Visualizing Galectin-3 Binding Protein Expression with ImmunoPET.

Galectin-3 binding protein (Gal-3BP) is a glycoprotein that is overexpressed and secreted by several cancers and has been implicated as a marker of both tumor progression and poor prognosis in melanoma, non-small cell lung cancer, head and neck squamous cell carcinoma, and breast cancer. The expression of Gal-3BP by a variety of neoplasms makes it an enticing target for both diagnostics and therapeutics, including immuno-positron emission tomography (immunoPET) probes and antibody-drug conjugates (ADCs). Herein, we report the development, in vitro characterization, and in vivo evaluation of a pair of Gal-3BP-targeting radioimmunoconjugates for 89Zr-immunoPET. A humanized anti-Gal-3BP antibody, 1959, and its corresponding ADC, 1959-sss/DM4 (DM4 = ravtansine), were modified with desferrioxamine (DFO) to yield DFO-1959 and DFO-1959-sss/DM4 immunoconjugates bearing 1-2 DFO/monoclonal antibody. Both DFO-modified immunoconjugates retained their affinity for Gal-3BP in enzyme-linked immunosorbent assay experiments. The chelator-bearing antibodies were radiolabeled with zirconium-89 (t1/2 ≈ 3.3 d) to produce radioimmunoconjugates ─ [89Zr]Zr-DFO-1959 and [89Zr]Zr-DFO-1959-sss/DM4 ─ with high specific activity (>444 MBq/mg, >12 mCi/mg) and stability (>80% intact after 168 h in human serum at 37 °C). In mice bearing subcutaneous Gal-3BP-secreting A375-MA1 xenografts, [89Zr]Zr-DFO-1959 clearly delineated tumor tissue, reaching a maximum tumoral activity concentration (54.8 ± 15.8%ID/g) and tumor-to-background contrast (tumor-to-blood = 8.0 ± 4.6) at 120 h post-injection. The administration of [89Zr]Zr-DFO-1959 to mice bearing subcutaneous Gal-3BP-expressing melanoma patient-derived xenografts produced similarly promising results. [89Zr]Zr-DFO-1959 and [89Zr]Zr-DFO-1959-sss/DM4 exhibited nearly identical pharmacokinetic profiles in the mice bearing A375-MA1 tumors, though the latter produced higher uptake in the spleen and kidneys. Both [89Zr]Zr-DFO-1959 and [89Zr]Zr-DFO-1959-sss/DM4 effectively visualized Gal-3BP-secreting tumors in murine models of melanoma. These results suggest that both probes could play a role in the clinical imaging of Gal-3BP-expressing malignancies, particularly as companion theranostics for the identification of patients likely to respond to Gal-3BP-targeted therapeutics such as 1959-sss/DM4.

Bio-Tailored Sensing at the Nanoscale: Biochemical Aspects and Applications.

The demonstration of the first enzyme-based electrode to detect glucose, published in 1967 by S. J. Updike and G. P. Hicks, kicked off huge efforts in building sensors where biomolecules are exploited as native or modified to achieve new or improved sensing performances. In this growing area, bionanotechnology has become prominent in demonstrating how nanomaterials can be tailored into responsive nanostructures using biomolecules and integrated into sensors to detect different analytes, e.g., biomarkers, antibiotics, toxins and organic compounds as well as whole cells and microorganisms with very high sensitivity. Accounting for the natural affinity between biomolecules and almost every type of nanomaterials and taking advantage of well-known crosslinking strategies to stabilize the resulting hybrid nanostructures, biosensors with broad applications and with unprecedented low detection limits have been realized. This review depicts a comprehensive collection of the most recent biochemical and biophysical strategies for building hybrid devices based on bioconjugated nanomaterials and their applications in label-free detection for diagnostics, food and environmental analysis.

Recent Advances on Affibody- and DARPin-Conjugated Nanomaterials in Cancer Therapy.

Affibodies and designed ankyrin repeat proteins (DARPins) are synthetic proteins originally derived from the Staphylococcus aureus virulence factor protein A and the human ankyrin repeat proteins, respectively. The use of these molecules in healthcare has been recently proposed as they are endowed with biochemical and biophysical features heavily demanded to target and fight diseases, as they have a strong binding affinity, solubility, small size, multiple functionalization sites, biocompatibility, and are easy to produce; furthermore, impressive chemical and thermal stability can be achieved. especially when using affibodies. In this sense, several examples reporting on affibodies and DARPins conjugated to nanomaterials have been published, demonstrating their suitability and feasibility in nanomedicine for cancer therapy. This minireview provides a survey of the most recent studies describing affibody- and DARPin-conjugated zero-dimensional nanomaterials, including inorganic, organic, and biological nanoparticles, nanorods, quantum dots, liposomes, and protein- and DNA-based assemblies for targeted cancer therapy in vitro and in vivo.

Taking Advantage of the Morpheein Behavior of Peroxiredoxin in Bionanotechnology.

Morpheeins are proteins that reversibly assemble into different oligomers, whose architectures are governed by conformational changes of the subunits. This property could be utilized in bionanotechnology where the building of nanometric and new high-ordered structures is required. By capitalizing on the adaptability of morpheeins to create patterned structures and exploiting their inborn affinity toward inorganic and living matter, "bottom-up" creation of nanostructures could be achieved using a single protein building block, which may be useful as such or as scaffolds for more complex materials. Peroxiredoxins represent the paradigm of a morpheein that can be applied to bionanotechnology. This review describes the structural and functional transitions that peroxiredoxins undergo to form high-order oligomers, e.g., rings, tubes, particles, and catenanes, and reports on the chemical and genetic engineering approaches to employ them in the generation of responsive nanostructures and nanodevices. The usefulness of the morpheeins' behavior is emphasized, supporting their use in future applications.

Distance education during COVID 19: an Italian survey on the university teachers' perspectives and their emotional conditions.

BACKGROUND: Following the COVID-19 pandemic, distance education (DE) replaced traditional "face-to-face" teaching and has become the main method of teaching. The aim of this study was to 1) evaluate the impact of DE by teachers in our department during the second semester of the 2019-20 academic year following the March-May 2020 Italian national lockdown and 2) evaluate the relationship between DE and the emotional well-being of teachers during the period of home confinement. METHODS: Ninety-seven university teachers (51.5% women; most represented age group 60-69 years range, 40.2%) responded to an anonymous online cross-sectional survey between July 15 - September 30, 2020, on the advantages and disadvantages of DE, developed by one online teacher focus group. The emotional conditions were assessed by a short version of the Beck Depression Inventory-II (BDI-II). The internal consistency reliability survey and the 10-item BDI-II were measured by Cronbach's alpha. A correlation analysis (r-Pearson) was conducted between the overall evaluation of the experience of DE and the variables included in the study. RESULTS: Teachers reported difficulties in technical aspects, and in psychological factors, as the discomfort of "speaking in the void" (64.7%). The absence of "face-to-face" eye contact with the students was complained by 81% of teachers. Significant impairments in sleep patterns and loss of energy were reported, with female teachers having greater difficulty concentrating than their male colleagues. A quarter of teachers showed depressive symptoms of varying severity. The most satisfied teachers were those most stimulated by DE (r = 0.752, p < 0.000), who showed a lower impact of depressive symptoms (r = - 0.289, p = 0.005). The teaching load in hours influenced the perception of disadvantages (r = 0.214, p = 0.035) and contributed to a lower appreciation of the challenges of DE. The more significant the manifestation of depressive symptoms during the lockdown was, the greater the subjective recovery of a good emotional condition once the domestic confinement was over (r = 0.344, p = 0.001), despite maintaining DE. CONCLUSIONS: Our study highlights the impact of technical, didactic, and psychological difficulties of DE, reported by our teachers. The appreciation of their new learning promoted by DE seemed related to better emotional well-being of university teachers accepting this "challenge" in their important role in the high-education system, influencing good learning and promoting students' professional success.

Local anesthetics counteract cell proliferation and migration of human triple-negative breast cancer and melanoma cells.

In different retrospective studies, a protective role of regional anesthetics in reducing cancer recurrence after surgery was indicated. Accordingly, it has been previously demonstrated a protective effect of anesthetics in breast cancer cells and in other types of cancer. On the other hand, how anesthetics influence cancer needs in-depth investigations. For this purpose, two different human cancer cell lines, MDA-MB-231, triple-negative breast cancer, and A375, melanoma, were used in this study. By means of Western blotting and immunofluorescence and terminal deoxynucleotidyl transferase dUTP nick end labeling analyses, the signal transduction pathways activated by the anesthetics, such as ropivacaine and levobupivacaine, were analyzed. The data obtained demonstrated that both anesthetics are able to counteract cell proliferation by positively modulating cell death signaling and by decreasing cell proliferation and survival pathways.

Antibody-Drug Conjugates: The New Frontier of Chemotherapy.

In recent years, antibody-drug conjugates (ADCs) have become promising antitumor agents to be used as one of the tools in personalized cancer medicine. ADCs are comprised of a drug with cytotoxic activity cross-linked to a monoclonal antibody, targeting antigens expressed at higher levels on tumor cells than on normal cells. By providing a selective targeting mechanism for cytotoxic drugs, ADCs improve the therapeutic index in clinical practice. In this review, the chemistry of ADC linker conjugation together with strategies adopted to improve antibody tolerability (by reducing antigenicity) are examined, with particular attention to ADCs approved by the regulatory agencies (the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA)) for treating cancer patients. Recent developments in engineering Immunoglobulin (Ig) genes and antibody humanization have greatly reduced some of the problems of the first generation of ADCs, beset by problems, such as random coupling of the payload and immunogenicity of the antibody. ADC development and clinical use is a fast, evolving area, and will likely prove an important modality for the treatment of cancer in the near future.

PPARγ and Cognitive Performance.

Recent findings have led to the discovery of many signaling pathways that link nuclear receptors with human conditions, including mental decline and neurodegenerative diseases. PPARγ agonists have been indicated as neuroprotective agents, supporting synaptic plasticity and neurite outgrowth. For these reasons, many PPARγ ligands have been proposed for the improvement of cognitive performance in different pathological conditions. In this review, the research on this issue is extensively discussed.

Lifestyle and Food Habits Impact on Chronic Diseases: Roles of PPARs.

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that exert important functions in mediating the pleiotropic effects of diverse exogenous factors such as physical exercise and food components. Particularly, PPARs act as transcription factors that control the expression of genes implicated in lipid and glucose metabolism, and cellular proliferation and differentiation. In this review, we aim to summarize the recent advancements reported on the effects of lifestyle and food habits on PPAR transcriptional activity in chronic disease.

Targeted therapy of human glioblastoma via delivery of a toxin through a peptide directed to cell surface nucleolin.

Targeted anticancer therapies demand discovery of new cellular targets to be exploited for the delivery of toxic molecules and drugs. In this perspective, in the last few years, nucleolin has been identified as an interesting surface marker to be used for the therapy of glioblastoma. In this study, we investigated whether a synthetic antagonist of cell-surface nucleolin known as N6L, previously reported to decrease both tumor growth and tumor angiogenesis in several cancer cell lines, including glioblastoma cells, as well as endothelial cells proliferation, could be exploited to deliver a protein toxin (saporin) to glioblastoma cells. The pseudopeptide N6L cross-linked to saporin-S6 induced internalization of the toxin inside glioblastoma cancer cells. Our results in vitro demonstrated the effectiveness of this conjugate in inducing cell death, with an ID50 four orders of magnitude lower than that observed for free N6L. Furthermore, the preliminary in vivo study demonstrated efficiency in reducing the tumor mass in an orthotopic mouse model of glioblastoma.

The Involvement of PPARs in the Peculiar Energetic Metabolism of Tumor Cells.

Energy homeostasis is crucial for cell fate, since all cellular activities are strongly dependent on the balance between catabolic and anabolic pathways. In particular, the modulation of metabolic and energetic pathways in cancer cells has been discussed in some reports, but subsequently has been neglected for a long time. Meanwhile, over the past 20 years, a recovery of the study regarding cancer metabolism has led to an increasing consideration of metabolic alterations in tumors. Cancer cells must adapt their metabolism to meet their energetic and biosynthetic demands, which are associated with the rapid growth of the primary tumor and colonization of distinct metastatic sites. Cancer cells are largely dependent on aerobic glycolysis for their energy production, but are also associated with increased fatty acid synthesis and increased rates of glutamine consumption. In fact, emerging evidence has shown that therapeutic resistance to cancer treatment may arise from the deregulation of glucose metabolism, fatty acid synthesis, and glutamine consumption. Cancer cells exhibit a series of metabolic alterations induced by mutations that lead to a gain-of-function of oncogenes, and a loss-of-function of tumor suppressor genes, including increased glucose consumption, reduced mitochondrial respiration, an increase of reactive oxygen species, and cell death resistance; all of these are responsible for cancer progression. Cholesterol metabolism is also altered in cancer cells and supports uncontrolled cell growth. In this context, we discuss the roles of peroxisome proliferator-activated receptors (PPARs), which are master regulators of cellular energetic metabolism in the deregulation of the energetic homeostasis, which is observed in cancer. We highlight the different roles of PPAR isotypes and the differential control of their transcription in various cancer cells.

Uric Acid Amplifies Aß Amyloid Effects Involved in the Cognitive Dysfunction/Dementia: Evidences From an Experimental Model In Vitro.

There is still a considerable debate concerning whether uric acid is neuroprotective or neurotoxic agent. To clarify this topic, we tested the effects of uric acid on neuronal cells biology by using differentiated SHSY5Y neuroblastoma cells incubated with amyloid ß to reproduce an in vitro model of Alzheimer's disease. The incubation of cells with uric acid at the dose of 40 µM or higher significantly reduced cell viability and potentiated the proapoptotic effect of amyloid ß. Finally, uric acid enhanced the generation of 4-hydroxynonenal and the expression of PPARß/δ promoted by amyloid ß, indicating a prooxidant effects. In conclusion, uric acid could exert a detrimental influence on neuronal biology being this influence further potentiated by the concomitant exposure to neurotoxic stimuli. This effect is evident for uric acid concentrations close to those achievable in cerebrospinal fluid in presence of mild hyperuricemia thus suggesting a potential role of uric acid in pathophysiology of cognitive dysfunction. These effects are influenced by the concentrations of uric acid and by the presence of favoring conditions that commonly occur in neurodegenerative disorders and well as in the aging brain, including increased oxidative stress and exposure to amyloid ß. J. Cell. Physiol. 232: 1069-1078, 2017. © 2016 Wiley Periodicals, Inc.

PPARα Antagonist AA452 Triggers Metabolic Reprogramming and Increases Sensitivity to Radiation Therapy in Human Glioblastoma Primary Cells.

Glioblastoma (GB) is the most common cancer in the brain and with an increasing incidence. Despite major advances in the field, there is no curative therapy for GB to date. Many solid tumors, including GB, experienced metabolic reprogramming in order to sustain uncontrolled proliferation, hypoxic conditions, and angiogenesis. PPARs, member of the steroid hormone receptor superfamily, are particularly involved in the control of energetic metabolism, particularly lipid metabolism, which has been reported deregulated in gliomas. PPARα was previously indicated by us as a potential therapeutic target for this neoplasm, due to the malignancy grade dependency of its expression, being particularly abundant in GB. In this work, we used a new PPARα antagonist on patient-derived GB primary cells, with particular focus on the effects on lipid metabolism and response to radiotherapy. The results obtained demonstrated that blocking PPARα results in cell death induction, increase of radiosensitivity, and decrease of migration. Therefore, AA452 is proposed as a new adjuvant for the gold standard therapies for GB, opening the possibility for preclinical and clinical trials for this class of compounds. J. Cell. Physiol. 232: 1458-1466, 2017. © 2016 Wiley Periodicals, Inc.

Flavopiridol: An Old Drug With New Perspectives? Implication for Development of New Drugs.

Glioblastoma, the most common brain tumor, is characterized by high proliferation rate, invasion, angiogenesis, and chemo- and radio-resistance. One of most remarkable feature of glioblastoma is the switch toward a glycolytic energetic metabolism that leads to high glucose uptake and consumption and a strong production of lactate. Activation of several oncogene pathways like Akt, c-myc, and ras induces glycolysis and angiogenesis and acts to assure glycolysis prosecution, tumor proliferation, and resistance to therapy. Therefore, the high glycolytic flux depends on the overexpression of glycolysis-related genes resulting in an overproduction of pyruvate and lactate. Metabolism of glioblastoma thus represents a key issue for cancer research. Flavopiridol is a synthetic flavonoid that inhibits a wide range of Cyclin-dependent kinase, that has been demonstrate to inactivate glycogen phosphorylase, decreasing glucose availability for glycolysis. In this work the study of glucose metabolism upon flavopiridol treatment in the two different glioblastoma cell lines. The results obtained point towards an effect of flavopiridol in glycolytic cells, thus suggesting a possible new use of this compound or flavopiridol-derived formulations in combination with anti-proliferative agents in glioblastoma patients. J. Cell. Physiol. 232: 312-322, 2017. © 2016 Wiley Periodicals, Inc.

The PPARß/δ Agonist GW0742 Induces Early Neuronal Maturation of Cortical Post-Mitotic Neurons: Role of PPARß/δ in Neuronal Maturation.

Increasing evidences support that signaling lipids participate in synaptic plasticity and cell survival, and that the lipid signaling is closely associated with neuronal differentiation, learning, and memory and with pathologic events, such as epilepsy and Alzheimer's disease. The Peroxisome Proliferator-Activated Receptors (PPAR) are strongly involved in the fatty acid cell signaling, as many of the natural lypophylic compounds are PPAR ligands. We have previously shown that PPARß/δ is the main isotype present in cortical neuron primary cultures and that during neuronal maturation, PPARß/δ is gradually increased and activated. To get more insight into the molecular mechanism by which PPARß/δ may be involved in neuronal maturation processes, in this work a specific PPARß/δ agonist, GW0742 was used administered alone or in association with a specific PPARß/δ antagonist, the GSK0660, and the parameters involved in neuronal differentiation and maturation were assayed. The data obtained demonstrated the strong involvement of PPARß/δ in neuronal maturation, triggering the agonist an anticipation of neuronal differentiation, and the antagonist abolishing the observed effects. These effects appear to be mediated by the activation of BDNF pathway.

Optimization of construct design and fermentation strategy for the production of bioactive ATF-SAP, a saporin based anti-tumoral uPAR-targeted chimera.

BACKGROUND: The big challenge in any anti-tumor therapeutic approach is represented by the development of drugs selectively acting on the target with limited side effects, that exploit the unique characteristics of malignant cells. The urokinase (urokinase-type plasminogen activator, uPA) and its receptor uPAR have been identified as preferential target candidates since they play a key role in the evolution of neoplasms and are associated with neoplasm aggressiveness and poor clinical outcome in several different tumor types. RESULTS: To selectively target uPAR over-expressing cancer cells, we prepared a set of chimeric proteins (ATF-SAP) formed by the human amino terminal fragments (ATF) of uPA and the plant ribosome inactivating protein saporin (SAP). Codon-usage optimization was used to increase the expression levels of the chimera in the methylotrophic yeast Pichia pastoris. We then moved the bioprocess to bioreactors and demonstrated that the fed-batch production of the recombinant protein can be successfully achieved, obtaining homogeneous discrete batches of the desired constructs. We also determined the cytotoxic activity of the obtained batch of ATF-SAP which was specifically cytotoxic for U937 leukemia cells, while another construct containing a catalytically inactive mutant form of SAP showed no activity. CONCLUSION: Our results demonstrate that the uPAR-targeted, saporin-based recombinant fusion ATF-SAP can be produced in a fed-batch fermentation with full retention of the molecules selective cytotoxicity and hence therapeutic potential.

PPARß/δ and γ in a rat model of Parkinson's disease: possible involvement in PD symptoms.

Parkinson's disease is one of the most common neurologic disorder, affecting about 1-4% of persons older than 60 years. Among the proposed mechanisms of PD generation, free radical damage is believed to play a pivotal role in the development and/or progression of the disease. Recently, PPARs, a class of transcription factors involved in several pathways both in physiological and pathological conditions, have been linked by us and others to neurodegeneration. Particularly, PPARγ and its ligands have been indicated as potential therapeutic targets for the treatment of several pathological conditions associated with neuroinflammation within the CNS. The anti-inflammatory function of PPARγ has attracted attention since agonists exert a broad spectrum of protective effects in several animal models of neurological diseases, including psychiatric diseases. On the other hand a detrimental role for PPARß/δ has been proposed in Alzheimer, being closely related to the decrease of BDNF and Trkfl. On these bases, in this work we used a 6-OHDA hemi-lesioned rat model, inducing loss of dopaminergic neurons, to study the effects of the lesion at three time points from the lesion (1, 2, and 3 weeks), in relevant areas of PD motor symptoms, such as substantia nigra and globus pallidus and in the area of reward and mood control, the nucleus accumbens. In particular, it was studied: (i) the expression of BDNF and its downstream signals; (ii) the modulation of PPARs levels. The results obtained indicate the possible use of a dual PPARß/δ antagonist/PPARγ agonist to counteract primary and secondary signs of PD neurodegeneration.

PDZ Domain in the Engineering and Production of a Saporin Chimeric Toxin as a Tool for targeting Cancer Cells.

In this paper we have studied a PDZ protein domain as a possible tool for cellular targeting of the ribosome inactivating protein Saporin, exploiting the ability of PDZ domains to recognize and bind short peptide sequences located at the C-terminus of a cognate protein. We have focused our attention on the PDZ domain from hCASK (Human calcium/calmodulin-dependent serine protein kinase) that binds extracellular CD98 in epithelial cells, being this antigen recognized as a marker for several human tumors and particularly considered a negative prognostic marker for human glioblastoma. We produced recombinant fusions of one or two hCASK-PDZ domains with the ribosome inactivating protein Saporin and assayed them on two human glioblastoma cell lines (GL15 and U87). These constructs proved to be toxic, with increasing activity as a function of the number of PDZ domains, and induce cell death by apoptotic mechanisms in a dose-dependent and/or time dependent manner.

Systematic comparison of single-chain Fv antibody-fusion toxin constructs containing Pseudomonas Exotoxin A or saporin produced in different microbial expression systems.

BACKGROUND: Antibodies raised against selected antigens over-expressed at the cell surface of malignant cells have been chemically conjugated to protein toxin domains to obtain immunotoxins (ITs) able to selectively kill cancer cells. Since latest generation immunotoxins are composed of a toxic domain genetically fused to antibody fragment(s) which confer on the IT target selective specificity, we rescued from the hydridoma 4KB128, a recombinant single-chain variable fragment (scFv) targeting CD22, a marker antigen expressed by B-lineage leukaemias and lymphomas. We constructed several ITs using two enzymatic toxins both able to block protein translation, one of bacterial origin (a truncated version of Pseudomonas exotoxin A, PE40) endowed with EF-2 ADP-ribosylation activity, the other being the plant ribosome-inactivating protein saporin, able to specifically depurinate 23/26/28S ribosomal RNA. PE40 was selected because it has been widely used for the construction of recombinant ITs that have already undergone evaluation in clinical trials. Saporin has also been evaluated clinically and has recently been expressed successfully at high levels in a Pichia pastoris expression system. The aim of the present study was to evaluate optimal microbial expression of various IT formats. RESULTS: An anti-CD22 scFv termed 4KB was obtained which showed the expected binding activity which was also internalized by CD22+ target cells and was also competed for by the parental monoclonal CD22 antibody. Several fusion constructs were designed and expressed either in E. coli or in Pichia pastoris and the resulting fusion proteins affinity-purified. Protein synthesis inhibition assays were performed on CD22+ human Daudi cells and showed that the selected ITs were active, having IC50 values (concentration inhibiting protein synthesis by 50% relative to controls) in the nanomolar range. CONCLUSIONS: We undertook a systematic comparison between the performance of the different fusion constructs, with respect to yields in E. coli or P. pastoris expression systems and also with regard to each constructs specific killing efficacy. Our results confirm that E. coli is the system of choice for the expression of recombinant fusion toxins of bacterial origin whereas we further demonstrate that saporin-based ITs are best expressed and recovered from P. pastoris cultures after yeast codon-usage optimization.

Anti-LGALS3BP antibody-drug conjugate treatment induces durable and potent antitumor response in a preclinical model of adenoid cystic carcinoma.

OBJECTIVES: Adenoid cystic carcinoma (ACC) is a rare type of cancer that typically arises from glandular tissues, most commonly in the salivary glands. Although relatively rare, it represents a serious clinical issue as the management of the disease is highly complex being the only therapeutic options represented by invasive surgery and/or radiotherapy. In the present study, we have explored the potential of galectin-3 binding protein (LGALS3BP) as a novel target for antibody-drug conjugate (ADC) therapy in ACC. MATERIALS AND METHODS: RNAseq was conducted on a panel of 10 ACC patient-derived xenografts (PDX)s tissues and 6 normal salivary glands to analyze LGALS3BP gene expression. Protein expression was assessed in ACC PDX and primary tumor tissues using immunohistochemistry. Anti-LGALS3BP ADC named 1959-sss/DM4, was tested in high LGALS3BP expressing ACC PDX model ST1502B. RESULTS: RNAseq analysis revealed that LGALS3BP expression was highly expressed in ACC PDX tissues compared to normal salivary gland tissues. As evaluated by immunohistochemical analysis, LGALS3BP protein was found to be heterogeneously expressed in 10 ACC PDX and in tumor tissues derived from a cohort of 37 ACC patients. Further, treatment with 1959-sss/DM4 ADC led to durable tumor growth inhibition (TGI) in 100% of animals without observed toxicity. CONCLUSIONS: Our study provides strong evidence that LGALS3BP is a promising therapeutic target for ACC, warranting further expedited preclinical and clinical investigation.