br Activating Effector Cells br Activated CD T
3.2 Activating Effector Cells
Activated CD8+ T-cells are the key effectors of tumor-specific (adaptive) immunity. Even if clonal antitumor T-cells that can recognize and attack tumor cells are generated, they must efficiently infiltrate the tumor stroma and remain activated in order to be effective. T-cell-mediated cytotoxicity is a tightly regulated process. Numerous receptors are expressed on the T-cell surface that can stimulate or inhibit the activity of activated T-cells77. Inhibitory receptors include CTLA-4 (CD152), PD-1 (CD279), TIM-2, and LAG-3 (CD223). Many tumors express ligands to these receptors, such as PDL-1, in an effort to reduce the activation and maturation of APCs and infiltrating CD8+ T-cells. Inhibitory antibodies to CTLA-4 and PD-L1 are far and away the most successful cancer immunotherapies to date. However, they are not
without limitations. Among immunogenic histologies like NSCLC, they are only effective in tumors that overexpress the PD-L1 ligand78. Durable response rates in PD-L1 overexpressing tumors are generally less than 50%. Many non-immunogenic tumors, including MSS colorectal tumors, express high levels of PDL-1 but are not responsive to anti-PD-1 therapies79,80. Natural killer (NK) cells are also important innate (antigen non-specific) cytotoxic effector cells. They harbor inhibitory surface receptors such as CD47 that can be overexpressed by tumors to avoid clearance by the innate immune system81. Intensive research has been dedicated to identifying novel methods of improving the activation of effector cells in the TME.
3.2.1 Enhancing T-cell Checkpoint Inhibition
NP delivery has been used to improve the efficacy of CTLA-4 and PDL-1 inhibitors by providing sustained release in tumors and co-delivery with other immune adjuvants. One group engineered photodegradable PLA NPs co-encapsulated with hollow gold nanoshells and anti-PD-1 peptide (APP)82. These particles enabled sustained release of APP for up to 40 days. Accelerated release could be stimulated with a near infrared laser. In a metastatic model system, laser-irradiation of “primary” tumors stimulated partial regression and growth delay in the laser-treated and distant tumor deposits indicating the activation of a systemic immune response. Zhen Gu and his group developed a cleavable nucleic acid-based NP comprised of the potent immune-stimulant CpG-ODN to deliver anti-PDL-1 antibodies83. The nucleic Ezatiostat shell was constructed with ssDNA and CpG repeats with cutting sites for the restriction enzyme HhaI. HhaI was caged in triglycerol monostearate NPs attached to the larger DNA NPs which could be enzymatically-cleaved by wound-responsive esterases and matrix metalloproteinases. Wound-associated inflammation (triggered by partial tumor resection) resulted in degradation of the DNA carrier and release of anti-PDL1 antibodies and CpG-ODN fragments. Systemic NP administration and subtotal tumor resection stimulated complete tumor rejection in 40% of mice, whereas no complete rejections were observed with systemic co-administration of CpG-ODN and anti-PDL-1 antibodies.
NP-delivered nucleic acid therapies have also been used for the targeted knockdown of inhibitory receptors in effector cells or inhibitory ligands in tumors that can enhance tumoral infiltration and activity of anti-tumor CD8+ lymphocytes. Wu et al. utilized lipid-coated calcium phosphate NPs to deliver siRNA targeted against PD-1 receptor and PDL-1 ligand84. These NPs efficiently knocked down PD-1 receptor in tumor-infiltrating lymphocytes (isolated from patient samples) and PDL-1 in MCF-7 breast cancer cells in vitro. Combined knockdown significantly enhanced CD8-mediated cytotoxicity assessed using ex vivo cytotoxic T-cell assays. They also observed significant increases in INFγ and TNFα excretion following co-culture of PD-1- TILs and PD-L1- MCF tumor cells. Lian et al. generated cationic lipid NPs to deliver siRNA against PDL-1 and CD47 in 13 tumor cell lines in vitro85. Tumor targeting and uptake was enhanced by decorating the particle surface with an EpCAM targeting peptide. Dual knockdown significantly enhanced CD8+ and NK-mediated cytotoxicity in vitro. They also demonstrated that their targeted NPs efficiently improved tumoral infiltration of CD8+ and NK cells in vivo using a 4T1 metastatic lung cancer model. This resulted in improved local tumor control and significantly decreased frequency of lung metastases. Similar improvements in tumor regression and CD8+ infiltration were observed in B16 melanoma tumors in a similar study utilizing lymphocyte-directed NP delivery of CTLA-4 siRNA86. One group recently compared the efficiency of PD-1 siRNA in lymphocytes using layered double hydroxide and lipid calcium phosphate NPs87. While both could facilitate decreased PD-1 receptor expression, cellular uptake and gene silencing were more effective with the lipid calcium phosphate particles suggesting that it may be the preferred platform for lymphocyte-directed siRNA delivery.