Detection of Treatment Response in Triple-Negative Breast Tumors to Paclitaxel Using MRI Cell Size Imaging.

BACKGROUND: Breast cancer treatment response evaluation using the response evaluation criteria in solid tumors (RECIST) guidelines, based on tumor volume changes, has limitations, prompting interest in novel imaging markers for accurate therapeutic effect determination. PURPOSE: To use MRI-measured cell size as a new imaging biomarker for assessing chemotherapy response in breast cancer. STUDY TYPE: Longitudinal; animal model. STUDY POPULATION: Triple-negative human breast cancer cell (MDA-MB-231) pellets (4 groups, n = 7) treated with dimethyl sulfoxide (DMSO) or 10 nM of paclitaxel for 24, 48, and 96 hours, and 29 mice with MDA-MB-231 tumors in right hind limbs treated with paclitaxel (n = 16) or DMSO (n = 13) twice weekly for 3 weeks. FIELD STRENGTH/SEQUENCE: Oscillating gradient spin echo and pulsed gradient spin echo sequences at 4.7 T. ASSESSMENT: MDA-MB-231 cells were analyzed using flowcytometry and light microscopy to assess cell cycle phases and cell size distribution. MDA-MB-231 cell pellets were MR imaged. Mice were imaged weekly, with 9, 6, and 14 being sacrificed for histology after MRI at weeks 1, 2, and 3, respectively. Microstructural parameters of tumors/cell pellets were derived by fitting diffusion MRI data to a biophysical model. STATISTICAL TESTS: One-way ANOVA compared cell sizes and MR-derived parameters between treated and control samples. Repeated measures 2-way ANOVA with Bonferroni post-tests compared temporal changes in MR-derived parameters. A P-value <0.05 was considered statistically significant. RESULTS: In vitro experiments showed that the mean MR-derived cell sizes of paclitaxel-treated cells increased significantly with a 24-hours treatment and decreased (P = 0.06) with a 96-hour treatment. For in vivo xenograft experiments, the paclitaxel-treated tumors showed significant decreases in cell size at later weeks. MRI observations were supported by flowcytometry, light microscopy, and histology. DATA CONCLUSIONS: MR-derived cell size may characterize the cell shrinkage during treatment-induced apoptosis, and may potentially provide new insights into the assessment of therapeutic response. LEVEL OF EVIDENCE: 2 TECHNICAL EFFICACY STAGE: 4.

Evaluation of contributors to amide proton transfer-weighted imaging and nuclear Overhauser enhancement-weighted imaging contrast in tumors at a high magnetic field.

PURPOSE: The purpose is to evaluate the relative contribution from confounding factors (T1 weighting and magnetization transfer) to the CEST ratio (CESTR)-quantified amide proton transfer (APT) and nuclear Overhauser enhancement (NOE) (-3.5) in tumors as well as whether the CESTR can reflect the distribution of the solute concentration (fs ). METHODS: We first provided a signal model that shows the separate dependence of CESTR on these confounding factors and the clean CEST/NOE effects quantified by an apparent exchange-dependent relaxation (AREX) method. We then measured the change in these effects in the 9-L tumor model in rats, through which we calculated the relative contribution of each confounding factor. fs was also fitted, and its correlations with the CESTR and AREX were assessed to evaluate their capabilities to reflect fs . RESULTS: The CESTR-quantified APT shows "positive" contrast in tumors, which arises primarily from R1w at low powers and both R1w and magnetization transfer at high powers. CESTR-quantified NOE (-3.5) shows no or weak contrast in tumors, which is due to the cancelation of R1w and NOE (-3.5), which have opposite contributions. CESTR-quantified APT has a stronger correlation with APT fs than AREX-quantified APT. CESTR-quantified NOE (-3.5) has a weaker correlation with NOE (-3.5) fs than AREX-quantified NOE (-3.5). CONCLUSION: CESTR reflects a combined effect of T1 weighting and CEST/NOE. Both factors depend on fs , which contributes positively to the dependence of CESTR on fs in APT imaging and enhances its correlation with fs . In contrast, these factors have opposite contributions to its dependence on fs in NOE (-3.5) imaging, thereby weakening the correlation.

Diffusion time dependency of extracellular diffusion.

PURPOSE: To quantify the variations of the power-law dependences on diffusion time t or gradient frequency f $$ f $$ of extracellular water diffusion measured by diffusion MRI (dMRI). METHODS: Model cellular systems containing only extracellular water were used to investigate the t / f $$ t/f $$ dependence of D ex $$ {D}_{ex} $$ , the extracellular diffusion coefficient. Computer simulations used a randomly packed tissue model with realistic intracellular volume fractions and cell sizes. DMRI measurements were performed on samples consisting of liposomes containing heavy water(D2 O, deuterium oxide) dispersed in regular water (H2 O). D ex $$ {D}_{ex} $$ was obtained over a broad t $$ t $$ range (∼1-1000 ms) and then fit power-law equations D ex ( t ) = D const + const · t - ϑ t $$ {D}_{ex}(t)={D}_{\mathrm{const}}+\mathrm{const}\cdotp {t}^{-{\vartheta}_t} $$ and D ex ( f ) = D const + const · f ϑ f $$ {D}_{ex}(f)={D}_{\mathrm{const}}+\mathrm{const}\cdotp {f}^{\vartheta_f} $$ . RESULTS: Both simulated and experimental results suggest that no single power-law adequately describes the behavior of D ex $$ {D}_{ex} $$ over the range of diffusion times of most interest in practical dMRI. Previous theoretical predictions are accurate over only limited t $$ t $$ ranges; for example, θ t = θ f = - 1 2 $$ {\theta}_t={\theta}_f=-\frac{1}{2} $$ is valid only for short times, whereas θ t = 1 $$ {\theta}_t=1 $$ or θ f = 3 2 $$ {\theta}_f=\frac{3}{2} $$ is valid only for long times but cannot describe other ranges simultaneously. For the specific t $$ t $$ range of 5-70 ms used in typical human dMRI measurements, θ t = θ f = 1 $$ {\theta}_t={\theta}_f=1 $$ matches the data well empirically. CONCLUSION: The optimal power-law fit of extracellular diffusion varies with diffusion time. The dependency obtained at short or long t $$ t $$ limits cannot be applied to typical dMRI measurements in human cancer or liver. It is essential to determine the appropriate diffusion time range when modeling extracellular diffusion in dMRI-based quantitative microstructural imaging.

Towards differentiation of brain tumor from radiation necrosis using multi-parametric MRI: Preliminary results at 4.7 T using rodent models.

BACKGROUND: It remains a clinical challenge to differentiate brain tumors from radiation-induced necrosis in the brain. Despite significant improvements, no single MRI method has been validated adequately in the clinical setting. METHODS: Multi-parametric MRI (mpMRI) was performed to differentiate 9L gliosarcoma from radiation necrosis in animal models. Five types of MRI methods probed complementary information on different scales i.e., T2 (relaxation), CEST based APT (probing mobile proteins/peptides) and rNOE (mobile macromolecules), qMT (macromolecules), diffusion based ADC (cell density) and SSIFT iAUC (cell size), and perfusion based DSC (blood volume and flow). RESULTS: For single MRI parameters, iAUC and ADC provide the best discrimination of radiation necrosis and brain tumor. For mpMRI, a combination of iAUC, ADC, and APT shows the best classification performance based on a two-step analysis with the Lasso and Ridge regressions. CONCLUSION: A general mpMRI approach is introduced to choosing candidate multiple MRI methods, identifying the most effective parameters from all the mpMRI parameters, and finding the appropriate combination of chosen parameters to maximize the classification performance to differentiate tumors from radiation necrosis.

Rodent Model of Brain Radionecrosis Using Clinical LINAC-Based Stereotactic Radiosurgery.

Purpose: Our purpose was to develop a rodent model of brain radionecrosis using clinical linear accelerator based stereotactic radiosurgery. Methods and Materials: Single fraction maximum prescription points in the mouse's left hemisphere were irradiated using linear accelerator-based stereotactic radiosurgery with multiple arcs at 60 (n = 5), 100 (n = 5), and 140 (n = 5) Gy. Rats (n = 6) were similarly treated with 140 Gy. Gadolinium (Gd)-enhanced magnetic resonance imaging (MRI) was used to track radiation injury in mice over weeks (100 and 140 Gy) or months (60 Gy). Target accuracy was measured by the distance from the prescription point to the center of the earliest Gd-MRI enhancement. Confirmation of necrosis via histology was performed at the subject endpoints. Results: Radiation injury as indicated by Gd-MRI was first identified at 2 weeks (140 Gy), 4 to 6 weeks (100 Gy), and 8 months (60 Gy). A volumetric time course showed rapid growth in the volume of Gd-MRI signal enhancement after the appearance of apparent necrosis. Histopathologic features were consistent with radionecrosis. Conclusions: The presented method uses a commonly available clinical linear accelerator to induce radiation necrosis in both mice and rats. The treatment is modeled after patient therapy for a more direct model of human tissue under a range of doses used in clinical neuro-ablation techniques. The short time to onset of apparent necrosis, accurate targeting of the prescription point, high incidence of necrosis, and similar pathologic features make this a suitable animal model for further research in radionecrosis.

Selective Cell Size MRI Differentiates Brain Tumors from Radiation Necrosis.

Brain metastasis is a common characteristic of late-stage lung cancers. High doses of targeted radiotherapy can control tumor growth in the brain but can also result in radiotherapy-induced necrosis. Current methods are limited for distinguishing whether new parenchymal lesions following radiotherapy are recurrent tumors or radiotherapy-induced necrosis, but the clinical management of these two classes of lesions differs significantly. Here, we developed, validated, and evaluated a new MRI technique termed selective size imaging using filters via diffusion times (SSIFT) to differentiate brain tumors from radiotherapy necrosis in the brain. This approach generates a signal filter that leverages diffusion time dependence to establish a cell size-weighted map. Computer simulations in silico, cultured cancer cells in vitro, and animals with brain tumors in vivo were used to comprehensively validate the specificity of SSIFT for detecting typical large cancer cells and the ability to differentiate brain tumors from radiotherapy necrosis. SSIFT was also implemented in patients with metastatic brain cancer and radiotherapy necrosis. SSIFT showed high correlation with mean cell sizes in the relevant range of less than 20 µm. The specificity of SSIFT for brain tumors and reduced contrast in other brain etiologies allowed SSIFT to differentiate brain tumors from peritumoral edema and radiotherapy necrosis. In conclusion, this new, cell size-based MRI method provides a unique contrast to differentiate brain tumors from other pathologies in the brain. SIGNIFICANCE: This work introduces and provides preclinical validation of a new diffusion MRI method that exploits intrinsic differences in cell sizes to distinguish brain tumors and radiotherapy necrosis.

Improving MR cell size imaging by inclusion of transcytolemmal water exchange.

The goal of the current study is to include transcytolemmal water exchange in MR cell size imaging using the IMPULSED model for more accurate characterization of tissue cellular properties (e.g., apparent volume fraction of intracellular space v in ) and quantification of indicators of transcytolemmal water exchange. We propose a heuristic model that incorporates transcytolemmal water exchange into a multicompartment diffusion-based method (IMPULSED) that was developed previously to extract microstructural parameters (e.g., mean cell size d and apparent volume fraction of intracellular space v in ) assuming no water exchange. For t diff ≤ 5 ms, the water exchange can be ignored, and the signal model is the same as the IMPULSED model. For t diff ≥ 30 ms, we incorporated the modified Kärger model that includes both restricted diffusion and exchange between compartments. Using simulations and previously published in vitro cell data, we evaluated the accuracy and precision of model-derived parameters and determined how they are dependent on SNR and imaging parameters. The joint model provides more accurate d values for cell sizes ranging from 10 to 12 microns when water exchange is fast (e.g., intracellular water pre-exchange lifetime τ in ≤ 100 ms) than IMPULSED, and reduces the bias of IMPULSED-derived estimates of v in , especially when water exchange is relatively slow (e.g., τ in > 200 ms). Indicators of transcytolemmal water exchange derived from the proposed joint model are linearly correlated with ground truth τ in values and can detect changes in cell membrane permeability induced by saponin treatment in murine erythroleukemia cancer cells. Our results suggest this joint model not only improves the accuracy of IMPULSED-derived microstructural parameters, but also provides indicators of water exchange that are usually ignored in diffusion models of tissues.

Noninvasive Capsulotomy for Refractory Depression by Frameless Stereotactic Radiosurgery.

PURPOSE: Effective treatment options for refractory depression are needed. Recent advancements permit both precise ablative radiation and functional neurologic connectome analysis using standard magnetic resonance imaging. We combined these innovations to perform stereotactic radiosurgical capsulotomy for the treatment of medically refractory major depressive disorder and study connectome response using a novel tractography-based approach. METHODS AND MATERIALS: Patients with medically refractory depression were enrolled on a prospective pilot single-arm observational trial from 2020 to 2021 at a single academic tertiary referral center. Bilateral ablation of the anterior limb of the internal capsule was accomplished by mask-based linear accelerator stereotactic radiosurgery. Beck's Depression Inventory measured efficacy. Montreal Cognitive Assessment evaluated cognition. RESULTS: Three patients were enrolled. Depression burden was improved by 88% at 12-month follow-up and by 55% at 18-month follow-up for patient 1 and 2, respectively. Patient 1 discontinued ketamine therapy, and patient 2 discontinued electroconvulsive therapy. Patient 3 reported global improvement in symptoms and function at 3 months. All 3 patients had reduction or resolution of suicidal ideation. No patient experienced cognitive decline or neurologic toxicity, and Montreal Cognitive Assessment score, as well as subjective patient-reported evaluations of concentration and attention, were superior after treatment. Tractography confirmed intended disruption of the cortico-striatal-thalamo-cortical loop with structural reorganization in the connectome. Connectome change was consistent between patients. Observed increases in caudate and putamen connectivity and decreases in thalamic connectivity may explain improved concentration, attention, and depression. The diversity and magnitude of connectome change may correlate with degree of clinical response. CONCLUSIONS: In 3 patients with refractory depression, radiosurgical capsulotomy significantly reduced the burden of depression. Functional connectome reorganization offers neurobiological evidence to support further investigations of the role of radiosurgery in depression.

Disposable point-of-care portable perfusion phantom for quantitative DCE-MRI.

PURPOSE: To develop a disposable point-of-care portable perfusion phantom (DP4) and validate its clinical utility in a multi-institutional setting for quantitative dynamic contrast-enhanced magnetic resonance imaging (qDCE-MRI). METHODS: The DP4 phantom was designed for single-use and imaged concurrently with a human subject so that the phantom data can be utilized as the reference to detect errors in qDCE-MRI measurement of human tissues. The change of contrast-agent concentration in the phantom was measured using liquid chromatography-mass spectrometry. The repeatability of the contrast enhancement curve (CEC) was assessed with five phantoms in a single MRI scanner. Five healthy human subjects were recruited to evaluate the reproducibility of qDCE-MRI measurements. Each subject was imaged concurrently with the DP4 phantom at two institutes using three 3T MRI scanners from three different vendors. Pharmacokinetic (PK) parameters in the regions of liver, spleen, pancreas, and paravertebral muscle were calculated based on the Tofts model (TM), extended Tofts model (ETM), and shutter speed model (SSM). The reproducibility of each PK parameter over three measurements was evaluated with the intraclass correlation coefficient (ICC) and compared before and after DP4-based error correction. RESULTS: The contrast-agent concentration in the DP4 phantom was linearly increased over 10 min (0.17 mM/min, measurement accuracy: 96%) after injecting gadoteridol (100 mM) at a constant rate (0.24 ml/s, 4 ml). The repeatability of the CEC within the phantom was 0.997 when assessed by the ICC. The reproducibility of the volume transfer constant, Ktrans , was the highest of the PK parameters regardless of the PK models. The ICCs of Ktrans in the TM, ETM, and SSM before DP4-based error correction were 0.34, 0.39, and 0.72, respectively, while those increased to 0.93, 0.98, and 0.86, respectively, after correction. CONCLUSIONS: The DP4 phantom is reliable, portable, and capable of significantly improving the reproducibility of qDCE-MRI measurements.

MR cell size imaging with temporal diffusion spectroscopy.

Cytological features such as cell size and intracellular morphology provide fundamental information on cell status and hence may provide specific information on changes that arise within biological tissues. Such information is usually obtained by invasive biopsy in current clinical practice, which suffers several well-known disadvantages. Recently, novel MRI methods such as IMPULSED (imaging microstructural parameters using limited spectrally edited diffusion) have been developed for direct measurements of mean cell size non-invasively. The IMPULSED protocol is based on using temporal diffusion spectroscopy (TDS) to combine measurements of water diffusion over a wide range of diffusion times to probe cellular microstructure over varying length scales. IMPULSED has been shown to provide rapid, robust, and reliable mapping of mean cell size and is suitable for clinical imaging. More recently, cell size distributions have also been derived by appropriate analyses of data acquired with IMPULSED or similar sequences, which thus provides MRI-cytometry. This review summarizes the basic principles, practical implementations, validations, and example applications of MR cell size imaging based on TDS and demonstrates how cytometric information can be used in various applications. In addition, the limitations and potential future directions of MR cytometry are identified including the diagnosis of nonalcoholic steatohepatitis of the liver and the assessment of treatment response of cancers.

MRI-cytometry: Mapping nonparametric cell size distributions using diffusion MRI.

PURPOSE: This report introduces and validates a new diffusion MRI-based method, termed MRI-cytometry, which can noninvasively map intravoxel, nonparametric cell size distributions in tissues. METHODS: MRI was used to acquire diffusion MRI signals with a range of diffusion times and gradient factors, and a model was fit to these data to derive estimates of cell size distributions. We implemented a 2-step fitting method to avoid noise-induced artificial peaks and provide reliable estimates of tumor cell size distributions. Computer simulations in silico, experimental measurements on cultured cells in vitro, and animal xenografts in vivo were used to validate the accuracy and precision of the method. Tumors in 7 patients with breast cancer were also imaged and analyzed using this MRI-cytometry approach on a clinical 3 Tesla MRI scanner. RESULTS: Simulations and experimental results confirm that MRI-cytometry can reliably map intravoxel, nonparametric cell size distributions and has the potential to discriminate smaller and larger cells. The application in breast cancer patients demonstrates the feasibility of direct translation of MRI-cytometry to clinical applications. CONCLUSION: The proposed MRI-cytometry method can characterize nonparametric cell size distributions in human tumors, which potentially provides a practical imaging approach to derive specific histopathological information on biological tissues.

MRI of tumor T cell infiltration in response to checkpoint inhibitor therapy.

BACKGROUND: Immune checkpoint inhibitors, the most widespread class of immunotherapies, have demonstrated unique response patterns that are not always adequately captured by traditional response criteria such as the Response Evaluation Criteria in Solid Tumors or even immune-specific response criteria. These response metrics rely on monitoring tumor growth, but an increase in tumor size and/or appearance after starting immunotherapy does not always represent tumor progression, but also can be a result of T cell infiltration and thus positive treatment response. Therefore, non-invasive and longitudinal monitoring of T cell infiltration are needed to assess the effects of immunotherapies such as checkpoint inhibitors. Here, we proposed an innovative concept that a sufficiently large influx of tumor infiltrating T cells, which have a smaller diameter than cancer cells, will change the diameter distribution and decrease the average size of cells within a volume to a degree that can be quantified by non-invasive MRI. METHODS: We validated our hypothesis by studying tumor response to combination immune-checkpoint blockade (ICB) of anti-PD-1 and anti-CTLA4 in a mouse model of colon adenocarcinoma (MC38). The response was monitored longitudinally using Imaging Microstructural Parameters Using Limited Spectrally Edited Diffusion (IMPULSED), a diffusion MRI-based method which has been previously shown to non-invasively map changes in intracellular structure and cell sizes with the spatial resolution of MRI, in cell cultures and in animal models. Tumors were collected for immunohistochemical and flow cytometry analyzes immediately after the last imaging session. RESULTS: Immunohistochemical analysis revealed that increased T cell infiltration of the tumors results in a decrease in mean cell size (eg, a 10% increase of CD3+ T cell fraction results a ~1 µm decrease in the mean cell size). IMPULSED showed that the ICB responders, mice with tumor volumes were less than 250 mm3 or had tumors with stable or decreased volumes, had significantly smaller mean cell sizes than both Control IgG-treated tumors and ICB non-responder tumors. CONCLUSIONS: IMPULSED-derived cell size could potentially serve as an imaging marker for differentiating responsive and non-responsive tumors after checkpoint inhibitor therapies, a current clinical challenge that is not solved by simply monitoring tumor growth.

Mapping hepatocyte size in vivo using temporal diffusion spectroscopy MRI.

PURPOSE: The goal of this study is to implement a noninvasive method for in vivo mapping of hepatocyte size. This method will have a broad range of clinical and preclinical applications, as pathological changes in hepatocyte sizes are relevant for the accurate diagnosis and assessments of treatment response of liver diseases. METHODS: Building on the concepts of temporal diffusion spectroscopy in MRI, a clinically feasible imaging protocol named IMPULSED (Imaging Microstructural Parameters Using Limited Spectrally Edited Diffusion) has been developed, which is able to report measurements of cell sizes noninvasively. This protocol acquires a selected set of diffusion imaging data and fits them to a model of water compartments in tissues to derive robust estimates of the cellular structures that restrict free diffusion. Here, we adapt and further develop this approach to measure hepatocyte sizes in vivo. We validated IMPULSED in livers of mice and rats and implemented it to image healthy human subjects using a clinical 3T MRI scanner. RESULTS: The IMPULSED-derived mean hepatocyte sizes for rats and mice are about 15-20 µm and agree well with histological findings. Maps of mean hepatocyte size for humans can be achieved in less than 15 minutes, a clinically feasible scan time. CONCLUSION: Our results suggest that this method has potential to overcome major limitations of liver biopsy and provide noninvasive mapping of hepatocyte sizes in clinical applications.

Relayed nuclear Overhauser enhancement sensitivity to membrane Cho phospholipids.

PURPOSE: Phospholipids are key constituents of cell membranes and serve vital functions in the regulation of cellular processes; thus, a method for in vivo detection and characterization could be valuable for detecting changes in cell membranes that are consequences of either normal or pathological processes. Here, we describe a new method to map the distribution of partially restricted phospholipids in tissues. METHODS: The phospholipids were measured by signal changes caused by relayed nuclear Overhauser enhancement-mediated CEST between the phospholipid Cho headgroup methyl protons and water at around -1.6 ppm from the water resonance. The biophysical basis of this effect was examined by controlled manipulation of head group, chain length, temperature, degree of saturation, and presence of cholesterol. Additional experiments were performed on animal tumor models to evaluate potential applications of this novel signal while correcting for confounding contributions. RESULTS: Negative relayed nuclear Overhauser dips in Z-spectra were measured from reconstituted Cho phospholipids with cholesterol but not for other Cho-containing metabolites or proteins. Significant contrast was found between tumor and contralateral normal tissue signals in animals when comparing both the measured saturation transfer signal and a more specific imaging metric. CONCLUSION: We demonstrated specific relayed nuclear Overhauser effects in partially restricted phospholipid phantoms and similar effects in solid brain tumors after correcting for confounding signal contributions, suggesting possible translational applications of this novel molecular imaging method, which we name restricted phospholipid transfer.

Magnetic resonance imaging of mean cell size in human breast tumors.

PURPOSE: Cell size is a fundamental characteristic of all tissues, and changes in cell size in cancer reflect tumor status and response to treatments, such as apoptosis and cell-cycle arrest. Unfortunately, cell size can currently be obtained only by pathological evaluation of tumor tissue samples obtained invasively. Previous imaging approaches are limited to preclinical MRI scanners or require relatively long acquisition times that are impractical for clinical imaging. There is a need to develop cell-size imaging for clinical applications. METHODS: We propose a clinically feasible IMPULSED (imaging microstructural parameters using limited spectrally edited diffusion) approach that can characterize mean cell sizes in solid tumors. We report the use of a combination of pulse sequences, using different gradient waveforms implemented on clinical MRI scanners and analytical equations based on these waveforms to analyze diffusion-weighted MRI signals and derive specific microstructural parameters such as cell size. We also describe comprehensive validations of this approach using computer simulations, cell experiments in vitro, and animal experiments in vivo and demonstrate applications in preoperative breast cancer patients. RESULTS: With fast acquisitions (~7 minutes), IMPULSED can provide high-resolution (1.3 mm in-plane) mapping of mean cell size of human tumors in vivo on clinical 3T MRI scanners. All validations suggest that IMPULSED provides accurate and reliable measurements of mean cell size. CONCLUSION: The proposed IMPULSED method can assess cell-size variations in tumors of breast cancer patients, which may have the potential to assess early response to neoadjuvant therapy.

In vivo magnetic resonance imaging of treatment-induced apoptosis.

Imaging apoptosis could provide an early and specific means to monitor tumor responses to treatment. To date, despite numerous attempts to develop molecular imaging approaches, there is still no widely-accepted and reliable method for in vivo imaging of apoptosis. We hypothesized that the distinct cellular morphologic changes associated with treatment-induced apoptosis, such as cell shrinkage, cytoplasm condensation, and DNA fragmentation, can be detected by temporal diffusion spectroscopy imaging (TDSI). Cetuximab-induced apoptosis was assessed in vitro and in vivo with cetuximab-sensitive (DiFi) and insensitive (HCT-116) human colorectal cancer cell lines by TDSI. TDSI findings were complemented by flow cytometry and immunohistochemistry. Cell cycle analysis and flow cytometry detected apoptotic cell shrinkage in cetuximab-treated DiFi cells, and significant apoptosis was confirmed by histology. TDSI-derived parameters quantified key morphological changes including cell size decreases during apoptosis in responsive tumors that occurred earlier than gross tumor volume regression. TDSI provides a unique measurement of apoptosis by identifying cellular characteristics, particularly cell shrinkage. The method will assist in understanding the underlying biology of solid tumors and predict tumor response to therapies. TDSI is free of any exogenous agent or radiation, and hence is very suitable to be incorporated into clinical applications.

Quantitative temporal diffusion spectroscopy as an early imaging biomarker of radiation therapeutic response in gliomas: A preclinical proof of concept.

PURPOSE: This study aims to test the ability of quantitative temporal diffusion spectroscopy (qTDS) to assess cellular changes associated with radiation-induced cell death in a rat glioma model. METHODS AND MATERIALS: Tumor response to a single fraction of 20 Gy of x-ray radiation was investigated in a rat glioma (9L) model. Tumor response was monitored longitudinally at postinoculation days 21, 23, and 25, using a specific implementation of qTDS with acronym IMPULSED (Imaging Microstructural Parameters Using Limited Spectrally Edited Diffusion), as well as conventional diffusion and high-resolution anatomic imaging. IMPULSED method combines diffusion-weighted signals acquired over a range of diffusion times that are then analyzed and interpreted using a theoretical model of water diffusion in tissues, which generates parametric maps depicting cellular and subcellular structural information on a voxel-wise basis. Results from different metrics were compared statistically. RESULTS: A single dose of 20 Gy x-ray radiation significantly prolonged survival of 9L-bearing rats. The mean cell sizes of irradiated tumors decreased (P < .005) after radiation treatment, which we associate with cell shrinkage and the formation of small cellular bodies during apoptosis and necrosis. A combination of IMPULSED-derived parameters (mean cell size d and extracellular structural parameter ß ex ) separated 90% of irradiated tumors from the nonirradiated cases at post inoculation day 23, whereas a combination of tumor growth and conventional apparent diffusion coefficient did not differentiate irradiated tumors from nonirradiated tumors. CONCLUSIONS: This proof-of-concept study demonstrates the IMPULSED method to be a new method for deriving quantitative microstructural parameters in a preclinical tumor model. The method provides unique information based on the diffusion time dependency of diffusion magnetic resonance imaging, which cannot be obtained by conventional diffusion weighted imaging methods, and the results have a close correlation with primary biologic markers of treatment efficacy, such as cell death and survival.

Translating preclinical MRI methods to clinical oncology.

The complexity of modern in vivo magnetic resonance imaging (MRI) methods in oncology has dramatically changed in the last 10 years. The field has long since moved passed its (unparalleled) ability to form images with exquisite soft-tissue contrast and morphology, allowing for the enhanced identification of primary tumors and metastatic disease. Currently, it is not uncommon to acquire images related to blood flow, cellularity, and macromolecular content in the clinical setting. The acquisition of images related to metabolism, hypoxia, pH, and tissue stiffness are also becoming common. All of these techniques have had some component of their invention, development, refinement, validation, and initial applications in the preclinical setting using in vivo animal models of cancer. In this review, we discuss the genesis of quantitative MRI methods that have been successfully translated from preclinical research and developed into clinical applications. These include methods that interrogate perfusion, diffusion, pH, hypoxia, macromolecular content, and tissue mechanical properties for improving detection, staging, and response monitoring of cancer. For each of these techniques, we summarize the 1) underlying biological mechanism(s); 2) preclinical applications; 3) available repeatability and reproducibility data; 4) clinical applications; and 5) limitations of the technique. We conclude with a discussion of lessons learned from translating MRI methods from the preclinical to clinical setting, and a presentation of four fundamental problems in cancer imaging that, if solved, would result in a profound improvement in the lives of oncology patients. Level of Evidence: 5 Technical Efficacy: Stage 3 J. Magn. Reson. Imaging 2019;50:1377-1392.

Measurement of APT using a combined CERT-AREX approach with varying duty cycles.

The goal is to develop an imaging method where contrast reflects amide-water magnetization exchange, with minimal signal contributions from other sources. Conventional chemical exchange saturation transfer (CEST) imaging of amides (often called amide proton transfer, or APT, and quantified by the metric MTRasym) is confounded by several factors unrelated to amides, such as aliphatic protons, water relaxation, and macromolecular magnetization transfer. In this work, we examined the effects of combining our previous chemical exchange rotation (CERT) approach with the non-linear AREX method while using different duty cycles (DC) for the label and reference scans. The dependencies of this approach, named AREXdouble,vdc, on tissue parameters, including T1, T2, semi-solid component concentration (fm), relayed nuclear Overhauser enhancement (rNOE), and nearby amines, were studied through numerical simulations and control sample experiments at 9.4T and 1µT irradiation. Simulations and experiments show that AREXdouble,vdc is sensitive to amide-water exchange effects, but is relatively insensitive to T1, T2, fm, nearby amine, and distant aliphatic protons, while the conventional metric MTRasym, as well as several other APT imaging methods, are significantly affected by at least some of these confounding factors.