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Abstract: Acute lymphoblastic leukemia (ALL) arises from the uncontrolled proliferation of B-cell precursors (BCP-ALL) or T cells (T-ALL). Current treatment protocols obtain high cure rates in children but are based on toxic polychemotherapy. Novel therapies are urgently needed, especially in relapsed/refractory (R/R) disease, high-risk (HR) leukemias and T-ALL, in which immunotherapy approaches remain scarce. Although the interleukin-7 receptor (IL-7R) plays a pivotal role in ALL development, no IL-7R-targeting immunotherapy has yet reached clinical application in ALL. The IL-7Rα chain (CD127)-targeting IgG4 antibody lusvertikimab (LUSV; formerly OSE-127) is a full antagonist of the IL-7R pathway, showing a good safety profile in healthy volunteers. Here, we show that ∼85% of ALL cases express surface CD127. We demonstrate significant in vivo efficacy of LUSV immunotherapy in a heterogeneous cohort of BCP- and T-ALL patient-derived xenografts (PDX) in minimal residual disease (MRD) and overt leukemia models, including R/R and HR leukemias. Importantly, LUSV was particularly effective when combined with polychemotherapy in a phase 2-like PDX study with CD127high samples leading to MRD-negativity in >50% of mice treated with combination therapy. Mechanistically, LUSV targeted ALL cells via a dual mode of action comprising direct IL-7R antagonistic activity and induction of macrophage-mediated antibody-dependent cellular phagocytosis (ADCP). LUSV-mediated in vitro ADCP levels significantly correlated with CD127 expression levels and the reduction of leukemia burden upon treatment of PDX animals in vivo. Altogether, through its dual mode of action and good safety profile, LUSV may represent a novel immunotherapy option for any CD127+ ALL, particularly in combination with standard-of-care polychemotherapy., (© 2024 American Society of Hematology. Published by Elsevier Inc. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

The activating receptor natural killer group 2, member D (NKG2D) represents an attractive target for immunotherapy as it exerts a crucial role in cancer immunosurveillance by regulating the activity of cytotoxic lymphocytes. In this study, a panel of novel NKG2D-specific single-chain fragments variable (scFv) were isolated from naïve human antibody gene libraries and fused to the fragment antigen binding (Fab) of rituximab to obtain [CD20×NKG2D] bibodies with the aim to recruit cytotoxic lymphocytes to lymphoma cells. All bispecific antibodies bound both antigens simultaneously. Two bibody constructs, [CD20×NKG2D#3] and [CD20×NKG2D#32], efficiently activated natural killer (NK) cells in co-cultures with CD20+ lymphoma cells. Both bibodies triggered NK cell-mediated lysis of lymphoma cells and especially enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) by CD38 or CD19 specific monoclonal antibodies suggesting a synergistic effect between NKG2D and FcγRIIIA signaling pathways in NK cell activation. The [CD20×NKG2D] bibodies were not effective in redirecting CD8+ T cells as single agents, but enhanced cytotoxicity when combined with a bispecific [CD19×CD3] T cell engager, indicating that NKG2D signaling also supports CD3-mediated T cell activation. In conclusion, engagement of NKG2D with bispecific antibodies is attractive to directly activate cytotoxic lymphocytes or to support their activation by monoclonal antibodies or bispecific T cell engagers. As a perspective, co-targeting of two tumor antigens may allow fine-tuning of antibody cancer therapies. Our proposed combinatorial approach is potentially applicable for many existing immunotherapies but further testing in different preclinical models is necessary to explore the full potential., Competing Interests: AF, MH, SD and TS are co-founders and shareholders of YUMAB GmbH. SL, CK, MG, MP and YUMAB GmbH submitted a patent application related to the described antibodies. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Lutz, Klausz, Albici, Ebinger, Sellmer, Teipel, Frenzel, Langner, Winterberg, Krohn, Hust, Schirrmann, Dübel, Scherließ, Humpe, Gramatzki, Kellner and Peipp.)

Competing Interests: LL and DMS received research funding from OSE Immunotherapeutics outside the submitted work. DMS was an advisory board member for Bayer, SOBI, and Jazz Pharmaceuticals. MS received research funding from Shire, and from Servier, as well as fees for Advisory Board functions from Jazz Pharmaceuticals and Servier. All the other authors have no conflicts of interest to disclose.

Targeting CD19 represents a promising strategy for the therapy of B-cell malignancies. Although non-engineered CD19 antibodies are poorly effective in mediating complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP), these effector functions can be enhanced by Fc-engineering. Here, we engineered a CD19 antibody with the aim to improve effector cell-mediated killing and CDC activity by exchanging selected amino acid residues in the Fc domain. Based on the clinically approved Fc-optimized antibody tafasitamab, which triggers enhanced ADCC and ADCP due to two amino acid exchanges in the Fc domain (S239D/I332E), we additionally added the E345K amino acid exchange to favor antibody hexamerization on the target cell surface resulting in improved CDC. The dual engineered CD19-DEK antibody bound CD19 and Fcγ receptors with similar characteristics as the parental CD19-DE antibody. Both antibodies were similarly efficient in mediating ADCC and ADCP but only the dual optimized antibody was able to trigger complement deposition on target cells and effective CDC. Our data provide evidence that from a technical perspective selected Fc-enhancing mutations can be combined (S239D/I332E and E345K) allowing the enhancement of ADCC, ADCP and CDC with isolated effector populations. Interestingly, under more physiological conditions when the complement system and FcR-positive effector cells are available as effector source, strong complement deposition negatively impacts FcR engagement. Both effector functions were simultaneously active only at selected antibody concentrations. Dual Fc-optimized antibodies may represent a strategy to further improve CD19-directed cancer immunotherapy. In general, our results can help in guiding optimal antibody engineering strategies to optimize antibodies' effector functions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gehlert, Rahmati, Boje, Winterberg, Krohn, Theocharis, Cappuzzello, Lux, Nimmerjahn, Ludwig, Lustig, Rösner, Valerius, Schewe, Kellner, Klausz and Peipp.)

Immunotherapy has evolved as a powerful tool for the treatment of B-cell malignancies, and patient outcomes have improved by combining therapeutic antibodies with conventional chemotherapy. Overexpression of antiapoptotic B-cell lymphoma 2 (Bcl-2) is associated with a poor prognosis, and increased levels have been described in patients with "double-hit" diffuse large B-cell lymphoma, a subgroup of Burkitt's lymphoma, and patients with pediatric acute lymphoblastic leukemia harboring a t(17;19) translocation. Here, we show that the addition of venetoclax (VEN), a specific Bcl-2 inhibitor, potently enhanced the efficacy of the therapeutic anti-CD20 antibody rituximab, anti-CD38 daratumumab, and anti-CD19-DE, a proprietary version of tafasitamab. This was because of an increase in antibody-dependent cellular phagocytosis by macrophages as shown in vitro and in vivo in cell lines and patient-derived xenograft models. Mechanistically, double-hit lymphoma cells subjected to VEN triggered phagocytosis in an apoptosis-independent manner. Our study identifies the combination of VEN and therapeutic antibodies as a promising novel strategy for the treatment of B-cell malignancies., (© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Acute lymphoblastic leukemia (ALL) is the most common malignant disease affecting children. Although therapeutic strategies have improved, T-cell acute lymphoblastic leukemia (T-ALL) relapse is associated with chemoresistance and a poor prognosis. One strategy to overcome this obstacle is the application of monoclonal antibodies. Here, we show that leukemic cells from patients with T-ALL express surface CD38 and CD47, both attractive targets for antibody therapy. We therefore investigated the commercially available CD38 antibody daratumumab (Dara) in combination with a proprietary modified CD47 antibody (Hu5F9-IgG2σ) in vitro and in vivo. Compared with single treatments, this combination significantly increased in vitro antibody-dependent cellular phagocytosis in T-ALL cell lines as well as in random de novo and relapsed/refractory T-ALL patient-derived xenograft (PDX) samples. Similarly, enhanced antibody-dependent cellular phagocytosis was observed when combining Dara with pharmacologic inhibition of CD47 interactions using a glutaminyl cyclase inhibitor. Phase 2-like preclinical in vivo trials using T-ALL PDX samples in experimental minimal residual disease-like (MRD-like) and overt leukemia models revealed a high antileukemic efficacy of CD47 blockade alone. However, T-ALL xenograft mice subjected to chemotherapy first (postchemotherapy MRD) and subsequently cotreated with Dara and Hu5F9-IgG2σ displayed significantly reduced bone marrow infiltration compared with single treatments. In relapsed and highly refractory T-ALL PDX combined treatment with Dara and Hu5F9-IgG2σ was required to substantially prolong survival compared with single treatments. These findings suggest that combining CD47 blockade with Dara is a promising therapy for T-ALL, especially for relapsed/refractory disease harboring a dismal prognosis in patients., (© 2022 by The American Society of Hematology.)

Integrin associated protein (CD47) is an important target in immunotherapy, as it is expressed as a "don't eat me" signal on many tumor cells. Interference with its counter molecule signal regulatory protein alpha (SIRPα), expressed on myeloid cells, can be achieved with blocking Abs, but also by inhibiting the enzyme glutaminyl cyclase (QC) with small molecules. Glutaminyl cyclase inhibition reduces N-terminal pyro-glutamate formation of CD47 at the SIRPα binding site. Here, we investigated the impact of QC inhibition on myeloid effector cell-mediated tumor cell killing by epidermal growth factor receptor (EGFR) Abs and the influence of Ab isotypes. SEN177 is a QC inhibitor and did not interfere with EGFR Ab-mediated direct growth inhibition, complement-dependent cytotoxicity, or Ab-dependent cell-mediated cytotoxicity (ADCC) by mononuclear cells. However, binding of a human soluble SIRPα-Fc fusion protein to SEN177 treated cancer cells was significantly reduced in a dose-dependent manner, suggesting that pyro-glutamate formation of CD47 was affected. Glutaminyl cyclase inhibition in tumor cells translated into enhanced Ab-dependent cellular phagocytosis by macrophages and enhanced ADCC by polymorphonuclear neutrophilic granulocytes. Polymorphonuclear neutrophilic granulocyte-mediated ADCC was significantly more effective with EGFR Abs of human IgG2 or IgA2 isotypes than with IgG1 Abs, proposing that the selection of Ab isotypes could critically affect the efficacy of Ab therapy in the presence of QC inhibition. Importantly, QC inhibition also enhanced the therapeutic efficacy of EGFR Abs in vivo. Together, these results suggest a novel approach to specifically enhance myeloid effector cell-mediated efficacy of EGFR Abs by orally applicable small molecule QC inhibitors., (© 2021 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most frequent malignancy in children and also occurs in adulthood. Despite high cure rates, BCP-ALL chemotherapy can be highly toxic. This type of toxicity can most likely be reduced by antibody-based immunotherapy targeting the CD19 antigen which is commonly expressed on BCP-ALL cells. In this study, we generated a novel Fc-engineered CD19-targeting IgG1 antibody fused to a single chain tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) domain (CD19-TRAIL). As TRAIL induces apoptosis in tumor cells but not in healthy cells, we hypothesized that CD19-TRAIL would show efficient killing of BCP-ALL cells. CD19-TRAIL showed selective binding capacity and pronounced apoptosis induction in CD19-positive (CD19 + ) BCP-ALL cell lines in vitro and in vivo. Additionally, CD19-TRAIL significantly prolonged survival of mice transplanted with BCP-ALL patient-derived xenograft (PDX) cells of different cytogenetic backgrounds. Moreover, simultaneous treatment with CD19-TRAIL and Venetoclax (VTX), an inhibitor of the anti-apoptotic protein BCL-2, promoted synergistic apoptosis induction in CD19 + BCP-ALL cells in vitro and prolonged survival of NSG-mice bearing the BCP-ALL cell line REH. Therefore, IgG1-based CD19-TRAIL fusion proteins represent a new potential immunotherapeutic agent against BCP-ALL.

Central nervous system (CNS) involvement remains a challenge in the diagnosis and treatment of acute lymphoblastic leukemia (ALL). In this study, we identify CD79a (also known as Igα), a signaling component of the preB cell receptor (preBCR), to be associated with CNS-infiltration and -relapse in B-cell precursor (BCP)-ALL patients. Furthermore, we show that downregulation of CD79a hampers the engraftment of leukemia cells in different murine xenograft models, particularly in the CNS.

Background: Native cluster of differentiation (CD) 19 targeting antibodies are poorly effective in triggering antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC), which are crucial effector functions of therapeutic antibodies in cancer immunotherapy. Both functions can be enhanced by engineering the antibody's Fc region by altering the amino acid sequence (Fc protein-engineering) or the Fc-linked glycan (Fc glyco-engineering). We hypothesized that combining Fc glyco-engineering with Fc protein-engineering will rescue ADCC and CDC in CD19 antibodies., Results: Four versions of a CD19 antibody based on tafasitamab's V-regions were generated: a native IgG1, an Fc protein-engineered version with amino acid exchanges S267E/H268F/S324T/G236A/I332E (EFTAE modification) to enhance CDC, and afucosylated, Fc glyco-engineered versions of both to promote ADCC. Irrespective of fucosylation, antibodies carrying the EFTAE modification had enhanced C1q binding and were superior in inducing CDC. In contrast, afucosylated versions exerted an enhanced affinity to Fcγ receptor IIIA and had increased ADCC activity. Of note, the double-engineered antibody harboring the EFTAE modification and lacking fucose triggered both CDC and ADCC more efficiently., Conclusions: Fc glyco-engineering and protein-engineering could be combined to enhance ADCC and CDC in CD19 antibodies and may allow the generation of antibodies with higher therapeutic efficacy by promoting two key functions simultaneously.

A 60-year-old man presented with a recurrent translucent nodule on the dorsum of the left thumb. After puncturing, a clear gelatinous fluid appeared. The diagnosis was digital mucous cyst. These cysts commonly result from joint fluid leaking from an osteoarthritic distal interphalangeal joint through a communicating canal.

A 12-year-old girl presented with a pruritic maculopapularpurpuric exanthema limited to the hands and feet, the cause of which was papular-purpuric gloves and socks syndrome.

A blister was noted on the dorsal side of the right hand of an otherwise healthy female neonate: a sucking blister, which disappeared in a few days spontaneously.

Kawasaki disease is an acute vasculitis that occurs especially in young children. Because there is no specific laboratory test available, diagnosis has to be made on the basis of clinical characteristics: prolonged fever, oropharyngeal changes, conjunctival injection, erythema and edema of hands and feet, rash, and cervical lymphadenopathy. Without treatment there is a 25% chance of cardiac complications, especially aneurysms of the coronary arteries. Early treatment with intravenous immunoglobulin reduces this risk to 5%. Accurate diagnosis and therapy is crucial. Kawasaki syndrome has been reported in all racial groups with the highest incidence in Japanese children. Together with the fact that the disease is more common in boys this indicates that genetic factors play an important role in determining susceptibility to a (probably infectious) trigger. In spite of 40 years of intensive scientific research, the cause of Kawasaki disease still remains unknown.

In a 4-year-old boy with left-sided inguinal lymphadenopathy and a papule on his left lower leg situated at the end of a scar of a probable cat scratch, cat scratch disease was confirmed serologically.

In a 2-year-old girl, transverse furrows of the fingernails were noticed: Beau's lines associated with Kawasaki's syndrome.

A diagnosis of intracranial actinomycosis was made in a 9-year-old boy with headache, cranial nerve dysfunction and ataxia. Poor dental hygiene leading to caries and an inflammation in his right upper jaw probably was the cause of cervicofacial actinomycosis with intracranial extension. A full recovery was achieved by treatment with benzylpenicillin. Actinomyces is a gram-positive bacterium belonging to the endogenous flora of the mucous membranes of the oropharynx, gastrointestinal tract, and female genital tract. Disruption of the mucous membrane is the portal of entry for an extremely destructive infective process that is effectively treated with penicillin. Good orodental hygiene is important for the prevention of actinomycosis.

A 3-year-and-7-month-old girl with Down's syndrome had papules and vesicles on hands, feet and buttocks, and vesicular lesions in the oral cavity. A positive culture of enterovirus in vesicular fluid confirmed the clinical diagnosis 'hand-foot-mouth disease'.

In two neonate girls with vesicular skin lesions, incontinentia pigmenti (Bloch-Sulzberger syndrome) was diagnosed. This rare X-linked dominant ectodermal disease can cause abnormalities in several organ systems. Most prominent are the dermatological abnormalities, developing in 4 stages: the vesicular stage, the verrucous stage, the hyperpigmentation phase, and the atrophic phase. In addition to the cutaneous manifestations, many patients have anomalies of nails, hair and teeth. Serious related abnormalities of the eyes (intraocular vasculopathy) and central nervous system (convulsions, mental retardation) may occur. In 1989 the locus for the incontinentia pigmenti mutation was shown to be present on Xq28. Recently it was shown that the causative mutation is located on the NEMO ('NF-kappa B essential modulator') gene. A NEMO knock-out mouse model shows a dermatopathy of a transient nature, resembling the skin lesions in patients with incontinentia pigmenti.

A 4-year-old Indonesian girl was seen with streaky haematomas on back and chest. Because of fever, throat pain, cough and vomiting, her parents had rubbed her with balm and a coin. Coin rubbing is a harmless, traditional treatment for various illnesses which is regularly used in Southeast Asia. The cutaneous manifestations can be mistaken for battering or a bleeding disorder.

Following a febrile illness with nasal discharge and vomiting a 1.5-year-old girl developed monomorphous erythematous papular lesions on the face, buttocks and extremities. A diagnosis of Gianotti-Crosti syndrome on the basis of an Epstein-Barr virus infection was made. The patient recovered in eight weeks without treatment. In the Netherlands the originally described relationship with a hepatitis B infection is very improbable in patients with the Gianotti-Crosti syndrome. Other viral agents like Epstein-Barr virus are much more likely.

The growing number of referrals to specialists is a matter of concern to the Government and the 'health insurance collective' (ziekenfonds). In spite of all kinds of measures this increase in referrals continues, also in pediatrics. An effective policy to reverse this trend depends on an understanding of its causes. This paper goes into the mechanisms of supply and demand in health care and the roles of both G.P. and patient in the decision to refer. Parents worry about their children's symptoms. These worries are not always understood by their general practitioner and may lead to referrals in which not medical questions but the worries of the parents take a central place. In treatment at the Out-Patient Clinic consideration should be given not only to the questions, worries and doubts of the G.P., but also to those of the patient and above all the patient's parents.