Immunotherapy-Based Neoadjuvant Treatment of Advanced Microsatellite Instability-High Gastric Cancer: A Case Series.

Despite the use of first-line therapies like fluoropyrimidine and platinum-based cytotoxic chemotherapy, gastric cancer (GC) continues to carry a poor prognosis. Recent subgroup analyses of first-line phase III trials have demonstrated that patients with microsatellite instability-high (MSI-H) metastatic GC derive significant improvement in survival rates when immune checkpoint inhibitors (ICIs) are combined with chemotherapy compared with chemotherapy alone. However, it remains to be seen whether the success of ICIs in the metastatic setting can be translated into earlier stages of GC with resectable disease. We report 6 cases of locally advanced, nonmetastatic MSI-H GC that all demonstrated favorable response following treatment with pembrolizumab in addition to neoadjuvant chemotherapy. With the exception of immune-related colitis in one patient, pembrolizumab was well-tolerated. To our knowledge, this is the first reported US case series of patients treated with an ICI in combination with neoadjuvant chemotherapy for advanced, nonmetastatic, resectable or unresectable MSI-H GC.

Regression of Glioblastoma after Chimeric Antigen Receptor T-Cell Therapy.

A patient with recurrent multifocal glioblastoma received chimeric antigen receptor (CAR)-engineered T cells targeting the tumor-associated antigen interleukin-13 receptor alpha 2 (IL13Rα2). Multiple infusions of CAR T cells were administered over 220 days through two intracranial delivery routes - infusions into the resected tumor cavity followed by infusions into the ventricular system. Intracranial infusions of IL13Rα2-targeted CAR T cells were not associated with any toxic effects of grade 3 or higher. After CAR T-cell treatment, regression of all intracranial and spinal tumors was observed, along with corresponding increases in levels of cytokines and immune cells in the cerebrospinal fluid. This clinical response continued for 7.5 months after the initiation of CAR T-cell therapy. (Funded by Gateway for Cancer Research and others; ClinicalTrials.gov number, NCT02208362 .).

Optimization of IL13Rα2-Targeted Chimeric Antigen Receptor T Cells for Improved Anti-tumor Efficacy against Glioblastoma.

T cell immunotherapy is emerging as a powerful strategy to treat cancer and may improve outcomes for patients with glioblastoma (GBM). We have developed a chimeric antigen receptor (CAR) T cell immunotherapy targeting IL-13 receptor α2 (IL13Rα2) for the treatment of GBM. Here, we describe the optimization of IL13Rα2-targeted CAR T cells, including the design of a 4-1BB (CD137) co-stimulatory CAR (IL13BBζ) and a manufacturing platform using enriched central memory T cells. Utilizing orthotopic human GBM models with patient-derived tumor sphere lines in NSG mice, we found that IL13BBζ-CAR T cells improved anti-tumor activity and T cell persistence as compared to first-generation IL13ζ-CAR CD8+ T cells that had shown evidence for bioactivity in patients. Investigating the impact of corticosteroids, given their frequent use in the clinical management of GBM, we demonstrate that low-dose dexamethasone does not diminish CAR T cell anti-tumor activity in vivo. Furthermore, we found that local intracranial delivery of CAR T cells elicits superior anti-tumor efficacy as compared to intravenous administration, with intraventricular infusions exhibiting possible benefit over intracranial tumor infusions in a multifocal disease model. Overall, these findings help define parameters for the clinical translation of CAR T cell therapy for the treatment of brain tumors.

Skin/nail infections with the addition of pertuzumab to trastuzumab-based chemotherapy.

We report a series of breast cancer patients with invasive skin and nail infections with Staphylococcus species that we attribute to the addition of pertuzumab to trastuzumab-based therapy. With the suspicion of an increased incidence of cutaneous infection in patients treated with pertuzumab and trastuzumab-based chemotherapy, treating medical oncologists identified patients receiving therapy who experienced infection. Between March and October 2014, 18 patients treated with pertuzumab and trastuzumab-based chemotherapy were found to have 21 separate skin/nail infections. Treatment was administered as neoadjuvant therapy in 12 (67%) patients, adjuvant therapy in four (22%) patients, and for metastatic disease in two (11%) patients. Granulocyte growth factors were administered in 11 (61%) patients and no patients were documented to be neutropenic. New skin and nail lesions developed as early as cycle 1 and as late as 8 months from initial therapy. The 21 separate infections documented were folliculitis and "bite-like" lesions (10), abscess (6), paronychia (3), and cellulitis (2). The appearance of these lesions was distinct from typical EGFR-associated skin changes. When cultures were obtained, Staphylococcus species were isolated. Quantitative immunoglobulins were assessed in 14 (78%) patients and were abnormally low in six (43%) of these patients. The skin infections resulted in treatment delay in two (11%) patients and premature discontinuation of therapy in one patient. We believe that the skin/nail infections reported here in patients treated with the combination of pertuzumab and trastuzumab represent a previously unrecognized toxicity of adding pertuzumab to trastuzumab-based therapies.

New approach based on compressive sampling for sample rate enhancement in DASs for low-cost sensing nodes.

The paper deals with the problem of improving the maximum sample rate of analog-to-digital converters (ADCs) included in low cost wireless sensing nodes. To this aim, the authors propose an efficient acquisition strategy based on the combined use of high-resolution time-basis and compressive sampling. In particular, the high-resolution time-basis is adopted to provide a proper sequence of random sampling instants, and a suitable software procedure, based on compressive sampling approach, is exploited to reconstruct the signal of interest from the acquired samples. Thanks to the proposed strategy, the effective sample rate of the reconstructed signal can be as high as the frequency of the considered time-basis, thus significantly improving the inherent ADC sample rate. Several tests are carried out in simulated and real conditions to assess the performance of the proposed acquisition strategy in terms of reconstruction error. In particular, the results obtained in experimental tests with ADC included in actual 8- and 32-bits microcontrollers highlight the possibility of achieving effective sample rate up to 50 times higher than that of the original ADC sample rate.

PSCA-CAR T cell therapy in metastatic castration-resistant prostate cancer: a phase 1 trial.

Despite recent therapeutic advances, metastatic castration-resistant prostate cancer (mCRPC) remains lethal. Chimeric antigen receptor (CAR) T cell therapies have demonstrated durable remissions in hematological malignancies. We report results from a phase 1, first-in-human study of prostate stem cell antigen (PSCA)-directed CAR T cells in men with mCRPC. The starting dose level (DL) was 100 million (M) CAR T cells without lymphodepletion (LD), followed by incorporation of LD. The primary end points were safety and dose-limiting toxicities (DLTs). No DLTs were observed at DL1, with a DLT of grade 3 cystitis encountered at DL2, resulting in addition of a new cohort using a reduced LD regimen + 100 M CAR T cells (DL3). No DLTs were observed in DL3. Cytokine release syndrome of grade 1 or 2 occurred in 5 of 14 treated patients. Prostate-specific antigen declines (>30%) occurred in 4 of 14 patients, as well as radiographic improvements. Dynamic changes indicating activation of peripheral blood endogenous and CAR T cell subsets, TCR repertoire diversity and changes in the tumor immune microenvironment were observed in a subset of patients. Limited persistence of CAR T cells was observed beyond 28 days post-infusion. These results support future clinical studies to optimize dosing and combination strategies to improve durable therapeutic outcomes. ClinicalTrials.gov identifier NCT03873805 .

Locoregional delivery of IL-13Rα2-targeting CAR-T cells in recurrent high-grade glioma: a phase 1 trial.

Chimeric antigen receptor T cell (CAR-T) therapy is an emerging strategy to improve treatment outcomes for recurrent high-grade glioma, a cancer that responds poorly to current therapies. Here we report a completed phase I trial evaluating IL-13Rα2-targeted CAR-T cells in 65 patients with recurrent high-grade glioma, the majority being recurrent glioblastoma (rGBM). Primary objectives were safety and feasibility, maximum tolerated dose/maximum feasible dose and a recommended phase 2 dose plan. Secondary objectives included overall survival, disease response, cytokine dynamics and tumor immune contexture biomarkers. This trial evolved to evaluate three routes of locoregional T cell administration (intratumoral (ICT), intraventricular (ICV) and dual ICT/ICV) and two manufacturing platforms, culminating in arm 5, which utilized dual ICT/ICV delivery and an optimized manufacturing process. Locoregional CAR-T cell administration was feasible and well tolerated, and as there were no dose-limiting toxicities across all arms, a maximum tolerated dose was not determined. Probable treatment-related grade 3+ toxicities were one grade 3 encephalopathy and one grade 3 ataxia. A clinical maximum feasible dose of 200 × 106 CAR-T cells per infusion cycle was achieved for arm 5; however, other arms either did not test or achieve this dose due to manufacturing feasibility. A recommended phase 2 dose will be refined in future studies based on data from this trial. Stable disease or better was achieved in 50% (29/58) of patients, with two partial responses, one complete response and a second complete response after additional CAR-T cycles off protocol. For rGBM, median overall survival for all patients was 7.7 months and for arm 5 was 10.2 months. Central nervous system increases in inflammatory cytokines, including IFNγ, CXCL9 and CXCL10, were associated with CAR-T cell administration and bioactivity. Pretreatment intratumoral CD3 T cell levels were positively associated with survival. These findings demonstrate that locoregional IL-13Rα2-targeted CAR-T therapy is safe with promising clinical activity in a subset of patients. ClinicalTrials.gov Identifier: NCT02208362 .

Peritumoral Immune-suppressive Mechanisms Impede Intratumoral Lymphocyte Infiltration into Colorectal Cancer Liver versus Lung Metastases.

Patients with microsatellite stable (MSS) colorectal cancer with liver metastases are resistant to immune checkpoint inhibitor (ICI) therapy, while about one-third of patients with colorectal cancer without liver metastases, particularly those with lung-only metastases, respond to ICI. We analyzed primary colorectal cancer tumors and major metastatic sites (liver, lung, peritoneal) using multiplex immunofluorescence and whole-slide spatial analyses to identify variations in immune contexture and regional localization within the tumor microenvironment. While levels of T and B cells within peritumoral regions were similar, their levels were significantly lower within the tumor core of liver and peritoneal metastases compared with lung metastases. In contrast, antigen-presenting cells (APC) and APC-T cell interactions were more abundant in all regions of lung metastases. We also identified an abundance of lymphoid aggregates throughout lung metastases, but these were present only within peritumoral regions of liver and peritoneal metastases. Larger lymphoid aggregates consistent with features of tertiary lymphoid structures were observed within or adjacent to primary tumors, but not metastatic lesions. Our findings were validated using NanoString GeoMx DSP, which further showed that liver metastases had higher expression of immune-suppressive markers, while lung metastases showed higher proinflammatory activity and T-cell activation markers. Peritoneal metastases demonstrated higher expression of cancer-associated fibroblast-related proteins and upregulated PD-1/PD-L1 signaling molecules. Our results demonstrate that functional status and spatial distribution of immune cells vary significantly across different metastatic sites. These findings suggest that metastatic site-dependent immune contexture may underlie discordant responses to ICI therapy in patients with MSS colorectal cancer. SIGNIFICANCE: Our results demonstrate that functional status and spatial distribution of immune cells vary significantly across different metastatic sites in MSS colorectal cancer. These findings suggest that metastatic site-dependent immune contexture may underlie discordant responses to ICI therapy in patients with MSS colorectal cancer.

Expansion of endogenous T cells in CSF of pediatric CNS tumor patients undergoing locoregional delivery of IL13R〿2-targeting CAR T cells: an interim analysis.

Outcomes for pediatric brain tumor patients remain poor, and there is optimism that chimeric antigen receptor (CAR) T cell therapy can improve prognosis. Here, we present interim results from the first six pediatric patients treated on an ongoing phase I clinical trial (NCT04510051) of IL13BBζ-CAR T cells delivered weekly into the lateral cerebral ventricles, identifying clonal expansion of endogenous CAR-negative CD8+ T cells in the cerebrospinal fluid (CSF) over time. Additionally, of the five patients evaluable for disease response, three experienced transient radiographic and/or clinical benefit not meeting protocol criteria for response. The first three patients received CAR T cells alone; later patients received lymphodepletion before the first infusion. There were no dose limiting toxicities (DLTs). Aside from expected cytopenias in patients receiving lymphodepletion, serious adverse events possibly attributed to CAR T cell infusion were limited to one episode of headache and one of liver enzyme elevation. One patient withdrew from treatment during the DLT period due to a Grade 3 catheter-related infection and was not evaluable for disease response, although this was not attributed to CAR T cell infusion. Importantly, scRNA- and scTCR-sequence analyses provided insights into CAR T cell interaction with the endogenous immune system. In particular, clonally expanded endogenous CAR- T cells were recovered from the CSF, but not the peripheral blood, of patients who received intraventricular IL13BBζ-CAR T cell therapy. Additionally, although immune infiltrates in CSF and post-therapy tumor did not generally correlate, a fraction of expanded T cell receptors (TCRs) was seen to overlap between CSF and tumor. This has important implications for what samples are collected on these trials and how they are analyzed. These initial findings provide support for continued investigation into locoregionally-delivered IL13BBζ-CAR T cells for children with brain tumors.

Spatial organization of heterogeneous immunotherapy target antigen expression in high-grade glioma.

High-grade (WHO grades III-IV) glioma remains one of the most lethal human cancers. Adoptive transfer of tumor-targeting chimeric antigen receptor (CAR)-redirected T cells for high-grade glioma has revealed promising indications of anti-tumor activity, but objective clinical responses remain elusive for most patients. A significant challenge to effective immunotherapy is the highly heterogeneous structure of these tumors, including large variations in the magnitudes and distributions of target antigen expression, observed both within individual tumors and between patients. To obtain a more detailed understanding of immunotherapy target antigens within patient tumors, we immunochemically mapped at single cell resolution three clinically-relevant targets, IL13Rα2, HER2 and EGFR, on tumor samples drawn from a 43-patient cohort. We observed that within individual tumor samples, expression of these antigens was neither random nor uniform, but rather that they mapped into local neighborhoods - phenotypically similar cells within regions of cellular tumor - reflecting not well understood properties of tumor cells and their milieu. Notably, tumor cell neighborhoods of high antigen expression were not arranged independently within regions. For example, in cellular tumor regions, neighborhoods of high IL13Rα2 and HER2 expression appeared to be reciprocal to those of EGFR, while in areas of pseudopalisading necrosis, expression of IL13Rα2 and HER2, but not EGFR, appeared to reflect the radial organization of tumor cells around hypoxic cores. Other structural features affecting expression of immunotherapy target antigens remain to be elucidated. This structured but heterogeneous organization of antigen expression in high grade glioma is highly permissive for antigen escape, and combinatorial antigen targeting is a commonly suggested potential mitigating strategy. Deeper understanding of antigen expression within and between patient tumors will enhance optimization of combination immunotherapies, the most immediate clinical application of the observations presented here being the importance of including (wild-type) EGFR as a target antigen.

A Phase 2 Trial Combining Pembrolizumab and Palliative Radiation Therapy in Gastroesophageal Cancer to Augment Abscopal Immune Responses.

PURPOSE: Single agent PD-1 inhibitors have yielded durable responses in a minority of gastroesophageal cancers. Radiation therapy has been recognized to promote antitumor immune responses and may synergize with anti-PD-1 agents. We sought to evaluate if combining palliative radiation therapy with pembrolizumab can augment antitumor immune responses in gastroesophageal cancer. METHODS AND MATERIALS: Patients had metastatic gastroesophageal cancer with indication for palliative radiation therapy with ≥2 disease sites outside of the radiation field assessable for abscopal response and biopsies for laboratory correlative analyses. Palliative radiation was delivered to a dose of 30 Gy over 10 fractions. Pembrolizumab, 200 mg, was administered concurrently intravenously every 3 weeks until disease progression, unacceptable toxicity, or study withdrawal, for up to 2 years. Endpoints included PD-L1 expression in pre- and posttreatment biopsies and abscopal objective response rate per Response Evaluation Criteria in Solid Tumors. RESULTS: Of 14 enrolled patients, the objective response rate was 28.6% (95% confidence interval, 8.4%-58.1%), and the median duration of response was not reached (95% confidence interval, 6.9-NR months). Overall, 2 patients had treatment-related grade 3 to 4 adverse events with no grade 5 events. One patient discontinued therapy due to grade 4 colitis. We did not observe an association between radiation and abscopal changes in PD-L1 expression via assessment of an analogous PD-L1 Combined Positive Score, Tumor Proportion Score, Mononuclear Immune Cell Density Score, or proportion of PD-L1-expressing immune cells between pre- and posttreatment tumor biopsies. CONCLUSIONS: Combining palliative radiation therapy and pembrolizumab provided promising durable responses in this patient population but we were unable to definitively distinguish abscopal biologic changes. Biomarker analyses beyond PD-L1 expression are needed to better understand putative mechanisms and identify patients who will benefit from this approach.

Off-the-shelf, steroid-resistant, IL13Rα2-specific CAR T cells for treatment of glioblastoma.

BACKGROUND: Wide-spread application of chimeric antigen receptor (CAR) T cell therapy for cancer is limited by the current use of autologous CAR T cells necessitating the manufacture of individualized therapeutic products for each patient. To address this challenge, we have generated an off-the-shelf, allogeneic CAR T cell product for the treatment of glioblastoma (GBM), and present here the feasibility, safety, and therapeutic potential of this approach. METHODS: We generated for clinical use a healthy-donor derived IL13Rα2-targeted CAR+ (IL13-zetakine+) cytolytic T-lymphocyte (CTL) product genetically engineered using zinc finger nucleases (ZFNs) to permanently disrupt the glucocorticoid receptor (GR) (GRm13Z40-2) and endow resistance to glucocorticoid treatment. In a phase I safety and feasibility trial we evaluated these allogeneic GRm13Z40-2 T cells in combination with intracranial administration of recombinant human IL-2 (rhIL-2; aldesleukin) in six patients with unresectable recurrent GBM that were maintained on systemic dexamethasone (4-12 mg/day). RESULTS: The GRm13Z40-2 product displayed dexamethasone-resistant effector activity without evidence for in vitro alloreactivity. Intracranial administration of GRm13Z40-2 in four doses of 108 cells over a two-week period with aldesleukin (9 infusions ranging from 2500-5000 IU) was well tolerated, with indications of transient tumor reduction and/or tumor necrosis at the site of T cell infusion in four of the six treated research subjects. Antibody reactivity against GRm13Z40-2 cells was detected in the serum of only one of the four tested subjects. CONCLUSIONS: This first-in-human experience establishes a foundation for future adoptive therapy studies using off-the-shelf, zinc-finger modified, and/or glucocorticoid resistant CAR T cells.

Feasibility of intracerebrally administering multiple doses of genetically modified neural stem cells to locally produce chemotherapy in glioma patients.

Neural stem cells (NSCs) are tumor tropic and can be genetically modified to produce anti-cancer therapies locally in the brain. In a prior first-in-human study we demonstrated that a single dose of intracerebrally administered allogeneic NSCs, which were retrovirally transduced to express cytosine deaminase (CD), tracked to glioma sites and converted oral 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU). The next step in the clinical development of this NSC-based anti-cancer strategy was to assess the feasibility of administering multiple intracerebral doses of CD-expressing NSCs (CD-NSCs) in patients with recurrent high-grade gliomas. CD-NSCs were given every 2 weeks using an indwelling brain catheter, followed each time by a 7-d course of oral 5-FC (and leucovorin in the final patient cohort). Fifteen evaluable patients received a median of 4 (range 2-10) intracerebral CD-NSC doses; doses were escalated from 50 × 106 to 150 × 106 CD-NSCs. Neuropharmacokinetic data confirmed that CD-NSCs continuously produced 5-FU in the brain during the course of 5-FC. There were no clinical signs of immunogenicity, and only three patients developed anti-NSC antibodies. Our results suggest intracerebral administration of serial doses of CD-NSCs is safe and feasible and identified a recommended dose for phase II testing of 150 × 106 CD-NSCs.

Desmoplasia and oncogene driven acinar-to-ductal metaplasia are concurrent events during acinar cell-derived pancreatic cancer initiation in young adult mice.

The five-year survival rate of patients diagnosed with advanced pancreatic ductal adenocarcinoma (PDAC) has remained static at <5% despite decades of research. With the exception of erlotinib, clinical trials have failed to demonstrate the benefit of any targeted therapy for PDAC despite promising results in preclinical animal studies. The development of more refined mouse models of PDAC which recapitulate the carcinogenic progression from non-neoplastic, adult exocrine subsets of pancreatic cells to invasive carcinoma in humans are needed to facilitate the accurate translation of therapies to the clinic. To study acinar cell-derived PDAC initiation, we developed a genetically engineered mouse model of PDAC, called KPT, utilizing a tamoxifen-inducible Cre recombinase/estrogen receptor (ESR1) fusion protein knocked into the Ptf1a locus to activate the expression of oncogenic KrasG12D and Trp53R270H alleles in mature pancreatic acinar cells. Oncogene-expressing acinar cells underwent acinar-to-ductal metaplasia, and formed pancreatic intraepithelial neoplasia lesions following the induction of oncogene expression. After a defined latency period, oncogene-expressing acinar cells initiated the formation of highly differentiated and fibrotic tumors, which metastasized to the lungs and liver. Whole-transcriptome analysis of microdissected regions of acinar-to-ductal metaplasia and histological validation experiments demonstrated that regions of acinar-to-ductal metaplasia are characterized by the deposition of the extracellular matrix component hyaluronan. These results indicate that acinar cells expressing KrasG12D and Trp53R270H can initiate PDAC development in young adult mice and implicate hyaluronan deposition in the formation of the earliest characterized PDAC precursor lesions (and the progression of pancreatic cancer). Further studies are necessary to provide a comprehensive characterization of PDAC progression and treatment response in KPT mice and to investigate whether the KPT model could be used as a tool to study translational aspects of acinar cell-derived PDAC tumorigenesis.

A brainstem mass of Müllerian type Epithelial Origin without any primary cancer source.

Brainstem tumors are rare, even rarer is a brainstem tumor containing tissues of an embryologic gynecologic origin. We report a very rare case of presence of a calcified heterogeneously contrast enhancing brainstem mass of Müllerian origin in a patient in a 38 year old female with no female genital tract cancer and past surgical history of ventriculoperitoneal (VP) shunt placement for congenital hydrocephalus. To our knowledge this is the very first and unusual case of a mass of gynecologic origin in the brainstem region especially in the setting of no history of gynecological tumor. The authors also reviewed the literature for all tumors reported for anterograde and retrograde dissemination of tumor cells through VP shunt. This case is a reaffirmation of the importance of brain tumor location and tissue diagnosis for the purpose of adjuvant treatment of neurosurgical lesions in the neurocritical care setting. It also highlights the role of catheters as potential routes of iatrogenic transmission not just in anterograde but also in a retrograde manner to the CNS, which is very unusual. This is the only second case to report retrograde flow of tumor cells from an extraneural source up the VP catheters. The authors suggest that intraperitoneal chemotherapy should be considered in the cases of known extraneural abdominal malignancies of high malignant potential with or without the presence of peritoneal infiltration in order to avoid dissemination through VP shunts.

Pituitary metastasis: a rare condition.

Tumor metastasis to the pituitary gland is a life-threatening condition associated with short life span. Pituitary metastasis is rare, however, and not well-documented. A better understanding of its clinical manifestations could lead to earlier diagnosis, appropriate therapy, and potentially improving quality of life. Therefore, we retrospectively studied the charts of patients with pituitary metastases who were treated at the City of Hope National Medical Center in Duarte, California, from 1984 to 2018. We reviewed and analyzed tumor origin, primary pituitary clinical manifestation, duration between primary tumor diagnosis and pituitary metastasis, type of treatment, and patient survival. A total 11 patients with a mean age of 59.2 years and median survival of 50.33 months were identified. Breast cancer and lymphoma were the most common primary origins in these cases, and diabetes insipidus and panhypopituitarism were the most common primary manifestations of their metastasis. We also compared our results with reports in the literature published between 1957 and 2018. A total 289 patients with pituitary metastasis have been reported in the literature. Breast cancer was the most frequent primary origin of the metastasis, and visual involvement was the most common primary manifestation. The posterior part of the pituitary is more susceptible than the anterior to metastasis. Pituitary metastasis may occur as a consequence of successful primary tumor treatment prolonging the chance of seeding. Future studies are needed to determine the molecular mechanism of metastasis to the pituitary.

Early Changes in Tumor Perfusion from T1-Weighted Dynamic Contrast-Enhanced MRI following Neural Stem Cell-Mediated Therapy of Recurrent High-Grade Glioma Correlate with Overall Survival.

BACKGROUND: The aim of this study was to correlate T1-weighted dynamic contrast-enhanced MRI- (DCE-MRI-) derived perfusion parameters with overall survival of recurrent high-grade glioma patients who received neural stem cell- (NSC-) mediated enzyme/prodrug gene therapy. METHODS: A total of 12 patients were included in this retrospective study. All patients were enrolled in a first-in-human study (NCT01172964) of NSC-mediated therapy for recurrent high-grade glioma. DCE-MRI data from all patients were collected and analyzed at three time points: MRI#1-day 1 postsurgery/treatment, MRI#2- day 7 ± 3 posttreatment, and MRI#3-one-month follow-up. Plasma volume (Vp), permeability (Ktr), and leakage (λtr) perfusion parameters were calculated by fitting a pharmacokinetic model to the DCE-MRI data. The contrast-enhancing (CE) volume was measured from the last dynamic phase acquired in the DCE sequence. Perfusion parameters and CE at each MRI time point were recorded along with their relative change between MRI#2 and MRI#3 (Δ32). Cox regression was used to analyze patient survival. RESULTS: At MRI#1 and at MRI#3, none of the parameters showed a significant correlation with overall survival (OS). However, at MRI#2, CE and λtr were significantly associated with OS (p < 0.05). The relative λtr and Vp from timepoint 2 to timepoint 3 (Δ32λtr and Δ32Vp) were each associated with a higher hazard ratio (p < 0.05). All parameters were highly correlated, resulting in a multivariate model for OS including only CE at MRI#2 and Δ32Vp, with an R2 of 0.89. CONCLUSION: The change in perfusion parameter values from 1 week to 1 month following NSC-mediated therapy combined with contrast-enhancing volume may be a useful biomarker to predict overall survival in patients with recurrent high-grade glioma.

TLR9 expression and secretion of LIF by prostate cancer cells stimulates accumulation and activity of polymorphonuclear MDSCs.

Proinflammatory signals promote prostate tumorigenesis and progression, but their origins and downstream effects remain unclear. We recently demonstrated that the expression of an innate immune receptor, TLR9, by prostate cancer cells is critical for their tumor-propagating potential. We investigated whether cancer cell-intrinsic TLR9 signaling alters composition of the prostate tumor microenvironment. We generated Ras/Myc (RM9) and Myc-driven (Myc-CaP) prostate cancer cells expressing the tetracycline-inducible gene Tlr9 (Tlr9ON ) or the control LacZ (LacZON ). When engrafted into mice and treated with tetracycline, Tlr9ON , but not LacZON , tumors showed accelerated growth kinetics compared with tumors in PBS-treated mice. Tlr9 upregulation in cancer cells triggered the selective accumulation of CD11b+Ly6GHILy6CLO myeloid cells, phenotypically similar to PMN-MDSCs. The PMN-MDSCs from tetracycline-treated RM9-Tlr9ON tumors increased the immunosuppressive activity of the STAT3 transcription factor, thereby more potently inhibiting T cell proliferation. We identified LIF, an IL-6-type cytokine and STAT3 activator, as a potential mediator of crosstalk between TLR9-expressing prostate cancer cells and PMN-MDSCs. Antibody-mediated LIF neutralization reduced the percentage of tumor-infiltrating PMN-MDSCs and inhibited tumor growth in mice. The clinical relevance of LIF is confirmed by the correlation between TLR9 and LIF expression in prostate cancer specimens. Furthermore, blood samples from patients with prostate cancer showed elevated levels of LIF and high LIFR expression on circulating PMN-MDSCs. Our results suggest that TLR9+ prostate cancers promote immune evasion via LIF-mediated expansion and activation of PMN-MDSCs. Finally, targeting TLR9/LIF/STAT3 signaling using oligonucleotide-based inhibitors, such as CpG-STAT3dODN, can offer new opportunities for prostate cancer immunotherapy.

Neural Stem Cell-Based Anticancer Gene Therapy: A First-in-Human Study in Recurrent High-Grade Glioma Patients.

Purpose: Human neural stem cells (NSC) are inherently tumor tropic, making them attractive drug delivery vehicles. Toward this goal, we retrovirally transduced an immortalized, clonal NSC line to stably express cytosine deaminase (HB1.F3.CD.C21; CD-NSCs), which converts the prodrug 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU).Experimental Design: Recurrent high-grade glioma patients underwent intracranial administration of CD-NSCs during tumor resection or biopsy. Four days later, patients began taking oral 5-FC every 6 hours for 7 days. Study treatment was given only once. A standard 3 + 3 dose escalation schema was used to increase doses of CD-NSCs from 1 × 107 to 5 × 107 and 5-FC from 75 to 150 mg/kg/day. Intracerebral microdialysis was performed to measure brain levels of 5-FC and 5-FU. Serial blood samples were obtained to assess systemic drug concentrations as well as to perform immunologic correlative studies.Results: Fifteen patients underwent study treatment. We saw no dose-limiting toxicity (DLT) due to the CD-NSCs. There was 1 DLT (grade 3 transaminitis) possibly related to 5-FC. We did not see development of anti-CD-NSC antibodies and did not detect CD-NSCs or replication-competent retrovirus in the systemic circulation. Intracerebral microdialysis revealed that CD-NSCs produced 5-FU locally in the brain in a 5-FC dose-dependent manner. Autopsy data indicate that CD-NSCs migrated to distant tumor sites and were nontumorigenic.Conclusions: Collectively, our results from this first-in-human study demonstrate initial safety and proof of concept regarding the ability of NSCs to target brain tumors and locally produce chemotherapy. Clin Cancer Res; 23(12); 2951-60. ©2016 AACR.