Immune checkpoint blockade (ICB) has demonstrated improvements in patients with soft tissue sarcoma (STS), however, response is limited to patients with pre-existing tumor-infiltrating T cells. Most STS harbor a myeloid-dominant infiltrate highlighting a therapeutic opportunity to leverage their ability to engage T cells. We hypothesized that BO-112, a nanoplexed poly I:C, will target myeloid cells to complement radiotherapy (RT) and ICB to generate T cell immunity in STS murine models and patients.
Methods
We tested combination hypofractionated RT ± intratumoral BO-112 ± anti-PD-1 in ICB-resistant murine models (autochthonous and transplantable KrasG12DP53-/- (KP) sarcoma). Tumor growth and survival were tracked. Target cells of BO-112 were identified using rhodamine-labeled BO-112. KP-bearing YFP-Cre mice were treated with intratumoral Cre-recombinase to track leukocytes. Tumor-infiltrating and peripheral immune cells were assessed longitudinally by flow cytometry. The contribution of lymphocyte subsets to anti-tumor effect was evaluated by antibody-mediated deletion. In a Phase I neoadjuvant trial, patients with STS received intratumoral BO-112, hypofractionated RT ± nivolumab followed by surgery. Longitudinal tumor (n=3–4 timepoints) and blood were evaluated by flow cytometry, IHC, TCR and whole exome DNA sequencing, and single cell and bulk RNA sequencing.
Results
Tumor control and survival were significantly improved with the local-only combination of RT and BO-112 in both KP models compared to untreated. Rhodamine-labeled BO-112 preferentially targeted myeloid cells, while RT depleted intratumoral lymphocytes and spared myeloid cells. Despite myeloid targeting and initial lymphocyte depletion, RT and BO-112 heightened IFNg production by CD8+ T cells in the tumor and spleen and resulted in CD8-dependent tumor control, which was augmented by anti-PD1. In 14 patients with high-risk STS, RT and BO-112 (± nivolumab) increased effector T cell infiltration and myeloid MHC II expression at surgery following neoadjuvant therapy. Longitudinal clinical specimens re-demonstrated initial depletion of intratumoral T cells following RT with subsequent clonal TCR replacement, with a shift toward effector phenotypes and a global interferon transcriptomic signature. Despite clonal replacement, highest frequency clones at baseline persisted, suggesting antigen-driven reaccumulation after RT-induced depletion.
Conclusions
Targeting myeloid cells with BO-112 and RT yields a lymphocyte-dependent, local and systemic anti-tumor immune response in ICB-resistant murine models of STS, which is augmented by anti-PD1 therapy. Similar remodeling of tumor immunity was observed in patients with STS leading to clonal TCR replacement and increased infiltration of effector T cells. Targeting myeloid cells with BO-112 may be a complementary strategy to promoting T cell responses in STS.
Trial Registration
NCT02828098.
Ethics Approval
Ethics approval was obtained for the Phase I clinical trial (NCT02828098 approved the UCLA Institutional Review Board #19-000419) and animal research (ARC-2017-088 approved by the UCLA Institutional Animal Care and Use Committee/Animal Research Committee (ARC)) Clinical trial participants gave informed consent before enrolling and participating in the trial.
A 43-year-old man with a growing mass in the right groin, concerned for liposarcoma, underwent MRI and 68 Ga-fibroblast activation protein inhibitor (FAPI)-46 PET/CT before surgery. Fibroblast activation protein inhibitor PET/CT demonstrated increased uptake (SUV max , 3.2) predominantly in the solid portion, where MRI showed gadolinium enhancement. The patient subsequently underwent surgery and was diagnosed with hibernoma. The immunohistochemistry of the tumor revealed the fibroblast activation protein expression in the fibrovascular network and myofibroblastic cells of the tumor. This case suggests that the FAPI uptake can be affected by the vascular cells, and thus, a careful interpretation of the FAPI PET signal may be needed.
<p>PDF file - 91K, Supplemental Figure 5: Knockdown of dCK expression does not change proliferation or alter glucose, lactate, and glutamine metabolism in liposarcoma cell lines.</p>
Abstract Background: Liposarcoma (LPS) is an understudied form of soft tissue sarcoma (STS). The well-differentiated (WD) and de-differentiated (DD) subtypes of LPS are associated with indolent and aggressive disease courses, respectively, but this histologic stratification fails to fully capture disease heterogeneity. Molecular approaches may help refine prognostication and inform treatment intensification. Insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3 or IMP3), is an RNA-binding protein that regulates gene expression by controlling mRNA stability and has been implicated in tumorigenesis and poor prognosis in many cancers. We hypothesized IGF2BP3 would refine the prognostication of LPS beyond its WD/DD histologic status. Methods: We examined the association between IGF2BP3 gene or protein expression and clinical data in four datasets: (1) patients with STS subtypes (n=206) in the cancer genome atlas (TCGA) database, (2) an in-house gene microarray of lipomatous tumors (n=71), LPS cell lines (n=3) and patient-derived xenografts (PDX, n=3), (3) an in-house tissue microarray (TMA) of lipomatous tumors (n=115), LPS cell lines (n=3) and PDXs (n=3) and (4) an in-house TMA of WD/DD LPS (n=71). IGF2BP3 protein expression in TMAs was quantified by immunohistochemistry (IHC). IGF2BP3 gene and protein expression values from identical samples were compared by Pearson correlation (n=43). The Kaplan-Meier method and log-rank test were used to compare survival outcomes. Results: In the TCGA cohort, which does not include WD LPS, IGF2BP3 expression was a poor prognostic factor solely in DD LPS (n=50, median overall survival (mOS): 1.6 vs 5.0 years, p=0.009). Among gene microarray samples, IGF2BP3 expression was highest in DD LPS (n=18) compared to WD LPS (n=29) and lipoma (n=7) by paired t-tests (p=0.03 and 0.002, respectively) and IGF2BP3 expression was associated with worse survival in WD/DD LPS (mOS 7.7 vs 21.5 years, p=0.02). In both TMAs, IGF2BP3 expression (>25% cell positivity/core) portended worse survival in WD/DD LPS (mOS (3): 3.7 vs 13.8 years, p<0.001 and mOS (4): 2.7 vs 14.9 years, p<0.001). IGF2BP3 was not expressed in myxoid LPS (n=21) or lipoma (n=8) samples. Gene and protein expression of IGF2BP3 were positively correlated in WD/DD LPS (r2 = 0.69). IGF2BP3 expression was more strongly associated with survival than LPS differentiation status (mOS: 7.0 (DD) vs 15.2 years (WD), p=0.02). Furthermore, all LPS cell lines and PDXs demonstrated high gene and protein expression of IGF2BP3. Conclusion: IGF2BP3 is highly expressed in a subset of LPS. Across independent datasets, IGF2BP3 is also a biomarker of disease progression and worse survival, and may stratify patients more effectively than histologic differentiation. Mechanistic studies of IGF2BP3 in LPS tumorigenesis and progression using aforementioned LPS cell lines and PDX models are ongoing. Citation Format: Kyle D. Klingbeil, Jack Pengfei Tang, Sarah M. Dry, Fritz C. Eilber, Dinesh S. Rao, Brian E. Kadera, Anusha Kalbasi. IGF2BP3 (IMP3) expression is associated with worse survival in well-differentiated/dedifferentiated (WD/DD) liposarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3490.
