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In situ vaccination at a peripheral tumor site augments response against melanoma brain metastases.

ABSTRACT: BACKGROUND:Immune checkpoint inhibition (ICI) alone is not efficacious for a large number of patients with melanoma brain metastases. We previously established an in situ vaccination (ISV) regimen combining radiation and immunocytokine to enhance response to ICIs. Here, we tested whether ISV inhibits the development of brain metastases in a murine melanoma model. METHODS:B78 (GD2+) melanoma 'primary' tumors were engrafted on the right flank of C57BL/6 mice. After 3-4 weeks, primary tumors were treated with ISV (radiation (12?Gy, day 1), ?-GD2 immunocytokine (hu14.18-IL2, days 6-10)) and ICI (?-CTLA-4, days 3, 6, 9). Complete response (CR) was defined as no residual tumor observed at treatment day 90. Mice with CR were tested for immune memory by re-engraftment with B78 in the left flank and then the brain. To test ISV efficacy against metastases, tumors were also engrafted in the left flank and brain of previously untreated mice. Tumors were analyzed by quantitative reverse transcription-PCR, immunohistochemistry, flow cytometry and multiplex cytokine assay. RESULTS:ISV+?-CTLA-4 resulted in immune memory and rejection of B78 engraftment in the brain in 11 of 12 mice. When B78 was engrafted in brain prior to treatment, ISV+?-CTLA-4 increased survival compared with ICI alone. ISV+?-CTLA-4 eradicated left flank tumors but did not elicit CR at brain sites when tumor cells were engrafted in brain prior to ISV. ISV+?-CTLA-4 increased CD8+ and CD4+ T cells in flank and brain tumors compared with untreated mice. Among ISV + ?-CTLA-4 treated mice, left flank tumors showed increased CD8+ infiltration and CD8+:FOXP3+ ratio compared with brain tumors. Flank and brain tumors showed minimal differences in expression of immune checkpoint receptors/ligands or Mhc-1. Cytokine productions were similar in left flank and brain tumors in untreated mice. Following ISV+?-CTLA-4, production of immune-stimulatory cytokines was greater in left flank compared with brain tumor grafts. CONCLUSION:ISV augmented response to ICIs in murine melanoma at brain and extracranial tumor sites. Although baseline tumor-immune microenvironments were similar at brain and extracranial tumor sites, response to ISV+?-CTLA-4 was divergent with reduced infiltration and activation of immune cells in brain tumors. Additional therapies may be needed for effective antitumor immune response against melanoma brain metastases.

SUBMITTER: Clark PA

PROVIDER: S-EPMC7371368 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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In situ vaccination at a peripheral tumor site augments response against melanoma brain metastases.

Clark Paul A PA Sriramaneni Raghava N RN Jin Won Jong WJ Jagodinsky Justin C JC Bates Amber M AM Jaquish Abigail A AA Anderson Bryce R BR Le Trang T Lubin Jonathan A JA Chakravarty Ishan I Arthur Ian S IS Heinze Clinton M CM Guy Emily I EI Kler Jasdeep J Klar Kelsey A KA Carlson Peter M PM Kim Kyung Mann KM Kuo John S JS Morris Zachary S ZS

Journal for immunotherapy of cancer 20200701 2

<h4>Background</h4>Immune checkpoint inhibition (ICI) alone is not efficacious for a large number of patients with melanoma brain metastases. We previously established an in situ vaccination (ISV) regimen combining radiation and immunocytokine to enhance response to ICIs. Here, we tested whether ISV inhibits the development of brain metastases in a murine melanoma model.<h4>Methods</h4>B78 (GD2<sup>+</sup>) melanoma 'primary' tumors were engrafted on the right flank of C57BL/6 mice. After 3-4 we ...[more]

PMID: 32690669

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Project description:Brain metastases develop in over 60% of advanced melanoma patients and negatively impact quality of life and prognosis. In a murine melanoma model, we previously showed that an in situ vaccination (ISV) regimen, combining radiation treatment and intratumoral (IT) injection of immunocytokine (IC: anti-GD2 antibody fused to IL2), along with the immune checkpoint inhibitor anti-CTLA-4, robustly eliminates peripheral flank tumors but only has modest effects on co-occurring intracranial tumors. In this study, we investigated the ability of low-dose radiation to the brain to potentiate anti-tumor immunity against a brain tumor when combined with ISV + anti-CTLA-4. B78 (GD2+, immunologically "cold") melanoma tumor cells were implanted into the flank and the right striatum of the brain in C57BL/6 mice. Flank tumors (50-150 mm3) were treated following a previously optimized ISV regimen [radiation (12 Gy × 1, treatment day 1), IT-IC (50 µg daily, treatment days 6-10), and anti-CTLA-4 (100 µg, treatment days 3, 6, 9)]. Mice that additionally received whole-brain radiation treatment (WBRT, 4 Gy × 1) on day 15 demonstrated significantly increased survival compared to animals that received ISV + anti-CTLA-4 alone, WBRT alone or no treatment (control) (P < 0.001, log-rank test). Timing of WBRT was critical, as WBRT administration on day 1 did not significantly enhance survival compared to ISV + anti-CTLA-4, suggesting that the effect of WBRT on survival might be mediated through immune modulation and not just direct tumor cell cytotoxicity. Modest increases in T cells (CD8+ and CD4+) and monocytes/macrophages (F4/80+) but no changes in FOXP3+ regulatory T cells (Tregs), were observed in brain melanoma tumors with addition of WBRT (on day 15) to ISV + anti-CTLA-4. Cytokine multiplex immunoassay revealed distinct changes in both intracranial melanoma and contralateral normal brain with addition of WBRT (day 15) to ISV + anti-CTLA-4, with notable significant changes in pro-inflammatory (e.g., IFNγ, TNFα and LIX/CXCL5) and suppressive (e.g., IL10, IL13) cytokines as well as chemokines (e.g., IP-10/CXCL10 and MIG/CXCL9). We tested the ability of the alkylphosphocholine analog, NM600, to deliver immunomodulatory radiation to melanoma brain tumors as a targeted radionuclide therapy (TRT). Yttrium-86 (86Y) chelated to NM600 was delivered intravenously by tail vein to mice harboring flank and brain melanoma tumors, and PET imaging demonstrated specific accumulation up to 72 h at each tumor site (∼12:1 brain tumor/brain and ∼8:1 flank tumor/muscle). When NM600 was chelated to therapeutic β-particle-emitting 90Y and administered on treatment day 13, T-cell infiltration and cytokine profiles were altered in melanoma brain tumor, like that observed for WBRT. Overall, our results demonstrate that addition of low-dose radiation, timed appropriately with ISV administration to tumors outside the brain, significantly increases survival in animals co-harboring melanoma brain tumors. This observation has potentially important translational implications as a treatment strategy for increasing the response of tumors in the brain to systemically administered immunotherapies.

Project description:BackgroundHigh tumor-infiltrating lymphocytes (TILs) and hemorrhage are important prognostic factors in patients who have undergone craniotomy for melanoma brain metastases (MBM) before 2011 at the University of Pittsburgh Medical Center (UPMC). We have investigated the prognostic or predictive role of these histopathologic factors in a more contemporary craniotomy cohort from the University of North Carolina at Chapel Hill (UNC-CH). We have also sought to understand better how various immune cell subsets, angiogenic factors, and blood vessels may be associated with clinical and radiographic features in MBM.MethodsBrain tumors from the UPMC and UNC-CH patient cohorts were (re)analyzed by standard histopathology, tumor tissue imaging, and gene expression profiling. Variables were associated with overall survival (OS) and radiographic features.ResultsThe patient subgroup with high TILs in craniotomy specimens and subsequent treatment with immune checkpoint inhibitors (ICIs, n=7) trended to have longer OS compared to the subgroup with high TILs and no treatment with ICIs (n=11, p=0.059). Bleeding was significantly associated with tumor volume before craniotomy, high melanoma-specific expression of basic fibroblast growth factor (bFGF), and high density of CD31+αSMA- blood vessels. Brain tumors with high versus low peritumoral edema before craniotomy had low (17%) versus high (41%) incidence of brisk TILs. Melanoma-specific expression of the vascular endothelial growth factor (VEGF) was comparable to VEGF expression by TILs and was not associated with any particular prognostic, radiographic, or histopathologic features. A gene signature associated with gamma delta (gd) T cells was significantly higher in intracranial than same-patient extracranial metastases and primary melanoma. However, gdT cell density in MBM was not prognostic.ConclusionsICIs may provide greater clinical benefit in patients with brisk TILs in MBM. Intratumoral hemorrhage in brain metastases, a significant clinical problem, is not merely associated with tumor volume but also with underlying biology. bFGF may be an essential pathway to target. VEGF, a factor principally associated with peritumoral edema, is not only produced by melanoma cells but also by TILs. Therefore, suppressing low-grade peritumoral edema using corticosteroids may harm TIL function in 41% of cases. Ongoing clinical trials targeting VEGF in MBM may predict a lack of unfavorable impacts on TIL density and/or intratumoral hemorrhage.

Project description:Cancer immunotherapy approaches have emerged as novel treatment regimens against cancer. A particularly interesting avenue is the concept of in situ vaccination, where immunostimulatory agents are introduced into an identified tumor to overcome local immunosuppression and, if successful, mount systemic antitumor immunity. We had previously shown that nanoparticles from cowpea mosaic virus (CPMV) are highly potent in inducing long-lasting antitumor immunity when used as an in situ vaccine in various tumor mouse models. Here we asked whether the nanoparticles from tobacco mosaic virus (TMV) could also be applied as an in situ vaccine and, if so, whether efficacy or mechanism of immune-activation would be affected by the nanoparticle size (300 × 18 nm native TMV vs 50 × 18 nm short TMV nanorods), shape (nanorods vs spherical TMV, termed SNP), or state of assembly (assembled TMV rod vs free coat protein, CP). Our studies indicate that CPMV, but less so TMV, elicits potent antitumor immunity after intratumoral treatment of dermal melanoma (B16F10 using C57BL/6 mice). TMV and TMVshort slowed tumor growth and increased survival time, however, at significantly lower potency compared to that of CPMV. There were no apparent differences between TMV, TMVshort, or the SNP indicating that the aspect ratio does not necessarily play a role in plant viral in situ vaccines. The free CPs did not elicit an antitumor response or immunostimulation, which may indicate that a multivalent assembly is required to trigger an innate immune recognition and activation. Differential potency of CPMV vs TMV can be explained with differences in immune-activation: data indicate that CPMV stimulates an antitumor response through recruitment of monocytes into the tumor microenvironment (TME), establishing signaling through the IFN-? pathway, which also leads to recruitment of tumor-infiltrated neutrophils (TINs) and natural killer (NK) cells. Furthermore, the priming of the innate immune system also mounts an adaptive response with CD4+ and CD8+ T cell recruitment and establishment of effector memory cells. While the TMV treatment also lead to the recruitment of innate immune cells as well as T cells (although to a lesser degree), key differences were noted in cyto/chemokine profiling with TMV inducing a potent immune response early on characterized by strong pro-inflammatory cytokines, primarily IL-6. Together, data indicate that some plant viral nanotechnology platforms are more suitable for application as in situ vaccines than others; understanding the intricate differences and underlying mechanism of immune-activation may set the stage for clinical development of these technologies.

Dataset Information

In situ vaccination at a peripheral tumor site augments response against melanoma brain metastases.

Publications

In situ vaccination at a peripheral tumor site augments response against melanoma brain metastases.

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