Randomized trial of neoadjuvant vaccination with tumor-cell lysate induces T cell response in low-grade gliomas

BACKGROUND. Long-term prognosis of WHO grade II low-grade gliomas (LGGs) is poor, with a high risk of recurrence and malignant transformation into high-grade gliomas. Given the relatively intact immune system of patients with LGGs and the slow tumor growth rate, vaccines are an attractive treatment strategy. METHODS. We conducted a pilot study to evaluate the safety and immunological effects of vaccination with GBM6-AD, lysate of an allogeneic glioblastoma stem cell line, with poly-ICLC in patients with LGGs. Patients were randomized to receive the vaccines before surgery (arm 1) or not (arm 2) and all patients received adjuvant vaccines. Coprimary outcomes were to evaluate safety and immune response in the tumor. RESULTS. A total of 17 eligible patients were enrolled — 9 in arm 1 and 8 in arm 2. This regimen was well tolerated with no regimen-limiting toxicity. Neoadjuvant vaccination induced upregulation of type-1 cytokines and chemokines and increased activated CD8+ T cells in peripheral blood. Single-cell RNA/T cell receptor sequencing detected CD8+ T cell clones that expanded with effector phenotype and migrated into the tumor microenvironment (TME) in response to neoadjuvant vaccination. Mass cytometric analyses detected increased tissue resident–like CD8+ T cells with effector memory phenotype in the TME after the neoadjuvant vaccination. CONCLUSION. The regimen induced effector CD8+ T cell response in peripheral blood and enabled vaccine-reactive CD8+ T cells to migrate into the TME. Further refinements of the regimen may have to be integrated into future strategies

Requirement for Estrogen Receptor ? in a Mouse Model for Human Papillomavirus–Associated Cervical Cancer

The majority of human cervical cancers are associated with the high-risk human papillomaviruses (HPV), which encode the potent E6 and E7 oncogenes. On prolonged treatment with physiologic levels of exogenous estrogen, K14E7 transgenic mice expressing HPV-16 E7 oncoprotein in their squamous epithelia succumb to uterine cervical cancer. Furthermore, prolonged withdrawal of exogenous estrogen results in complete or partial regression of tumors in this mouse model. In the current study, we investigated whether estrogen receptor ? (ER?) is required for the development of cervical cancer in K14E7 transgenic mice. We show that exogenous estrogen fails to promote either dysplasia or cervical cancer in K14E7/ER??/? mice despite the continued presence of the presumed cervical cancer precursor cell type, reserve cells, and evidence for E7 expression therein. We also observed that cervical cancers in our mouse models are strictly associated with atypical squamous metaplasia (ASM), which is believed to be the precursor for cervical cancer in women. Consistently, E7 and exogenous estrogen failed to promote ASM in the absence of ER?. We conclude that ER? plays a crucial role at an early stage of cervical carcinogenesis in this mouse model. [Cancer Res 2008;68(23):9928–34

BIOM-38. PI3K/AKT/mTOR SIGNALING PATHWAY ACTIVITY IN IDH-MUTANT DIFFUSE GLIOMA

Abstract PI3K/AKT/mTOR signaling pathway activation is a common mechanism of tumor progression in diffuse lower grade glioma. Robust and accurate biomarkers are needed to stratify patients for therapies targeting this pathway. To investigate the potential of phosphoprotein quantification to provide a direct and functional pathway readout, we analyzed 90 tumors from 83 patients with IDH-mutant diffuse glioma. The cohort comprised 50 IDH-mutant astrocytomas, 40 IDH-mutant and 1p/19q-codeleted oligodendrogliomas, 7 of whom had paired samples from initial diagnosis and recurrence. We developed and validated a pipeline using multiplex immunofluorescence to quantify tumor cell-specific phospho-protein expression of 3 pathway nodes, ribosomal protein S6 (RPS6), PRAS40, and 4E-BP1. In oligodendroglioma the fraction of tumor cells expressing each of the three phosphorylated proteins increased with tumor grade (p< 0.05). Comparing tumors at initial diagnosis (n=48) and at recurrence (n=42), p-RPS6 and p-PRAS40 increased in tumor cells (p< 0.05) and there was an overall increase in intertumoral heterogeneity of signaling activity at recurrence (p< 0.04). Analysis of paired samples demonstrated increased signaling pathway activity in a subset at recurrence. Robust signaling activity, defined as a phospho-positive tumor cell fraction ≥ median for all three phosphoproteins, was identified in 71.4% of grade 3 IDH-mutant astrocytoma(5/7) and 45.4% of grade 3 IDH-mutant, 1p/19q-codeleted oligodendroglioma(5/11). In a subset of cases analyzed by targeted NGS, robust signaling pathway activity was identified in 38%(11/29) at the protein level while genetic alterations predicted to activate the pathway were present in only 17.2% (5/29). Our results demonstrate robust PI3K/AKT/mTOR signaling activity in a significant fraction of IDH-mutant diffuse glioma, an association with increasing tumor grade in oligodendroglioma, and an increase at recurrence in both oligodendroglioma and astrocytoma. Overall, our data suggest that quantitative evaluation of phosphoproteins may be a sensitive method to detect PI3K/AKT/mTOR pathway activity and may be useful for patient stratification

MicroRNA dynamics in the stages of tumorigenesis correlate with hallmark capabilities of cancer

While altered expression of microRNAs (miRs) in tumors has been well documented, it remains unclear how the miR transcriptome intersects neoplastic progression. By profiling the miR transcriptome we identified miR expression signatures associated with steps in tumorigenesis and the acquisition of hallmark capabilities in a prototypical mouse model of cancer. Metastases and a rare subset of primary tumors shared a distinct miR signature, implicating a discrete lineage for metastatic tumors. The miR-200 family is strongly down-regulated in metastases and met-like primary tumors, thereby relieving repression of the mesenchymal transcription factor Zeb1, which in turn suppresses E-cadherin. Treatment with a clinically approved angiogenesis inhibitor normalized angiogenic signature miRs in primary tumors, while altering expression of metastatic signature miRs similarly to liver metastases, suggesting their involvement in adaptive resistance to anti-angiogenic therapy via enhanced metastasis. Many of the miR changes associated with specific stages and hallmark capabilities in the mouse model are similarly altered in human tumors, including cognate pancreatic neuroendocrine tumors, implying a generality

PI3K/AKT/mTOR signaling pathway activity in IDH-mutant diffuse glioma and clinical implications.

BackgroundIDH-mutant diffuse gliomas are heterogeneous, and improved methods for optimal patient therapeutic stratification are needed. PI3K/AKT/mTOR signaling activity can drive disease progression and potential therapeutic inhibitors of the pathway are available. Yet, the prevalence of PI3K/AKT/mTOR signaling pathway activity in IDH-mutant glioma is unclear and few robust strategies to assess activity in clinical samples exist.MethodsPI3K/AKT/mTOR signaling pathway activity was evaluated in a retrospective cohort of 132 IDH-mutant diffuse glioma (91 astrocytoma and 41 oligodendroglioma, 1p/19q-codeleted) through quantitative multiplex immunoprofiling using phospho-specific antibodies for PI3K/AKT/mTOR pathway members, PRAS40, RPS6, and 4EBP1, and tumor-specific anti-IDH1 R132H. Expression levels were correlated with genomic evaluation of pathway intrinsic genes and univariate and multivariate Cox proportional hazard regression models were used to evaluate the relationship with outcome.ResultsTumor-specific expression of p-PRAS40, p-RPS6, and p-4EBP1 was common in IDH-mutant diffuse glioma and increased with CNS WHO grade from 2 to 3. Genomic analysis predicted pathway activity in 21.7% (13/60) while protein evaluation identified active PI3K/AKT/mTOR signaling in 56.6% (34/60). Comparison of expression in male versus female patients suggested sexual dimorphism. Of particular interest, when adjusting for clinical prognostic factors, the level of phosphorylation of RPS6 was strongly associated with PFS (P < .005). Phosphorylation levels of both PRAS40 and RPS6 showed an association with PFS in univariate analysis.ConclusionsOur study emphasizes the value of proteomic assessment of signaling pathway activity in tumors as a means to identify relevant oncogenic pathways and potentially as a biomarker for identifying aggressive disease