Rapid Manufacturing Process Research Articles

20 Seconds to fabricate high-performance NiFe-based anode for seawater electrolysis via bidirectional pulse current method

Direct electrolysis of seawater to produce green hydrogen has attracted great attention. While the development of efficient and rapid manufacturing processes for high-performance electrodes, particularly for the oxygen evolution reaction (OER), remains trail. Here, we introduce an innovative approach to fabricate high-performance NiFe-based OER electrodes using an ultrafast and eco-friendly bidirectional pulse current (BPC) method, which remarkably shortens the production time to only 20 s. Our BPC method alternates oxidation and reduction currents within a single period, promoting the in-situ formation of porous NiFe(oxy)hydroxide nanosheets through dissolution/deposition processes. Moreover, this methodology creatively utilizes the “Cl− induced effect”, typically a hindrance in other electrochemical systems involving seawater or brine, to enhance the synthesis of electrodes from NaCl-based electrolyte devoid of additional Ni/Fe cations. The prepared NiFe-based electrode (NFF O-R 20 s) demonstrates remarkable OER catalytic activity, with optimal overpotentials (ηj) of 272, 308 and 367 mV at current densities (j) of 10, 100 and 1000 mA cm−2, respectively. Impressively, it maintains a low cell voltage of 1.59 V after enduring 1000 h of operation under the industrially current density of 300 mA cm−2, with a direct current energy consumption of 3.8 kWh Nm−3 H2 for overall alkaline seawater electrolysis. Our BPC preparation approach provides a promising path to construction high-catalytic activity, good-stability electrode, and low-energy consumption direct seawater electrolyzer.

Additive manufacturing of Invar 36 alloy

Invar 36 alloy, renowned for its ultra-low coefficient of thermal expansion (CTE), is a functional Fe–Ni alloy that finds wide applications in aerospace and precision instruments. Additive manufacturing (AM), as a rapid digital manufacturing process, can efficiently and flexibly fabricate Invar 36 alloy parts with complex geometric and exceptional properties. Considering the advantages of AM and the increasing interest and demand for Invar 36 alloy parts fabrication by AM in recent years, it is imperative to provide a comprehensive summary of the current research status and technological advancements pertaining to AM Invar 36 alloy. Firstly, an overview of the AM processes currently employed for fabricating Invar 36 alloy is provided, including laser powder bed fusion, selective laser sintering, directed energy deposition, cold spray additive manufacturing, wire arc additive manufacturing, and binder jetting. Additionally, the microstructure, mechanical properties, and CTE of Invar 36 alloy manufactured by AM are summarized. Even eliminating the post heat treatment, the as-printed Invar 36 alloy can achieve excellent low CTE and mechanical properties, comparable to or exceeding traditional manufacturing processes. Moreover, the advantages and research progress of AM Invar 36 alloy composites are introduced. Enhancing mechanical properties while reducing CTE is a common challenge for both AM Invar 36 alloy and Invar composites. Finally, the technical gaps, research trends, and potential applications of AM Invar 36 alloy are summarized and prospected.

Triplet Upconversion under Ambient Conditions Enables Digital Light Processing 3D Printing.

The rapid photochemical conversion of materials from liquid to solid (i.e., curing) has enabled the fabrication of modern plastics used in microelectronics, dentistry, and medicine. However, industrialized photocurables remain restricted to unimolecular bond homolysis reactions (Type I photoinitiations) that are driven by high-energy UV light. This narrow mechanistic scope both challenges the production of high-resolution objects and restricts the materials that can be produced using emergent manufacturing technologies (e.g., 3D printing). Herein we develop a photosystem based on triplet-triplet annihilation upconversion (TTA-UC) that efficiently drives a Type I photocuring process using green light at low power density (<10 mW/cm2) and in the presence of ambient oxygen. This system also exhibits a superlinear dependence of its cure depth on the light exposure intensity, which enhances spatial resolution. This enables for the first-time integration of TTA-UC in an inexpensive, rapid, and high-resolution manufacturing process, digital light processing (DLP) 3D printing. Moreover, relative to traditional Type I and Type II (photoredox) strategies, the present TTA-UC photoinitiation method results in improved cure depth confinement and resin shelf stability. This report provides a user-friendly avenue to utilize TTA-UC in ambient photochemical processes and paves the way toward fabrication of next-generation plastics with improved geometric precision and functionality.

A Self-Healing System for Polydicyclopentadiene Thermosets.

Self-healing offers promise for addressing structural failures, increasing lifespan, and improving durability in polymeric materials. Implementing self-healing in thermoset polymers faces significant manufacturing challenges, especially due to the elevated temperature requirements of thermoset processing. To introduce self-healing into structural thermosets, the self-healing system must be thermally stable and compatible with the thermoset chemistry. This article demonstrates a self-healing microcapsule-based system stable to frontal polymerization (FP), a rapid and energy-efficient manufacturing process with a self-propagating exothermic reaction (≈200 °C). A thermally latent Grubbs-type complex bearing two N-heterocyclic carbene ligands addresses limitations in conventional G2-based self-healing approaches. Under FP's elevated temperatures, the catalyst remains dormant until activated by a Cu(I) co-reagent, ensuring efficient polymerization of the dicyclopentadiene (DCPD) upon damage to the polyDCPD matrix. The two-part microcapsule system consists of one capsule containing the thermally latent Grubbs-type catalyst dissolved in the solvent, and another capsule containing a Cu(I) coagent blended with liquid DCPD monomer. Using the same chemistry for both matrix fabrication and healing results in strong interfaces as demonstrated by lap-shear tests. In an optimized system, the self-healing system restores the mechanical properties of the tough polyDCPD thermoset. Self-healing efficiencies greater than 90% via tapered double cantilever beam tests are observed.

Senza5 TM CART5: An Autologous CD5-Deleted Anti-CD5 CART Product with Enhanced Anti-T-Cell Lymphoma Activity

Introduction: Treatment with adoptive T cell therapies expressing Chimeric Antigen Receptors (CART) produces unprecedented clinical responses in relapsed or refractory (r/r) B-cell acute lymphoblastic leukemia (B-ALL), non-Hodgkin lymphomas (NHL), and multiple myeloma. Despite the transformative potential of these therapies for cancer, most patients treated with CART therapy experience incomplete responses or eventual relapse. Moreover, CART therapy is not yet effective for solid cancers or other hematological malignancies such as T-cell neoplasms. Thus, there is a dire need to enhance CART efficacy and extend this approach to more cancer types. We previously presented preliminary preclinical results of CD5-directed CAR-T cells (CART5) that had CD5 knocked out (KO) [ Blood (2020) 136 (Supplement 1): 51-52]. CD5 KO not only prevented CART fratricide but also dramatically enhanced CART function in multiple in vivo models. Enhanced CART function reflected disinhibition of early T-cell activation by abrogating CD5 signaling. Here, we combine the functional advantage of CD5 KO CART5 with a clinical-grade “rapid” 5-day manufacturing process, characterize the resulting drug product, and evaluate its in vivo antitumor efficacy in a human xenograft model of T-cell lymphoma/leukemia. Methods and Results: Using normal healthy donor T cells, we developed a proprietary, “rapid” 5-day GMP-validated manufacturing process for CD5 KO CART5, completing the process in 5 days (CART5 rapid) instead of the conventional 14+ days (CART5 conv). At the end of manufacturing, the two products possessed similar ratios of CD8 + and CD4 + T cells, CD5 editing, and CAR5 expression. The CART5 rapid product did not exhibit any genetic or functional abnormalities characteristic of tumorigenic risk, demonstrated by low off-target gene editing in low-risk genes, no in vitro soft agar colony formation, and normal karyotypes. Importantly, the CART5 rapid manufacturing process did not skew TCRβ profiles, indicating that the CART5 rapid product retains a diverse T-cell repertoire. CD5 KO CART5 rapid cells possessed high specificity for T-cell leukemia/lymphoma, reflected by the absence of recognition of ex vivo normal human cells (cardiomyocytes, hepatocytes, neurons, monocytes, etc). Interestingly, CD5 KO CART5 rapid cells possessed a more central memory-like phenotype than CD5 KO CART5 conv cells which has been correlated with improved clinical efficacy. To compare their antitumor efficacy, we performed an in vivo experiment with CD5 + Jurkat cells utilizing a suboptimal dose of CART5 cells. Immunodeficient NSG mice were engrafted with 10 6 Jurkat cells on day -7 and treated intravenously on day 0 with a suboptimal dose (0.5x10 6 CAR + cells) of CD5 KO CART5 rapid or CD5 KO CART5 conv cells. Controls received an equivalent total number of Mock KO UTD conv. Mice treated with CD5 KO CART5 rapid cells demonstrated superior antitumor responses compared to CD5 KO CART5 conv. Improved tumor control correlated with dramatically increased overall survival of mice treated with CD5 KO CART5 rapid (median overall survival Mock KO UTD = 21 days; CD5 KO CART5 conv = 44 days vs. CD5 KO CART5 rapid = not reached, p=0.0019, Mantel-Cox; Figure 1). Conclusion: This novel CD5 KO anti-CD5 CART product manufactured in a 5-day GMP process shows high specificity and significantly enhanced antitumor efficacy resulting from the synergistic effect of improved early T-cell activation due to CD5 deletion and preservation of T-cell memory phenotypes conferred by a novel, 5-day manufacturing processes. Autologous Senza5 TM CART5 for the treatment of CD5 + nodal T-cell lymphomas is expected to enter the clinic in a Phase I study later this year.

T-Charge™ Manufacturing of the Anti-BCMA CAR-T, Durcabtagene Autoleucel (PHE885), Promotes Expansion and Persistence of CAR-T Cells with High TCR Repertoire Diversity

BCMA targeted CAR-T cells are an effective therapy for patients with relapsed or refractory multiple myeloma (r/r MM). However, autologous CAR-T cell products are highly heterogeneous and the functional roles of various T cell populations within these products have not been established. Stem-like memory T cells (Tscm) are a rare T cell subset which maintain high capacity for self-renewal and multipotency. Previously, we demonstrated that the T-Charge TM platform, a novel rapid manufacturing process that reduces manufacturing time to &amp;lt;2 days, preserves less differentiated CAR-T cells which exhibit potent anti-tumor activity and robust expansion in preclinical models (Dexiu Bu, ASH 2021). We conducted a phase 1 clinical trial in r/r MM (NCT04318327) of durcabtagene autoleucel (PHE885), a fully human product manufactured using the T-Charge platform. Here we present a detailed analysis of CAR-T cell products and subsequent expansion of CAR-T cells in 32 patients enrolled in this clinical trial at the Dana-Farber Cancer Institute. Apheresis products (APH), final products (FP) and post-infusion peripheral blood mononuclear cells (PBMC) were characterized using flow cytometry, mass cytometry (CyTOF) and TCR sequencing. We previously reported a 98% overall response rate (ORR) across all dose levels (2.5-20x10 6 CAR-T cells) and 100% ORR at doses &amp;gt;5x10 6 cell dose (Sperling, ASCO 2023). CAR-T cells expanded rapidly after infusion reaching median peak levels of 3,118 cells/ul (range 373 to 17,865) with a median 87.4% (range, 46.9-97.8) of CD3+ T cells expressing the CAR at a median of 14 days (range 10 to 27) after infusion. CAR-T cells persisted at high levels with transgene detectable by qPCR in 67% of patients at 6 months. Among 28 evaluable patients, 13 (43%) had &amp;gt;20% CAR positive T cells detectable by flow cytometry at 3 months. Phenotypic analysis of APH and FP samples showed that less-differentiated T cell subsets, including Tscm and central memory T cells were maintained in FP. CyTOF evaluation of functional markers revealed high expression of proliferation and activation markers in Tscm in FP and subsequently in CAR-T cells at the time of peak expansion in vivo. Subsequently, more differentiated CAR-T cells increased, accompanied by a decline in activation markers, while no significant changes were observed in inhibitory receptors. The proportion of Tscm in the FP positively correlated with early in vivo CAR-T cell expansion. TCR repertoire diversity and TCR clone tracking were used to characterize product manufacturing and CAR-T cells in peripheral blood after infusion. We sorted naïve/Tscm CD4 T cells, naïve/Tscm CD8 T cells, memory CD4 T cells, and memory CD8 T cells from APH and FP, and isolated CAR-positive CD4 T cells, CAR-positive CD8 T cells, CAR-negative CD4 cells, and CAR-negative CD8 T cells from post-infusion PBMC by fluorescence-activated cell sorting and ran TCR sequence from the extracted gDNA from each T cell subpopulation. Measurement of TCR diversity post-infusion revealed higher TCR repertoire diversity in CAR-T cells than in non-CAR-T cells. Notably, post-infusion CAR-T cells shared significantly more TCR clonotypes with T SCM than with memory T cells in the FP, suggesting that the highly heterogeneous post-infusion CAR-T clones were preferentially derived from T SCM clones in the FP. In three evaluable patients with long-term persistence, a highly diverse TCR repertoire in CAR T cells was maintained 1 year after treatment. Our findings demonstrate that the T-Charge™ manufacturing platform successfully maintains highly heterogeneous transduced Tscm clones with self-renewal potential in durcabtagene autoleucel products. Maintenance of Tscm in manufactured products contributes to robust CAR-T expansion and long-term persistence of CAR-T cells with a highly diverse TCR repertoire after infusion.

Rapid multiscale surface texture manufacturing process using hybrid laser belt machining

Rapid multiscale surface texture manufacturing process using hybrid laser belt machining

A study on the prospects of vacuum gamma irradiation to enhance crosslinking for 3D-Printing PLA/MCC biocomposite filaments

3D-printing or Additive Manufacturing (AM) has been growing as a rapid manufacturing process for many different applications, with Poly (Lactic Acid) as one of the most used materials for 3D-printing. PLA shows great promise for many applications to achieve the goals of the SDGs due to its biodegradability and biocompatibility but lacks when it comes to mechanical strength and thermal resistance. In this study, microcrystalline cellulose (MCC) fibers were introduced as a reinforcement to PLA. The biocomposite filaments were irradiated in a vacuum to enhance the crosslinking. Gamma-ray irradiation in a vacuum has successfully shown signs of crosslinking by increasing the tensile strength and thermal stability of the biocomposite, indicating an enhancement for PLA/MCC for various applications. On the other hand, changes in thermal properties also indicated that irradiation may reduce the processability of the composite, so it is necessary to study the conditions under which the mechanical properties and processability are compatible.

Grayscale-patterned integrated multilayer-metal-dielectric microcavities for on-chip multi/hyperspectral imaging in the extended visible bandwidth.

Pixelated filter arrays of Fabry-Perot (FP) cavities are widely integrated with photodetectors to achieve a WYSIWYG ("what you see is what you get") on-chip spectral measurements. However, FP-filter-based spectral sensors typically have a trade-off between their spectral resolution and working bandwidth due to design limitations of conventional metal or dielectric multilayer microcavities. Here, we propose a new idea of integrated color filter arrays (CFAs) consisting of multilayer metal-dielectric-mirror FP microcavities that, enable a hyperspectral resolution over an extended visible bandwidth (∼300 nm). By introducing another two dielectric layers on the metallic film, the broadband reflectance of the FP-cavity mirror was greatly enhanced, accompanied by as-flat-as-possible reflection-phase dispersion. This resulted in balanced spectral resolution (∼10 nm) and spectral bandwidth from 450 nm to 750 nm. In the experiment, we used a one-step rapid manufacturing process by using grayscale e-beam lithography. A 16-channel (4 × 4) CFA was fabricated and demonstrated on-chip spectral imaging with a CMOS sensor and an impressive identification capability. Our results provide an attractive method for developing high-performance spectral sensors and have potential commercial applications by extending the utility of low-cost manufacturing process.

Binocular fringe projection profilometry for the metrology of meter-scale optical surfaces

Demand for better quality, larger quantity, and size of astronomical telescopes from visible to radio frequencies is increasing. More rapid, efficient, and adaptable manufacturing processes are needed to support the needs of growing science and engineering communities in these fields. To aid the development and execution of these new processes, a flexible, accurate, and low-cost metrology system is needed. This paper outlines a variety of fringe projection profilometry (FPP) that has demonstrated high accuracy over large areas, making it a critical tool for manufacturing steel molds for forming primary reflectors and shape verification of the reflectors themselves used for radio astronomy.

Out-of-oven rapid synthesis of entropy stabilized oxides using radio frequency heating

Entropy stabilized oxides (ESOs) are a new class of stable hybrids and single phase metal oxides made from multiple ions with material properties somewhere between the constituent oxides, or occasionally entirely new properties. One of the limitations of ESOs is their energy-intensive fabrication process, which has resulted in slow development and scale-up of new ESOs. In this work, we present a novel energy-efficient ESO synthesis method that uses the ability of carbonaceous materials to heat rapidly in response to radio frequency (RF) fields in 1–200 MHz range. Using carbon fibers and graphene as RF susceptors, synthesis of (Mg0·2Co0·2Ni0·2Cu0·2Zn0.2)O is achieved through RF-initiated combustion synthesis with heating rates of 203 °C/s at 20 W of input power. This method reduces the formation time of ESOs to less than a minute, allowing for much more efficient fabrication. The corresponding morphology and composition of the as-synthesized ESO-carbon fiber and ESO-graphene were studied using extensive spectroscopy and characterization. Additionally, single carbon fibers coated with ESO were tested for tensile strength and modulus; little change in mechanical properties was observed as compared to pristine fibers. This work opens an exciting frontier for the rapid synthesis of ESO-carbon composites using RF heating as a non-contact, rapid, and efficient manufacturing process.

YTB323 (Rapcabtagene Autoleucel) Demonstrates Durable Efficacy and a Manageable Safety Profile in Patients with Relapsed/Refractory Diffuse Large B-Cell Lymphoma: Phase I Study Update

Background: CD19-directed CAR-T cell therapies (tx) have shown efficacy in patients (pts) with B-cell malignancies. However, many pts fail to respond or experience disease relapse after initial response. YTB323 (rapcabtagene autoleucel) is an autologous CD19-directed CAR-T cell tx generated by the innovative T-Charge™ platform, which produces CAR-T cells by a rapid manufacturing process (2 days) preserving T-cell stemness. This methodology is expected to increase CAR-T cell persistence and expansion and yield higher efficacy and a manageable safety profile. Here we present efficacy and safety, dose exposure, and biomarker analyses for rapcabtagene autoleucel in adults with r/r DLBCL in the ongoing Phase I, multicenter, dose-escalation study (NCT03960840). Methods: Eligible pts had r/r DLBCL after ≥2 lines of prior tx, measurable disease at enrollment, and ECOG 0-1. Pts received single-dose rapcabtagene autoleucel at targeted dose level (DL) 1 (2.5×106 CAR+ cells), DL2 (12.5×106 CAR+ cells), DL3 (25×106 CAR+ cells), or DL4 (40×106 CAR+ cells). Primary endpoints characterized safety and dose-limiting toxicities to identify a recommended dose. Secondary endpoints include response rate by local investigator assessment and cellular kinetics. Results: As of March 31, 2022, 45 pts with r/r DLBCL received rapcabtagene autoleucel: 4 at DL1, 28 at DL2, 7 at DL3, and 6 at DL4. Median age was 64.8 y, 67% received 2 prior lines of tx, and 29% had prior autologous stem cell transplant. Median (range) time since most recent relapse/progression was 2.8 (1.4-81.8) mo. Pts were followed for a median (range) of 10 (0.3-29) mo. Complete response (CR) rate at DL2 was 65%. CR at DL2, excluding pts in CR before rapcabtagene autoleucel (due to either a late effect of prior tx or bridging chemotherapy), was 61%. Responses at DL2 were durable, with CR rates of 63% (12/19) at 3 mo and 69% (11/16) at 6 mo (Figure 1). Median duration of response at DL2 was not reached. Of 45 pts evaluable for safety, 96% reported 1 adverse event (AE) of any grade (Gr) and 89% had ≥1 AE Gr ≥3. Cytokine release syndrome (CRS) was experienced by 15 pts (33%): 1 at DL1, 10 at DL2, 2 at DL3, and 2 at DL4. One DL2 pt (4%) experienced Gr 4 CRS. CRS was managed with tocilizumab, corticosteroids, and vasopressors in 7 (70%), 3 (30%), and 1 (10%) pts at DL2, respectively, and 1 DL3 pt (50%) received tocilizumab. Median (range) time to CRS onset was 9 (1-36) d and resolution was 5 (1-18) d. Five pts (11%) had immune effector cell-associated neurotoxicity syndrome (ICANS) events: 3 at DL2 and 2 DL4. Two DL2 pts (7%) experienced Gr 3 ICANS (no pts experienced Gr 4-5). Median (range) time to onset and resolution of ICANS was 16 (6-28) and 16 (1-36) d, respectively. ICANS management was based on dexamethasone, methylprednisolone, and anakinra in 2 (67%), 1 (33%), and 1 (33%) pts at DL2, respectively, and 1 DL4 pt (50%) received dexamethasone. Median time to peak rapcabtagene autoleucel expansion was delayed compared with tisagenlecleucel: 16 d at DL2 vs 9 d in JULIET. At a 25-fold lower dose, rapcabtagene autoleucel expansion at DL2 was comparable by qPCR to tisagenlecleucel expansion in JULIET. Expansion increased from DL1 to DL2 with no further increases at higher doses. Data from DL2 show CAR transgene was detectable by qPCR in 1 out of 2 pts with 12 mo follow-up. scRNAseq analysis shows the rapcabtagene autoleucel manufacturing process preserves T-cell fitness by retaining the composition of cell populations from apheresis to final product through the short manufacturing time. scRNAseq analysis also suggests that certain characteristics of the final product composition may lead to improved CR rates at 3 mo. B-cell aplasia (≤50 CD19+ cells/μL) was established before/after rapcabtagene autoleucel in all pts. In responders, B-cell aplasia persisted for ≥8 mo. Conclusions: Rapcabtagene autoleucel is a potent new CD19-directed CAR-T cell tx with distinct cellular kinetics, durable efficacy, and a manageable safety profile. DL2 (12.5×106) CAR+ viable T cells is the recommended dose for Phase III studies, based on the CR rate, favorable safety profile, and cellular kinetics. Updated results with expanded cellular kinetics and biomarker analyses will be presented at the meeting. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Spatial-Division Multiplexing Approach for Simultaneous Detection of Fiber-Optic Ball Resonator Sensors: Applications for Refractometers and Biosensors.

Fiber-optic ball resonators are an attractive technology for refractive index (RI) sensing and optical biosensing, as they have good sensitivity and allow for a rapid and repeatable manufacturing process. An important feature for modern biosensing devices is the multiplexing capacity, which allows for interrogating multiple sensors (potentially, with different functionalization methods) simultaneously, by a single analyzer. In this work, we report a multiplexing method for ball resonators, which is based on a spatial-division multiplexing approach. The method is validated on four ball resonator devices, experimentally evaluating both the cross-talk and the spectral shape influence of one sensor on another. We show that the multiplexing approach is highly efficient and that a sensing network with an arbitrary number of ball resonators can be designed with reasonable penalties for the sensing capabilities. Furthermore, we validate this concept in a four-sensor multiplexing configuration, for the simultaneous detection of two different cancer biomarkers across a widespread range of concentrations.

Comparison of process mass intensity (PMI) of continuous and batch manufacturing processes for biologics

Biologics encompasses a wide variety of therapeutics including monoclonal antibodies, fusion proteins, and enzymes, among others. The biologics market is growing at a rapid pace and different manufacturing processes, including continuous manufacturing processes, are being increasingly adopted. There is a strong drive to assess the sustainability of such processes. Here, we calculated the process mass intensity (PMI) of a continuous manufacturing process and compared it to the PMI of batch processes for monoclonal antibodies (mAbs). Results show that the PMI of continuous manufacturing process is comparable to that of batch processes. Sensitivity analysis was performed to assess the impact of different process strategies on the material usage efficiency of continuous processes. Although PMI is a useful benchmarking metric of sustainability, it does not account for factors such as energy consumption which is a key driver of sustainability for biologics manufacturing. Comparison of a higher PMI continuous process with a lower PMI batch process operating at the same bioreactor scale shows that since the productivity (in g of drug substance, DS) per unit time is multifold higher for the continuous process, the overall energy consumption per unit of DS produced might be lower leading to a more environmentally sustainable process. This study highlights some of these key aspects that would require additional metrics and models to be developed to assess the overall sustainability of biologics processes.

Engineered-Skin of Single Dermal Layer Containing Printed Hybrid Gelatin-Polyvinyl Alcohol Bioink via 3D-Bioprinting: In Vitro Assessment under Submerged vs. Air-Lifting Models.

Three-dimensional (3D) in vitro skin models are frequently employed in cosmetic and pharmaceutical research to minimize the demand for animal testing. Hence, three-dimensional (3D) bioprinting was introduced to fabricate layer-by-layer bioink made up of cells and improve the ability to develop a rapid manufacturing process, while maintaining bio-mechanical scaffolds and microstructural properties. Briefly, gelatin-polyvinyl alcohol (GPVA) was mixed with 1.5 × 106 and 3.0 × 106 human dermal fibroblast (HDF) cell density, together with 0.1% genipin (GNP), as a crosslinking agent, using 3D-bioprinting. Then, it was cultured under submerged and air-lifting conditions. The gross appearance of the hydrogel's surface and cross-section were captured and evaluated. The biocompatibility testing of HDFs and cell-bioink interaction towards the GPVA was analyzed by using live/dead assay, cell migration activity, cell proliferation assay, cell morphology (SEM) and protein expression via immunocytochemistry. The crosslinked hydrogels significantly demonstrated optimum average pore size (100-199 μm). The GPVA crosslinked with GNP (GPVA_GNP) hydrogels with 3.0 × 106 HDFs was proven to be outstanding, compared to the other hydrogels, in biocompatibility testing to promote cellular interaction. Moreover, GPVA-GNP hydrogels, encapsulated with 3.0 × 106 HDFs under submerged cultivation, had a better outcome than air-lifting with an excellent surface cell viability rate of 96 ± 0.02%, demonstrated by 91.3 ± 4.1% positively expressed Ki67 marker at day 14 that represented active proliferative cells, an average of 503.3 ± 15.2 μm for migration distance, and maintained the HDFs' phenotypic profiles with the presence of collagen type I expression. It also presented with an absence of alpha-smooth muscle actin positive staining. In conclusion, 3.0 × 106 of hybrid GPVA hydrogel crosslinked with GNP, produced by submerged cultivation, was proven to have the excellent biocompatibility properties required to be a potential bioinks for the rapid manufacturing of 3D in vitro of a single dermal layer for future use in cosmetic, pharmaceutic and toxicologic applications.

Experimental investigations of warm incremental sheet forming process on magnesium AZ31 and aluminium 6061 alloy

Aerospace, automobile and biomedical sector focuses on innovative rapid manufacturing processes, which can enable product customization, reduction in cost and manufacturing lead time. Aerospace, automobile and biomedical industries mostly use magnesium AZ31 and aluminium 6061 alloy materials for manufacturing of products because of their lower elastic modulus, excellent corrosion and fatigue resistance. The conventional sheet metal forming processes often require additional tools like punch and die and would not be economically viable to produce the customized structure for applications. An emerging forming technique known as warm incremental sheet forming process is evolved, which is capable of forming intricate, asymmetrical components at elevated temperature with localized deformation method. The characteristics of warm incremental sheet forming process suggest the need for the investigation of magnesium AZ31 and aluminium 6061 alloy materials for wide application of the process. Experimental investigation on the influence of process parameters on warm incremental sheet metal forming of magnesium AZ31 and aluminium 6061 alloy materials using electric heating technique was studied. Experimental results revealed that magnesium AZ31 and aluminium 6061 alloy sheet metal component formed with incremental depth of 0.1 mm, wall angle of 50° and temperature of 300°C showed maximum formability and thickness reduction with minimum geometrical deviation.

Creep behavior of polyamide 12, produced by selective laser sintering with different build orientations

The successful use of components produced by selective laser sintering as a rapid manufacturing process requires a comprehensive understanding of the material. In this study, the effect of specimen build orientation on mechanical properties of selective laser-sintered polyamide 12 was investigated in detail. Samples were printed with an orientation of 0°, 15°, 45°, and 90° to the build platform. In addition to quasi-static tensile tests, creep tests under different loads (5 MPa, 10 MPa, 15 MPa, and 20 MPa) and for different times (10 h and 1000 h) with and without relaxation were performed. Creep behavior was analyzed using the Burgers model. Therefore, the elastic strain, the relaxant strain, the viscous strain, and the total deformation were determined. Results show that the build orientation has no significant influence on the long-term creep behavior, at small stresses. Short-term creep and relaxation tests show that the elastic and viscous strain are only slightly influenced by the build orientation. However, the viscoelastic strain is affected by the build orientation. Furthermore, the deformations resulting from creep and relaxation have no significant influence on the mechanical behavior as shown by tensile tests.

Abstract 2821: Incorporation of intrinsic checkpoint blockade enhances functionality of multigenic autologous UltraCAR-T® cells manufactured using non-viral gene delivery and rapid manufacturing process

Abstract Current chimeric antigen receptor (CAR)-T manufacturing utilizes viral vectors and extensive ex vivo expansion at large central facilities leading to an exhausted CAR-T phenotype, high costs and long vein-to-vein times. While allogeneic CAR-T can reduce delays in patient treatment, they require extensive manipulation of donor cells, severe lymphodepletion and demonstrate short persistence limiting their therapeutic window. The UltraCAR-T® platform is designed to overcome these limitations by utilizing non-viral multigene delivery and a rapid, decentralized manufacturing process without ex vivo activation or expansion of T cells. Patient’s own T cells are collected and manufactured at the medical center and re-infused one day after gene transfer. Here we describe the next generation UltraCAR-T platform that addresses the inhibitory tumor microenvironment by incorporating a novel mechanism for intrinsic downregulation of one or more checkpoint inhibitor (CPI) genes. Our design achieves intrinsic CPI blockade without gene editing and is aimed at avoiding systemic toxicity and the high cost of combining CPI antibodies. Next generation UltraCAR-T cells simultaneously express CAR, membrane bound IL-15 (mbIL15), kill switch, and incorporate intrinsic CPI blockade. To illustrate the ability of this platform, we designed exemplary non-viral transposons to generate UltraCAR-T cells against multiple tumor targets incorporating intrinsic blockade of either one (PD-1) or two (PD-1 and TIGIT) CPI genes. Healthy donor T cells were transfected using the UltraPorator™ electroporation system to manufacture UltraCAR-T cells without ex vivo activation or expansion. The co-expression of CAR, mbIL15 and kill switch was confirmed by flow cytometry and western blot. Activated UltraCAR-T showed significant reduction in CPI gene expression compared to control CAR-T cells lacking the CPI blockade and did not show unintended off-target activity. Downregulation of CPI gene(s) on UltraCAR-T enhanced cytotoxicity and inflammatory cytokine production, especially at low effector to target (E:T) cell ratios, when co-cultured with PD-L1+/CD155+ tumor cells. Single-cell cytokine proteomics showed significant increase in polyfunctionality of UltraCAR-T with intrinsic downregulation of CPI gene(s). In vivo, a single infusion of receptor tyrosine kinase-like orphan receptor 1 (ROR1)-specific UltraCAR-T with intrinsic PD-1 blockade resulted in rapid expansion, an increase in preferred T cell memory (TSCM/TCM) populations, and significantly improved overall survival of ROR1+ PD-L1+ tumor bearing mice. These preclinical data highlight the improved efficacy of incorporating intrinsic CPI blockade in UltraCAR-T cells using non-viral gene delivery and an established rapid, decentralized manufacturing process. Citation Format: Tim Chan, Cheryl Bolinger, Sean Scott, Mengyan Du, Carol Poortman, Byron Koenitzer, Taranjit Athwal, Lindsey Shepard, R. Daniel Slone, Shourik Dutta, Steven Zilko, James M. Dunleavey, Giorgio Zenere, Jacques Plummer, Bernward Klocke, Christian Zinser, Shamim Ahmad, Douglas E. Brough, Rutul R. Shah, Helen Sabzevari. Incorporation of intrinsic checkpoint blockade enhances functionality of multigenic autologous UltraCAR-T® cells manufactured using non-viral gene delivery and rapid manufacturing process [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2821.

Identification and Development of PHE885: A Novel and Highly Potent Fully Human Anti-BCMA CAR-T Manufactured with a Novel T-Charge TM Platform for the Treatment of Multiple Myeloma

Identification and Development of PHE885: A Novel and Highly Potent Fully Human Anti-BCMA CAR-T Manufactured with a Novel T-Charge TM Platform for the Treatment of Multiple Myeloma

IoT device fabrication using roll-to-roll printing process

With the development of technology, wireless and IoT devices are increasingly used from daily life to industry, placing demands on rapid and efficient manufacturing processes. This study demonstrates the fabrication of an IoT device using a roll-to-roll printing process, which could shorten the device fabrication time and reduce the cost of mass production. Here, the fabricated IoT device is designed to acquire data through the sensor, process the data, and communicate with end-user devices via Bluetooth communication. For fabrication, a four-layer circuit platform consisting of two conductive layers, an insulating layer including through holes, and a solder resist layer is directly printed using a roll-to-roll screen printing method. After the printing of the circuit platform, an additional layer of solder paste is printed to assemble the electrical components into the device, inspiring the fully roll-to-roll process for device fabrication. Successful IoT device deployment opens the chance to broaden the roll-to-roll fabrication process to other flexible and multilayer electronic applications.