Liposomal Rifabutin-A Promising Antibiotic Repurposing Strategy against Methicillin-Resistant Staphylococcus aureus Infections.

Methicillin-resistant Staphylococcus aureus (M RSA) infections, in particular biofilm-organized bacteria, remain a clinical challenge and a serious health problem. Rifabutin (RFB), an antibiotic of the rifamycins class, has shown in previous work excellent anti-staphylococcal activity. Here, we proposed to load RFB in liposomes aiming to promote the accumulation of RFB at infected sites and consequently enhance the therapeutic potency. Two clinical isolates of MRSA, MRSA-C1 and MRSA-C2, were used to test the developed formulations, as well as the positive control, vancomycin (VCM). RFB in free and liposomal forms displayed high antibacterial activity, with similar potency between tested formulations. In MRSA-C1, minimal inhibitory concentrations (MIC) for Free RFB and liposomal RFB were 0.009 and 0.013 µg/mL, respectively. Minimum biofilm inhibitory concentrations able to inhibit 50% biofilm growth (MBIC50) for Free RFB and liposomal RFB against MRSA-C1 were 0.012 and 0.008 µg/mL, respectively. Confocal microscopy studies demonstrated the rapid internalization of unloaded and RFB-loaded liposomes in the bacterial biofilm matrix. In murine models of systemic MRSA-C1 infection, Balb/c mice were treated with RFB formulations and VCM at 20 and 40 mg/kg of body weight, respectively. The in vivo results demonstrated a significant reduction in bacterial burden and growth index in major organs of mice treated with RFB formulations, as compared to Control and VCM (positive control) groups. Furthermore, the VCM therapeutic dose was two fold higher than the one used for RFB formulations, reinforcing the therapeutic potency of the proposed strategy. In addition, RFB formulations were the only formulations associated with 100% survival. Globally, this study emphasizes the potential of RFB nanoformulations as an effective and safe approach against MRSA infections.

Mapping the contributions of dyadic approaches to couples' psychosocial adaptation to prostate cancer: a scoping review.

INTRODUCTION: Prostate cancer is the second-most prevalent cancer diagnosis worldwide among males. Although prostate cancer affects the physical, sexual, and mental health of patients, the impact of prostate cancer on partners has also been increasingly recognized. Hence, taking a dyadic approach is of relevance. Moreover, there is evidence of the utility of dyadic approaches to the study of relational stress that chronic diseases such as prostate cancer can bring to couples, even though knowledge is sparse about prostate cancer. OBJECTIVES: This scoping review aimed to map existing dyadic studies on the psychosocial adaptation of couples to prostate cancer. METHODS: A systematic search of studies published from 2005 to November 2022 was conducted on electronic databases (PubMed, Cochrane Library, EBSCOHost, Scopus, and Web of Science) following PRISMA-ScR (Preferred Reporting Items for Systematic Reviews and Meta-analyses-Extension for Scoping Reviews). RESULTS: The review included 25 eligible studies from the initial 2514 articles retrieved. Overall, the results emphasized the interdependency between couple members and suggested how partners' adaptation influences patients' adaptation to prostate cancer and vice versa, regarding several psychosocial dimensions (eg, intimacy, quality of life). CONCLUSIONS: This work can bring awareness to health care professionals to adopt a couples approach when managing prostate cancer whenever there is a partner, due to these interdependent influences. For researchers and future studies, this work can strengthen the relevance of dyadic approaches on how couples adapt to prostate cancer and explore which other dimensions influence these complex dynamics.

Panobinostat-loaded folate targeted liposomes as a promising drug delivery system for treatment of canine B-cell lymphoma.

Introduction: Cancer is a major public health problem with over 19 million cases reported in 2020. Similarly to humans, dogs are also largely affected by cancer, with non-Hodgkin's lymphoma (NHL) among the most common cancers in both species. Comparative medicine has the potential to accelerate the development of new therapeutic options in oncology by leveraging commonalities between diseases affecting both humans and animals. Within this context, in the present study, we investigated the potential of panobinostat (Pan)-loaded folate-targeted PEGylated liposomes (FA-PEG-Pan-Lip) for the treatment of canine B-cell lymphoma, while contributing to new perspectives in comparative oncology. Methods and results: Two formulations were developed, namely: PEG-Pan-Lip and FA-PEG-Pan-Lip. Firstly, folate receptor expression in the CLBL-1 canine B-cell lymphoma cell line was assessed. After confirming receptor expression, both Pan-loaded formulations (PEG-Pan-Lip, FA-PEG-Pan-Lip) demonstrated dose-dependent inhibitory effects on CLBL-1 cell proliferation. The FA-PEG-Pan-Lip formulation (IC50 = 10.9 ± 0.03 nM) showed higher cytotoxicity than the non-targeted PEG-Pan-Lip formulation (IC50 = 12.9 ± 0.03 nM) and the free panobinostat (Pan) compound (IC50 = 18.32±0.03 nM). Moreover, mechanistically, both Pan-containing formulations induced acetylation of H3 histone and apoptosis. Flow cytometry and immunofluorescence analysis of intracellular uptake of rhodamine-labeled liposome formulations in CLBL-1 cells confirmed cellular internalization of PEG-Lip and FA-PEG-Lip formulations and higher uptake profile for the latter. Biodistribution studies of both radiolabeled formulations in CD1 and SCID mice revealed a rapid clearance from the major organs and a 1.6-fold enhancement of tumor uptake at 24 h for 111In-FA-PEG-Pan-Lip (2.2 ± 0.1 %ID/g of tumor) compared to 111In-PEG-Pan-Lip formulation (1.2±0.2 %ID/g of tumor). Discussion: In summary, our results provide new data validating Pan-loaded folate liposomes as a promising targeted drug delivery system for the treatment of canine B-cell lymphoma and open innovative perspectives for comparative oncology.

Short-term Clinical and Microbiological Performance of Resin-modified Glass Ionomer Cement Containing Chlorhexidine for Atraumatic Restorative Treatment.

Aim: This study evaluated the short-term clinical and microbiological performance of resin-modified glass ionomer cement (RM-GIC) cement containing chlorhexidine (CHX) for atraumatic restorative treatment (ART) in primary teeth. Materials and methods: The clinical trial was conducted in 36 children that received ART in primary molars either with GIC (group I, n = 18) or GIC containing 1.25% CHX (group II, n = 18). The survival rate of restorations was checked 7 days, 3, and 6 months after their application when saliva and biofilm were collected for microbiological assessment of mutans streptococci (MS) counts. Data were analyzed using the Kruskal-Wallis/Mann-Whitney U tests for clinical analysis and microbiological evaluations (p < 0.05). Results: The survival rate of restorations was similar comparing groups I with II. Microbiological analysis showed a significant reduction in MS levels 7 days after the treatment in both saliva and biofilm of children treated with RM-GIC containing CHX (group II); however, MS counts at 3 and 6 months did not differ from the initial counts. Conclusion: A total of 1.25% CHX improved the microbiological properties of GIC in the short term without impairing the clinical performance of ART restorations. Clinical significance: Glass ionomer cement (GIC) containing CHX could be an alternative in ART procedures with the objective of promoting an additional antimicrobial effect, which is interesting for children with high counts of MS during the initial phase of adaptation to dental treatment. How to cite this article: da Silva ME, de Sena MD, Colombo NH, et al. Short-term Clinical and Microbiological Performance of Resin-modified Glass Ionomer Cement Containing Chlorhexidine for Atraumatic Restorative Treatment. Int J Clin Pediatr Dent 2023;16(S-1):S27-S32.

A systematic review on the efficacy of CBT interventions for the mental and sexual health of survivors of prostate cancer.

INTRODUCTION: Prostate cancer (PCa) is the second-most frequently diagnosed oncologic condition among biological men, affecting physical and psychological well-being, as well as sexual health and quality of life. Prior research has shown that cognitive-behavioral therapy (CBT) can be effective in addressing a range of psychological and sexual problems but also in improving the sexual and mental health of survivors of PCa. OBJECTIVES: This systematic review aimed to methodically research and summarize results concerning the efficacy of CBT in the mental and sexual health of survivors of PCa. METHODS: A systematic search was carried out via electronic databases until August 2022 (EBSCO, MEDLINE, Cochrane Library, and Web of Science). By combining specific search words and following the PRISMA checklist, we identified 15 eligible articles among 8616 initial records. RESULTS: Four studies showed evidence of intervention efficacy for the improvement of sexual health, specifically for overall sexual function, erectile function, sexual desire, and sexual satisfaction. Eight studies found intervention efficacy for the improvement of mental health dimensions, such as psychological distress, depressive symptoms, anxiety, and quality of life. CONCLUSION: There is evidence that CBT interventions have the potential to effectively promote mental and sexual health in survivors of PCa, but further research comprising larger and more diverse populations is needed. Future studies should focus on understanding mechanisms of change through CBT interventions to ensure the mental and sexual health of survivors of PCa.

Rabbit derived VL single-domains as promising scaffolds to generate antibody-drug conjugates.

Antibody-drug conjugates (ADCs) are among the fastest-growing classes of therapeutics in oncology. Although ADCs are in the spotlight, they still present significant engineering challenges. Therefore, there is an urgent need to develop more stable and effective ADCs. Most rabbit light chains have an extra disulfide bridge, that links the variable and constant domains, between Cys80 and Cys171, which is not found in the human or mouse. Thus, to develop a new generation of ADCs, we explored the potential of rabbit-derived VL-single-domain antibody scaffolds (sdAbs) to selectively conjugate a payload to Cys80. Hence, a rabbit sdAb library directed towards canine non-Hodgkin lymphoma (cNHL) was subjected to in vitro and in vivo phage display. This allowed the identification of several highly specific VL-sdAbs, including C5, which specifically target cNHL cells in vitro and present promising in vivo tumor uptake. C5 was selected for SN-38 site-selective payload conjugation through its exposed free Cys80 to generate a stable and homogenous C5-DAB-SN-38. C5-DAB-SN-38 exhibited potent cytotoxicity activity against cNHL cells while inhibiting DNA-TopoI activity. Overall, our strategy validates a platform to develop a novel class of ADCs that combines the benefits of rabbit VL-sdAb scaffolds and the canine lymphoma model as a powerful framework for clinically translation of novel therapeutics for cancer.

Automatic attention to sexual stimuli: exploring the role of neuroticism and sexual excitation/inhibition through event-related potentials.

BACKGROUND: Previous findings have shown that neuroticism is a higher-order vulnerability factor in the development and maintenance of sexual dysfunctions and can have an impact on the attentional processing of sexual stimuli; however, the influence of psychosexual dimensions on the early automatic phases of such cognitive processes has not been established yet. AIM: To examine the mediating role of sexual inhibition/excitation propensity in the relationship between neuroticism and automatic attention to visual erotica and to identify the neuroelectric correlates of such a process. METHODS: We analyzed the answers provided by 58 individuals on the Neuroticism subscale of the NEO Personality Inventory-Revised and the Sexual Inhibition/Excitation Scales. Event-related potentials (ERPs) were recorded during a modified oddball paradigm containing romantic and sexually explicit pictures. Parallel mediations were performed to simultaneously test the mediating role of sexual inhibition/excitation in the relationship between neuroticism and each ERP. OUTCOMES: Three early attention ERP components (P1, P2, and N2) were assessed. RESULTS: Findings revealed an indirect effect of neuroticism on automatic attention, via sexual inhibition due to threat of performance failure (SIS1), for romantic and sexually explicit stimuli. This effect was significant only for component N2, which showed increased amplitudes and earlier latencies in participants with high SIS1. CLINICAL IMPLICATIONS: Sexual stimuli, due to their emotional valence and arousal potential, might be perceived as virtually threatening by individuals with neuroticism, who may benefit from strategies that decrease hyperarousal and sympathetic activation. STRENGTHS AND LIMITATIONS: This was one of the first studies to analyze neuroelectric activity associated with automatic attention toward sexual stimuli in relation to personality and sexual excitation/inhibition propensity. Nevertheless, the limited number of participants demands caution in generalizing the results. CONCLUSION: These results provide a better understanding of the relationship between personality and sexual cognition and open new avenues of research in relation to other automatic cognitive phenomena related to human sexual behavior.

Occurrence of MDR1 1-delta mutation in herding dog breeds in Portugal.

The impact of drug transporters in veterinary medicine has been recognized in recent years. One of the most well-characterized is the product of the MDR1 gene, P-gp. A 4-bp deletion in the MDR1 gene known since 2001 has been described to affect herding dog breeds. Since many used drugs in veterinary medicine are substrates for P-gp, including the macrocyclic lactones, such as avermectins, this 4-bp deletion causes a pathological condition known as "ivermectin toxicosis." For this reason, it is important to determine the animal status concerning this mutation. In Portugal, the information of the occurrence of this mutation in our breeds is limited. The aim of the present study was to determine the occurrence of this mutation and evaluate its association with Portuguese and non-Portuguese dog breeds in Portugal. To achieve this, a total of 105 animals were studied for the presence of the MDR1 4-bp deletion, 23 of which were from Barbado da Terceira, 10 from Cão da Serra d'Aires, 55 belonging to breeds known to carry the mutation (Australian Shepperd, Border Collie and others) and 17 to other breeds (Labrador Retriever, Jack Russel, and others). Despite the small sample size, we observed the presence of the MDR1 1-delta mutation in previously described breeds and identified this mutation in Barbado da Terceira breed for the first time.

Characterization of the canine CD20 as a therapeutic target for comparative passive immunotherapy.

Anti-CD20 therapies have revolutionized the treatment of B-cell malignancies. Despite these advances, relapsed and refractory disease remains a major treatment challenge. The optimization of CD20-targeted immunotherapies is considered a promising strategy to improve current therapies. However, research has been limited by the scarcity of preclinical models that recapitulate the complex interaction between the immune system and cancers. The addition of the canine lymphoma (cNHL) model in the development of anti-CD20 therapies may provide a clinically relevant approach for the translation of improved immunotherapies. Still, an anti-CD20 therapy for cNHL has not been established stressing the need of a comprehensive target characterization. Herein, we performed an in-depth characterization on canine CD20 mRNA transcript and protein expression in a cNHL biobank and demonstrated a canine CD20 overexpression in B-cell lymphoma samples. Moreover, CD20 gene sequencing analysis identified six amino acid differences in patient samples (C77Y, L147F, I159M, L198V, A201T and G273E). Finally, we reported the use of a novel strategy for the generation of anti-CD20 mAbs, with human and canine cross-reactivity, by exploring our rabbit derived single-domain antibody platform. Overall, these results support the rationale of using CD20 as a target for veterinary settings and the development of novel therapeutics and immunodiagnostics.

Diazaborines Are a Versatile Platform to Develop ROS-Responsive Antibody Drug Conjugates*.

Antibody-drug conjugates (ADCs) are a new class of therapeutics that combine the lethality of potent cytotoxic drugs with the targeting ability of antibodies to selectively deliver drugs to cancer cells. In this study we show for the first time the synthesis of a reactive-oxygen-species (ROS)-responsive ADC (VL-DAB31-SN-38) that is highly selective and cytotoxic to B-cell lymphoma (CLBL-1 cell line, IC50 value of 54.1 nM). The synthesis of this ADC was possible due to the discovery that diazaborines (DABs) are a very effective ROS-responsive unit that are also very stable in buffer and in plasma. DFT calculations performed on this system revealed a favorable energetic profile (ΔGR=-74.3 kcal mol-1 ) similar to the oxidation mechanism of aromatic boronic acids. DABs' very fast formation rate and modularity enabled the construction of different ROS-responsive linkers featuring self-immolative modules, bioorthogonal functions, and bioconjugation handles. These structures were used in the site-selective functionalization of a VL antibody domain and in the construction of the homogeneous ADC.

Highly Specific Blood-Brain Barrier Transmigrating Single-Domain Antibodies Selected by an In Vivo Phage Display Screening.

A major bottleneck in the successful development of central nervous system (CNS) drugs is the discovery and design of molecules that can cross the blood-brain barrier (BBB). Nano-delivery strategies are a promising approach that take advantage of natural portals of entry into the brain such as monoclonal antibodies (mAbs) targeting endogenous BBB receptors. However, the main selected mAbs rely on targeting broadly expressed receptors, such as the transferrin and insulin receptors, and in selection processes that do not fully mimic the native receptor conformation, leading to mistargeting and a low fraction of the administered dose effectively reaching the brain. Thus, there is an urgent need to identify new BBB receptors and explore novel antibody selection approaches that can allow a more selective delivery into the brain. Considering that in vitro models fail to completely mimic brain structure complexity, we explored an in vivo cell immunization approach to construct a rabbit derived single-domain antibody (sdAb) library towards BBB endothelial cell receptors. The sdAb antibody library was used in an in vivo phage display screening as a functional selection of novel BBB targeting antibodies. Following three rounds of selections, next generation sequencing analysis, in vitro brain endothelial barrier (BEB) model screenings and in vivo biodistribution studies, five potential sdAbs were identified, three of which reaching >0.6% ID/g in the brain. To validate the brain drug delivery proof-of-concept, the most promising sdAb, namely RG3, was conjugated at the surface of liposomes encapsulated with a model drug, the pan-histone deacetylase inhibitor panobinostat (PAN). The translocation efficiency and activity of the conjugate liposome was determined in a dual functional in vitro BEB-glioblastoma model. The RG3 conjugated PAN liposomes enabled an efficient BEB translocation and presented a potent antitumoral activity against LN229 glioblastoma cells without influencing BEB integrity. In conclusion, our in vivo screening approach allowed the selection of highly specific nano-antibody scaffolds with promising properties for brain targeting and drug delivery.

Chemokine-Directed Tumor Microenvironment Modulation in Cancer Immunotherapy.

Chemokines are a large family of small chemotactic cytokines that coordinates immune cell trafficking. In cancer, they have a pivotal role in the migration pattern of immune cells into the tumor, thereby shaping the tumor microenvironment immune profile, often towards a pro-tumorigenic state. Furthermore, chemokines can directly target non-immune cells in the tumor microenvironment, including cancer, stromal and vascular endothelial cells. As such, chemokines participate in several cancer development processes such as angiogenesis, metastasis, cancer cell proliferation, stemness and invasiveness, and are therefore key determinants of disease progression, with a strong influence in patient prognosis and response to therapy. Due to their multifaceted role in the tumor immune response and tumor biology, the chemokine network has emerged as a potential immunotherapy target. Under the present review, we provide a general overview of chemokine effects on several tumoral processes, as well as a description of the currently available chemokine-directed therapies, highlighting their potential both as monotherapy or in combination with standard chemotherapy or other immunotherapies. Finally, we discuss the most critical challenges and prospects of developing targeted chemokines as therapeutic options.

Immunotherapeutic Strategies for Canine Lymphoma: Changing the Odds Against Non-Hodgkin Lymphoma.

The new era of immune-oncology has brought complexities and challenges that emphasize the need to identify new strategies and models to develop successful and cost-effective therapies. The inclusion of a canine model in the drug development of cancer immunotherapies is being widely recognized as a valid solution to overcome several hurdles associated with conventional preclinical models. Driven by the success of immunotherapies in the treatment of human non-Hodgkin lymphoma (NHL) and by the remarkable similarities of canine NHL to its human counterpart, canine NHL has been one of the main focus of comparative research. Under the present review, we summarize a general overview of the challenges and prospects of today's cancer immunotherapies and the role that comparative medicine might play in solving the limitations brought by this rapidly expanding field. The state of art of both human and canine NHL and the rationale behind the use of the canine model to bridge the translational gap between murine preclinical studies and human clinical trials are addressed. Finally, a review of currently available immunotherapies for canine NHL is described, highlighting the potential of these therapeutic options.

Liposomes as a Nanoplatform to Improve the Delivery of Antibiotics into Staphylococcus aureus Biofilms.

Staphylococcus aureus biofilm-associated infections are a major public health concern. Current therapies are hampered by reduced penetration of antibiotics through biofilm and low accumulation levels at infected sites, requiring prolonged usage. To overcome these, repurposing antibiotics in combination with nanotechnological platforms is one of the most appealing fast-track and cost-effective approaches. In the present work, we assessed the potential therapeutic benefit of three antibiotics, vancomycin, levofloxacin and rifabutin (RFB), through their incorporation in liposomes. Free RFB displayed the utmost antibacterial effect with MIC and MBIC50 below 0.006 µg/mL towards a methicillin susceptible S. aureus (MSSA). RFB was selected for further in vitro studies and the influence of different lipid compositions on bacterial biofilm interactions was evaluated. Although positively charged RFB liposomes displayed the highest interaction with MSSA biofilms, RFB incorporated in negatively charged liposomes displayed lower MBIC50 values in comparison to the antibiotic in the free form. Preliminary safety assessment on all RFB formulations towards osteoblast and fibroblast cell lines demonstrated that a reduction on cell viability was only observed for the positively charged liposomes. Overall, negatively charged RFB liposomes are a promising approach against biofilm S. aureus infections and further in vivo studies should be performed.

Liposomes as Antibiotic Delivery Systems: A Promising Nanotechnological Strategy against Antimicrobial Resistance.

Antimicrobial drugs are key tools to prevent and treat bacterial infections. Despite the early success of antibiotics, the current treatment of bacterial infections faces serious challenges due to the emergence and spread of resistant bacteria. Moreover, the decline of research and private investment in new antibiotics further aggravates this antibiotic crisis era. Overcoming the complexity of antimicrobial resistance must go beyond the search of new classes of antibiotics and include the development of alternative solutions. The evolution of nanomedicine has allowed the design of new drug delivery systems with improved therapeutic index for the incorporated compounds. One of the most promising strategies is their association to lipid-based delivery (nano)systems. A drug's encapsulation in liposomes has been demonstrated to increase its accumulation at the infection site, minimizing drug toxicity and protecting the antibiotic from peripheral degradation. In addition, liposomes may be designed to fuse with bacterial cells, holding the potential to overcome antimicrobial resistance and biofilm formation and constituting a promising solution for the treatment of potential fatal multidrug-resistant bacterial infections, such as methicillin resistant Staphylococcus aureus. In this review, we aim to address the applicability of antibiotic encapsulated liposomes as an effective therapeutic strategy for bacterial infections.

Loss of α-gal during primate evolution enhanced antibody-effector function and resistance to bacterial sepsis.

Most mammals express a functional GGTA1 gene encoding the N-acetyllactosaminide α-1,3-galactosyltransferase enzyme, which synthesizes Gal-α1-3Gal-ß1-4GlcNAc (α-gal) and are thus tolerant to this self-expressed glycan. Old World primates including humans, however, carry loss-of-function mutations in GGTA1 and lack α-gal. Presumably, fixation of such mutations was propelled by natural selection, favoring the emergence of α-gal-specific immunity, conferring resistance to α-gal-expressing pathogens. Here, we show that loss of Ggta1 function in mice enhances resistance to bacterial sepsis, irrespectively of α-Gal-specific immunity. Rather, the absence of α-gal from IgG-associated glycans increases IgG effector function via a mechanism associated with enhanced IgG-Fc gamma receptor (FcγR) binding. The ensuing survival advantage against sepsis comes alongside a cost of accelerated reproductive senescence in Ggta1-deleted mice. Mathematical modeling of this trade-off suggests that high exposure to virulent pathogens exerts sufficient selective pressure to fix GGTA1 loss-of-function mutations, as likely occurred during the evolution of primates toward humans.

Lipid-based nanosystems for targeting bone implant-associated infections: current approaches and future endeavors.

Bone infections caused by Staphylococcus aureus are a major concern in medical care, particularly when associated with orthopedic-implant devices. The ability of the bacteria to form biofilms and their capacity to invade and persist within osteoblasts turn the infection eradication into a huge challenge. The reduction of antibiotic penetration through bacterial biofilms associated with the presence of persistent cells, ability to survive in the host, and high tolerance to antibiotics are some of the reasons for the difficult treatment of these infections. Effective therapeutic approaches are urgently needed. In this sense, lipid-based nanosystems, such as liposomes, have been investigated as an innovative and alternative strategy for the treatment of implant-associated S. aureus infections, due to their preferential accumulation at infected sites and interaction with S. aureus. This review highlights the recent advances on antibiotic-loaded liposome formulations both in vitro and in vivo and how the interaction with S. aureus biofilms may be improved by modulating the liposomal external surface. Graphical Abstract.

Dominance of vaccine serotypes in pediatric invasive pneumococcal infections in Portugal (2012-2015).

We evaluated the impact of continued 13-valent pneumococcal conjugate vaccine (PCV13) use in the private market (uptake of 61%) in pediatric invasive pneumococcal disease (pIPD) in Portugal (2012-2015). The most frequently detected serotypes were: 3 (n = 32, 13.8%), 14 (n = 23, 9.9%), 1 (n = 23, 9.9%), 7F (n = 15, 6.4%), 19A (n = 13, 5.6%), 6B and 15B/C (both n = 12, 5.2%), and 24F, 10A and 12B (all with n = 10, 4.3%). Taken together, non-PCV13 serotypes were responsible for 42.2% of pIPD with a known serotype. The use of PCR to detect and serotype pneumococci in both pleural and cerebrospinal fluid samples contributed to 18.1% (n = 47) of all pIPD. Serotype 3 was mostly detected by PCR (n = 21/32, 65.6%) and resulted from a relevant number of vaccine failures. The incidence of pIPD varied in the different age groups but without a clear trend. There were no obvious declines of the incidence of pIPD due to serotypes included in any of the PCVs, and PCV13 serotypes still accounted for the majority of pIPD (57.8%). Our study indicates that a higher vaccination uptake may be necessary to realize the full benefits of PCVs, even after 15 years of moderate use, and highlights the importance of using molecular methods in pIPD surveillance, since these can lead to substantially increased case ascertainment and identification of particular serotypes as causes of pIPD.

Establishment of a bioluminescent canine B-cell lymphoma xenograft model for monitoring tumor progression and treatment response in preclinical studies.

Canine diffuse large B-cell lymphoma (DLBCL) is one of the most common cancers in dogs which shares remarkable similarities with its human counterpart, making the dog an excellent model for the investigation of novel therapeutic agents. However, the integration of canine lymphoma in comparative studies has been limited due in part to the lack of suitable xenograft mouse models for preclinical studies. To overcome these limitations, we established and characterized a localized subcutaneous bioluminescent canine DLBCL xenograft mouse model. The canine CLBL-1 cell line stably expressing the luciferase and green fluorescent protein reporters was generated and used to establish the xenograft tumor model. A pilot study was first conducted with three different cell densities (0.1×10(6), 0.5×10(6) and 1×10(6) cells) in SCID mice. All mice presented homogeneous tumor induction within eight days after subcutaneous injection, with a 100% engraftment efficiency and no significant differences were observed among groups. The tumors were highly aggressive and localized at the site of inoculation and reproduced histological features and immunophenotype consistent with canine DLBCL. Importantly, xenograft tumors were detected and quantified by bioluminescent imaging. To assess response to therapy, a therapeutic study with a histone deacetylase inhibitor, panobinostat, was performed. The results demonstrated that panobinostat (20 mg/kg) efficiently inhibited tumor growth and that bioluminescent imaging allowed the monitorization and quantification of tumor response to therapy. In summary, this study provides a bioluminescence canine DLBCL model that offers high engraftment efficiency, preservation of tumor features, and noninvasive monitoring of tumor progression, validating the model as a promising preclinical tool for both veterinary and human medicine.

The histone deacetylase inhibitor panobinostat is a potent antitumor agent in canine diffuse large B-cell lymphoma.

Non-Hodgkin lymphoma (NHL) is one of the most common causes of cancer-related death in the United States and Europe. Although the outcome of NHL patients has improved over the last years with current therapies, the rate of mortality is still high. A plethora of new drugs is entering clinical development for NHL treatment; however, the approval of new treatments remains low due in part to the paucity of clinically relevant models for validation. Canine lymphoma shares remarkable similarities with its human counterpart, making the dog an excellent animal model to explore novel therapeutic molecules and approaches. Histone deacetylase inhibitors (HDACis) have emerged as a powerful new class of anti-cancer drugs for human therapy. To investigate HDACi antitumor properties on canine diffuse large B-cell lymphoma, a panel of seven HDACi compounds (CI-994, panobinostat, SBHA, SAHA, scriptaid, trichostatin A and tubacin) was screened on CLBL-1 canine B-cell lymphoma cell line. Our results demonstrated that all HDACis tested exhibited dose-dependent inhibitory effects on proliferation of CLBL-1 cells, while promoting increased H3 histone acetylation. Amongst all HDACis studied, panobinostat proved to be the most promising compound and was selected for further in vitro and in vivo evaluation. Panobinostat cytotoxicity was linked to H3 histone and α-tubulin acetylation, and to apoptosis induction. Importantly, panobinostat efficiently inhibited CLBL-1 xenograft tumor growth, and strongly induced acetylation of H3 histone and apoptosis in vivo. In conclusion, these results provide new data validating HDACis and, especially, panobinostat as a novel anti-cancer therapy for veterinary applications, while contributing to comparative oncology.