RNA aggregates harness the danger response for potent cancer immunotherapy.

Cancer immunotherapy remains limited by poor antigenicity and a regulatory tumor microenvironment (TME). Here, we create "onion-like" multi-lamellar RNA lipid particle aggregates (LPAs) to substantially enhance the payload packaging and immunogenicity of tumor mRNA antigens. Unlike current mRNA vaccine designs that rely on payload packaging into nanoparticle cores for Toll-like receptor engagement in immune cells, systemically administered RNA-LPAs activate RIG-I in stromal cells, eliciting massive cytokine/chemokine response and dendritic cell/lymphocyte trafficking that provokes cancer immunogenicity and mediates rejection of both early- and late-stage murine tumor models. In client-owned canines with terminal gliomas, RNA-LPAs improved survivorship and reprogrammed the TME, which became "hot" within days of a single infusion. In a first-in-human trial, RNA-LPAs elicited rapid cytokine/chemokine release, immune activation/trafficking, tissue-confirmed pseudoprogression, and glioma-specific immune responses in glioblastoma patients. These data support RNA-LPAs as a new technology that simultaneously reprograms the TME while eliciting rapid and enduring cancer immunotherapy.

CD19/22 CAR T cells in children and young adults with B-ALL: phase 1 results and development of a novel bicistronic CAR.

Remission durability following single-antigen targeted chimeric antigen receptor (CAR) T-cells is limited by antigen modulation, which may be overcome with combinatorial targeting. Building upon our experiences targeting CD19 and CD22 in B-cell acute lymphoblastic leukemia (B-ALL), we report on our phase 1 dose-escalation study of a novel murine stem cell virus (MSCV)-CD19/CD22-4-1BB bivalent CAR T-cell (CD19.22.BBζ) for children and young adults (CAYA) with B-cell malignancies. Primary objectives included toxicity and dose finding. Secondary objectives included response rates and relapse-free survival (RFS). Biologic correlatives included laboratory investigations, CAR T-cell expansion and cytokine profiling. Twenty patients, ages 5.4 to 34.6 years, with B-ALL received CD19.22.BBζ. The complete response (CR) rate was 60% (12 of 20) in the full cohort and 71.4% (10 of 14) in CAR-naïve patients. Ten (50%) developed cytokine release syndrome (CRS), with 3 (15%) having ≥ grade 3 CRS and only 1 experiencing neurotoxicity (grade 3). The 6- and 12-month RFS in those achieving CR was 80.8% (95% confidence interval [CI]: 42.4%-94.9%) and 57.7% (95% CI: 22.1%-81.9%), respectively. Limited CAR T-cell expansion and persistence of MSCV-CD19.22.BBζ compared with EF1α-CD22.BBζ prompted laboratory investigations comparing EF1α vs MSCV promoters, which did not reveal major differences. Limited CD22 targeting with CD19.22.BBζ, as evaluated by ex vivo cytokine secretion and leukemia eradication in humanized mice, led to development of a novel bicistronic CD19.28ζ/CD22.BBζ construct with enhanced cytokine production against CD22. With demonstrated safety and efficacy of CD19.22.BBζ in a heavily pretreated CAYA B-ALL cohort, further optimization of combinatorial antigen targeting serves to overcome identified limitations (www.clinicaltrials.gov #NCT03448393).

Cutaneous Lagenidium deciduum infection in a patient with relapsed acute myeloid leukemia.

BACKGROUND: Lagenidium deciduum is an oomycete that can cause infections in mammals that present similarly to pythiosis and mucormycosis. Most of the existing case reports have occurred in canines and have been fatal. In animals, medical therapy has not been successful, so surgical excision is the mainstay of treatment. Lagenidium sp. infections in humans are rare. There is only one case of a human Lagenidium sp. infection in the literature, and it presented as an ocular infection. The human ocular infection was resistant to medical therapy and required a penetrating keratoplasty for cure. Additional reports of effective therapy are needed to guide management of this emerging pathogen. We present the first case of a cutaneous Lagenidium deciduum infection in a human patient, which is also the first documented case of a Lagenidium deciduum infection in an immunocompromised host of any species. CASE PRESENTATION: An 18-year-old female with relapsed acute myeloid leukemia, awaiting a haploidentical stem cell transplant, presented with erythematous cutaneous lesions on her left hip and bilateral buttocks that enlarged and blackened over several days. About 1 week later, boil-like lesions appeared on her bilateral buttocks. The skin lesions were initially presumed to be bacterial in origin, so the patient was treated with clindamycin and cefepime with little improvement. Upon further investigation, fungal cultures and skin biopsies revealed aseptate hyphae, so the patient was switched to isavuconazole and amphotericin B due to concern for mucormycosis. Phenotypic characterization and DNA sequencing were performed by the Fungus Testing Laboratory, University of Texas Health Science Center at San Antonio, which identified the causal fungal organism as Lagenidium deciduum. All of her cutaneous lesions were surgically excised, and the patient was treated with micafungin, terbinafine, doxycycline, and azithromycin. Micafungin and terbinafine were continued until she achieved engraftment post-transplant. CONCLUSIONS: We report the first successful treatment of a human Lagenidium infection in an immunocompromised host through a combination of aggressive surgical excision and prolonged antifungal therapy during the prolonged neutropenia associated with allogeneic stem cell transplant. Prompt diagnosis and management may prevent disseminated oomycosis.

The role of donor type and pre-transplant immunosuppression on outcomes of hematopoietic stem cell transplantation in children and young adults with severe aplastic anemia.

BACKGROUND: The goal of this study was to assess the effect of donor type and pre-transplant immunotherapy (IST) on outcomes of hematopoietic stem cell transplantation (HSCT) for children and young adults with severe aplastic anemia (SAA). METHODS: This retrospective, multi-center study included 52 SAA patients, treated in 5 pediatric transplant programs in Florida, who received HSCT between 2010 and 2020 as the first- or second-line treatment. RESULTS: The median age at HSCT for all 52 patients was 15 years (range 1-25). The 3-year overall survival (OS) by donor type were as follows: 95% [95% CI 85.4-99] for matched related donors (MRD) (N = 24), 84% [95% CI 63.5-99] for haploidentical (N = 13), and 71% [95% CI 36-99] for matched unrelated donors (MUD) (N = 7). The 3-year OS was 81% [95% CI 69.7-99] for all patients, 90.5% [95% CI 79.5-99] for non-IST patients (N = 27), and 70% [95% CI 51-99] for IST patients (N = 24) (log-rank p = .04). Survival of haploidentical HSCT (haplo-HSCT) recipients with post-transplant cyclophosphamide (PTCy) (N = 13) was excellent for both groups: 100% for non-IST patients (N = 3) and 80% for IST patients (N = 10). The 3-year OS for patients with previous IST by donor type in groups where >5 patients were available was 78.8% [95% CI 52.3-99] for haplo-HSCT (N = 10) and 66.7% [95% CI 28.7-99] for MUD (N = 6). Although it appears that patients receiving HSCT ≥6 months after the start of IST had worse survival, the number of patients in each category was small and log-rank was not significant(p = .65). CONCLUSIONS: Patients receiving MUD and haplo-HSCT with PTCy had similar outcomes, suggesting that haplo-HSCT with PTCy could be included in randomized trials of upfront IST versus alternative donor HSCT.

Emerging frontiers in immuno- and gene therapy for cancer.

BACKGROUND AIMS: The field of cell and gene therapy in oncology has moved rapidly since 2017 when the first cell and gene therapies, Kymriah followed by Yescarta, were approved by the Food and Drug Administration in the United States, followed by multiple other countries. Since those approvals, several new products have gone on to receive approval for additional indications. Meanwhile, efforts have been made to target different cancers, improve the logistics of delivery and reduce the cost associated with novel cell and gene therapies. Here, we highlight various cell and gene therapy-related technologies and advances that provide insight into how these new technologies will speed the translation of these therapies into the clinic. CONCLUSIONS: In this review, we provide a broad overview of the current state of cell and gene therapy-based approaches for cancer treatment - discussing various effector cell types and their sources, recent advances in both CAR and non-CAR genetic modifications, and highlighting a few promising approaches for increasing in vivo efficacy and persistence of therapeutic drug products.

Sampling Multiple Catheter Lumens to Improve Detection of Bloodstream Infection in Pediatric Oncology Patients.

Current guidelines recommend sampling each central-access lumen during the initial evaluation of febrile pediatric oncology patients. We investigated this recommendation's validity at centers implementing a diagnostic stewardship program to reduce blood cultures in critically ill children. Among 146 oncology patients admitted to the intensive care unit, there were 34 eligible blood culture-sets. Eleven (34%) sets yielded discordant results, most commonly cultivating a likely pathogen from one lumen and no growth from another. As hospitals move toward reducing testing overuse, these results emphasize the continued importance of culturing each central-access lumen to optimize the detection of bacteremia in the initial evaluation of critically ill pediatric oncology patients.

Consequences of hemophagocytic lymphohistiocytosis-like cytokine release syndrome toxicities and concurrent bacteremia.

Serious bacterial infections (SBI) can lead to devastating complications with CD19 CAR T cells and cytokine release syndrome (CRS). Little is known about consequences of and risk factors for SBI with novel CAR T-cell constructs or with CRS complicated by HLH-like toxicities. We report on three patients with B-cell acute lymphoblastic leukemia treated with CD22 CAR T cells who developed SBI and CRS-associated HLH. Serum cytokine profiling revealed sustained elevations well beyond CRS resolution, suggesting ongoing systemic inflammation. Heightened inflammatory states converging with SBI contribute to poor outcomes, and recognition and prevention of extended inflammation may be needed to improve outcomes.

Desmoid tumor mimics local recurrence of extremity sarcoma on MRI.

BACKGROUND AND OBJECTIVES: The development of desmoid fibromatosis after tumor resection may mimic local recurrence. To our knowledge, this phenomenon has not been reported after extremity sarcoma resection. We report four cases of desmoid-type fibromatosis ("desmoid tumors") mimicking local recurrence after extremity sarcoma resection. METHODS: We retrospectively reviewed the records of patients treated for extremity sarcoma by our orthopedic oncology service from 2014 to 2019 and identified four patients with biopsy-proven desmoid tumors. We extracted clinical, pathologic, radiographic, and operative data for the primary neoplasms and desmoid tumors. RESULTS: Four patients with postresection surveillance magnetic resonance imaging suspicious for local recurrence underwent further analysis showing desmoid tumors. Patients underwent image-guided needle biopsy, with specimens demonstrating fibromatosis-type histologic characteristics. Two cases were ß-catenin positive. Desmoid tumors were managed with observation. No patient had experienced local or distant recurrence of the primary tumor at a mean follow-up of 30 months after resection (range, 23-34 months); none underwent surgery for symptoms of desmoid tumors. CONCLUSIONS: Desmoid tumors should be considered part of the differential diagnosis when assessing patients with radiographic concern for postresection local recurrence of extremity bone and soft-tissue sarcoma. An image-guided needle biopsy can inform diagnosis and management.

Invasive fusariosis masquerading as extramedullary disease in rapidly progressive acute lymphoblastic leukemia.

Invasive fusariosis (IF) most commonly occurs in patients with hematologic malignancies and severe neutropenia, particularly during concomitant corticosteroid use. Breakthrough infections can occur in high-risk patients despite Aspergillus-active antifungal prophylaxis. We describe a patient with rapid acute lymphoblastic leukemia (ALL) progression who presented with multifocal skin nodules thought to be choloromatous disease. These lesions were ultimately diagnosed as IF and the patient had two simultaneously active disease processes. This case highlights the importance of pathologic diagnosis of new skin lesions in ALL patients, even during leukemia progression, and demonstrates that IF can occur despite normal neutrophil counts and Aspergillus-active prophylaxis.

Differentiation of Deep Venous Thrombosis Among Children With or Without Osteomyelitis.

BACKGROUND: Children with osteomyelitis are at risk for deep venous thrombosis (DVT). This study evaluates the characteristics of DVT among children to differentiate between those with and without osteomyelitis. METHODS: Children with DVT of any cause were studied between 2008 and 2016. Children with DVT and osteomyelitis were compared with those with DVT without osteomyelitis. Another comparison cohort included children with osteomyelitis but without DVT. Comorbidities, severity of illness (SOI), and clinical course were compared between cohorts. RESULTS: DVT was identified in 224 children, a prevalence of 2.5 per 10,000 children. Among those with DVT, 28 (12.1%) had osteomyelitis. The DVT rate among 466 children with osteomyelitis was 6.0%. Children with osteomyelitis and DVT had greater SOI (9.1 vs. 2.7), bacteremia rate (82.1% vs. 38.4%), methicillin-resistant Staphylococcus aureus rate (89.3% vs. 21.2%), surgeries per child (2.1 vs. 0.7), and intensive care unit admission rate (67.9% vs. 5.9%) than that of children without DVT (P<0.00001). Of 196 children who had DVT without osteomyelitis, 166 (84.7%) had comorbidities including defined hypercoagulability (27 or 13.8%). Children with DVT due to osteomyelitis were without comorbidities or hypercoagulability (P<0.00001). The rate of pulmonary embolism was similar for children with DVT with or without osteomyelitis (3/28, or 10.7% vs. 18/196, or 9.2%). CONCLUSIONS: Children with DVT and osteomyelitis differ substantially from other children with DVT by the absence of comorbidities or post-thrombotic syndrome. They also differ from children with osteomyelitis without DVT by higher SOI, methicillin-resistant S. aureus rate, and occurrence of intensive care. Awareness of for the characteristics of DVT among children with osteomyelitis will reduce delay to diagnostic ultrasound and improve anticoagulation management which must be carefully coordinated given the high rate of surgery of these children. LEVEL OF EVIDENCE: Level II-prognostic, retrospective cohort comparison.

Pediatric Cancer Immunotherapy and Potential for Impact on Fertility: A Need for Evidence-Based Guidance.

The use of immunotherapies for the treatment of cancer in children, adolescents, and young adults has become common. As the use of immunotherapy has expanded, including in earlier lines of therapy, it has become evident that several aspects of how these immunotherapies impact longer-term outcomes among survivors are understudied. Traditional cancer therapies like alkylating and platin agents carry the greatest risk of infertility, but little is known about the impact of novel immunotherapies on fertility. This topic is of great interest to patients, patient advocates, and clinicians. In this article, we review immunotherapeutic agents used to treat childhood and young adult cancers and discuss potential mechanisms by which they may impact fertility based on the known interplay between the immune system and reproductive organs. We highlight the relative paucity of high-quality literature examining these late effects. We discuss interventions to optimize fertility preservation (FP) for our patients. Conducting longitudinal, collaborative, and prospective research on the fertility outcomes of pediatric and young adult patients with cancer who receive immunotherapy is critical to learn how to effectively counsel our patients on long-term fertility outcomes and indications for FP procedures. Collection of patient-level data will be necessary to draft evidence-based guidelines on which providers can make therapy recommendations.

INSPIRED Symposium Part 4B: Chimeric Antigen Receptor T Cell Correlative Studies-Established Findings and Future Priorities.

Chimeric antigen receptor (CAR) T cell therapy has revolutionized the treatment of B cell malignancies, with multiple CAR T cell products approved for numerous indications by regulatory agencies worldwide. However, significant work remains to be done to enhance these treatments. In March 2023, a group of experts in CAR T cell therapy assembled at the National Institutes of Health in Bethesda, Maryland at the Insights in Pediatric CAR T Cell Immunotherapy: Recent Advances and Future Directions (INSPIRED) Symposium to identify key areas for research for the coming years. In session 4B, correlative studies to be incorporated into future clinical trials and real-world settings were discussed. Active areas of research identified included (1) optimizing CAR T cell product manufacturing; (2) ensuring adequate lymphodepletion prior to CAR T cell administration; (3) overcoming immunoregulatory cells and tumor stroma present in the tumor microenvironment, particularly in solid tumors; (4) understanding tumor intrinsic properties that lead to CAR T cell immunotherapy resistance; and (5) uncovering biomarkers predictive of treatment resistance, treatment durability, or immune-related adverse events. Here we review the results of previously published clinical trials and real-world studies to summarize what is currently known about each of these topics. We then outline priorities for future research that we believe will be important for improving our understanding of CAR T cell therapy and ultimately leading to better outcomes for patients.

A multidisciplinary pediatric oncofertility team improves fertility preservation and counseling across 7 years.

BACKGROUND: Oncofertility is a developing field of increasing importance, particularly in pediatric oncology, where most patients are likely to survive long-term and have not yet had the opportunity to have children. AIMS: We performed a quality improvement initiative to increase our rates of fertility preservation counseling and referral through the implementation of a pediatric oncofertility team, and we report outcomes 7 years following implementation of our initiative. METHODS AND RESULTS: We compare our baseline oncofertility survey to 44 post-intervention survey respondents and electronic medical record documentation for 149 patients treated in 2019. Ninety-five percent of post-intervention survey respondents recalled fertility counseling (baseline 70%, p = .004) and 89.3% were appropriately referred for fertility preservation (baseline 50%, p = .017). Counseling was documented in 60.4% of charts; 81% of patients analyzed by chart review were appropriately referred for fertility preservation. Fertility preservation outcomes differed by sex assigned at birth. CONCLUSION: Creation of an oncofertility team produced improvements in fertility counseling and fertility preservation referral across an extended period of time.

Three-Dimensional Bioconjugated Liquid-Like Solid (LLS) Enhance Characterization of Solid Tumor - Chimeric Antigen Receptor T cell interactions.

Cancer immunotherapy offers lifesaving treatments for cancers, but the lack of reliable preclinical models that could enable the mechanistic studies of tumor-immune interactions hampers the identification of new therapeutic strategies. We hypothesized 3D confined microchannels, formed by interstitial space between bio-conjugated liquid-like solids (LLS), enable CAR T dynamic locomotion within an immunosuppressive TME to carry out anti-tumor function. Murine CD70-specific CAR T cells cocultured with the CD70-expressing glioblastoma and osteosarcoma demonstrated efficient trafficking, infiltration, and killing of cancer cells. The anti-tumor activity was clearly captured via longterm in situ imaging and supported by upregulation of cytokines and chemokines including IFNg, CXCL9, CXCL10, CCL2, CCL3, and CCL4. Interestingly, target cancer cells, upon an immune attack, initiated an "immune escape" response by frantically invading the surrounding microenvironment. This phenomenon however was not observed for the wild-type tumor samples which remained intact and produced no relevant cytokine response. Single cells collection and transcriptomic profiling of CAR T cells at regions of interest revealed feasibility of identifying differential gene expression amongst the immune subpopulations. Complimentary 3D in vitro platforms are necessary to uncover cancer immune biology mechanisms, as emphasized by the significant roles of the TME and its heterogeneity.

mRNA challenge predicts brain cancer immunogenicity and response to checkpoint inhibitors.

To prospectively determine whether brain tumors will respond to immune checkpoint inhibitors (ICIs), we developed a novel mRNA vaccine as a viral mimic to elucidate cytokine release from brain cancer cells in vitro. Our results indicate that cytokine signatures following mRNA challenge differ substantially from ICI responsive versus non-responsive murine tumors. These findings allow for creation of a diagnostic assay to quickly assess brain tumor immunogenicity, allowing for informed treatment with ICI or lack thereof in poorly immunogenic settings.

Bioconjugated liquid-like solid enhances characterization of solid tumor - chimeric antigen receptor T cell interactions.

Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable success as an immunotherapy for hematological malignancies, and its potential for treating solid tumors is an active area of research. However, limited trafficking and mobility of T cells within the tumor microenvironment (TME) present challenges for CAR T cell therapy in solid tumors. To gain a better understanding of CAR T cell function in solid tumors, we subjected CD70-specific CAR T cells to a challenge by evaluating their immune trafficking and infiltration through a confined 3D microchannel network in a bio-conjugated liquid-like solid (LLS) medium. Our results demonstrated successful CAR T cell migration and anti-tumor activity against CD70-expressing glioblastoma and osteosarcoma tumors. Through comprehensive analysis of cytokines and chemokines, combined with in situ imaging, we elucidated that immune recruitment occurred via chemotaxis, and the effector-to-target ratio plays an important role in overall antitumor function. Furthermore, through single-cell collection and transcriptomic profiling, we identified differential gene expression among the immune subpopulations. Our findings provide valuable insights into the complex dynamics of CAR T cell function in solid tumors, informing future research and development in this promising cancer treatment approach. STATEMENT OF SIGNIFICANCE: The use of specialized immune cells named CAR T cells to combat cancers has demonstrated remarkable success against blood cancers. However, this success is not replicated in solid tumors, such as brain or bone cancers, mainly due to the physical barriers of these solid tumors. Currently, preclinical technologies do not allow for reliable evaluation of tumor-immune cell interactions. To better study these specialized CAR T cells, we have developed an innovative in vitro three-dimensional model that promises to dissect the interactions between tumors and CAR T cells at the single-cell level. Our findings provide valuable insights into the complex dynamics of CAR T cell function in solid tumors, informing future research and development in this promising cancer treatment approach.

A Phase II Trial of Guadecitabine in Children and Adults with SDH-Deficient GIST, Pheochromocytoma, Paraganglioma, and HLRCC-Associated Renal Cell Carcinoma.

PURPOSE: Succinate dehydrogenase (dSDH)-deficient tumors, including pheochromocytoma/paraganglioma, hereditary leiomyomatosis and renal cell cancer-associated renal cell carcinoma (HLRCC-RCC), and gastrointestinal stromal tumors (GIST) without KIT or platelet-derived growth factor receptor alpha mutations are often resistant to cytotoxic chemotherapy, radiotherapy, and many targeted therapies. We evaluated guadecitabine, a dinucleotide containing the DNA methyltransferase inhibitor decitabine, in these patient populations. PATIENTS AND METHODS: Phase II study of guadecitabine (subcutaneously, 45 mg/m2/day for 5 consecutive days, planned 28-day cycle) to assess clinical activity (according to RECISTv.1.1) across three strata of patients with dSDH GIST, pheochromocytoma/paraganglioma, or HLRCC-RCC. A Simon optimal two-stage design (target response rate 30% rule out 5%) was used. Biologic correlates (methylation and metabolites) from peripheral blood mononuclear cells (PBMC), serum, and urine were analyzed. RESULTS: Nine patients (7 with dSDH GIST, 1 each with paraganglioma and HLRCC-RCC, 6 females and 3 males, age range 18-57 years) were enrolled. Two patients developed treatment-limiting neutropenia. No partial or complete responses were observed (range 1-17 cycles of therapy). Biologic activity assessed as global demethylation in PBMCs was observed. No clear changes in metabolite concentrations were observed. CONCLUSIONS: Guadecitabine was tolerated in patients with dSDH tumors with manageable toxicity. Although 4 of 9 patients had prolonged stable disease, there were no objective responses. Thus, guadecitabine did not meet the target of 30% response rate across dSDH tumors at this dose, although signs of biologic activity were noted.

mRNA aggregates harness danger response for potent cancer immunotherapy.

Messenger RNA (mRNA) has emerged as a remarkable tool for COVID-19 prevention but its use for induction of therapeutic cancer immunotherapy remains limited by poor antigenicity and a regulatory tumor microenvironment (TME). Herein, we develop a facile approach for substantially enhancing immunogenicity of tumor-derived mRNA in lipid-particle (LP) delivery systems. By using mRNA as a molecular bridge with ultrapure liposomes and foregoing helper lipids, we promote the formation of 'onion-like' multi-lamellar RNA-LP aggregates (LPA). Intravenous administration of RNA-LPAs mimics infectious emboli and elicits massive DC/T cell mobilization into lymphoid tissues provoking cancer immunogenicity and mediating rejection of both early and late-stage murine tumor models. Unlike current mRNA vaccine designs that rely on payload packaging into nanoparticle cores for toll-like receptor engagement, RNA-LPAs stimulate intracellular pathogen recognition receptors (RIG-I) and reprogram the TME thus enabling therapeutic T cell activity. RNA-LPAs were safe in acute/chronic murine GLP toxicology studies and immunologically active in client-owned canines with terminal gliomas. In an early phase first-in-human trial for patients with glioblastoma, we show that RNA-LPAs encoding for tumor-associated antigens elicit rapid induction of pro-inflammatory cytokines, mobilization/activation of monocytes and lymphocytes, and expansion of antigen-specific T cell immunity. These data support the use of RNA-LPAs as novel tools to elicit and sustain immune responses against poorly immunogenic tumors.

Mediport use as an acceptable standard for CAR T cell infusion.

Introduction: Mediport use as a clinical option for the administration of chimeric antigen receptor T cell (CAR T cell) therapy in patients with B-cell malignancies has yet to be standardized. Concern for mediport dislodgement, cell infiltration, and ineffective therapy delivery to systemic circulation has resulted in variable practice with intravenous administration of CAR T cell therapy. With CAR T cell commercialization, it is important to establish practice standards for CAR T cell delivery. We conducted a study to establish usage patterns of mediports in the clinical setting and provide a standard of care recommendation for mediport use as an acceptable form of access for CAR T cell infusions. Methods: In this retrospective cohort study, data on mediport use and infiltration rate was collected from a survey across 34 medical centers in the Pediatric Real-World CAR Consortium, capturing 504 CAR T cell infusion routes across 489 patients. Data represents the largest, and to our knowledge sole, report on clinical CAR T cell infusion practice patterns since FDA approval and CAR T cell commercialization in 2017. Results: Across 34 sites, all reported tunneled central venous catheters, including Broviac® and Hickman® catheters, as accepted standard venous options for CAR T cell infusion. Use of mediports as a standard clinical practice was reported in 29 of 34 sites (85%). Of 489 evaluable patients with reported route of CAR T cell infusion, 184 patients were infused using mediports, with no reported incidences of CAR T cell infiltration. Discussion/Conclusion: Based on current clinical practice, mediports are a commonly utilized form of access for CAR T cell therapy administration. These findings support the safe practice of mediport usage as an accepted standard line option for CAR T cell infusion.