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CD19-redirected chimeric antigen receptor (CAR) T cells have shown remarkable activity against B-cell cancers. While second-generation CARs induce complete remission in >80% of patients with acute lymphoblastic leukemia, similar monotherapy induces long-term remissions in only 26% of patients with chronic lymphocytic leukemia (CLL). This disparity is attributed to cell-intrinsic effector defects in autologous CLL-derived T cells. However, the mechanisms by which leukemic cells impact CAR T-cell potency are poorly understood. Herein we describe an in vitro assay that recapitulates endogenous CLL-mediated T-cell defects in healthy donor CAR T cells. Contact with CLL cells insufficiently activates, but does not irreversibly impair, CAR T-cell function. This state is rescuable by strong antigenic stimulation or IL2, and is not driven by immune suppression. Rather, this activation defect is attributable to low levels of costimulatory molecules on CLL cells, and exogenous costimulation enhanced CAR T-cell activation. We next assessed the stimulatory phenotype of CLL cells derived from different niches within the same patient. Lymph node (LN)-derived CLL cells had a strong costimulatory phenotype and promoted better CAR T-cell degranulation and cytokine production than matched peripheral blood CLL cells. Finally, in vitro CD40L-activated CLL cells acquired a costimulatory phenotype similar to the LN-derived tumor and stimulated improved CAR T-cell proliferation, cytokine production, and cytotoxicity. Together, these data identify insufficient activation as a driver of poor CAR T-cell responses in CLL. The costimulatory phenotype of CLL cells drives differential CAR T-cell responses, and can be augmented by improving costimulatory signaling. Significance: CLL cells insufficiently activate CAR T cells, driven by low levels of costimulatory molecules on the tumor. LN-derived CLL cells are more costimulatory and mediate enhanced CAR T-cell killing. This costimulatory phenotype can be modeled via CD40 L activation, and the activated tumor promotes stronger CAR T-cell responses.

Prostate cancer cell lines from diverse backgrounds are important to addressing disparities in prostate cancer incidence and mortality rates among Black men. ACRJ-PC28 was developed from a transrectal needle biopsy and established via inactivation of the CDKN2A locus and simultaneous expression of human telomerase. Characterization assays included growth curve analysis, immunoblots, IHC, three-dimensional cultures, immunofluorescence imaging, confocal microscopy, flow cytometry, whole-genome sequencing (WGS), and RNA sequencing (RNA-seq). ACRJ-PC28 has been passaged more than 40 times in vitro over 10 months with a doubling time of 45 hours. Short tandem repeat profiling confirmed the novelty and human origin of the cell line. RNA-seq confirmed the expression of prostate specific genes alpha-methylacyl-CoA racemase (AMACR) and NKX3.1 and neuroendocrine specific markers synaptophysin and enolase 2 (ENO2) and IHC confirmed the presence of AMACR. Immunoblots indicated the cell line is of basal-luminal type; expresses p53 and pRB and is androgen receptor (AR) negative. WGS confirmed the absence of exonic mutations and the presence of intronic variants that appear to not affect function of AR, p53, and pRB. RNA-seq data revealed numerous TP53 and RB1 mRNA splice variants and the lack of AR mRNA expression. This is consistent with retention of p53 function in response to DNA damage and pRB function in response to contact inhibition. Soft agar anchorage-independent analysis indicated that the cells are transformed, confirmed by principal component analysis where ACRJ-PC28 cells cluster alongside other prostate cancer tumor tissues, yet was distinct. The novel methodology described should advance prostate cell line development, addressing the disparity in prostate cancer among Black men.Significance:Cell line development continues to attract less than 10% success rate. More than 98% of prostate cancer cell lines are from White men. This may contribute to the poorer response by Black men with prostate cancer to therapy compared with White men with prostate cancer, increasing overall survivorship among White men. The methodology described here to develop ACRJ-PC28, should advance the presence of Black prostate cancer cell lines thereby addressing prostate cancer disparity.

In these prior attempts, culture conditions such as media, extra-cellular matrices and culture flasks were varied to determine the optimal culture conditions for generation of prostate cancer explants.

CD19-redirected chimeric antigen receptor (CAR) T cells have shown remarkable activity against B-cell cancers. While second-generation CARs induce complete remission in >80% of patients with acute lymphoblastic leukemia, similar monotherapy induces long-term remissions in only 26% of patients with chronic lymphocytic leukemia (CLL). This disparity is attributed to cell-intrinsic effector defects in autologous CLL-derived T cells. However, the mechanisms by which leukemic cells impact CAR T-cell potency are poorly understood. Herein we describe an in vitro assay that recapitulates endogenous CLL-mediated T-cell defects in healthy donor CAR T cells. Contact with CLL cells insufficiently activates, but does not irreversibly impair, CAR T-cell function. This state is rescuable by strong antigenic stimulation or IL2, and is not driven by immune suppression. Rather, this activation defect is attributable to low levels of costimulatory molecules on CLL cells, and exogenous costimulation enhanced CAR T-cell activation. We next assessed the stimulatory phenotype of CLL cells derived from different niches within the same patient. Lymph node (LN)-derived CLL cells had a strong costimulatory phenotype and promoted better CAR T-cell degranulation and cytokine production than matched peripheral blood CLL cells. Finally, in vitro CD40L-activated CLL cells acquired a costimulatory phenotype similar to the LN-derived tumor and stimulated improved CAR T-cell proliferation, cytokine production, and cytotoxicity. Together, these data identify insufficient activation as a driver of poor CAR T-cell responses in CLL. The costimulatory phenotype of CLL cells drives differential CAR T-cell responses, and can be augmented by improving costimulatory signaling.Significance:CLL cells insufficiently activate CAR T cells, driven by low levels of costimulatory molecules on the tumor. LN-derived CLL cells are more costimulatory and mediate enhanced CAR T-cell killing. This costimulatory phenotype can be modeled via CD40 L activation, and the activated tumor promotes stronger CAR T-cell responses.

Cell viability was determined using the MTS assay and OD readings were taken a 409nm. Plots were generated in Microsoft Excel and each data point represents the average of three replicates. Experiments shown are representative of two independent experiments.

Data from RNA-Seq analysis. Noteworthy were the gene ontologies with a high proportion of expressed genes, such as the nuclear compartment (28%), extracellular exosome compartment (15%), regulation of transcription (8%), apoptotic process (3%) and oxidation-reduction process (3%).

Supplemental Figure 1: An overview of study design choices. Supplemental Figure 2: IL-2 Supplementation Enhances the APC phenotype of CLL cells. Supplemental Figure 3: Exogenous Co-stimulation Improves Second-generation CAR T cell Activation. Supplemental Figure 4: aCLL Stimulation Shows Inconsistent Cytokine Production After a 6hr Incubation. Supplemental Table 1: Full Antibody List. Supplemental Table 2: CLL Donor List. Supplemental Table 3: Full list of adjusted P-values from Holm-Sidak multiple comparisons.

STR profile of ACRJ-PC28 generated by IDEXX using PC indicates the novelty of the cell line (A) as the cell line senctic profile was compared to the profiles of other cell lines in the DSMZ STR database and did not match anv other reported profiles here: (B) that the cclls were purely of humar origin and (C) the absence of mycoplasma intection

Tissue sample was obtained from a transrectal needle biopsy of the prostate gland of a patient with PSA

IHC staining on ACRJ-PC28 cells compared to positive controls.

Numerous studies have clarified the involvement of circular RNAs (circRNAs) in modulating malignant behavior of non-small cell lung cancer (NSCLC), while the concrete mechanism is not completely elucidated. The aim of the study was to figure out the latent functions and molecular mechanisms of circRNA Eps15-homology domain containing protein 2 (EHD2) on NSCLC proliferation, glycolysis and autophagy. The results clarified in NSCLC elevated expression of circEHD2 and declined expression of microRNA (miR)-3186-3p. Repressive circEHD2 or enhancive miR-3186-3p facilitated cell apoptosis rate and autophagy substrates LC3BII and Beclin-1, but curbed the colony-formation and DNA replication ability of NSCLC, glucose consumption, lactic acid production, glycolytic rate-limiting enzyme HK-2 and glutamine hydrolase GLS1 and P62, while overexpressed circEHD2 was adverse. Meanwhile, the impacts of repressive and elevated circEHD2 on NSCLC were turned around via reduced miR-3186-3p or forkhead box k1 (FOXK1) separately. Mechanically, FOXK1 was augmented via circEHD2's competitive integration of miR-3186-3p. Depressive circEHD2 refrained NSCLC tumor growth, which was accelerated via enhancive one. All in all, circEHD2 accelerates the proliferation and glycolysis of NSCLC, but refrains autophagy and apoptosis via strengthening FOXK1 via the adsorption of miR-3186-3p, which is supposed to be a latent molecular target for NSCLC therapy later.

Background: Radiotherapy is widely applied to breast cancer treatment, while the resistance to radiotherapy is inevitable. TGF-β1 has been considered to be an endogenous factor for RT resistance. As a large portion of TGF-β1 is secreted associating with the extracellular vesicles (TGF-β1EV), to understand the regulation mechanisms and the immunosuppressive function of TGF-β1EV in the radiated tumors will pay a way for overcoming radiotherapy resistance. Methods: Based on TCGA database of 958 samples, the intratumoral TGF-β1 expression and the Tregs production were compared between patients received and unreceived radiotherapy. The superoxide-Zinc-PKC-ζ-TGF-β1EV pathway in breast cancer cells were identified through sequence alignments of different PKC isoforms, speculation and experimental confirmation. A series of functional and molecular studies were performed by quantitative real-time PCR, western blot and flow cytometry analysis. Mice survival and tumor growth was determined using observation of the animals and tumor growth measurement. Student’s t test or type II ANOVA with correction was used for comparisons of groups. Results: The radiotherapy brought the increased intratumoral TGF-β1 expression and the Tregs production in the breast cancer tissues, and patients with higher TGF-β1 expression are associated with a poor survival. The increased intratumoral TGF-β mainly exists in the extracellular vesicles associated form both in the murine breast cancer model and in the human lung cancer tissues. Furthermore, radiation induced the more secretion of the TGF-β1EV and the higher percentage of Tregs by promoting protein kinase C zeta (PKC-ζ) expression and phosphorylation. Importantly, we found that naringenin rather than 1D11 significantly improve radiotherapy efficacy with low side effects. The underlying mechanism of naringenin is via downregulating radiation activated the superoxide-Zinc-PKC-ζ-TGF-β1EV pathway, which is distinct from TGF-β1 neutralizing antibody 1D11. Conclusions: The superoxide-zinc-PKC-ζ-TGF-β1EV release pathway was elucidated to induce the accumulation of Tregs resulting in radiotherapy resistance in the TME. Therefore, targeting PKC-ζ to counteract TGF-β1EV function could represent a novel strategy to overcome radioresistance in breast cancer treatment or other cancers. Trial registration: Using of tissues from patients with malignant Non-Small Cell Lung Cancer (NSCLC) was approved by the ethics committees at Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China (NCC2022C-702, from June 8th, 2022).

Prostate cancer cell lines from diverse backgrounds are important to addressing disparities in prostate cancer incidence and mortality rates among Black men. ACRJ-PC28 was developed from a transrectal needle biopsy and established via inactivation of the CDKN2A locus and simultaneous expression of human telomerase. Characterization assays included growth curve analysis, immunoblots, IHC, three-dimensional cultures, immunofluorescence imaging, confocal microscopy, flow cytometry, whole-genome sequencing (WGS), and RNA sequencing (RNA-seq). ACRJ-PC28 has been passaged more than 40 times in vitro over 10 months with a doubling time of 45 hours. Short tandem repeat profiling confirmed the novelty and human origin of the cell line. RNA-seq confirmed the expression of prostate specific genes alpha-methylacyl-CoA racemase (AMACR) and NKX3.1 and neuroendocrine specific markers synaptophysin and enolase 2 (ENO2) and IHC confirmed the presence of AMACR. Immunoblots indicated the cell line is of basal-luminal type; expresses p53 and pRB and is androgen receptor (AR) negative. WGS confirmed the absence of exonic mutations and the presence of intronic variants that appear to not affect function of AR, p53, and pRB. RNA-seq data revealed numerous TP53 and RB1 mRNA splice variants and the lack of AR mRNA expression. This is consistent with retention of p53 function in response to DNA damage and pRB function in response to contact inhibition. Soft agar anchorage-independent analysis indicated that the cells are transformed, confirmed by principal component analysis where ACRJ-PC28 cells cluster alongside other prostate cancer tumor tissues, yet was distinct. The novel methodology described should advance prostate cell line development, addressing the disparity in prostate cancer among Black men. Significance: Cell line development continues to attract less than 10% success rate. More than 98% of prostate cancer cell lines are from White men. This may contribute to the poorer response by Black men with prostate cancer to therapy compared with White men with prostate cancer, increasing overall survivorship among White men. The methodology described here to develop ACRJ-PC28, should advance the presence of Black prostate cancer cell lines thereby addressing prostate cancer disparity.

The frequency spectra and polarization states of terahertz waves can convey significant information about physical interactions and material properties. Compact and miniaturized on-chip platforms for effective capturing of these quantities are being extensively investigated because of their promising potential for paramount applications of terahertz technology such as in situ sensing and characterization. Here, we present a metamaterial-graphene hybrid device that integrates the functions of photodetection, wavelength, and polarization selectivity into a monolithic architecture. Leveraging the ultrahigh design freedom of metamaterial optical properties and the electronically controllable hot-carrier-assisted photothermoelectric effect in graphene, our detector shows resonantly enhanced photoresponse at two specific target wavelengths with orthogonal polarizations. We demonstrate its versatile capabilities for spectrally selective and polarization resolved imaging on a single-chip platform that is free from advanced optical components. Our strategy is beneficial to the future development of multifunctional, compact, and low-cost terahertz sensors.

A notable number of acute lymphoblastic leukemia (ALL) patients develop CD19-positive relapse within 1 year after receiving chimeric antigen receptor (CAR) T cell therapy. It remains unclear if the long-term response is associated with the characteristics of CAR T cells in infusion products, hindering the identification of biomarkers to predict therapeutic outcomes. Here, we present 101,326 single-cell transcriptomes and surface protein landscape from the infusion products of 12 ALL patients. We observed substantial heterogeneity in the antigen-specific activation states, among which a deficiency of T helper 2 function was associated with CD19-positive relapse compared with durable responders (remission, >54 months). Proteomic data revealed that the frequency of early memory T cells, rather than activation or coinhibitory signatures, could distinguish the relapse. These findings were corroborated by independent functional profiling of 49 patients, and an integrative model was developed to predict the response. Our data unveil the molecular mechanisms that may inform strategies to boost specific T cell function to maintain long-term remission.

Non-small-cell lung cancer (NSCLC) accounts for most new diagnoses of lung cancer, with high morbidity and mortality worldwide. Immune checkpoint inhibitor (ICI) therapy has transformed the treatment of metastatic and advanced NSCLC. For resectable NSCLC, while surgery is the cornerstone of standard treatment, a number of clinical trials of neoadjuvant immunotherapy have been conducted. To perform a systematic review on the safety and efficacy of neoadjuvant ICI therapy in patients with resectable NSCLC. This systematic review was performed according to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. We undertook a comprehensive literature search of PubMed, Embase, Cochrane Library, and abstracts and posters from annual meetings of the major oncology societies, American Society of Clinical Oncology (ASCO), European Society for Medical Oncology (ESMO), American Association for Cancer Research (AACR) up until 29 April 2021. A total of 399 patients were identified from six articles and four meeting abstracts. 229, 140, and 30 patients received anti-programmed cell death ligand 1 therapy (anti-PD-L1, atezolizumab and durvalumab), anti-programmed cell death 1 therapy (anti-PD-1, nivolumab, pembrolizumab, sintilimab), and anti-PD-1/anti-CTLA-4 combination therapy (nivolumab and ipilimumab), respectively. 255 patients received only ICI therapy before surgery, and 144 patients received ICI and chemotherapy. While ICI therapy was generally well tolerated, grade 3 or higher immune-related adverse events were observed in 13 of 144 patients (9.0%) in the five studies that reported such adverse events data. Patients displayed an overall mean surgical resection rate of 87.5% (349/399, range, 66.7–100%), a surgical delay rate of 1.4%, and an incidence of surgical complications of 21%. On average, 45.6% (159/349), (range 17–83%) of patients exhibited major pathological response (MPR), while 76/349 (21.8%) patients achieved pathological complete response (pCR). In the studies with patients undergoing ICI and chemotherapy, the MPR rate was 66.7% and pCR rate was 35.4%. ICI neoadjuvant therapy may be a safe and efficacious treatment option in patients with resectable NSCLC. Combined with chemotherapy, ICI appears to be more efficacious, but displays more adverse events. More ongoing clinical trials will shed further light on the safety and efficacy of ICI neoadjuvant therapy in patients with resectable NSCLC.

Passive millimeter-wave (PMMW) imaging has been used for several close-range applications such as personal security checks, scene monitoring, and so on. PMMW images contain a variety of types of image edges which represent intensity discontinuities. As a special edge, an object contour edge is an important feature for object detection and recognition, which denotes the external shape of the object. In this article, a physically based contour edge display method using adjustable linear polarization ratio (ALPR) by multipolarization imaging is proposed. Polarization brightness temperature (TB) models of the object and radiometer observation are built. According to the physical principle of edge generation, the ALPR properties of the contour edge are investigated. By fusing multipolarization information, the simple feature parameter ALPR is sensitive to the object contour edge. Then, the specific operation process of the proposed method based on multipolarization images is presented. Simulations and measurements of polarimetric imaging for personal security inspection scenes were conducted to verify the contour edge display performance. Finally, the applicability of the method is discussed and summarized.

In millimeter-wave imaging of a hidden target, the effect of the dielectric cover before the target is typically ignored. This results in ripple-corrupted images that pose challenges for target recognition. In this paper, we provide a perspective for understanding the image of the hidden target, which clearly reveals the origin of the ripples, and propose a separation method that not only gets rid of ripples, but also obtains the target’s depth map. Reflections and transmissions during imaging are considered and decoupled to separately form images corresponding to each real or virtual object. An algorithm based on the range-direction spread function is developed to iteratively estimate the depth and reflectivity of the target. Imaging experiments with and without a cover are conducted to demonstrate the formation and influence of ripples and to verify the proposed algorithm. Our work deepens the comprehension of covered target imaging. Benefited fields might include non-destructive testing, through-wall imaging, subsurface imaging, and security screening.

Chimeric antigen receptor (CAR) T cell therapy has been promising in chronic lymphocytic leukemia (CLL) treatment showing durable remission. However, only a third of CLL patients can achieve complete remission with persistent effect featured by early memory T cells. This disparity is attributed to T cell-intrinsic defects or tumor-mediated immunosuppression. The mechanism how CLL cells impact CAR T cell potency still remains poorly understood. Here we used second-generation CD19- and ROR1-directed CAR T cells with a 4-1BB intracellular signaling domain. To recapitulate CLL-derived T cell defects, we developed an in vitro model of chronic antigen stimulation using CAR T cells recursively exposed to primary CLL cells. Upon each round of CLL stimulation, both types of CAR T cells were lack of proliferation and differentiation, accompanied with cytokine production failure in IL-2, suggesting hypofunction of CAR T cell induced by CLL. We next found that this lack of CAR T cell response is not permanent but can be rescued by strong antigen stimulus or IL-2 administration. On the other hand, IL-2 affects phenotype of CLL cells by enhancing expression of co-stimulatory molecules CD80 and CD86, suggesting that IL-2 supplementation or auxiliary co-stimulation via CD80/CD86 can rescue CAR T cell reactivity against these tumor cells. To test this hypothesis, we employed the immunomodulatory drugs (IMiDs) lenalidomide and pomalidomide that have previously been demonstrated to upregulate co-stimulatory molecules on CLL and restore T-cell function via IL-2 upregulation. We sought to test if IMiDs can improve CAR T cell response to CLL. CAR T cells repeatedly exposed to primary CLL cells were co-treated with IMiDs at each round, showing significantly increased proliferation compared to the group without drug treatment. Furthermore, IMiDs can enhance the functions of these CLL stimulated CAR T cells, including differentiation (higher frequency of effector memory T cells), activation (HLA-DR, CTLA-4), proliferation (Ki67), and effector function (Granzyme B). In addition, IMiDs can increase cytotoxic efficacy of CD8+ CAR19 T cells on CLL cells. Overall, our study unveiled that in CLL, activation defect of 4-1BB, CD3-signaling anti-CD19 CAR T cells was attributable to low levels of co-stimulatory molecules on CLL cells and can be effectively overcome via upregulation of co-stimulation. Our data suggested that CLL resistance to CAR T cell can be successfully rescued by concurrent treatment with IMiDs. This study provides a potential translational approach to overcome the bottleneck of CLL treatment by CAR T cells. Citation Format: Ziran Zhao, McKensie Collins, Clare Sun, Adrian Wiestner, J. Joseph Melenhorst. Co-stimulatory signaling boosts CAR T cell efficacy against chronic lymphocytic leukemia. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4100.

Programmed death receptor-1 (PD-1) inhibitors have shown efficacy in first-line treatment of NSCLC; however, evidence of PD-1 inhibitor as neoadjuvant treatment is limited. This is a phase 1b study to evaluate the safety and outcome of PD-1 inhibitor in neoadjuvant setting.Treatment-naive patients with resectable NSCLC (stage IA-IIIB) received two cycles of sintilimab (200 mg, intravenously, day 1 out of 22). Operation was performed between day 29 and 43. Positron emission tomography-computed tomography scans were obtained at baseline and before the operation. The primary end point was safety. Efficacy end points included rate of major pathologic response (MPR) and objective response rate. Expression of programmed cell death ligand 1 was also evaluated (registration number: ChiCTR-OIC-17013726).A total of 40 patients enrolled, all of whom received two doses of sintilimab and 37 underwent radical resection. A total of 21 patients (52.5%) experienced neoadjuvant treatment-related adverse events (TRAEs). Four patients (10.0%) experienced grade 3 or higher neoadjuvant TRAEs, and one patient had grade 5 TRAE. Eight patients achieved radiological partial response, resulting in an objective response rate of 20.0%. Among 37 patients, 15 (40.5%) achieved MPR, including six (16.2%) with a pathologic complete response in primary tumor and three (8.1%) in lymph nodes as well. Squamous cell NSCLC exhibited superior response compared with adenocarcinoma (MPR: 48.4% versus 0%). Decrease of maximum standardized uptake values after sintilimab treatment correlated with pathologic remission (p0.00001). Baseline programmed cell death ligand 1 expression of stromal cells instead of tumor cells was correlated with pathologic regression (p = 0.0471).Neoadjuvant sintilimab was tolerable for patients with NSCLC, and 40.5% MPR rate is encouraging. The decrease of maximum standardized uptake values after sintilimab might predict pathologic response.

Ultra-broadband photodetection is crucial for various applications like imaging and sensing and has become a hot research topic in recent years. However, most of the reported ultra-broadband photod...