ABSTRACT

The PDGF family members are potent mitogens for cells of mesenchymal origin and serve as important regulators of cell migration, survival, apoptosis, and transformation. Tumor-derived PDGF ligands are thought to function in both autocrine and paracrine manners, activating receptors on tumor and surrounding stromal cells. PDGF-C and -D are secreted as latent dimers, unlike PDGF-A and -B. Cleavage of the CUB domain from the PDGF-C and -D dimers is required for their biological activity. At present, little is known about the proteolytic processing of PDGF-C, the rate-limiting step in the regulation of PDGF-C activity. Here we show that the breast carcinoma cell line, MCF7, engineered to overexpress PDGF-C, produces proteases capable of cleaving PDGF-C to its active form. Increased PDGF-C expression enhances cell proliferation, anchorage independent cell growth, and tumor cell motility by autocrine signaling. In addition, MCF7-produced PDGF-C induces fibroblast cell migration in a paracrine manner. Interestingly, PDGF-C enhances tumor cell invasion in the presence of fibroblast, suggesting a role of tumor-derived PDGF-C in tumor-stromal interactions. In the present study, we identify tissue plasminogen activator (tPA) and matriptase as major proteases for processing of PDGF-C in MCF7 cells. In in vitro studies, we also show that urokinase plasminogen activator (uPA) is able to process PDGF-C. Furthermore, by site-directed mutagenesis, we identify the cleavage site for these proteases in PDGF-C. Lastly, we provide evidence suggesting a 2-step proteolytic processing of PDGF-C involving creation of a hemidimer, followed by growth factor domain dimer (GFD-D) generation.

Purpose The acquisition of multiparametric quantitative magnetic resonance imaging (qMRI) is becoming increasingly important for functional characterization of cancer prior to‐ and throughout the course of radiation therapy. The feasibility of a qMRI method known as magnetic resonance fingerprinting (MRF) for rapid T1 and T2 mapping was assessed on a low‐field MR‐linac system. Methods A three‐dimensional MRF sequence was implemented on a 0.35T MR‐guided radiotherapy system. MRF‐derived measurements of T1 and T2 were compared to those obtained with gold standard single spin echo methods, and the impacts of the radiofrequency field homogeneity and scan times ranging between 6 and 48 min were analyzed by acquiring between 1 and 8 spokes per time point in a standard quantitative system phantom. The short‐term repeatability of MRF was assessed over three measurements taken over a 10‐h period. To evaluate transferability, MRF measurements were acquired on two additional MR‐guided radiotherapy systems. Preliminary human volunteer studies were performed. Results The phantom benchmarking studies showed that MRF is capable of mapping T1 and T2 values within 8% and 10% of gold standard measures, respectively, at 0.35T. The coefficient of variation of T1 and T2 estimates over three repeated scans was < 5% over a broad range of relaxation times. The T1 and T2 times derived using a single‐spoke MRF acquisition across three scanners were near unity and mean percent errors in T1 and T2 estimates using the same phantom were < 3%. The mean percent differences in T1 and T2 as a result of truncating the scan time to 6 min over the large range of relaxation times in the system phantom were 0.65% and 4.05%, respectively. Conclusions The technical feasibility and accuracy of MRF on a low‐field MR‐guided radiation therapy device has been demonstrated. MRF can be used to measure accurate T1 and T2 maps in three dimensions from a brief 6‐min scan, offering strong potential for efficient and reproducible qMRI for future clinical trials in functional plan adaptation and tumor/normal tissue response assessment.

Purpose: Magnetic resonance-guided radiation therapy (MRgRT) has shown great promise for localization and real-time tumor monitoring. However, to date, quantitative imaging has been limited for low field MRgRT. This work benchmarks quantitative T1, R2*, and Proton Density (PD)mapping in a phantom on a 0.35 T MR-linac and implements a novel acquisition method, STrategically Acquired Gradient Echo (STAGE). To further validate STAGE in a clinical setting, a pilot study was undertaken in a cohort of brain tumor patients to elucidate opportunities for longitudinal functional imaging with an MR-linac in the brain. Methods: STAGE (two triple-echo gradient echo (GRE) acquisitions) was optimized for a 0.35T low-field MR-linac. Simulations were performed to choose two flip angles to optimize signal-tonoise ratio (SNR) and T1-mapping precision. Tradeoffs between SNR, scan time, and spatial resolution for whole-brain coverage were evaluated in healthy volunteers. Data were inputted into a STAGE processing pipeline to yield four qualitative images (T1-weighted, enhanced T1-weighted, protondensity (PD) weighted, and simulated FLuid-Attenuated Inversion Recovery (sFLAIR)), and three quantitative datasets (T1, PD, and R2*). A benchmarking ISMRM/NIST phantom consisting of vials with variable NiCl 2 and MnCl 2 concentrations was scanned using variable flip angles (VFA) (2-60 degrees) and inversion recovery (IR) methods at 0.35 T. STAGE and VFA T1 values of vials were compared to IR T1 values. As measures of agreement with reference values and repeatability, relative error (RE) and coefficient of variability (CV) were calculated, respectively, for quantitative MR values within the phantom vials (spheres). To demonstrate feasibility, longitudinal STAGE data (pretreatment, weekly, and~2 months post-treatment) were acquired in an IRB-approved pilot study of brain tumor cases via the generation of temporal and differential quantitative MRI maps. Results: In the phantom, RE of measured VFA T1 and STAGE relative to IR reference values were 7.0 AE 2.5% and 9.5 AE 2.2% respectively. RE for the PD vials was 8.1 AE 6.8% and CV for phantom R2* measurements was 10.1 AE 9.9%. Simulations and volunteer experiments yielded final STAGE parameters of FA = 50°/10°, 1 9 1 9 3 mm 3 resolution, TR = 40 ms, TE = 5/20/34 ms in 10 min (64 slices). In the pilot study of brain tumor patients, differential maps for R2* and T1 maps were sensitive to local tumor changes and appeared similar to 3 T follow-up MRI datasets. Conclusion: Quantitative T1, R2*, and PD mapping are promising at 0.35 T agreeing well with reference data. STAGE phantom data offer quantitative representations comparable to traditional methods in a fraction of the acquisition time. Initial feasibility of implementing STAGE at 0.35 T in a patient brain tumor cohort suggests that detectable changes can be observed over time. With confirmation in a larger cohort, results may be implemented to identify areas of recurrence and facilitate adaptive radiation therapy.

Conclusion: The interobserver variation of target volumes delineation of OPC patient based on MR is large. For the GTV, volume differences up to a factor of 10 were observed. For the CTV, differences up to of factor of 8 were observed.This demonstrates the need for international MR delineation guidelines for oropharyngeal cancer patients treated using MR-guided radiotherapy.