18 research outputs found
IND-Enabling Studies for a Clinical Trial to Genetically Program a Persistent Cancer-Targeted Immune System
PURPOSE:
To improve persistence of adoptively transferred T-cell receptor (TCR)-engineered T cells and durable clinical responses, we designed a clinical trial to transplant genetically-modified hematopoietic stem cells (HSCs) together with adoptive cell transfer of T cells both engineered to express an NY-ESO-1 TCR. Here, we report the preclinical studies performed to enable an investigational new drug (IND) application.
EXPERIMENTAL DESIGN:
HSCs transduced with a lentiviral vector expressing NY-ESO-1 TCR and the PET reporter/suicide gene HSV1-sr39TK and T cells transduced with a retroviral vector expressing NY-ESO-1 TCR were coadministered to myelodepleted HLA-A2/Kb mice within a formal Good Laboratory Practice (GLP)-compliant study to demonstrate safety, persistence, and HSC differentiation into all blood lineages. Non-GLP experiments included assessment of transgene immunogenicity and in vitro viral insertion safety studies. Furthermore, Good Manufacturing Practice (GMP)-compliant cell production qualification runs were performed to establish the manufacturing protocols for clinical use.
RESULTS:
TCR genetically modified and ex vivo-cultured HSCs differentiated into all blood subsets in vivo after HSC transplantation, and coadministration of TCR-transduced T cells did not result in increased toxicity. The expression of NY-ESO-1 TCR and sr39TK transgenes did not have a detrimental effect on gene-modified HSC's differentiation to all blood cell lineages. There was no evidence of genotoxicity induced by the lentiviral vector. GMP batches of clinical-grade transgenic cells produced during qualification runs had adequate stability and functionality.
CONCLUSIONS:
Coadministration of HSCs and T cells expressing an NY-ESO-1 TCR is safe in preclinical models. The results presented in this article led to the FDA approval of IND 17471
VIP deficient mice exhibit resistance to lipopolysaccharide induced endotoxemia with an intrinsic defect in proinflammatory cellular responses.
Vasoactive intestinal peptide (VIP) is a pleiotropic neuropeptide with immunomodulatory properties. The administration of this peptide has been shown to have beneficial effects in murine models of inflammatory diseases including septic shock, rheumatoid arthritis, multiple sclerosis (MS) and Crohn's disease. However, the role of the endogenous peptide in inflammatory disease remains obscure because VIP-deficient mice were recently found to exhibit profound resistance in a model of MS. In the present study, we analyzed the response of female VIP deficient (KO) mice to intraperitoneal lipopolysaccharide (LPS) administration. We observed significant resistance to LPS in VIP KO mice, as evidenced by lower mortality and reduced tissue damage. The increased survival was associated with decreased levels of proinflammatory cytokines (TNFα, IL-6 and IL-12) in sera and peritoneal suspensions of these mice. Moreover, the expression of TNFα and IL-6 mRNA was reduced in peritoneal cells, spleens and lungs from LPS-treated VIP KO vs. WT mice, suggesting that the resistance might be mediated by an intrinsic defect in the responsiveness of immune cells to endotoxin. In agreement with this hypothesis, peritoneal cells isolated from VIP KO naive mice produced lower levels of proinflammatory cytokines in response to LPS in vitro. Finally, decreased NF-κB pathway activity in peritoneal cells was observed both in vivo and in vitro, as determined by assay of phosphorylated I-κB. The results demonstrate that female VIP KO mice exhibit resistance to LPS-induced shock, explainable in part by the presence of an intrinsic defect in the responsiveness of inflammatory cells to endotoxin
VIP KO mice exhibit reduced mortality and lung histopathology in response to LPS injection.
<p>Female WT (C57BL6) and VIP KO mice were injected i.p. with LPS (40 mg/Kg). A, Kaplan Meier curve analysis of survival cumulative of four experiments (total WT n = 29; VIP KO n = 28) (Curve comparison Logrank test **p<0.01). B, Representative sections of lungs from control (noninjected) or LPS-injected WT and VIP KO mice (24 hours post injection) stained with H&E. C, Histological scores of LPS-injected WT vs. VIP KO mice (mean of two experiments; total WT n = 7; VIP KO n = 9), 24 hours after LPS injection, scored from 0 to 3 according to the level of lung inflammation as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036922#s2" target="_blank"><i>Materials and Methods</i></a>. (Student's <i>t</i>-test *p<0.05).</p
VIP KO mice exhibit reduced levels of proinflammatory cytokines in the peritoneal fluid and serum.
<p>Female WT (C57BL6) (n = 6) and VIP KO mice (n = 6) were injected i.p. with LPS (40 mg/Kg), and serum and peritoneal suspensions were collected 0, 3 and 6 (and also 24 for IL-10) hours post-injection. The levels of TNFα, IL-6, IL-12p40 and IL-10 were assessed by sandwich ELISA as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036922#s2" target="_blank"><i>Materials and Methods</i></a>. Student's <i>t</i>-test *p<0.05; **p<0.01. One of three representative experiments is shown.</p
Peritoneal cells from VIP KO mice exhibit an intrinsic defect in cytokine response to LPS- administration.
<p>Peritoneal cells were collected from WT (n = 3) and VIP KO mice (n = 3), and cultured in complete RPMI in triplicate in the presence or absence of LPS (10 ng/ml). Supernatants were collected 2 (A) and 16 h (B) later, and stored at −20°C for analysis of TNFα and IL-6 levels by ELISA. Student's <i>t</i>-test *p<0.05; **p<0.01; ***p<0.001. Representative data are shown of four independent experiments.</p
Recommended from our members
Antiestrogens in combination with immune checkpoint inhibitors in breast cancer immunotherapy.
Breast cancers (BCs) with expression of estrogen receptor-alpha (ERα) occur in more than 70% of newly-diagnosed patients in the U.S. Endocrine therapy with antiestrogens or aromatase inhibitors is an important intervention for BCs that express ERα, and it remains one of the most effective targeted treatment strategies. However, a substantial proportion of patients with localized disease, and essentially all patients with metastatic BC, become resistant to current endocrine therapies. ERα is present in most resistant BCs, and in many of these its activity continues to regulate BC growth. Fulvestrant represents one class of ERα antagonists termed selective ER downregulators (SERDs). Treatment with fulvestrant causes ERα down-regulation, an event that helps overcome several resistance mechanisms. Unfortunately, full antitumor efficacy of fulvestrant is limited by its poor bioavailability in clinic. We have designed and tested a new generation of steroid-like SERDs. Using ERα-positive BC cells in vitro, we find that these compounds suppress ERα protein levels with efficacy similar to fulvestrant. Moreover, these new SERDs markedly inhibit ERα-positive BC cell transcription and proliferation in vitro even in the presence of estradiol-17β. In vivo, the SERD termed JD128 significantly inhibited tumor growth in MCF-7 xenograft models in a dose-dependent manner (P < 0.001). Further, our findings indicate that these SERDs also interact with ER-positive immune cells in the tumor microenvironment such as myeloid-derived suppressor cells (MDSC), tumor infiltrating lymphocytes and other selected immune cell subpopulations. SERD-induced inhibition of MDSCs and concurrent actions on CD8+ and CD4 + T-cells promotes interaction of immune checkpoint inhibitors with BC cells in preclinical models, thereby leading to enhanced tumor killing even among highly aggressive BCs such as triple-negative BC that lack ERα expression. Since monotherapy with immune checkpoint inhibitors has not been effective for most BCs, combination therapies with SERDs that enhance immune recognition may increase immunotherapy responses in BC and improve patient survival. Hence, ERα antagonists that also promote ER downregulation may potentially benefit patients who are unresponsive to current endocrine therapies