Immunotherapy: SYNERGISTIC PERITONEAL ANTI-TUMOR EFFECT OF ALLOCETRAOTS, A CELLULAR IMMUNE-THERAPY, IN COMBINATION WITH IMMUNE CHECKPOINT INHIBITORS/CHEMOTHERAPY, THROUGH IN-VIVO REPROGRAMMING OF MACROPHAGES AND DENDRITIC CELLS

Background & Aim: Immune checkpoint inhibitors (ICI) revolutionized solid tumor treatment, however, in many tumors only partial response is achieved. Allocetra-OTS has an immune modulating effect on macrophages and dendritic cells and showed an excellent safety profile in patients including patients with sepsis and Covid-19. Here we investigated the anti-tumoral effect of Allocetra-OTS cellular therapy, in peritoneal solid tumor animal models. Methods, Results & Conclusion(s): Allocetra-OTS is manufactured from enriched mononuclear fractions and induced to undergo early apoptosis. Balb/c mice were inoculated intraperitoneally (IP) with AB12 (mesothelioma) with pLenti-PGK-V5-Luc-Neo and treated with anti- CTLA4 with or without Allocetra-OTS. Mice were monitored daily for clinical score and weekly using IVIS (Fig.1). Kaplan-Meier log rank test was done for survival. For Allocetra-OTS preparation, enriched mononuclear fractions were collected by leukapheresis from healthy eligible human donors and induced to undergo early apoptosis. Anti- CTLA4 standalone therapy significantly improved survival (Fig.2) from mean 34+/-9 to 44.9 +/-20 days. However, OTS standalone therapy was non-inferior and improved survival to 52.3 +/-20 days. Anti-CTLA4 + Allocetra-OTS combination therapy, ameliorated survival to 86.7+/-20 days with complete cancer remission in 60-100% of mice. Similar anti- tumoral effects of Allocetra-OTS were seen in mesothelioma model in a combination therapy with either anti-PD1 or cisplatin and using anti-PD1 in ID8 ovary cancer model. Based on single cell analysis confirmed by flow cytometry and pathology, the mechanism of action seems to be related or at least associated with an increase in f/480high peritoneal macrophages and a decrease in recruited macrophages, and to f/480high infiltration of the tumor. However, further studies are needed to confirm these observations. During IP tumor progression, Allocetra-OTS as a standalone therapy or in combination with ICI, or cisplatin, significantly reduced tumor size and resulted in complete remission in up to 100% treated mice. Similar results were obtained in ID8 ovary cancer. Based on excellent safety profile in > 50 patients treated in prior clinical trials for sepsis and Covid-19, Phase I/II clinical trial of Allocetra-OTS plus chemotherapy has started and three patient already recruited. A second phase I/II clinical trial of Allocetra- OTS plus anti-PD1, as a second- and third-line therapy in various cancers, was initiated in Q1 2023. [Figure presented]Copyright © 2023 International Society for Cell & Gene Therapy

Trained innate immunity and diseases: Bane with the boon

Emerging research shows that innate immunity can also keep the memory of prior experiences, challenging the long-held notion that immunological memory is only the domain of the adaptive immune cells. However, the absence of immunological memory in innate immune responses has recently been brought into question. Now it is known that after a few transient activations, innate immune cells may acquire immunological memory phenotype, resulting in a stronger response to a subsequent secondary challenge. When exposed to particular microbial and/or inflammatory stimuli, trained innate immunity is characterized by the enhanced non-specific response, which is regulated by substantial metabolic alterations and epigenetic reprogramming. Trained immunity is acquired by two main reprogramming, namely, epigenetic reprogramming and metabolic adaptation/reprogramming. Epigenetic reprogramming causes changes in gene expression and cell physiology, resulting in internal cell signaling and/or accelerated and amplified cytokine release. Metabolic changes due to trained immunity induce accelerated glycolysis and glutaminolysis. As a result, trained immunity can have unfavorable outcomes, such as hyper inflammation and the development of cardiovascular diseases, autoinflammatory diseases, and neuroinflammation. In this review, the current scenario in the area of trained innate immunity, its mechanisms, and its involvement in immunological disorders are briefly outlined.Copyright © 2022

Patterns of Progression and Treatment Discontinuation in a Real Life Latin American Prospective Cohort Study of Intermediate-Advanced Hepatocellular Carcinoma: Second Interim Analysis

Introduction and Objectives: Previously published regional real-world results of overall survival (OS) in Barcelona Clinic Liver Cancer (BCLC) B and C patients demanded a prospective cohort study nested in a systematic and continuous medical educational networking group. This study aimed to describe and evaluate the treatment decisions in patients with hepatocellular carcinoma (HCC) within BCLC B and C stages. Material(s) and Method(s): A multicenter prospective cohort study, conducted in different Latin American centers from Argentina, Brazil and Colombia, started on 15th May 2018 (delayed recruitment during COVID locked-down period). Patients within BCLC B or C stages were included. Survival, tumor progression and patterns of treatment suspension were evaluated. Result(s): At this second interim analysis (projected final analysis March 2023), 390 HCC BCLC-B or C patients were included (n=15 excluded);mean age 65 years, 75.6% males and 89.5% cirrhotic. Median OS since HCC diagnosis was 27.2 months. Among BCLC-B patients, the most frequent therapy was transarterial chemoembolization (TACE, 42.3%);51.8% using drug-eluting beads and 47.4% conventional TACE;with a median OS since 1st TACE of 41.9 months. Similar radiological responses after 1st TACE were observed between both modalities. Overall, 48.2% of the cohort received systemic therapy for HCC (n=188), 23.7% still on BCLC-B stage. The most frequent systemic treatments were Sorafenib (74.5%), atezolizumab bevacizumab (17.5%), and lenvatinib (12.2%), with a median OS since systemic therapy of 15.7 months. Lenvatinib or atezolizumab bevacizumab was used as the second line following sorafenib in 5 and 3 patients, respectively. The most common causes of systemic treatment discontinuation were tumor progression and liver function deterioration (15% to 36.4%). Patterns of tumor progression were not specifically associated with prognosis or treatment discontinuation. Conclusion(s): Liver function deterioration occurs in a third of patients following systemic therapies. The complexity of treatment decisions underly the need for a multidisciplinary team and the role of hepatologists.Copyright © 2023

Landscape Analysis of Quercetin: A Potential Candidate Against SARS-CoV-2

Fruit, vegetables, and green tea contain quercetin (a flavonoid). Some of the diet's most signifi-cant sources of quercetin are apples, onions, tomatoes, broccoli, and green tea. Antioxidant, anticancer, anti-inflammatory, antimicrobial, antibacterial, and anti-viral effects have been studied of quercetin. The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) virus, ribonucleic acid (RNA) polymer-ase, and other essential viral life-cycle enzymes are all prevented from entering the body by quercetin. Despite extensive in vitro and in vivo investigations on the immune-modulating effects of quercetin and vitamin C treatment. 3-methyl-quercetin has been shown to bind to essential proteins necessary to convert minus-strand RNA into positive-strand RNAs, preventing the replication of viral RNA in the cytoplasm. Quercetin has been identified as a potential SARS-CoV-2 3C-like protease (3CLpro) suppressor in recent molecular docking studies and in silico assessment of herbal medicines. It has been demonstrated that quercetin increases the expression of heme oxygenase-1 through the nuclear factor erythroid-related factor 2 (Nrf2) signal network. Inhibition of heme oxygenase-1 may increase bilirubin synthesis, an endoge-nous antioxidant that defends cells. When human gingival fibroblast (HGF) cells were exposed to lipo-polysaccharide (LPS), inflammatory cytokine production was inhibited. The magnesium (Mg+2) cation complexation improves quercetin free radical scavenging capacity, preventing oxidant loss and cell death. The main objective of this paper is to provide an overview of the pharmacological effects of quercetin, its protective role against SARS-CoV-2 infection, and any potential molecular processes.Copyright © 2022 Bentham Science Publishers.

Surgical oncology during the post-COVID-19 era: What is next?

During first outburst of COVID-19, several strategies had been applied for surgical oncology patients to minimize COVID-19 transmission. COVID-19 infection seemed to compromise survival and major complication rates of surgical oncology patients. However, survival, tumor progression and recurrence rates of surgical oncology patients were associated to the consequences of COVID-19 pandemic on their management. In addition, the severity of COVID-19 infections has been downgraded. Therefore, management of surgical oncology patients should be reconsidered. © 2022 Wiley Periodicals LLC.

Infiltracao Medular Por Leucemia Mieloide Aguda (Lma) E Tumor Solido (Ts) Metastatico: Relato De Caso

Objetivo: Descrever um caso com duas neoplasias concomitantes infiltrando medula ossea (MO), sendo uma delas hematologica e outra, TS. Material e metodos: Relato de caso por meio de coleta de dados de prontuario de paciente acompanhado no HAC. Resultados: Paciente 67 anos, masculino, tabagista (100 anos/maco), ex-etilista, historia de inalacao de tintas com benzeno (pintor de carros por 30 anos), com astenia e anemia ha 4 meses, associado a perda de 25kg. Procurou atendimento em marco/22 com diagnostico de COVID-19 e necessidade de internacao hospitalar. Evoluiu com bicitopenia sendo transferido ao HAC para investigacao. Ao exame fisico: ECOG 3. Hemograma: bicitopenia (Hb 5.8 mg/dL;plaquetas 40000/mm3;leucocitos 13000/mm3;69% blastos). TC: derrame pleural a direita;raros micronodulos pulmonares;enfisema pulmonar difuso acentuado com areas de fibrose associadas;linfonodomegalias mediastinais e hilares, lesoes osteoliticas esparsas. Mielograma: dispoese das tres linhagens, com 24% de celulas blasticas, alem de grumos de celulas nao-hematologicas (TS). Em Citometria de Fluxo (Euroflow): 16% de mieloblastos patologicos, expressando CD7, CD13, CD33, CD34, CD38, CD44, CD71, CD97, CD123. Citogenetica: 47,XY,+8. Nao foi realizada avaliacao molecular. Biopsia de MO: infiltracao macica da medula ossea por tumor solido. Imunohistoquimica com positividade para: CK7 e TTF-1 e negatividade para: CK20, PSA, PAX-8, CD10, CDX-2, GATA-3. Diagnostico final: infiltracao da MO por adenocarcinoma, favorecido sitio pulmonar, e em area residual, infiltracao por Leucemia mieloide aguda, com alteracao citogenetica associada a mielodisplasia (ELN 2022). Como tratava-se de idoso fragil, sem possibilidade de tratamento intensivo das doencas leucemica e oncologica, proposto tratamento paliativo exclusivo, com medidas de suporte. Paciente faleceu apos 2 meses do diagnostico. Discussao: A associacao de LMA com TS nao e rara. O risco de um paciente previamente tratado para TS desenvolver LMA pode ser ate 10 vezes maior que a populacao normal. Relatos de casos de TS recidivados em MO e LMA concomitantes foram descritos, com desfechos sombrios. Contudo, nao ha na literatura a descricao de LMA de novo com TS infiltrando MO concomitantes ao diagnostico. Estudo chines de 2020, analisou 12 casos de LMA associada a TS primario sincronicos, sendo: 3 etiologia pulmonar, 1 mama, 1 mola, e 7 com neoplasia de TGI. O intervalo de tempo mediano para o diagnostico das duas neoplasias foi de 4 meses. O tratamento da LMA foi realizado com quimioterapia (QT) de alta intensidade ou agente hipometilante. A sobrevida global mediana foi de 12.5 meses (porem somente 4/12 tinham TS avancado). Um estudo americano em 2009 analisou 5 pacientes com LMA e tumor de pulmao (2 com doenca avancada) sincronicos. Todos foram tratados com QT de alta intensidade, e a sobrevida media foi de 5 meses (3-21). Conclusao: Este e o primeiro relato de caso de LMA e TS metastatico infiltrando MO concomitantemente ao diagnostico. A LMA deve ser tratada prioritariamente por ser doenca altamente agressiva, levando-se em conta as condicoes clinicas do paciente, com avaliacao individual. E mesmo com tratamento intensivo, o prognostico e reservado. Copyright © 2022

UTILIZATION OF CONTINUOUS REMOTE MONITORING FOR A HIGH-RISK PATIENT REQUIRING INTERVAL TUMOR DEBULKING PALLIATIVE THERAPY

SESSION TITLE: Lung Cancer Case Report Posters 2 SESSION TYPE: Case Report Posters PRESENTED ON: 10/17/2022 12:15 pm - 01:15 pm INTRODUCTION: The outbreak of the SARS-CoV-2 virus identified a need for healthcare systems to transform in order to accommodate the large volume of patients. As a result, innovative new methods to monitor patients have emerged. One type of innovation are remote patient monitoring (RPM) devices, which allow for home vital sign (VS) measurements and telemonitoring. We present a case utilizing this technology to monitor a middle-aged male with metastatic colon cancer to the lung, who required regular debulking therapy as a means of palliation. CASE PRESENTATION: A 59 year-old male with a history of stage IV colon adenocarcinoma with metastasis to the lungs status post lung wedge resection and radiation therapy 7 years previously was found to have an enlarging left lower lobe (LLL) mass. Fiberoptic bronchoscopy revealed resurgence of his metastasis. While undergoing palliative chemotherapy, the patient became increasingly dyspneic. Serial PET CTs showed evolution of his left lung mass with left upper and lower lobe collapse due to endobronchial disease prompting bronchoscopy with argon plasma coagulation (APC) for tumor debulking within the left mainstem bronchus and dilation of the LLL airways. While the patient's symptoms improved, he became dyspneic over several months, and interval CT scans demonstrated invasion of the left mainstem bronchus with complete collapse of the left lung. Repeat dilation and APC were performed with improvement in symptoms. Due to rapid tumor growth, he was enrolled in the continuous RPM (CRPM) program for 24/7 nursing-led telemonitoring. He completed daily questionnaires on a vendor-provided digital tablet, and his VS, composed of heart rate (HR), respiratory rate (RR), SpO2, and temperature, were automatically uploaded to a network using an FDA-approved wearable device. Intermittent readings using peripheral devices to measure blood pressure and spirometry were gathered. His VS mirrored his tumor progression, indicated by elevation in his mean RR and HR while his SpO2 declined necessitating 2L of oxygen. Further evaluation showed tumor invasion into the left mainstem bronchus and began to invade his right mainstem. Successive APC and cryotherapy were performed every 2-3 months with a total of 8 debulking bronchoscopies. Once his disease progressed to obstruct his entire left mainstem, the patient unenrolled from the CRPM program and enrolled in hospice care. DISCUSSION: Several RPM devices have previously been used, but require self-reported VS rather than automated, continuous oximetry. Our CRPM program was piloted as a means to monitor COVID-19 patients following hospital discharge. However, our patient displayed benefit from his 180 day CRPM enrollment while receiving palliative tumor debulking procedures in order to fulfill his wish to maximize time at home. CONCLUSIONS: RPM devices offer a novel method of monitoring patients outside of healthcare facilities. Reference #1: Gordon WJ, Henderson D, DeSharone A, et al. Remote Patient Monitoring Program for Hospital Discharged COVID-19 Patients. Appl Clin Inform. 2020;11(05). doi:10.1055/s-0040-1721039 Reference #2: O'Carroll O, MacCann R, O'Reilly A, et al. Remote monitoring of oxygen saturation in individuals with COVID-19 pneumonia. Eur Respir J. 2020;56(2). doi:10.1183/13993003.01492-2020 Reference #3: Grutters LA, Majoor KI, Mattern ESK, Hardeman JA, van Swol CFP, Vorselaars ADM. Home telemonitoring makes early hospital discharge of COVID-19 patients possible. J Am Med Informatics Assoc. 2020;27(11). doi:10.1093/jamia/ocaa168 DISCLOSURES: No relevant relationships by Kevin Loudermilk Speaker/Speaker's Bureau relationship with Janssen Please note: $1001 - $5000 by Michael Morris, value=Honoraria Speaker/Speaker's Bureau relationship with GSK Please note: $1001 - $5000 by Michael Morris, value=Honoraria Removed 03/29/2022 by Michael Morris No relevant relationships by Michal Sobieszczyk No relevant relations ips by Robert Walter No relevant relationships by Whittney Warren

Lung Cancer Surgery in the Early Phase After Acute COVID-19 Pneumonitis

As the number of post-COVID-19 patients requiring surgery increases, it becomes pressing to develop guidelines outlining time requirements between active COVID-19 infection and surgery. We present a case of successful pulmonary segmentectomy 6 weeks following an acute COVID-19 infection in a 65-year-old female. The case patient was scheduled for a robotic assisted left upper lobectomy for radiologically diagnosed early-stage lung cancer. Unfortunately, prior to surgery she contracted Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-COV-2), resulting in the operation being rescheduled for 6 weeks' time. She was managed in the community for COVID pneumonitis and developed significant shortness of breath. At admission, with resolution of breathlessness, a repeat chest computed tomography scan showed the nodule had increased in size from 2 to 2.5cm, and widespread interstitial pneumonitis. The patient was saturating at 91% on air with no respiratory compromise. On balance of risk, surgery went ahead as planned due to concerns over tumour progression. A smaller lung resection was undertaken, with robot-assisted left upper division segmentectomy preferred to lobectomy. Post-operatively the patient received aggressive physiotherapy and high flow nasal oxygen to aid sputum expectoration. Chest tube was removed on day 2 post-operatively and the patient discharged 5 days following surgery without complication. Final histology confirmed a fully resected stage T1cN0M0 adenocarcinoma of the lung. This case highlights the importance of timing surgery correctly in post-COVID-19 patients to achieve the most favourable outcomes. We must balance clinical priority and the risk of disease progression against the severity of COVID-19 infection and the patient's comorbid status.

Synergistic anti-tumor effect of Allocetra-OTS in combination with immune checkpoint inhibitors (ICI)/chemotherapy/CAR T, through in-vivo reprogramming of macrophages

Background: ICI revolutionized solid tumor treatment, however, in many tumors only partial response is achieved. Here we investigated the anti-tumoral effect of Allocetra—OTS cellular therapy, in solid tumor models. Methods: Allocetra-OTS is manufactured from enriched mononuclear fractions and induced to undergo early apoptosis.Balb/c mice were inoculated in the peritoneal cavity with AB12 (mesothelioma) stably transduced with pLenti-PGK-V5-Luc-Neo for IVIS visualization and treated with anti-CTLA4, anti-PD1, or cisplatin, with or without Allocetra-OTS (also administered as monotherapy). Kaplan-Meier log rank test was done for survival. CAR T model was induced in SCID-Bg mice were inoculated intraperitoneally with human HeLa-CD19, followed by treatment with 10×109 cells of Allocetra-OTS or vehicle, and 1×107 CD19-CAR T cells or mock T cells. Results: Anti-CTLA4 therapy significantly improved survival from mean 34±9 to 44.9 ±20 days (p<0.05). However, Allocetra-OTS monotherapy improved survival to 52.3 ±20 days (p<0.02) and anti-CTLA4 + Allocetra-OTS combination therapy to 86.7±20 days (p<0.0001) with complete cancer remission in 60-100% of mice. Similar results were seen in combination therapy with either anti-PD1 or cisplatin. In the CAR-T model, SCID-Bg mice survived 30±5 days (range 27–37) and were sacrificed or died from tumor progression. Results were verified using IVIS of intraperitoneal HeLaCD19-Luc cells. CAR T cell therapy significantly improved survival to 55±11 days (p < 0.05 vs MOCK) but Alloctra-OTS further improved survival to 75±10 (p < 0.001) with 20-40% complete remission. Conclusions: During intraperitoneal tumor progression, Allocetra-OTS as monotherapy or in combination with ICI, cisplatin or CAR-T therapy, significantly reduced tumor size and resulted in complete remission in up to100% treated mice. Based on excellent safety profile in > 40 patients treated in prior clinical trials for sepsis and COVID-19, phase I/II clinical trial of Allocetra-OTS plus chemotherapy is planned for Q3 2022, and a second phase I/II clinical trial of Allocetra-OTS plus anti-PD1, as a second- and third-line therapy in various cancers, is planned for Q4 2022. Legal entity responsible for the study: The authors. Funding: Enlivex Therapeutics Ltd. Disclosure: D. Mevorach: Financial Interests, Personal, Royalties, Founder & CSO: Enlivex Therapeutics LTD. E. Yalon, E. Regev, C. Ankri, O. Hershkovitz, Y. Shabat, B. Reicher: Financial Interests, Personal, Full or part-time Employment: Enlivex Therapeutics LTD.

In-vivo reprogramming macrophages and dendritic cells with Allocetra-OTS: Successful mono- and combination-antitumor therapy

Background: Chimeric antigen receptor (CAR) T cells can activate an immune response to a cancer-specific antigen but is less effective in solid tumors. Immune check point inhibitors (ICI) revolutionized the treatment of solid tumors, however, in many tumors only partial response is achieved. Here we questioned the role of synergistic effect of Allocetra-OTS (cellular therapy for in-vivo reprogramming macrophages and dendritic cells, Enlivex Therap.) on solid tumor progression. Methods: To follow tumor growth in vivo, HeLa-CD19 cells were stably transduced with pLenti-PGK-V5-Luc-Neo. For CAR preparation, fresh mononuclear cells (MNC) were transfected with CD19-CAR plasmids. For the intraperitoneal solid tumor model, SCID-Bg mice were injected intraperitoneally (IP) with human HeLa- CD19 or HeLa-CD19-luciferase cells, 10×106 allocetra-OTS or vehicle, and 10×106 CD19-CAR T cells or mock T cells. In an immune-competent model, Balb/c mice were treated IP with AB12 (mesothelioma) with pLenti-PGK-V5-Luc-Neo and treated with anti-CTLA4 with or without Allocetra-OTS. Mice were monitored daily for clinical signs and peritoneal fluid accumulation and weekly for tumor growth. Kaplan-Meier log rank test was done for survival. Peritoneal cells were evaluated using single cell analysis and flow cytometry. Tumors were examined for bacterial presence by immunohistochemistry staining with antilipoteichoic acid (LTA) and antilipopolysaccharide (LPS). For allocetra-OTS preparation, enriched mononuclear fractions were collected by leukapheresis from healthy eligible human donors and induced to undergo early apoptosis. Results: SCID mice survived 30±5 days (range 27-37) and were sacrificed or died from solid tumor in the peritoneal cavity after accumulation of bloody peritoneal fluid and clinical deterioration. Results were verified using IVIS of intraperitoneal HeLaCD19- Luc cells. CAR T cell therapy significantly ameliorated survival to 55±11 days (p < 0.05 vs MOCK) but Alloctra-OTS further ameliorated survival to 75±10 (p < 0.001) with 20-40% complete remission. In AB12 model, anti CTLA4 therapy significantly ameliorated survival from 26±5 to 38 ±9 days (p < 0.05). However, Allocetra-OTS monotherapy ameliorated survival to 45 ±12 days (p < 0.02) and combinational therapy to 75±9 days (p < 0.0001) with complete remission in 60-75% of mice. Single cell analysis revealed that restoration of large peritoneal macrophages (LPM), were associated with antitumor activity. Conclusions: During intraperitoneal tumor progression, allocetra-OTS as monotherapy or combinational therapy with CAR or anti-CTLA4 significantly reduced tumor size and enable complete remission in up to 75% treated mice. Based on excellent safety profile in > 30 patients treated for sepsis and Covid19, human phase I/II of allocetra-OTS plus ICI, for peritoneal metastases, is planned for 2022.

Mesenchymal Stem/Stromal Cells: Late Breaking Abstract: FIRST EVIDENCE FOR THE TRANSLATIONAL POTENTIAL OF MESENCURE™, A PROFESSIONALIZED ALLOGENEIC MESENCHYMAL CELL THERAPY, FOR TREATING IMMUNOTHERAPY-RELATED CYTOKINE RELEASE SYNDROME

Background & Aim: Cytokine Release Syndrome (CRS) and Immune effector Cell-Associated Neurotoxicity Syndrome (ICANS) are related side effects of immunotherapies seen in up to 76% of patients treated with CAR-T and 48% of those treated with BiTEs. In up to 27% of the patients, these syndromes may lead to severe consequences. Current treatments for severe CRS are ineffective in >30% of the cases and can worsen ICANS prognosis, calling for novel treatments, especially in light of the expanding use of immunotherapies. Despite their obvious potential, mesenchymal cell (MSC) therapies were seldom investigated in this context. In the present study, Bonus BioGroup has set to assess the potential for treating CRS with MesenCure™, our allogeneic MSC platform, professionalized to enhance the cells’ potency and shown safe and effective in severe COVID patients. Methods, Results & Conclusion: A highly translational and validated CRS model was established in humanized NSG mice bearing human PBMCs, B-cell lymphoma, and CAR-T cells. CAR-T introduction significantly increased the serum levels of proinflammatory cytokines in model animals, indicative of CRS (Fig. 1A). Two IV MesenCure injections were well-tolerated in this model (Fig. 1B) and did not obstruct the CAR-Ts’ ability to inhibit tumor growth by 89% (Fig. 1C, p<0.0001). Remarkably, significant reductions in all proinflammatory cytokines tested (excluding IL-6) were measured in model animals treated with MesenCure, substantiating its potential to treat CRS (Fig. 1A). Interestingly, the magnitudes of these reductions resembled those observed in 50 severe COVID patients treated with MesenCure. MesenCure’s robust immunomodulatory capacity was further demonstrated in vitro by its ability to inhibit the proliferation of activated CD4 T cells with an IC50 of 6k MSC/200k PBMCs, twice more effectively than non-professionalized MSCs. Comparable results were also obtained with CD8 T cells. Similarly, MesenCure inhibited neutrophils’ ROS production by up to 80% within an hour following activation (IC50 19k MSC/200k neutrophils). These effects are likely mediated, in part, by IDO, whose RNA levels were found to be 6.8-fold higher in MesenCure cells than in non-professionalized MSCs (p<0.05), two hours after activation with IFNγ. Moreover, IDO inhibition by 1-MT (1 mM) reduced MesenCure’s (Figure Presented) Fig. 1 (A) The levels of serum proinflammatory cytokines measured in tumor-bearing NSG mice after CRS induction by injection of human PBMCs/CAR-Ts (or saline control) and MesenCure treatment (or saline control). Experimental groups’ designation: Control – not injected with PBMCs/CAR-Ts and not treated by MesenCure;CAR-T – CRS model animals, injected with PBMCs/CAR-Ts but not treated with MesenCure;MesenCure – treated with MesenCure but not injected with PBMCs/CARTs;and CAR-T + MesenCure – CRS model animals treated with MesenCure. (B) Relative change in body weight from the day of tumor induction (Day 0) and (C) IVIS analysis of tumor burden (dorsal aspect) in the above four experimental groups. Statistical significance indicators: ns – not significant, * p<0.05, *** p<0.001, **** p<0.0001. Statistical tests: Holm-Šídák’s multiple comparisons test (A) and two- sided t-test (C). ability to inhibit T cells’ proliferation by 73%. In conclusion, we provide the first evidence for the potential of MSCs and MesenCure, in particular, for treating immunotherapy-related CRS.

Inhibition of estrogen receptor signaling as a strategy for radiosensitization of ER+ breast cancers

Purpose: The estrogen receptor (ER) is expressed in over 80% of breast tumors and has been shown to be a significant driver of breast cancer (BC) pathogenesis and therefore a target of effective first-line therapies. While both ionizing radiation (RT) and endocrine therapies (ET) are used for the treatment of ER+ BC, the effect of ET on tumor radiosensitization remains unclear, with concerns it may be radioprotective based on G1 cell arrest with ET treatment. Here we assessed the efficacy and mechanism of ER-mediated radiosensitization using various pharmacologic approaches in ER+ BC. Methods: Radiosensitization with ER inhibitors (tamoxifen [TAM], fulvestrant [FULV], AZD9496) was assessed using clonogenic survival assays. DNA damage was assessed by the neutral comet assay. Efficiency of homologous recombination (HR) or non-homologous end joining (NHEJ) as well as changes in cell cycle, apoptosis, and senescence were assessed. The efficacy of TAM with RT in vivo was assessed with an MCF-7 xenograft model. Results: The selective estrogen receptor modulator TAM radiosensitized ER+ MCF-7 (enhancement ratio [enhR]: 1.14-1.50) and T47D (enhR: 1.33-1.60) cells but not ER-negative SUM-159 cells (enhR: 0.99-1.02). The selective estrogen receptor degrader (SERD) FULV had similar radiosensitization effects in MCF-7 (enhR: 1.33-1.76) and T47D cells (enhR: 0.97-2.81) with no radiosensitization observed in SUM-159 cells (enhR: 1.01-1.03). The novel oral SERD AZD9496 radiosensitized MCF-7 cells (enhR: 1.36-1.56). MCF-7 cells treated with TAM and RT had an increase in dsDNA breaks compared to RT alone as measured by the comet assay (p<0.05) and a decrease in NHEJ-mediated repair with TAM (p<0.05). No changes were observed in HR-mediated repair by Rad51 foci or a reporter (p=NS). RT alone and in combination with TAM or FULV induced similar levels of cell cycle arrest, suggesting that radiosensitization with the combination therapy is cell-cycle independent. There were no significant changes in apoptosis with TAM, FULV, RT, or the combination (p=NS). Although TAM or FULV did induce senescence, ET with RT increased senescence induction (p<0.05). In vivo, combination RT and TAM led to a significant delay in days to tumor doubling (control: 17, TAM: 40, RT: 32, TAM+RT: undefined;p<0.0001), and a significant difference in tumor growth between mice treated with TAM or RT alone compared combination treatment, with no increased toxicities or skin lesions from the combination treatment. Conclusion: Our data suggest that TAM, FULV, or AZD9496 can radiosensitize ER+ breast tumors, and these agents with RT may be more effective for radiosensitization. This work also supports further clinical investigation of the timing of RT for patients receiving ET, including using ET during RT, especially as initiating ET prior to RT has been increasingly utilized as a bridging therapy followed by concurrent ET+RT during the COVID-19 pandemic.

IMPACT OF THE COVID-19 PANDEMIC ON 30-DAY MORTALITY AND UNPLANNED REOPERATION RATES FOR PEDIATRIC CENTRAL NERVOUS SYSTEM TUMORS

INTRODUCTION: Unplanned reoperations and mortality within 30 days are important indicators when evaluating the quality of care provided by surgical systems. We reviewed these outcomes among children with primary central nervous system (CNS) tumors treated during the COVID- 19 pandemic. METHODS: This is a retrospective study of all pediatric patients who underwent neurosurgery for primary CNS tumors at the Philippine General Hospital, the national university hospital, from January 1, 2020 until December 31, 2021. Their clinical presentation, perioperative course, and outcomes were analyzed. During this time, our hospital concurrently served as a COVID-19 referral center, thus, the workforce was restructured, and resources were reallocated to care for COVID-19 patients. RESULTS: A total of 92 pediatric patients with CNS tumors underwent 140 neurosurgical operations during the study period. Two-thirds of the patients were males, and mean age was 9.3 ± 5.0 years (range: 3 months to 18 years). Average preoperative length of stay was 3.9 ± 2.6 days. Tumor resection was performed in 73 patients (79%). Most common histologic diagnoses were medulloblastoma (20%) and low-grade glioma including pilocytic astrocytoma (20%). Overall, the 30-day mortality and unplanned reoperation rates were 12% and 22%, respectively. Eight patients died from brain herniation and/or tumor progression. Reasons for unplanned reoperations were postoperative hydrocephalus (20%), infection (9%), hematoma (7%), and tumor residual (3%). DISCUSSION: Worldwide, the COVID-19 pandemic has altered hospital protocols and shifted resources considerably. The observed high rates of death and reoperation are likely due to delays in seeking care leading to worse neurologic status at presentation, delays in performing essential surgery within the hospital, and shortage of health workers providing specialist care. It is important to periodically assess perioperative outcomes to improve the quality of surgical care given to children with CNS tumors, who remain a vulnerable population during the COVID-19 pandemic.

CHALLENGES IN THE NEUROSURGICAL CARE OF PEDIATRIC PATIENTS WITH CENTRAL NERVOUS SYSTEM TUMORS DURING THE COVID-19 PANDEMIC

INTRODUCTION: Central nervous system (CNS) tumors account for 20 - 30% of all childhood cancers. The Philippines is a lower-middle income country, wherein brain centers are located mostly in urban areas. We aimed to identify challenges that pediatric patients with CNS tumors encountered during the COVID-19 pandemic, which aggravated delays in their diagnosis and treatment. METHODS: This is a retrospective review of all pediatric patients who underwent neurosurgery for CNS tumors at the Jose R. Reyes Memorial Medical Center, a tertiary referral center, from January 2020 until December 2021. We summarized patients' demographic data, clinical course, and perioperative outcomes. RESULTS: A total of 38 pediatric patients underwent neuro-oncologic surgery in our center during the study period. There were 18 males and 20 females, with a mean age of 7.5 ± 4.9 years. Tumor was biopsied and/or resected in 35 cases (92%). The most common histologic diagnoses were medulloblastoma (n=8, 21%) and high-grade glioma/glioblastoma (n=5, 13%). Median preoperative length of stay and total length of stay were 10 (IQR: 17) and 28 (IQR 33.75), respectively. There was a high perioperative mortality rate in 2020 (71%), but this decreased to 20% in 2021. Six patients (16%) developed COVID-19 infection during the perioperative period. There were nine patients (24%) who had documented tumor progression because of delays in adjuvant therapy. DISCUSSION: Aside from geographic barriers and catastrophic health expenditure, the major challenges that disrupted the care of pediatric patients with CNS tumors in our center during the COVID-19 pandemic were delays in neuroimaging for diagnosis, unavailability of operating room slots, deficiency in critical care beds, and workforce shortage due to COVID-19 infection among health workers. Health care systems must adapt to the changes brought about by the pandemic, so that children with CNS tumors are not neglected.

A CASE SERIES OF PEDIATRIC PATIENTS WITH BRAIN TUMORS AND PERIOPERATIVE COVID-19 INFECTION

INTRODUCTION: Surgery in patients diagnosed with COVID-19 is associated with increased risk of morbidity and mortality, especially within 6 weeks of SARSCoV- 2 infection. Furthermore, most studies have focused on adults, and little is known about perioperative outcomes in children with COVID-19. METHODS: We reviewed the operative census of the Division of Neurosurgery of Philippine General Hospital from March 2020 until December 2021. We identified all pediatric patients with brain tumors and confirmed COVID-19 infection within two weeks of their neuro-oncologic surgery. Their clinical course and outcomes are described herein. RESULTS: Four patients were included in this case series: three had tumors in the cerebellum, one in the pineal region. All of them were boys, with ages ranging from 4 months to 13 years. All tumors were malignant, and two were confirmed to be medulloblastoma after tumor resection. COVID-19 infection was diagnosed by the presence of SARS-CoV-2 RNA through a nasopharyngeal swab. Three patients acquired the virus post-operatively, likely from nosocomial transmission. In the remaining patient, it was community-acquired. All the patients had chest radiographs consistent with pneumonia but none had marked elevation of serum inflammatory markers. No patient received remdesivir or tocilizumab. At the time of their presentation, either the COVID-19 vaccine was not yet available in the country, or the patient was not yet eligible for vaccination. One patient died because of brain herniation from tumor progression, two were discharged and eventually underwent adjuvant therapy, and one remained in-hospital as of this writing. DISCUSSION: COVID-19 infection resulted in delays in the management of patients with pediatric CNS tumors. Given the high risk of these patients for potential complications, consensus guidelines must be established to achieve good outcomes and prolong survival.

Immunoregulatory role of nucleated erythroid cells

Nucleated erythroid cells (NEC) are the precursors of the most massive population of human cells – erythrocytes, for which functions of hemo- and immunoregulation have been shown at various stages of ontogenesis and in various organs and tissues of the human body. NEC perform this function by secreting cytokine proteins, growth factors, enzymes such as arginase-2, ROS, and by surface molecules PD-L1 and PD-L2. Their important regulatory role has been shown for the formation of fetoplacental immunosuppression, immunosuppression during pregnancy, suppression of the response against commensals in the gastrointestinal tract, in the pathogenesis of bacterial and viral infections in adults, in the pathogenesis of tumor growth and autoimmune diseases, as well as participation in the recognition of pathogen-associated molecular patterns using Toll-like receptors in fish and birds. Such qualities, together with their number and width of distribution, represent NEC as active participants in hemo- and immunoregulation, which makes it important to study their regulatory role in health and disease.

The elevated level of the main markers of neutrophil extracellular traps in metastatic colorectal cancer plasma highlights the enhanced risk of severe forms of COVID-19 in cancer patients

Several recent reports have described the involvement of neutrophil extracellular traps (NETs) formation in cancer progression. Moreover, we ourselves were amongst the first to link NETs dysregulation to the COVID-19 pathogenesis1 . In response to infection, stimulated neutrophils play an important role as the first line of innate immune defense, by producing extensive extracellular structures composed of granular protein assembled on a scaffold of released chromatin. In addition to mechanically trapping microorganisms, and thus impeding their dissemination in blood, NETs exploits coagulant function to segregate them within the circulation2 , while NET by-products such as circulating DNA, histones, granule enzymes (Elastase, Myeloperoxidase,.) also contribute to the triggering of an inflammatory process. As most of the sterile or infectious diseases associated with NET imbalance, cancer is established as a COVID-19 comorbidity. We hypothese that NET formation may result in higher COVID-19 severity. Our objective is to highlight that high NETs formation result in higher COVID-19 severity in cancer patients infected with SARS-Cov2. We are involved in two concurrent clinical studies in which we analyze blood samples for testing NET by-products and formation: (i) in newly diagnosed mCRC patients;and (ii) in patients with intermediate and severe forms of COVID-19. Initial data has shown that the level of the main markers of NETs formation, such as Elastase (ELA2), Myeloperoxidase (MPO), or circulating DNA (cirDNA), are (i) highly correlated in more than 221 mCRC patients, and (ii) statistically (P<0.0001) different than in healthy individuals (N=76). Median concentration values in healthy individuals (n=76): cirDNA: 5,16ng/mL;ELA2: 11,83ng/mL and MPO conc: 12,37ng/mL. Median concentration values in mCRC patients (n=221): cirDNA: 18,36ng/mL;ELA2: 34,70ng/mL and MPO: 38,90ng/mL. CirDNA and MPO or ELA2 concentration do not statistically correlate in healthy individuals, while cirDNA, ELA2 and MPO highly correlate each other (p<0.0001). Since malignant cell derived mutant cirDNA and total cirDNA are similarly correlated with increased level of NET by-products, we speculate that increased tumor burden and increased NET formation are correlated. Thus, NET by-products might be exploited as tumor progression biomarkers which could also be used to guide the choice of immunological therapeutic options. Lastly, our initial data support the hypothesis that cancer-associated NET exacerbation further contributes to COVID-19 severity.

Physical and visual (social) isolation increases tumor aggressiveness in thermoneutral housin temperatures in murine mammary cancer models

Correct interpretation of tumor progression data, including the influence of host biology, in mouse models of breast cancer requires models and conditions that faithfully recapitulate human disease and human host status. Our previous attempts to investigate the effects of social isolation have proven inconclusive due to premature mortality in tumor-bearing animals. Those studies were completed in standard temperature (ST), which commonly is 70-72°F (21-22°C) for in vivo murine research based on laboratory animal care and use guidelines. Previous work from the Repasky lab (Kokulus, 2013), which we have validated (Gaymon, 2020), demonstrates that ST housing results in chronic cold stress and immune suppression mediated by an increase in norepinrephrine (NE) levels, leading to increased tumor aggressiveness. Based on these findings, we investigated the effects of social isolation on BALB/cJ-4T1-luc and C57BL/6J/E0771 tumor progression and metastasis in thermoneutral housing conditions (84-85°F). Mice were first acclimatized to thermoneutral temperature and/or isolation for two weeks in cages that were unilaterally draped to provide physical and visual isolation. In BALB/cJ mice, 4T1-luc tumors were significantly larger in isolated mice compared to group-housed (GH) mice at day 18 (p<.0001). Statistically larger tumors were observed in isolated mice compared to GH mice through day 24 and final tumor masses were Salso significantly different (p=.004). Spleen masses were not statistically different. In C57BL/6J mice, E0771 tumors were significantly larger in isolates at Day 25 (p=.002). Final tumor masses were statistically (p=.002) different while no difference in spleen sizes were observed. Data on metastasis will be presented at the meeting. We hypothesized that social isolation may perturb immune function and next investigated the growth of 4T1-luc xenograft tumors in NSG mice. 4T1-luc/NSG tumor progression and metastasis data will also be presented at the meeting. We conclude that syngeneic breast tumor growth in immunocompetent BALB/cJ and C57BL/6J mice demonstrates that social isolation is a bona fide stress with sufficient influence to exacerbate breast cancer growth. These data are potentially clinically important due the known relationship of social support to survivorship outcomes in patients and the high-risk of depression and isolation in patients following breast cancer diagnosis. The data may provide additional insight into possible effects of COVID-19 isolation on breast cancer progression.

Endocrine therapy treatment radiosensitizes estrogen receptor-positive breast cancers

Purpose: Estrogen receptor (ER) expression is present in over 80% of breast tumors and has been shown to be a significant driver of breast cancer (BC) pathogenesis and therefore a target of first-line therapies for ER-positive (ER+) BC patients. While both ionizing radiation (RT) and endocrine therapies (ET) are used for the treatment of ER+ BC, the sequencing of therapy and the effect of ET on tumor radiosensitization remain unclear. Recently, this question has become much more clinically relevant when many physicians started offering ET as a bridging strategy to surgery and RT during the COVID-19 pandemic. Here we assessed the efficacy and mechanism of ER inhibition in ER+ BC in combination with RT in preclinical models. Methods: Clonogenic survival assays were used to assess radiosensitization. Inhibition of ER signaling was accomplished by treating ER+ MCF-7 and T47D cells with the selective ER modulator (SERM), tamoxifen, or the selective ER degrader (SERD), fulvestrant. The ER-negative SUM-159 cells were used as a negative control. DNA damage was assessed by the neutral comet assay. Efficiency of homologous recombination (HR) was measured by Rad51 foci or a GFP reporter system. Non-homologous end joining (NHEJ) efficiency was assessed with a pEYFP reporter. Cell cycle effects were measured using flow cytometry with propidium iodide (PI) staining. Apoptosis was assessed by annexin V/PI via flow Scytometry. Senescence was measured using β-galactosidase staining. Western blotting was used to quantify expression of proteins and phospho-proteins involved in cell cycle and apoptosis. An MCF-7 xenograft model was used to assess the efficacy of tamoxifen with RT in vivo. Synergy was determined using the fractional tumor volume (FTV) method. Results: ER inhibition with tamoxifen radiosensitized ER+ MCF-7 (10-250 nM, enhR: 1.14-1.50) and T47D (500 nM-2.0 μ M, enhR: 1.33-1.60) cells but not ER-negative SUM-159 cells (500 nM-2.0 μ M, enhR: 0.99-1.02). ER degradation with fulvestrant had similar radiosensitization effects in MCF-7 (1-25 nM, enhR: 1.33-1.76) and T47D cells (0.5-5 nM, enhR: 0.97-2.81) with no radiosensitization observed in SUM-159 cells (1-25 nM, enhR: 1.01-1.03). MCF-7 cells treated with 500 nM tamoxifen and 4 Gy RT had an increase in dsDNA breaks compared to RT alone as measured by the comet assay (p<0.05), and there was a decrease in NHEJ-mediated repair with tamoxifen treatment (p<0.05). No changes were observed in HR-mediated repair by Rad51 foci or an HR reporter (p=NS). RT alone and in combination with tamoxifen and fulvestrant induced similar levels of cell cycle arrest, suggesting that radiosensitization with the combination therapy is a cell-cycle independent effect. In addition, there were no significant changes in apoptosis in MCF-7 or T47D cells with endocrine therapy, RT, or the combination (p=NS). Although treatment with ET did induce senescence in ER+ MCF-7 and T47D cells, the combination treatment of ET with RT induced senescence to a much greater level suggesting this mechanism may contribute to radiosensitization (p<0.05). In vivo, combination RT and tamoxifen led to a significant delay in time to tumor doubling (17 days in control, 40 days with tamoxifen alone, 32 days with RT alone, and undefined with combination;p<0.0001) and a significant difference in tumor growth between mice treated with tamoxifen or RT alone compared to mice treated with tamoxifen and RT with synergy noted with combination treatment (FTV 1.297). Conclusion: Our data suggest that ET can radiosensitize ER+ breast tumors, and ET with RT may be more effective for radiosensitization. Ongoing studies will address concurrent versus sequential ET with RT. This work also supports further clinical investigation of the timing of RT for patients receiving ET, especially as ET prior to RT is increasingly used as a bridging therapy during the COVID-19 pandemic.

First line aromatase inhibitor (AI) + pa l bocicl i b with randomized switch to fulvestrant + palbociclib upon detection of circulating ESR1 mutation in HR+ HER2-metastatic breast cancer patients: Global safety results of PADA-1, a UCBG-GINECO phase III trial

Background: Aromatase inhibitors (AI) in combination with a CDK 4/6 inhibitor have been established as the standard first line treatment of non AI-resistant hormone receptor-positive (HR+) HER2-metastatic breast cancer (mBC) patients (pts). ESR1 mutations are known drivers of resistance to AIs in the metastatic setting but their actionability remains unknown. The phase 3 PADA-1 trial aimed both at refining the global safety of palbociclib combined to any AI as first line treatment of HR+ HER2-mBC pts, and at evaluating the clinical benefit associated with a switch to fulvestrant-palbociclib upon detection of a rising ESR1 mutation in blood (bESR1mut). Methods: PADA-1 (NCT03079011), a multicenter, randomized, open-label, phase 3 trial, enrolled HR+ HER2-mBC pts with no prior therapy for mBC, in the absence of AI-resistance. In the first step, pts received a combination of any AI and palbociclib at standard recommended doses and underwent centralized bESR1mut screening every two months. In the second step, bESR1mut+ pts with no S clinical/imaging concomitant disease progression were randomized between continuing the same therapy (standard arm) or switching to fulvestrant-palbociclib (experimental arm). The third step consisted in an optional cross-over after tumor progression for patients randomized in the standard arm. PADA-1 co-primary endpoints were global safety of the combination of palbociclib + endocrine therapy in the whole population of patients, throughout the study, with focus on hematological toxicities;and PFS in the second step. We present here the results of the global safety co-primary endpoint. Results: From 3/2017 to 01/2019, 1017 pts were accrued in 83 sites. As per 05/2021, 272 pts were still in step 1, 35 in step 2, and 8 in step 3. The overall follow-up was 33.7 months. 232 pts have deceased. 333 SAEs have been reported, including 21 grade 5, 35 grade 4, 183 grade 3, 53 grade 2, 26 grade 1 and 15 unknown grade. Among the grade 5 cases, 2 have been declared as potentially related to the underlying treatment (Death of unknown cause, pulmonary embolism). No pt died of SARS-CoV2 infection. The main hematological toxicities encountered, as well as selected non-hematological events are described in Table 1. Permanent discontinuation of the treatment due to toxicity occurred in 39 pts/1017 (3.8%). Palbociclib dose decreases occurred in 419 (41.2%) pts. Conclusion: By the number of included patients, PADA-1 is the largest prospective trial with 1st line AI and palbociclib. Data confirm the favorable safety profile of palbociclib when combined to any AI +/-switch to fulvestrant. Hematological toxicity appears limited and is mostly restricted to non-clinically significant neutropenia. Permanent discontinuation was exceptional. Detailed per-step analyses will be presented.