Daniel A. Pryma

Radiation and dual checkpoint blockade activate non-redundant immune mechanisms in cancer

Immune checkpoint inhibitors result in impressive clinical responses, but optimal results will require combination with each other and other therapies. This raises fundamental questions about mechanisms of non-redundancy and resistance. Here we report major tumour regressions in a subset of patients with metastatic melanoma treated with an anti-CTLA4 antibody (anti-CTLA4) and radiation, and reproduced this effect in mouse models. Although combined treatment improved responses in irradiated and unirradiated tumours, resistance was common. Unbiased analyses of mice revealed that resistance was due to upregulation of PD-L1 on melanoma cells and associated with T-cell exhaustion. Accordingly, optimal response in melanoma and other cancer types requires radiation, anti-CTLA4 and anti-PD-L1/PD-1. Anti-CTLA4 predominantly inhibits T-regulatory cells (Treg cells), thereby increasing the CD8 T-cell to Treg (CD8/Treg) ratio. Radiation enhances the diversity of the T-cell receptor (TCR) repertoire of intratumoral T cells. Together, anti-CTLA4 promotes expansion of T cells, while radiation shapes the TCR repertoire of the expanded peripheral clones. Addition of PD-L1 blockade reverses T-cell exhaustion to mitigate depression in the CD8/Treg ratio and further encourages oligoclonal T-cell expansion. Similarly to results from mice, patients on our clinical trial with melanoma showing high PD-L1 did not respond to radiation plus anti-CTLA4, demonstrated persistent T-cell exhaustion, and rapidly progressed. Thus, PD-L1 on melanoma cells allows tumours to escape anti-CTLA4-based therapy, and the combination of radiation, anti-CTLA4 and anti-PD-L1 promotes response and immunity through distinct mechanisms.

The nuclear receptor Rev-erbα controls circadian thermogenic plasticity

Circadian oscillation of body temperature is a basic, evolutionarily conserved feature of mammalian biology. In addition, homeostatic pathways allow organisms to protect their core temperatures in response to cold exposure. However, the mechanism responsible for coordinating daily body temperature rhythm and adaptability to environmental challenges is unknown. Here we show that the nuclear receptor Rev-erbα (also known as Nr1d1), a powerful transcriptional repressor, links circadian and thermogenic networks through the regulation of brown adipose tissue (BAT) function. Mice exposed to cold fare considerably better at 05:00 (Zeitgeber time 22) when Rev-erbα is barely expressed than at 17:00 (Zeitgeber time 10) when Rev-erbα is abundant. Deletion of Rev-erbα markedly improves cold tolerance at 17:00, indicating that overcoming Rev-erbα-dependent repression is a fundamental feature of the thermogenic response to cold. Physiological induction of uncoupling protein 1 (Ucp1) by cold temperatures is preceded by rapid downregulation of Rev-erbα in BAT. Rev-erbα represses Ucp1 in a brown-adipose-cell-autonomous manner and BAT Ucp1 levels are high in Rev-erbα-null mice, even at thermoneutrality. Genetic loss of Rev-erbα also abolishes normal rhythms of body temperature and BAT activity. Thus, Rev-erbα acts as a thermogenic focal point required for establishing and maintaining body temperature rhythm in a manner that is adaptable to environmental demands.

PET/CT imaging in cancer: current applications and future directions.

Positron emission tomography (PET) is a radiotracer imaging method that yields quantitative images of regional in vivo biology and biochemistry. PET, now used in conjunction with computed tomography (CT) in PET/CT devices, has had its greatest impact to date on cancer and is now an important part of oncologic clinical practice and translational cancer research. In this review of current applications and future directions for PET/CT in cancer, the authors first highlight the basic principles of PET followed by a discussion of the biochemistry and current clinical applications of the most commonly used PET imaging agent, 18F‐fluorodeoxyglucose (FDG). Then, emerging methods for PET imaging of other biologic processes relevant to cancer are reviewed, including cellular proliferation, tumor hypoxia, apoptosis, amino acid and cell membrane metabolism, and imaging of tumor receptors and other tumor‐specific gene products. The focus of the review is on methods in current clinical practice as well as those that have been translated to patients and are currently in clinical trials. Cancer 2014;120:3433–3445.

Localization of Parathyroid Adenomas by Sonography and Technetium Tc 99m Sestamibi Single-Photon Emission Computed Tomography Before Minimally Invasive Parathyroidectomy Are Both Studies Really Needed?

Objective. The purpose of this study was to determine the utility of radiologist‐performed sonography as the principal modality for parathyroid localization before minimally invasive parathyroidectomy. Methods. Both sonography and technetium Tc 99m sestamibi single‐photon emission computed tomography (SPECT) are commonly performed during imaging evaluation of patients with primary hyperparathyroidism (HPTH). Sonographic examinations ordered during the study period were performed by 1 author (M.E.T.), and results were immediately reported. Findings of a subsequent Tc 99m sestamibi study were recorded blinded to the sonographic results. The sensitivity and specificity of sonography and Tc 99m sestamibi SPECT were assessed with the use of surgery and pathology reports as a reference standard. The 2007 global Medicare reimbursement rates were used to assess the costs of preoperative localization. Results. Parathyroidectomy was performed in 144 of 172 patients evaluated by both modalities. The sensitivity, specificity, and positive predictive value of sonography for identifying abnormal parathyroid glands were 74%, 96%, and 90%, respectively. Sonography correctly localized a single adenoma or suggested multiglandular disease in 112 of 144 patients (78%). The sensitivity, specificity, and positive predictive value of SPECT were 58%, 96%, and 89%. Technetium 99m sestamibi SPECT correctly predicted an adenoma or multiglandular disease in 88 of 144 patients (61%). Five patients with negative sonographic findings were shown to have uniglandular disease on Tc 99m sestamibi SPECT. Selective use of Tc 99m sestamibi SPECT (ie, when sonographic findings were negative or equivocal) would have decreased the cost of imaging by 53%. Conclusions. Radiologist‐performed sonography may potentially be used as a principal imaging modality for patients with HPTH. Selective use of Tc 99m sestamibi in cases with negative or equivocal sonographic findings can decrease the cost of imaging before parathyroid resection considerably.

124 I-huA33 Antibody PET of Colorectal Cancer

Humanized A33 (huA33) is a promising monoclonal antibody that recognizes A33 antigen, which is present in more than 95% of colorectal cancers and in normal bowel. In this study, we took advantage of quantitative PET to evaluate 124I huA33 targeting, biodistribution, and safety in patients with colorectal cancer. We also determined the biodistribution of 124I-huA33 when a large dose of human intravenous IgG (IVIG) was administered to manipulate the Fc receptor or when 124I-huA33 was given via hepatic arterial infusion (HAI). Methods: We studied 25 patients with primary or metastatic colorectal cancer; 19 patients had surgical exploration or resection. Patients received a median of 343 MBq (44.4–396 MBq) and 10 mg of 124I-huA33. Nineteen patients received the antibody intravenously and 6 patients via HAI, and 5 patients also received IVIG. Results: Ten of 12 primary tumors were visualized in 11 patients. The median concentration in primary colon tumors was 0.016% injected dose per gram, compared with 0.004% in normal colon. The PET-based median ratio of hepatic tumor uptake to normal-liver uptake was 3.9 (range, 1.8–22.2). Quantitation using PET, compared with well counting of serum and tissue, showed little difference. Prominent uptake in bowel hindered tumor identification in some patients. Pharmacokinetics showed that patients receiving IVIG had a significantly shorter serum half-time (41.6 ± 14.0 h) than those without (65.2 ± 9.8 h). There were no differences in clearance rates among the intravenous group, IVIG group, and HAI group, nor was there any difference in serum area under the curve, maximum serum concentration, or volume of distribution. Weak titers of human–antihuman antibodies were observed in 6 of 25 patients. No acute side effects or significant toxicities were associated with huA33. Conclusion: Good localization of 124I-huA33 in colorectal cancer with no significant toxicity has been observed. PET-derived 124I concentrations agreed well with those obtained by well counting of surgically resected tissue and blood, confirming the quantitative accuracy of 124I-huA33 PET. The HAI route had no advantage over the intravenous route. No clinically significant changes in blood clearance were induced by IVIG.

A Phase I Trial of the HIV Protease Inhibitor Nelfinavir with Concurrent Chemoradiotherapy for Unresectable Stage IIIA/IIIB Non-small Cell Lung Cancer A Report of Toxicities and Clinical Response

Background: The objective of this phase I trial was to determine dose-limiting toxicities (DLT) and the maximally tolerated dose of the radiosensitizer Nelfinavir in combination with concurrent chemoradiotherapy in locally advanced non-small cell lung cancer (NSCLC). Methods: Nelfinavir (dose level 1: 625 mg orally [PO] twice a day; dose level 2: 1250 mg PO twice a day) was administered for 7 to 14 days before and concurrently with concurrent chemoradiotherapy to patients with biopsy confirmed IIIA or IIIB unresectable NSCLC. Five patients were treated at dose level 1; eight patients were treated at dose level 2. Patients were treated with concurrent chemoradiotherapy to a dose of 66.6 Gy. DLTs were defined as any treatment-related grade 4 hematologic toxicity requiring a break in therapy or nonhematologic grade 3 or higher toxicity except esophagitis and pneumonitis. Results: Sixteen patients were enrolled and 13 patients received at least one dose of nelfinavir. Twelve patients were treated with nelfinavir and concurrent chemoradiotherapy. No DLTs have been observed at either dose level. The maximum tolerated dose of nelfinavir was therefore 1250 mg PO twice a day. Six patients experienced grade 4 leukopenia. One patient experienced grade 4 thromobcytopenia. Median follow-up for all 12 response-evaluable patients was 31.6 months and for survivors is 23.5 months. Nine of the 12 patients had evaluable posttreatment positron emission tomography/computed tomography with metabolic response as follows: overall response: 9/9 (100%); complete response: 5/9 (56%); and partial response: 4/9 (44%). Conclusion: Nelfinavir administered with concurrent chemoradiotherapy is associated with acceptable toxicity in stage IIIA/IIIB NSCLC. The metabolic response and tumor response data suggest that nelfinavir has promising activity in this disease.

Molecular Imaging Biomarkers for Oncology Clinical Trials

Biomarkers can be used to characterize disease status or predict disease behavior. Cancer biomarkers have typically relied on assays of blood or tissue; however, molecular imaging has a promising and complementary role as a cancer biomarker. This “Focus on Molecular Imaging” article reviews the current role of biomarkers to direct cancer clinical trials and clinical practice, along with current and future cancer biomarker applications of molecular imaging.

Correlation of In Vivo and In Vitro Measures of Carbonic Anhydrase IX Antigen Expression in Renal Masses Using Antibody 124I-cG250

The goal of this study was to determine whether there is a potential correlation between quantification of radiolabeled macromolecular uptake in tumors determined in vivo using PET/CT and in vitro using autoradiography and γ-counting of tumor tissue. Methods: Twenty-six patients with renal masses scheduled for surgical resection received 124I-labeled antibody cG250. Tumor specimens obtained from resection were studied. Fifteen of these patients had clear cell cancer demonstrated by positive findings on PET/CT images and histopathology. Radioactivity in tumors was measured on PET/CT images and expressed as percentage injected dose per gram. These values were then normalized to measurements of known serum radioactivity from a venous blood sample obtained at the time of PET/CT. Comparable measurements were obtained in vitro using γ-well counting and digital autoradiography of tumor tissue. Results: There was a significant correlation between tumor radioactivity estimated in vivo and in vitro (Spearman correlation coefficient comparing normalized PET measurements with well counting of 0.84, P < 0.000001, and with autoradiography of 0.88, P < 0.000001). PET/CT measurements of tumor uptake were lower than measurements obtained with either of the in vitro methods, and digital autoradiography resulted in the highest measurements. Conclusion: PET/CT can be reliably used to quantify radiolabeled macromolecular uptake in vivo, suggesting important implications for quantitative pharmacokinetic estimates of macromolecular biodistribution.

Targeted delivery of antibody-based therapeutic and imaging agents to CNS tumors: crossing the blood--brain barrier divide

Introduction: Brain tumors are inherently difficult to treat in large part due to the cellular blood–brain barriers (BBBs) that limit the delivery of therapeutics to the tumor tissue from the systemic circulation. Virtually no large molecules, including antibody-based proteins, can penetrate the BBB. With antibodies fast becoming attractive ligands for highly specific molecular targeting to tumor antigens, a variety of methods are being investigated to enhance the access of these agents to intracranial tumors for imaging or therapeutic applications. Areas covered: This review describes the characteristics of the BBB and the vasculature in brain tumors, described as the blood–brain tumor barrier (BBTB). Antibodies targeted to molecular markers of central nervous system (CNS) tumors will be highlighted, and current strategies for enhancing the delivery of antibodies across these cellular barriers into the brain parenchyma to the tumor will be discussed. Noninvasive imaging approaches to assess BBB/BBTB permeability and/or antibody targeting will be presented as a means of guiding the optimal delivery of targeted agents to brain tumors. Expert opinion: Preclinical and clinical studies highlight the potential of several approaches in increasing brain tumor delivery across the BBB divide. However, each carries its own risks and challenges. There is tremendous potential in using neuroimaging strategies to assist in understanding and defining the challenges to translating and optimizing molecularly targeted antibody delivery to CNS tumors to improve clinical outcomes.

124I-huA33 Antibody Uptake Is Driven by A33 Antigen Concentration in Tissues from Colorectal Cancer Patients Imaged by Immuno-PET

The primary aim of this analysis was to examine the quantitative features of antibody–antigen interactions in tumors and normal tissue after parenteral administration of antitumor antibodies to human patients. Methods: Humanized anti-A33 antibody (10 mg) labeled with the positron-emitting radionuclide 124I (124I-huA33) was injected intravenously in 15 patients with colorectal cancer. Clinical PET/CT was performed approximately 1 wk later, followed by a detailed assay of surgically removed tissue specimens including radioactivity counting, autoradiography, immunohistochemistry, and antigen density determination. Results: PET/CT showed high levels of antibody targeting in tumors and normal bowel. In tissue specimens, the spatial distribution of 124I-huA33 conformed to that of A33 antigen, and there was a linear relationship between the amount of bound antibody and antigen concentration. Antibody uptake was high in 1- to 2-mm regions of antigen-positive tumor cells (mean, ∼0.05 percentage injected dose per gram) and in antigen-positive normal colonic mucosa (mean, ∼0.03 percentage injected dose per gram). The estimated binding site occupancy for tumor and normal colon was 20%–50%. Conclusion: The in vivo biodistribution of 124I-huA33 in human patients 1 wk after antibody administration was determined by A33 antigen expression. Our data imply that the optimal strategy for A33-based radioimmunotherapy of colon cancer will consist of a multistep treatment using a radionuclide with short-range (α- or β-particle) emissions.