(555i) Targeting Tumor Associated Macrophages with PAMAM Dendrimers Improves Therapeutic Efficacy in Glioblastoma

Introduction: Cancer
is the second leading cause of death in the United States, and brain cancers
are among the most severe types of cancers with poor patient outcomes due to
barriers associated with delivering therapeutics to the brain in addition to challenges
associated with delivering drugs to solid tumors.¹ Many anti-cancer
therapies fail to fully penetrate tumor tissue and access tumor cells at relevant
concentrations due to ill-developed vasculature and high interstitial pressure
within the tumor.² Additionally, the blood brain barrier (BBB),
which under healthy regimes is a natural defense mechanism stopping foreign
substances from reaching the brain, prevents therapeutically relevant
quantities of treatments from reaching the brain tumor.³ As a result,
patients with central nervous system cancers have failed to see improved
outcomes despite the discovery of powerful new anti-cancer drugs because these
therapies 1) have poor BBB penetration and 2) fail to distribute and accumulate
within the tumor. This two-pronged barrier necessitates the administration of
high doses of systemic therapies to achieve effective doses in the brain tumor,
which cause significant toxicity in the body. Therefore, a vehicle that carries
therapies to the brain, into the brain tumor, and to the tumor associated
macrophages (TAMs) while remaining inactive in the body provides a promising
clinical strategy.

Methods: Our
group has identified and extensively characterized hydroxyl polyamidoamine
(PAMAM) dendrimers as promising nano-vehicles for targeted drug delivery in
several small and large models of neurodegenerative disorders.^4,5 Here,
we present these dendrimers for targeting brain tumors in a mouse model of
glioblastoma (GBM). C57BL/6 mice 6-8 weeks old were inoculated with 100,000
GL261 murine GBM cells by intracranial injection. To assess tumor targeting, generation
4 PAMAM dendrimers were administered systemically 14 days post-inoculation and
analyzed via confocal microscopy and fluorescence spectrometry. In addition, two
therapies were evaluated: BLZ945, an immune-modulating agent, and triptolide,
an anti-proliferative drug. These therapies were conjugated to dendrimers via
pH-sensitive linkers for triggered release. With BLZ945, mice were administered
with a single systemic dose of free or dendrimer-conjugated drug 10 days
post-inoculation at 100 mg/kg on a drug basis. With triptolide, mice were
systemically administered daily at 0.5 mg/kg free or dendrimer-conjugated drug
starting 5 days post-inoculation. Mice were monitored daily and their disease
progression assessed with kyphosis scoring and in open field trials.

Results: We
demonstrate that upon systemic administration, PAMAM dendrimers (without
any targeting ligands) are able to cross the BBB, diffuse freely through the
brain parenchyma, and distribute uniformly throughout the brain tumor. Confocal
imaging shows that dendrimers selectively localize to Iba1-labelled (TAMs) while
exhibiting minimal uptake by resting microglia/macrophages in healthy brain
tissue (Fig.1). Quantitatively, dendrimers accumulated specifically
within the tumor at >5-fold greater levels compared to the contralateral
hemisphere.

We next leveraged this tumor targeting capacity to
deliver therapeutics for enhanced efficacy, particularly in the context of favorably
manipulating TAMs. We find that a single systemic dose of dendrimer-BLZ945
immunotherapy halfway through disease progression prolongs survival by >25%
compared to BLZ945 and control treatment groups. Additionally, dendrimer-BLZ945
treatment delayed the onset and severity of kyphosis, as well as improved motor
function in open field tests. This effect is mediated by a reduction in
expression of Arginase-1, a marker of anti-inflammatory tumor promoting
phenotype, indicating an enhancement in immune-modulating effect with dendrimer
delivery. For triptolide, targeted delivery with the dendrimer resulted in
significantly reduced tumor size compared to free triptolide, as well as
improvements to motor function and kyphosis.

Conclusions: PAMAM
dendrimers (without any targeting ligands) selectively target TAMs within the
GBM tumor upon systemic administration and show minimal accumulation in healthy
brain tissue. This targeting capacity results in improved efficacy of
immunotherapy BLZ945 and anti-proliferative triptolide compared to their free
drug counterparts. We are currently further assessing and optimizing these
dendrimer-mediated therapies, with the plan for combination therapies in the
future. Therefore, PAMAM dendrimers have significant clinical applications for
the treatment of GBM by selectively delivering therapies to the tumor,
particularly in the context of localized manipulation of the immune profile in
GBM tumors.

References:

¹Siegel, R.L., Miller,
K.D, Jemal, A. CA: A Cancer Journal for Clinicians. 2018. 68(1):7-30

²Tredan, O., Galmarini,
C.M., et al. J of the Nat Cancer Inst. 2007. 99(19): 1441-54

³van Tellingen, O.,
Yetkin-Arik, B., et al. Drug Resistance Updates. 2015. 19: 1-12

⁴Kannan, S., Dai, H.,
Navath, R.S., et al. Sci Trans Med. 2012. 4(130): 130ra46

⁵Mishra, M.K., Beaty,
C.A., et al. ACS Nano. 2014. 8(3): 2134-47