV-LacZ, SFV-IL-12 or PBS were i.t. injected as in Fig. 2. Three days later (i.e. day 16 after 4T1 cells inoculation), treated tumors were surgically removed (S-16). b Kaplan–Meier plot shows the survival rate of treated and control mice (n = 5-10). Non-significant (n.s), p< 0.001 (***)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4562361&req=5

Fig3: Neoadjuvant effect of SFV-IL-12. a Schematic representation of the therapeutic protocol performed in this study. Two doses of SFV-LacZ, SFV-IL-12 or PBS were i.t. injected as in Fig. 2. Three days later (i.e. day 16 after 4T1 cells inoculation), treated tumors were surgically removed (S-16). b Kaplan–Meier plot shows the survival rate of treated and control mice (n = 5-10). Non-significant (n.s), p< 0.001 (***)
Mentions: In order to mimic frequent management of patients with LABC, we have implemented a therapeutic protocol that included surgical removal of primary tumors after a preliminary treatment. Unlike conventional neadjuvant systemic approaches, here we performed local administration of the therapeutic agent (Fig. 3a). Two doses of SFV-IL-12 were injected inside the tumor mass at days 10 and 13 after 4T1 cells inoculation. Tumors were surgically removed 3 days later (day 16) and survival outcome was compared to SFV-LacZ or PBS treated mice. As shown in Fig. 3b, the overall survival of SFV-IL-12 treated animals was significantly higher than that of controls. In addition, 20 % of long-term survivors were observed in the IL-12-based neoadjuvant post-operatory setting. All of the non-survivor animals experienced breathing difficulties before death and showed numerous macroscopic lung metastases as was previously observed in untreated mice (see Fig. 1c). In contrast, mice that survived at the end of this study did not present any visible metastasis. However, the fact that long-term survival was only achieved in a small percentage of treated mice indicated that our strategy to treat this type of cancer needed to be improved. Based on several studies that report the benefit obtained when combining IL-12-based gene therapy with complementary strategies or drugs [36–40], we next decided to combine SFV-IL-12 with another biological agent that could collaborate in inducing a more efficient response against the metastatic disease.Fig. 3

Bottom Line: Moreover, pre-treatment with LVR01 seemed to suppress SFV-IL-12 antiangiogenic effects associated to lower IL-12 expression in this group.To our knowledge, these are the most encouraging results obtained to date in a post-operatory setting using the highly aggressive 4T1 animal model.SFV-IL-12-based gene therapy combined with Salmonella LVR01 neoadjuvant administration has a synergic antitumor effect and may be a promising therapeutic option to prevent and/or eradicate pre-operatory metastasis in locally advanced breast cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology, Instituto de Higiene, Facultad de Medicina, Universidad de la República, (UdelaR), Av. A. Navarro 3051, 11600, Montevideo, Uruguay. mgkramer@higiene.edu.uy.

ABSTRACT

Background: Metastatic breast cancer is a major cause of death among women worldwide; therefore efficient therapeutic strategies are extremely needed. In this work we have developed a gene therapy- and bacteria-based combined neoadjuvant approach and evaluated its antitumor effect in a clinically relevant animal model of metastatic breast cancer.

Methods: 2×10(8) particles of a Semliki Forest virus vector expressing interleukin-12 (SFV-IL-12) and/or 2×10(7) units of an aroC (-) Samonella Typhimurium strain (LVR01) were injected into 4T1 tumor nodules orthotopically implanted in mice. Tumors were surgically resected and long-term survival was determined. IL-12 and interferon-γ were quantified by Enzyme-Linked ImmunoSorbent Assay, bacteria was visualized by inmunohistochemistry and the number of lung metastasis was calculated with a clonogenic assay.

Results: SFV-IL-12 and LVR01 timely inoculated and followed by surgical resection of tumors succeeded in complete inhibition of lethal lung metastasis and long-term survival in 90% of treated mice. The combined therapy was markedly synergistic compared to each treatment alone, since SFV-IL-12 monotherapy showed a potent antiangiogenic effect, being able to inhibit tumor growth and extend survival, but could not prevent establishment of distant metastasis and death of tumor-excised animals. On the other hand, LVR01 alone also showed a significant, although limited, antitumor potential, despite its ability to invade breast cancer cells and induce granulocyte recruitment. The efficacy of the combined therapy depended on the order in which both factors were administered; inasmuch the therapeutic effect was only observed when SFV-IL-12 was administered previous to LVR01, whereas administration of LVR01 before SFV-IL-12 had negligible antitumor activity. Moreover, pre-treatment with LVR01 seemed to suppress SFV-IL-12 antiangiogenic effects associated to lower IL-12 expression in this group. Re-challenged mice were unable to reject a second 4T1 tumor; however 100% of them could be totally cured by applying the same neoadjuvant combined regimen. To our knowledge, these are the most encouraging results obtained to date in a post-operatory setting using the highly aggressive 4T1 animal model.

Conclusions: SFV-IL-12-based gene therapy combined with Salmonella LVR01 neoadjuvant administration has a synergic antitumor effect and may be a promising therapeutic option to prevent and/or eradicate pre-operatory metastasis in locally advanced breast cancer.

No MeSH data available.


Related in: MedlinePlus