Page 46 - WNS 2022 E-Program Booklet

Reprogramming the glioblastoma immune microenvironment with con-
vection enhanced gene therapy reveals intratumor IL6 drives glioblasto-
ma immunosuppression and growth
Jacob S. Young, MD, Resident, Department of Neurological Surgery
University of California, San Francisco
INTRODUCTION: The glioblastoma microenvironment is an immunosuppres-
sive barrier to therapeutic innovation. Here, we utilize intratumor convection
enhanced delivery (CED) of gene therapy vectors to reprogram the glioblasto-
ma immune microenvironment and elucidate therapeutic vulnerabilities.
METHODS: SB28 or GL261 glioblastoma allografts were implanted into immu-
nocompetent mice and treated with CED of attenuated adeno-associated virus
9 vectors (AAV9) encoding experimental cytokines (Il1b, Ccl4, or Apoa1) under-
lying infiltration or activation of anti-tumor immune cells in other intracranial
tumors. Serial intracranial bioluminescence was used to assess glioblastoma
growth and animals were monitored for survival. The impact of gene therapy
perturbations on the glioblastoma immune microenvironment was assessed
using histology, immunohistochemistry, single-cell mass cytometry (CyTOF),
and multiplexed cytokine assays. CYTOF and single-cell RNA sequencing of
human glioblastomas was performed for comparison with mouse glioblastoma
samples.
RESULTS: Serial body weight and systemic multiplexed cytokine measure-
ments showed no evidence of treatment toxicity. AAV9-APOA1 or AAV9-IL1B
gene therapy CED treatments attenuated SB28 growth and prolonged survival,
decreasing immunosuppressive macrophage infiltration and increasing CD8 T
cell and microglia infiltration of the glioblastoma microenvironment compared to
control AAV9 vectors. Gene therapy CED treatments did not attenuate GL261
growth or prolong survival, but CyTOF of human glioblastomas (n=6) in com-
parison to preclinical models revealed untreated GL261 glioblastomas were
endogenously enriched in CD8 T cells and other lymphoid lineages compared
to untreated SB28 or human glioblastomas. Multiplexed cytokine assays
demonstrated suppression of intratumor IL6 is a conserved mechanism of ac-
tion underlying glioblastoma gene therapy responses (SFig. 1). Analysis of
single-cell RNA sequencing of 32,877 cells from human glioblastomas (n=11)
showed IL6 is predominantly produced by radial glial like cancer stem cells or
endothelial cells in the tumor microenvironment. In support of these findings,
survival from intracranial SB28 glioblastomas was prolonged in Il6 knockout
C57BL/6J mice compared to wildtype mice.
CONCLUSIONS: Gene therapy and CED to reprogram the glioblastoma im-
mune microenvironment reveals IL6 drives glioblastoma immunosuppression
and growth.
Supplementary Figure 1. Multiplex cytokine array from SB28 tumors treated
with CED gene therapy shows conserved downregulation of IL-6 and LIF
across treatment conditions.

46