Promyelomonocytic leukemia (PML) is a prominent oncosuppressor whose inactivation is involved in the pathogenesis of hematological and epithelial cancers. Here, we report that PML aggregated in nuclear bodies in syncytia elicited by the envelope glycoprotein complex (Env) of human immunodeficiency virus-1 (HIV-1) in vitro. PML aggregation occurred after the fusion of nuclei (karyogamy) within syncytia but before the apoptotic program was activated. The aggregation of PML was detectable in syncytia present in the brain or lymph nodes from patients with HIV-1 infection, as well as in a fraction of blood leukocytes, correlating with viral status. Using a range of specific inhibitors of PML (the oncogenic PML/RARα fusion product or specific small interfering RNAs), we demonstrated that, in Env-elicited syncytia, PML was required for activating phosphorylation of ataxia telangiectasia mutated (ATM), which colocalized with PML in nuclear bodies, in a molecular complex that also involved topoisomerase IIβ-binding protein 1. PML knockdown thus inhibited the ATM-dependent DNA damage response that culminates in the activation of p53, p53-dependent transcription of pro-apoptotic genes and cell death. Infection of CD4-expressing cells with HIV-1 also induced syncytial apoptosis, which could be suppressed by inhibiting PML. Altogether, these data indicate that PML activation is a critical early event that participates in the apoptotic demise of HIV-1-elicited syncytia.

Background: Immune checkpoint blockade (ICB) therapies demonstrate low efficacy in microsatellite stable (MSS) colorectal cancer (CRC) due to an immune-desert tumor microenvironment (TME) characterized by low antigen presentation and limited tumor-infiltrating lymphocytes (TILs). Harmine, a natural small-molecule and its promising derivatives ACB1801 have shown anti-tumor potential in preclinical models; however, their potential to reprogram the TME and overcome ICB resistance in MSS CRC remains unexplored. This study investigates whether and how ACB1801 can reshape TME to sensitize MSS CRC to ICB therapies. Methods: We used the CT26 MSS colorectal cancer mouse model to evaluate the ability of the harmine derivative ACB1801 to enhance the efficacy of anti-PD-1 therapy. To characterize its mode of action, we performed immune landscape analysis and transcriptomic profiling of both CD45- and CD45+ tumor-derived cells. In parallel, mechanistic studies were conducted in vitro using mouse and human MSS CRC cell lines. Results We demonstrate that the harmine derivative ACB1801 enhances the effectiveness of anti-PD-1 therapy in an MSS CRC mouse model. Combination therapy significantly increased CD8+ T cell infiltration and reduced regulatory T-cell (Treg) density in the TME. Transcriptomic profiling of CRC cells isolated from tumors treated with either anti-PD-1 alone or in combination with ACB1801 revealed significant enrichment of metabolic pathways in the combination group, characterized by reduced glycolysis and enhanced ferroptosis signatures. These findings were supported by in vitro data showing that ACB1801 reduces tumor cell glycolytic activity and promotes ferroptotic vulnerability. Mechanistically, ACB1801 induced STAT1 signaling, promoted CXCL10 release, and enhanced major histocompatibility complex class I (MHC-I)-dependent antigen presentation on tumor cells, thereby increasing tumor susceptibility to anti-PD-1 therapy. Conclusion: Collectively, our findings indicate that combination therapy with harmine derivatives and ICBs represents a promising strategy for treating MSS CRC patients.

Abstract Colorectal cancer (CRC) is the most common gastrointestinal cancer and the third leading cause of cancer-related death in both sexes. Mismatch repair deficient/microsatellite instability-high (MSI) colorectal cancer patients benefit from immune checkpoint blockades (ICBs)-based immunotherapy; however, this benefit has not been translated into microsatellite stable (MSS) colorectal cancer. It has been proposed that the loss of major histocompatibility complex class I (MHC-I) in tumor cells limits the use of ICB in colorectal cancer. Harmine displays a number of biological and pharmacological properties affecting the immune response and tumor phenotype. Here, we assessed the impact of combining harmine, also referred to as ACB1801 molecule, on improving the responsiveness of ICB in the mouse MSS CRC model. Our results showed that combining ACB1801 significantly improves the therapeutic benefit of anti-PD-1 in the CT26 CRC mouse model by decreasing tumor growth and improving the survival of tumor-bearing mice. The improvement of anti-PD-1-based therapy by combining ACB-1801 was associated with a modification of the immune landscape of tumors as evidenced by an increase in the CD8+ T cells and a decrease in the Treg infiltrations into the tumor microenvironment. By profiling the cytokine/chemokine network, we showed that ACB1801 induced the expression and the release of the proinflammatory chemokine CXCL10 by CRC tumor cells in vitro, which could be responsible for driving CD8+ T cells to the tumor microenvironment. Mechanistically, we showed that ACB1801 increased the expression of MHC-I genes, including TAP1 and TAPASIN, and improved the antigen loading on MHC-I in CRC cells in vitro. Overall, our study highlights the value of combining ACB1801 and anti-PD-1 as a therapeutic opportunity to convert MSS colorectal cancer into an “immune hot” cancer, which may define the future treatment paradigm of colorectal cancer for which there is a great unmet need. Citation Format: Ruize Gao, Kris Van Moer, Coralie Pulido, Anaïs Oudin, Diane Murera, Teresa L. Ramos, Margaux Poussard, Andreas Schläpfer, Annette Ives, Christian Auclair, Bassam Janji. The beta-carboline Harmine sensitives microsatellite stable colorectal cancer mouse model to anti-PD-1 through upregulating MHC-I dependent antigen presentation machinery and improving the infiltration of CD8 T cells into the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4049.

Figure S3: Characterization of UT-7/Epo, HMC1.1 and HMC1.2 cell lines. Figure S4: 2D1 and 3G1-Fc do not compete with 104D2 mAb binding to KIT.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may cause severe respiratory illness with high mortality. SARS-CoV-2 infection results in a massive inflammatory cell infiltration into the infected lungs accompanied by excessive pro-inflammatory cytokine production. The lung histology of dead patients shows that some areas are severely emphysematous, with enormously dilated blood vessels and micro-thromboses. The inappropriate inflammatory response damaging the pulmonary interstitial arteriolar walls suggests that the respiratory distress may come in a large part from lung vasculature injuries. It has been recently observed that low plasmatic sphingosine-1-phosphate (S1P) is a marker of a worse prognosis of clinical outcome in severe coronavirus disease (COVID) patients. S1P is an angiogenic molecule displaying anti-inflammatory and anti-apoptotic properties, that promote intercellular interactions between endothelial cells and pericytes resulting in the stabilization of arteries and capillaries. In this context, it can be hypothesized that the benefit of a normal S1P level is due to its protective effect on lung vasculature functionality. This paper provides evidence supporting this concept, opening the way for the design of a pharmacological approach involving the use of an S1P lyase inhibitor to increase the S1P level that in turn will rescue the lung vasculature functionality.

Monitoring of phage antibody selection by titration, ELISA and scFv sequencing.

Materials and methods describing the production of scFv-Fc in HEK-T cells and KIT domains and scFv display on yeast. Table S2: Primers for PCR amplification of KIT and cloning into pYD vector by Gap repair. Figure S1: Selection and initial characterization of a panel of anti-KIT scFv. Figure S2: Anti-KIT D5 epitope characterization using yeast display.

Increasing evidence indicates that tumor vasculature normalization could be an appropriate strategy to increase therapies' efficacy in solid tumors by decreasing hypoxia and improving drug delivery. We searched for a novel approach that reduces hypoxia and enhances chemotherapy efficacy in pancreatic adenocarcinoma which is characterized by disrupted blood vasculature associated with poor patient survival. Clinical significance of plasma levels of the angiogenic lipid sphingosine-1-phosphate (S1P) was assessed at baseline in 175 patients. High plasma S1P concentration was found to be a favorable prognostic/predictive marker in advanced/metastatic pancreatic adenocarcinoma patients treated by gemcitabine alone but not in patients receiving a combination gemcitabine and PDGFR-inhibitor. In pancreatic adenocarcinoma PDX models, oral administration of an S1P lyase inhibitor (LX2931) significantly increased plasma S1P levels, decreased tumor expression of the hypoxia marker (CA IX), and enhanced chemotherapy efficacy when combined with gemcitabine treatment. The direct effect of S1P on tumor oxygenation was assessed by administration of S1P onto tumor-grafted CAM model and measuring intra-tumoral pO2 using a tissue oxygen monitor. S1P increased pO2 in a tumor-CAM model. Thus, increasing plasma S1P is a promising strategy to decrease tumor hypoxia and enhance therapy efficacy in solid tumors. S1P may act as a tumor vasculature normalizer.

Harmine is a dual-specificity tyrosine-regulated kinase 1A (DYRK1A) inhibitor that displays a number of biological and pharmacological properties. Also referred to as ACB1801 molecule, we have previously reported that harmine increases the presentation of major histocompatibility complex (MHC)-I-dependent antigen on melanoma cells. Here, we show that ACB1801 upregulates the mRNA expression of several proteins of the MHC-I such as Transporter Associated with antigen Processing TAP1 and 2, Tapasin and Lmp2 (hereafter referred to as MHC-I signature) in melanoma cells. Treatment of mice bearing melanoma B16-F10 with ACB1801 inhibits the growth and weight of tumors and induces a profound modification of the tumor immune landscape. Strikingly, combining ACB1801 with anti-PD1 significantly improves its therapeutic benefit in B16-F10 melanoma-bearing mice. These results suggest that, by increasing the MHC-I, ACB1801 can be combined with anti-PD1/PD-L1 therapy to improve the survival benefit in cancer patients displaying a defect in MHC-I expression. This is further supported by data showing that i) high expression levels of TAP1, Tapasin and Lmp2 was observed in melanoma patients that respond to anti-PD1; ii) the survival is significantly improved in melanoma patients who express high MHC-I signature relative to those expressing low MHC-I signature; and iii) high expression of MHC-I signature in melanoma patients was correlated with increased expression of CD8 and NK cell markers and overexpression of proinflammatory chemokines involved in the recruitment of CD8+ T cells.

Immune escape is one of the key mechanisms mediating cancer progression and metastatic invasion. This creates significant obstacles to successful cancer treatment, especially those reliant on the immune response such as T cell adoptive immunotherapy and immune checkpoint inhibitors (ICI) therapy. Despite a functioning host immune system, tumor cells often acquire either poorly immunogenic or immunosuppressive phenotypes, which impair immune-dependent resolution.

Numerous studies have shown that alteration of actin remodeling plays a pivotal role in the regulation of morphologic and phenotypic changes leading to malignancy. In the present study, we searched for drugs that can regulate actin polymerization and reverse the malignant phenotype in cancer cells. We developed a cell-free high-throughput screening assay for the identification of compounds that induce the actin polymerization in vitro, by fluorescence anisotropy. Then, the potential of the hit compound to restore the actin cytoskeleton and reverse the malignant phenotype was checked in EWS-Fli1-transformed fibroblasts and in B16-F10 melanoma cells. A β-carboline extracted from Peganum harmala (i.e., harmine) is identified as a stimulator of actin polymerization through a mechanism independent of actin binding and requiring intracellular factors involved in a process that regulates actin kinetics. Treatment of malignant cells with non-cytotoxic concentrations of harmine induces the recovery of a non-malignant cell morphology accompanied by reorganization of the actin cytoskeleton, rescued cell–cell adhesion, inhibition of cell motility and loss of anchorage-independent growth. In conclusion, harmine induces the reversion of the malignant phenotype by a process involving the modulation of actin dynamics and is a potential anti-tumor agent acting principally through a non-cytotoxic process.

AC BioScience is a Swiss biotech company based at the EPFL Innovation Park and Biopôle, dedicated to developing groundbreaking therapies to fight a range of cancers and infectious diseases. We are about to start clinical trials with two of four leading-edge cancer drugs mainly focusing on immune-oncology and tumor vascular normalization with multi-billion $ sales potential. Here, we present our strategy and one of our pioneering drug candidates that has already shown exceptional results with tumor cell conditioning to improve the efficacy of immune checkpoint inhibitors.

Abstract Objective: To assess masitinib in the treatment of ALS. Methods: Double-blind study, randomly assigning 394 patients (1:1:1) to receive riluzole (100 mg/d) plus placebo or masitinib at 4.5 or 3.0 mg/kg/d. Following a blinded transition from phase 2 to phase 2/3, a prospectively defined two-tiered design was implemented based on ALSFRS-R progression rate from disease-onset to baseline (ΔFS). This approach selects a more homogeneous primary efficacy population (“Normal Progressors”, ΔFS < 1.1 points/month) while concurrently permitting secondary assessment of the broader population. Primary endpoint was decline in ALSFRS-R at week-48 (ΔALSFRS-R), with the high-dose “Normal Progressor” cohort being the prospectively declared primary efficacy population. Missing data were imputed via last observation carried forward (LOCF) methodology with sensitivity analyses performed to test robustness. Results: For the primary efficacy population, masitinib (n = 99) showed significant benefit over placebo (n = 102) with a ΔALSFRS-R between-group difference (ΔLSM) of 3.4 (95% CI 0.65–6.13; p = 0.016), corresponding to a 27% slowing in rate of functional decline (LOCF methodology). Sensitivity analyses were all convergent, including the conservative multiple imputation technique of FCS-REGPMM with a ΔLSM of 3.4 (95% CI 0.53–6.33; p = 0.020). Secondary endpoints (ALSAQ-40, FVC, and time-to-event analysis) were also significant. Conversely, no significant treatment-effect according to ΔALSFRS-R was seen for the broader “Normal and Fast Progressor” masitinib 4.5 mg/kg/d cohort, or either of the low-dose (masitinib 3.0 mg/kg/d) cohorts. Rates of treatment-emergent adverse events (AEs) (regardless of causality or post-onset ΔFS) were 88% with masitinib 4.5 mg/kg/d, 85% with 3.0 mg/kg/d, and 79% with placebo. Likewise, rates of serious AE were 31, 23, and 18%, respectively. No distinct event contributed to the higher rate observed for masitinib and no deaths were related to masitinib. Conclusions: Results show that masitinib at 4.5 mg/kg/d can benefit patients with ALS. A confirmatory phase 3 study will be initiated to substantiate these data.

On the basis of pulse radiolysis data, the one-electron reduction of dioxygen was estimated to measure −0.33 V in aqueous media. Superoxide anion (O2 −) acts as a one-electron reducing agent in several reactions including reduction of quinones, tetranitromethane, cytochrome c and nitro-substituted aromatics such as nitroblue tetrazolium (NBT). 1 - 3 The reduction of NBT to blue formazan has been widely used as a probe of O2 − generation in chemical and biological systems. The reliability of the technique is based on the following assumptions:

Protein Kinase D1 (PKD1) is a serine/threonine kinase encoded by the PRKD1 gene. PKD1 has been previously shown to be a prognostic factor in ERα+ tamoxifen-resistant breast tumors and PKD1 overexpression confers estrogen independence to ERα+ MCF7 cells. In the present study, our goal was to determine whether PKD1 is a prognostic factor and/or a relevant therapeutic target in breast cancer. We analyzed PRKD1 mRNA levels in 527 primary breast tumors. We found that high PRKD1 mRNA levels were significantly and independently associated with a low metastasis-free survival in the whole breast cancer population and in the triple-negative breast cancer (TNBC) subtype specifically. High PRKD1 mRNA levels were also associated with a low overall survival in TNBC. We identified novel PKD1 inhibitors and assessed their antitumor activity in vitro in TNBC cell lines and in vivo in a TNBC patient-derived xenograft (PDX) model. Pharmacological inhibition and siRNA-mediated depletion of PKD1 reduced colony formation in MDA-MB-436 TNBC cells. PKD1 inhibition also reduced tumor growth in vivo in a TNBC PDX model. Together, these results establish PKD1 as a poor prognostic factor and a potential therapeutic target in TNBC.

We have initially demonstrated in knocking down experiments that decreasing TCTP in cancer cells leads in some tissues to cell death while in others to a complete reorganization of the tumor into architectural structures reminiscent of normal ones. Based on these experiments and a series of other findings confirming the key role of TCTP in cancer, it became important to find pharmacological compounds to inhibit its function, and this became for us a priority. In the present text, we explain in detail the experiments that were performed and the perspectives of sertraline in cancer treatment, as this became today a reality with a clinical study that started in collaboration with Columbia University and Johns Hopkins University.

Abstract Tamoxifen-resistant ERα positive (ERα+) breast cancers and triple-negative breast cancers represent two aggressive subgroups of mammary tumors with limited treatment options and a poor prognosis. Thus, there is still an urgent need to develop and improve targeted pharmacological treatments against these estrogen-independent cancers. Recent data from our group suggest that the protein kinase D1 (PKD1) participates in the development and/or maintenance of the estrogen-independent phenotype in breast cancer. Indeed, up-regulating PKD1 conferred anchorage-independent growth to estrogen-dependent MCF-7 cells in the absence of β-estradiol. In addition, we found that high PKD1 mRNA levels were associated with a worse prognosis in tamoxifen-treated tumors. PKD1 is a serine-threonine kinase encoded by the prkd1 gene and belongs to the protein kinase D (PKD) family, a novel subgroup of the calcium and calmodulin-regulated kinases (CAMK). The PKD family also includes PKD2 and PKD3 which share a high homology with PKD1. The objective of the present study was to determine whether PKD1 could be a prognostic factor and/or a pharmacological target for the treatment of breast cancer. Expression of prkd1, prkd2 and prkd3 was analyzed by RT-qPCR in 527 breast cancer specimens (181 ERα- tumors including 102 triple negative tumors and 346 ERα+ tumors). We showed that higher prkd1 mRNA levels were associated with a lower metastasis-free survival in the overall breast cancer population (p=0.015). prkd1 prognostic value was even stronger in ERα- tumors (p=0.00042) and in triple negative tumors (p=0.0043). Conversely, prkd1 expression was not associated with prognosis in ERα+ tumors. Interestingly, high prkd2 and prkd3 mRNA levels were associated with a better prognosis in the entire cohort (p=0.0074 and p=0.028 respectively). We have developed PKD1 inhibitors in collaboration with the AB Science pharmaceutical company and screened the ability of thirty-five compounds to inhibit anchorage-independent growth of MCF-7 cells in the absence of estrogens. The antitumor activity of two PKD1 inhibitors is currently being evaluated in vivo against patient-derived breast cancer xenografts. In conclusion, our results show that high prkd1 expression is a bad-prognosis factor in breast cancer and that PKD1 could be a relevant therapeutic target for the treatment of breast cancer, in particular those harboring an estrogen-independent phenotype. Citation Format: Caroline Spasojevic, Marie Regairaz, Sophie Vacher, Franck Assayag, Jean Marc Ricort, Elisabetta Marangoni, Ivan Bièche, Christian Auclair. Validation of protein kinase D1 as a prognostic factor and pharmacological target for the treatment of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3121. doi:10.1158/1538-7445.AM2017-3121

BACKGROUND:Melanoma is a highly metastatic type of cancer that is resistant to all standard anticancer therapies and thus has a poor prognosis. Therefore, metastatic melanoma represents a significant clinical problem and requires novel and effective targeted therapies. The protein kinase C (PKC) family comprises multiple isoforms of serine/threonine kinases that possess distinct roles in cancer development and progression. In this study, we determined whether inhibition of PKC could revert a major process required for melanoma progression and metastasis; i.e. the E- to N-cadherin switch.METHODS:The cadherin switch was analyzed in different patient-derived primary tumors and their respective metastatic melanoma cells to determine the appropriate cellular model (aggressive E-cadherin-negative/N-cadherin-positive metastasis-derived melanoma cells). Next, PKC inhibition in two selected metastatic melanoma cell lines, was performed by using either pharmacological inhibitors (Gö6976 and Gö6983) or stable lentiviral shRNA transduction. The expression of E-cadherin and N-cadherin was determined by western blot. The consequences of cadherin switch reversion were analyzed: cell morphology, intercellular interactions, and β-catenin subcellular localization were analyzed by immunofluorescence labeling and confocal microscopy; cyclin D1 expression was analyzed by western blot; cell metastatic potential was determined by anchorage-independent growth assay using methylcellulose as semi-solid medium and cell migration potential by wound healing and transwell assays.RESULTS:Gö6976 but not Gö6983 reversed the E- to N-cadherin switch and as a consequence induced intercellular interactions, profound morphological changes from elongated mesenchymal-like to cuboidal epithelial-like shape, β-catenin translocation from the nucleus to the plasma membrane inhibiting its oncogenic function, and reverting the metastatic potential of the aggressive melanoma cells. Comparison of the target spectrum of these inhibitors indicated that these observations were not the consequence of the inhibition of conventional PKCs (cPKCs), but allowed the identification of a novel serine/threonine kinase, i.e. protein kinase Cμ, also known as protein kinase D1 (PKD1), whose specific inhibition allows the reversion of the metastatic phenotype in aggressive melanoma.CONCLUSION:In conclusion, our study suggests, for the first time, that while cPKCs don't embody a pertinent therapeutic target, inhibition of PKD1 represents a novel attractive approach for the treatment of metastatic melanoma.

L’essai clinique international AB06006 a évalué l’efficacité et la tolérance du masitinib par voie orale (6 mg/kg/j pendant 24 semaines avec extension possible) contre placebo dans les patients (pts) atteints de mastocytose systémique indolente (MSI) ou mastocytose systémique latente/smouldering (MSL) non-répondeur aux traitements symptomatiques optimisés. Les pts avec symptômes graves (n = 135) ont étés définis comme ayant au moins une caractéristiques de base parmi : score de prurit ≥ 9 ; nombre de bouffées vasomotrice/semaine ≥ 8 ; dépression (score HAMD-17) ≥ 19 ; asthénie ≥ 75 selon le score total du Fatigue Impact Scale (FIS). L’effet du traitement a été évalué selon une méthode de mesures répétées pour maladies rares via le modèle d’équation d’estimation généralisé dans la population en intention de traiter modifié. Une réponse était définie comme cliniquement significative à ≥ 75 % d’amélioration par rapport aux caractéristiques de base des symptômes. Le critère d’évaluation principal (4H75 %) était la réponse cumulative (période S8–S24) sur au moins un des symptômes graves. La sécurité à long terme a été analysée sur la période S8-S96. Les critères secondaires comprenaient la sécurité, les symptômes rapportés par les pts et les critères d’évaluation objectifs représentatifs de l’activité des mastocytes ou du fardeau associé à la maladie. Le masitinib a montré une amélioration significative par rapport au placebo sur le critère d’évaluation primaire, 18,7 vs. 7,4 %, rapport de cotes (RC) de 3,6 (IC95 % 1,2–10,8, p = 0,008). Le masitinib a maintenu une réponse significative 4H75 % sur le long terme avec un RC de 3,5 (IC95 % 1,3–9,7, p = 0,016). Ce résultat est corroboré par la variation moyenne de la tryptase et l’urticaire pigmentaire. Les effets indésirables (EI) étaient principalement des gastro-intestinaux ou cutanés et généralement gérables via une réduction de dose. Les EIs graves rapportés avec une différence > 4 % entre masitinib et placebo étaient la diarrhée (9,8 %), éruption cutanée (5,7 %) et asthénie (4,1 %). Il n’y a pas eu de décès ou de risque mortel dans le bras masitinib. La sécurité à long terme a révélé une incidence d’EIs comparable entre masitinib et placebo. Le masitinib a généré un bénéfice thérapeutique chez les pts avec MSI/MSL avec symptômes graves. Le masitinib était capable de maintenir cet effet pendant plus de 2 ans avec une incidence similaire des effets indésirables par rapport au placebo. Le masitinib a montré un rapport bénéfice/risque positif et peut être considéré comme une option thérapeutique pour les pts atteints de MSI/MSL avec symptômes graves qui ne répondent pas aux traitements symptomatiques optimaux.