Chimeric antigen receptor T (CAR-T) cell therapy has produced impressive results in clinical trials for B-cell malignancies. However, safety concerns related to the inability to control CAR-T cells ...once infused into the patient remain a significant challenge. Here we report the engineering of recombinant antibody-based bifunctional switches that consist of a tumor antigen-specific Fab molecule engrafted with a peptide neo-epitope, which is bound exclusively by a peptide-specific switchable CAR-T cell (sCAR-T). The switch redirects the activity of the bio-orthogonal sCAR-T cells through the selective formation of immunological synapses, in which the sCAR-T cell, switch, and target cell interact in a structurally defined and temporally controlled manner. Optimized switches specific for CD19 controlled the activity, tissue-homing, cytokine release, and phenotype of sCAR-T cells in a dose-titratable manner in a Nalm-6 xenograft rodent model of B-cell leukemia. The sCAR–T-cell dosing regimen could be tuned to provide efficacy comparable to the corresponding conventional CART-19, but with lower cytokine levels, thereby offering a method of mitigating cytokine release syndrome in clinical translation. Furthermore, we demonstrate that this methodology is readily adaptable to targeting CD20 on cancer cells using the same sCAR-T cell, suggesting that this approach may be broadly applicable to heterogeneous and resistant tumor populations, as well as other liquid and solid tumor antigens.
The adoptive transfer of autologous T cells engineered to express a chimeric antigen receptor (CAR) has emerged as a promising cancer therapy. Despite impressive clinical efficacy, the general ...application of current CAR–T-cell therapy is limited by serious treatment-related toxicities. One approach to improve the safety of CAR-T cells involves making their activation and proliferation dependent upon adaptor molecules that mediate formation of the immunological synapse between the target cancer cell and T-cell. Here, we describe the design and synthesis of structurally defined semisynthetic adaptors we refer to as “switch” molecules, in which anti-CD19 and anti-CD22 antibody fragments are site-specifically modified with FITC using genetically encoded noncanonical amino acids. This approach allows the precise control over the geometry and stoichiometry of complex formation between CD19- or CD22-expressing cancer cells and a “universal” anti-FITC–directed CAR-T cell. Optimization of this CAR–switch combination results in potent, dose-dependent in vivo antitumor activity in xenograft models. The advantage of being able to titrate CAR–T-cell in vivo activity was further evidenced by reduced in vivo toxicity and the elimination of persistent B-cell aplasia in immune-competent mice. The ability to control CAR-T cell and cancer cell interactions using intermediate switch molecules may expand the scope of engineered T-cell therapy to solid tumors, as well as indications beyond cancer therapy.
The treatment of patients with acute myeloid leukemia (AML) with targeted immunotherapy is challenged by the heterogeneity of the disease and a lack of tumor-exclusive antigens. Conventional ...immunotherapy targets for AML such as CD33 and CD123 have been proposed as targets for chimeric antigen receptor (CAR)-engineered T-cells (CAR-T-cells), a therapy that has been highly successful in the treatment of B-cell leukemia and lymphoma. However, CD33 and CD123 are present on hematopoietic stem cells, and targeting with CAR-T-cells has the potential to elicit long-term myelosuppression. C-type lectin-like molecule-1 (CLL1 or CLEC12A) is a myeloid lineage antigen that is expressed by malignant cells in more than 90% of AML patients. CLL1 is not expressed by healthy Hematopoietic Stem Cells (HSCs), and is therefore a promising target for CAR-T-cell therapy. Here, we describe the development and optimization of an anti-CLL1 CAR-T-cell with potent activity on both AML cell lines and primary patient-derived AML blasts in vitro while sparing healthy HSCs. Furthermore, in a disseminated mouse xenograft model using the CLL1-positive HL60 cell line, these CAR-T-cells completely eradicated tumor, thus supporting CLL1 as a promising target for CAR-T-cells to treat AML while limiting myelosuppressive toxicity.
Contrast-enhanced US (CEUS), similar to other radiologic modalities, requires specific technical considerations and is subject to image artifacts. These artifacts may affect examination quality, ...negatively impact diagnostic accuracy, and decrease user comfort when using this emerging technique. Some artifacts are related to commonly known gray-scale US artifacts that can also appear on the contrast-only image (tissue-subtracted image obtained with the linear responses from background tissues nulled). These may include acoustic shadowing and enhancement; reverberation, refraction, and reflection; and poor penetration. Other artifacts are exclusive to CEUS owing to the techniques used for contrast mode image generation and the unique properties of the microbubbles that constitute ultrasound-specific contrast agents (UCAs). UCA-related artifacts may appear on the contrast-only image, the gray-scale image, or various Doppler mode images. Artifacts related to CEUS may include nonlinear artifacts and unintentional microbubble destruction resulting in pseudowashout. The microbubbles themselves may result in specific artifacts such as pseudoenhancement, signal saturation, and attenuation and shadowing and can confound the use of color and spectral Doppler US. Identifying and understanding these artifacts and knowing how to mitigate them may improve the quality of the imaging study, increase user confidence, and improve patient care. The authors review the principles of UCAs and the sound-microbubble interaction, as well as the technical aspects of image generation. Technical considerations, including patient positioning, depth, acoustic window, and contrast agent dose, also are discussed. Specific artifacts are described, with tips on how to identify and, if necessary, apply corrective measures, with the goal of improving examination quality.
RSNA, 2022
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The US Liver Imaging Reporting and Data System (LI-RADS) was released in 2017 and is the newest of the four American College of Radiology (ACR) LI-RADS algorithms. US LI-RADS provides standardized ...terminology, technical recommendations, and a reporting framework for US examinations performed for screening or surveillance in patients at risk for developing hepatocellular carcinoma (HCC). The appropriate patient population for screening and surveillance includes individuals who are at risk for developing HCC but do not have known or suspected cancer. This includes patients with cirrhosis from any cause and subsets of patients with chronic hepatitis B virus infection in the absence of cirrhosis. In an HCC screening or surveillance study, US LI-RADS recommends assigning two scores that apply to the entire study: the US category, which determines follow-up, and a visualization score, which communicates the expected level of sensitivity of the examination but does not affect management. Three US categories are possible: US-1 negative, a study with no evidence of HCC; US-2 subthreshold, a study in which an observation less than 10 mm is depicted that is not definitely benign; and US-3 positive, a study in which an observation greater than or equal to 10 mm or a new thrombus in vein is identified, for which diagnostic contrast material-enhanced imaging is recommended. Three visualization scores are possible: A (no or minimal limitations), B (moderate limitations), and C (severe limitations).
RSNA, 2019.
Ultrasound plays an essential role in the initial evaluation of patients with suspected or confirmed acute pancreatitis. In addition to evaluation of the pancreatic parenchyma, ultrasound is used for ...assessment of the gallbladder, biliary tree, peripancreatic tissues, and regional vascular structures. While enlarged and edematous pancreas are classic sonographic features of acute pancreatitis, the pancreas may appear sonographically normal in the setting of acute pancreatitis. Nonetheless, sonographic evaluation in this setting is valuable because assessment for etiologic factors such as gallstones or evidence of biliary obstruction are best performed with ultrasound. Complications of pancreatitis such as peripancreatic fluid collections, venous thrombosis, or arterial pseudoaneurysm can be identified with careful and focused ultrasound examination. Knowledge of various scanning techniques can help to mitigate some of the commonly encountered barriers to sonographic visualization of the pancreas and right upper quadrant structures. Ultrasound can also be used for guidance of percutaneous treatment such as drainage of fluid collections or pseudoaneurysm thrombosis. Difficulty in differentiating edematous from necrotizing pancreatitis can be mitigated with the use of contrast-enhanced ultrasound to assess pancreatic parenchymal enhancement.
Objective
The parasitic worm–derived immunomodulator ES‐62 protects against disease in the mouse collagen‐induced arthritis (CIA) model of rheumatoid arthritis (RA) by suppressing pathogenic ...interleukin‐17 (IL‐17) responses. The Th17‐associated cytokine IL‐22 also appears to have a pathogenic role in autoimmune arthritis, particularly in promoting proinflammatory responses by synovial fibroblasts and osteoclastogenesis. The present study was undertaken to investigate whether the protection against joint damage afforded by ES‐62 also reflects suppression of IL‐22.
Methods
The role(s) of IL‐22 was assessed by investigating the effects of neutralizing anti–IL‐22 antibodies and recombinant IL‐22 (rIL‐22) on proinflammatory cytokine production, synovial fibroblast responses, and joint damage in mice with CIA in the presence or absence of ES‐62.
Results
Neutralization of IL‐22 during the initiation phase abrogated CIA, while administration of rIL‐22 enhanced synovial fibroblast responses and exacerbated joint pathology. In contrast, after disease onset anti–IL‐22 did not suppress progression, whereas administration of rIL‐22 promoted resolution of inflammation. Consistent with these late antiinflammatory effects, the protection afforded by ES‐62 was associated with elevated levels of IL‐22 in the serum and joints that reflected a desensitization of the synovial fibroblast responses. Moreover, neutralization of IL‐22 during the late effector stage of disease prevented ES‐62–mediated desensitization of synovial fibroblast responses and protection against CIA.
Conclusion
IL‐22 plays a dual role in CIA, being pathogenic during the initiation phase while acting to resolve inflammation and joint damage during established disease. Harnessing of the tissue repair properties of IL‐22 by ES‐62 highlights the potential for joint‐targeted therapeutic modulation of synovial fibroblast responses and consequent protection against bone damage in RA.
Chimeric antigen receptor T (CAR‐T) cells have demonstrated promising results against hematological malignancies, but have encountered significant challenges in translation to solid tumors. To ...overcome these hurdles, we have developed a switchable CAR‐T cell platform in which the activity of the engineered cell is controlled by dosage of an antibody‐based switch. Herein, we apply this approach to Her2‐expressing breast cancers by engineering switch molecules through site‐specific incorporation of FITC or grafting of a peptide neo‐epitope (PNE) into the anti‐Her2 antibody trastuzumab (clone 4D5). We demonstrate that both switch formats can be readily optimized to redirect CAR‐T cells (specific for the corresponding FITC or PNE) to Her2‐expressing tumor cells, and afford dose‐titratable activation of CAR‐T cells ex vivo and complete clearance of the tumor in rodent xenograft models. This strategy may facilitate the application of immunotherapy to solid tumors by affording comparable efficacy with improved safety owing to switch‐based control of the CAR‐T response.
CAR‐T control: Chimeric antigen receptor T (CAR‐T) cells were engineered to be controlled by exogenous switch molecules. Site‐specific incorporation of the small molecule FITC or a short peptide neo‐epitope in the anti‐Her2 4D5 Fab allowed activation of corresponding switchable CAR‐T cells towards Her2‐expressing solid tumor cells, and displayed significant anti‐cancer effects both in vitro and in vivo.
In the context of chronic liver disease (CLD), sonographic features of hepatic steatosis, cirrhosis, and portal hypertension are discussed and examples are provided. The impact of CLD and ...hepatocellular carcinoma (HCC) is introduced, providing the rationale for a robust HCC screening and surveillance program for at-risk patients. The American College of Radiology Liver Imaging Reporting and Data System algorithms for screening and surveillance by ultrasound and for the definitive diagnosis of HCC by contrast-enhanced ultrasound are explained, with imaging examples provided. Contrast-enhanced ultrasound technique, limitations, and pitfalls also are introduced.
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