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AbstractThe transcription factor runt-related transcription factor 1 (Runx1) is essential for the establishment of definitive hematopoiesis during embryonic development. In adult blood homeostasis, Runx1 plays a pivotal role in the maturation of lymphocytes and megakaryocytes. Furthermore, Runx1 is required for the regulation of hematopoietic stem and progenitor cells. However, how Runx1 orchestrates self-renewal and lineage choices in combination with other factors is not well understood. In the present study, we describe a genome-scale RNA interference screen to detect genes that cooperate with Runx1 in regulating hematopoietic stem and progenitor cells. We identify the polycomb group protein Pcgf1 as an epigenetic regulator involved in hematopoietic cell differentiation and show that simultaneous depletion of Runx1 and Pcgf1 allows sustained self-renewal while blocking differentiation of lineage marker–negative cells in vitro. We found an up-regulation of HoxA cluster genes on Pcgf1 knock-down that possibly accounts for the increase in self-renewal. Moreover, our data suggest that cells lacking both Runx1 and Pcgf1 are blocked at an early progenitor stage, indicating that a concerted action of the transcription factor Runx1, together with the epigenetic repressor Pcgf1, is necessary for terminal differentiation. The results of the present study uncover a link between transcriptional and epigenetic regulation that is required for hematopoietic differentiation.

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Background: We developed a highly sensitive cardiomyocyte based screening system for the non-destructive electronic detection of chronotropic drugs and tissue-secreted factors involved in AT₁ receptor-mediated cardiovascular diseases. Methods: For this purpose we cultured spontaneously beating neonatal rat cardiomyocytes on microelectrode arrays (MEAs), and tested the optimised, stable culture parameters for a reproducible real-time recording of alterations in contraction frequency. After the evaluation of culture parameters, computer-based electronic measurement systems were used for counting of contractions by recording of the field potential of cardiomyocytes. Results: Using the biosensor, angiotensin II, the predominant ligand of the AT₁ receptor, was detected at very low concentrations of 10^-11 M via altered contractions of cardiomyocytes. Moreover, we demonstrated that cardiomyocyte coupled microarrays allow the detection of blood-derived low concentrated anti-AT₁ receptor autoimmune antibodies of pregnant women suffering from preeclampsia. Conclusion: This study demonstrates the first well-suited electrophysiological recording of cardiomyocytes on multielectrode arrays as a benefit for functional biomonitoring for the detection of AT₁ receptor/ligand interactions and other marker proteins in sera directed to cardiovascular diseases.

Severe neutropenia induced by chemotherapy or conditioning for hematopoietic cell transplantation often results in morbidity and mortality due to infection by opportunistic pathogens. A system has been developed to generate ex vivo-expanded mouse myeloid progenitor cells (mMPCs) that produce functional neutrophils in vivo upon transplantation in a pathogen challenge model. It has previously been demonstrated that transplantation of large numbers of freshly isolated myeloid progenitors from a single donor provides survival benefit in radiation-induced neutropenic mice. In the present work, an ex vivo-expanded and cryopreserved mMPC product generated from an allogeneic donor pool retains protective activity in vivo in a lethal fungal infection model. Infusion of the allogeneic pooled mMPC product is effective in preventing death from invasive Aspergillus fumigatus in neutropenic animals, and protection is dose dependent. Cell progeny from the mMPC product is detected in the bone marrow, spleen, blood, and liver by flow cytometry 1 week postinfusion but is no longer evident in most animals 4 weeks posttransplant. In this model, the ex vivo-generated pooled allogeneic mMPC product (i) expands and differentiates in vivo; (ii) is functional and prevents death from invasive fungal infection; and (iii) does not permanently engraft or cause allosensitization. These data suggest that an analogous ex vivo-expanded human myeloid progenitor cell product may be an effective off-the-shelf bridging therapy for the infectious complications that develop during hematopoietic recovery following hematopoietic cell transplantation or intensive chemotherapy.

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Background: ATRC-101 is a fully human, engineered IgG1 version of an antibody discovered through a target-agnostic process designed to identify patient-derived, tumor-targeting antibodies. The parental antibody of ATRC-101 was discovered from a patient with metastatic non-small cell lung cancer (NSCLC) undergoing an active immune response while receiving a checkpoint inhibitor. ATRC-101 binds selectively to human tumor specimens, including a majority of NSCLC, acral melanoma, breast, colorectal, and ovarian cancer samples. The target of ATRC-101 appears to be a tumor-associated ribonucleoprotein complex containing a form of polyadenylate-binding protein 1 (PABP-1) that can be found on the surface of tumor cells. Studies to further elucidate the tumor selectivity of ATRC-101 and the extracellular presentation of the target are ongoing. Preclinical data suggest that ATRC-101 stimulates an adaptive immune response against tumors via the innate immune system. ATRC-101 displays dose-dependent, single-agent activity in syngeneic mouse tumor models, including the EMT6 breast cancer model, which displays a T cell-excluded microenvironment and in which PD-1/PD-L1 inhibitors exhibit limited activity. ATRC-101 plus an anti-PD-1 antibody demonstrated significantly greater antitumor activity and longer survival in the EMT6 model compared with either agent alone, consistent with the mechanism of action proposed for ATRC-101. Methods: ATRC-101-A01 is an open-label, 3+3, dose-escalation phase 1b safety study of ATRC-101 as monotherapy (mono) or in combination (combo) with pembrolizumab (pembro) to treat patients with certain solid tumor types. Expansion cohorts of ≤12 patients may be enrolled at doses and frequencies at which biological activity is observed. The primary objective is to determine the safety of ATRC-101. Secondary objectives include evaluation of the following: recommended dose and schedule of ATRC-101 for expansion, pharmacokinetic profile of ATRC-101, immunogenicity, antitumor activity by RECIST v1.1, and lymphocytic infiltration in the tumor microenvironment. ATRC-101 is administered every 2 (q2w) or 3 (q3w) weeks up to 24 months or until disease progression. ATRC-101 dosing is increasing from the initial 0.3 mg/kg according to the following schedule, pending dose-limiting toxicity data: 1, 3, 10, and 30 mg/kg. In the combo cohort, ATRC-101 will be given q3w, and pembro will be administered at 200 mg q3w or 400 mg q6w to patients with disease progression while receiving prior pembro treatment or those with stable disease who were deemed by their physician to potentially benefit from the addition of ATRC-101. The q3w mono cohort is currently accruing patients at the 10-mg/kg dose, and the combo and q2w cohorts will begin at an ATRC-101 dose one level below the most recent cleared q3w mono dose. Trial registration NCT04244552. The study was approved by the institutional review board of each participating site. Citation Format: Jonathan E. Benjamin, Nicholas Higgins, Carl Millward, Jeff DeFalco, Amy Manning-Bog, Iraz T. Aydin, Danhui Zhang, Shaun M. Lippow, Alexander Scholz, Yvonne Leung, Yanhong Zhu, Anna Sedello, Zane Rogers, Ish K. Dhawan, Daniel Emerling, William H. Robinson, Tito A. Serafini, Norman M. Greenberg. First-in-human phase 1b study of ATRC-101, a patient-derived antibody with a tumor-specific target, as monotherapy or in combination with pembrolizumab, in patients with solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT203.

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