B7-H3-Targeting Chimeric Antigen Receptors Epstein Barr Virus Specific T Cells Provides a Tumor Agnostic Off-The-Shelf Therapy Against B7-H3 Positive Solid Tumors.

Encouraged by our observations of pronounced B7-H3 protein over-expression in many human solid tumors compared to healthy tissues, we focused on targeting B7-H3 with CAR T cells. We utilized a nanobody as the t B7-H3 targeting domain in our CAR construct to circumvent the stability issues associated with scFv-based domains. In efforts to expand patient access to CAR T cell therapy, we engineered our nanobody-based CAR into human Epstein-Barr Virus Specific T Cells (EBVSTs), offering a readily available off-the-shelf treatment. B7H3.CAR-armored EBVSTs demonstrated potent in vitro and in vivo activities against multiple B7-H3-positive human tumor cell lines and patient-derived xenograft models. Murine T cells expressing a murine equivalent of our B7H3.CAR exhibited no life-threatening toxicities in immunocompetent mice bearing syngeneic tumors. Further in vitro evaluation revealed that while human T, B and NK cells were unaffected by B7H3.CAR EBVSTs, monocytes were targeted due to upregulation of B7-H3. Such targeting of myeloid cells, which are key mediators of cytokine release syndrome (CRS), contributed to a low incidence of CRS in humanized mice after B7H3.CAR EBVST treatment. Notably, we showed that B7H3.CAR EBVSTs can target B7-H3 expressing myeloid-derived suppressor cells (MDSCs), thereby mitigating MDSC-driven immune suppression. In summary, our data demonstrate that our nanobody-based B7H3.CAR EBVSTs are effective as an off-the-shelf therapy for B7-H3 positive solid tumors. These cells also offer an avenue to modulate the immunosuppressive tumor microenvironment, highlighting their promising clinical potential in targeting solid tumors.

Searching for common patterns in parasite ecology: species and host contributions to beta-diversity in helminths of South African ungulates and fleas of South American rodents.

We searched for common patterns in parasite ecology by investigating species and host contributions to the beta-diversity of infracommunities (=assemblages of parasites harboured by a host individual) in helminths of three species of South African ungulates and fleas of 11 species of South American rodents, assuming that a comparison of patterns in distinctly different parasites and hosts would allow us to judge the generality or, at least, commonness of these patterns. We used data on species' composition and numbers of parasites and asked whether (i) parasite species' attributes (life cycle, transmission mode, and host specificity in helminths; possession of sclerotized combs, microhabitat preference, and host specificity in fleas) or their population structure (mean abundance and/or prevalence) and (ii) host characteristics (sex and age) affect parasite and host species' contributions to parasite beta-diversity (SCBD and HCBD, respectively). We found that parasite species' morphological and ecological attributes were mostly not associated with their SCBD. In contrast, parasite SCBD, in both ungulates and rodents, significantly increased with either parasite mean abundance or prevalence or both. The effect of host characteristics on HCBD was detected in a few hosts only. In general, parasite infracommunities' beta-diversity appeared to be driven by variation in parasite species rather than the uniqueness of the assemblages harboured by individual hosts. We conclude that some ecological patterns (such as the relationships between SCBD and parasite abundance/prevalence) appear to be common and do not differ between different host-parasite associations in different geographic regions, whereas other patterns (the relationships between SCBD and parasite species' attributes) are contingent and depend on parasite and host identities.

Nestedness and beta diversity of gastrointestinal helminth communities in common warthogs, Phacochoerus africanus (Suidae), at 2 localities in South Africa.

Few studies have investigated the ecological interactions between wild species of Suidae and their parasites, leaving our knowledge concerning this host­parasite system fragmented. In the present study, we applied network studies to analyse community nestedness in helminth assemblages of common warthogs, Phacochoerus africanus (Gmelin) (Suidae). Helminth data were compiled from 95 warthogs, including young and adult males and females, from 2 different conservation areas in Mpumalanga and Limpopo Provinces, South Africa, collected monthly over a period of 1 year each. The aim was to study the effect of host sex, age and season of sampling on the structure of helminth infracommunities harboured by the warthogs and to search for non-random structural patterns in the warthog­helminth interaction networks. Furthermore, we investigated the influence of a warthog's age, sex and season of sampling on beta diversity and dark diversity of their helminth infracommunities. Lastly, we asked whether the effects of host sex, age and sampling season on helminth communities differed between the 2 localities. We found that helminth communities of warthogs were nested and host­parasite interactions were influenced by all 3 factors as well as combinations thereof. However, the resulting patterns differed at the 2 localities, indicating that local environmental processes are important drivers of community structure.

Sympatry of Amblyomma eburneum and Amblyomma variegatum on African buffaloes and prevalence of pathogens in ticks.

The Amblyomma genus is represented on the African continent by 24 species, out of which 17 are known to occur in different ecological niches of southern Africa. Amblyomma, known for their aggressive hunting behaviour and aptitude as pathogen vectors, are of main concern to travellers, mainly in rural and conservation areas of Africa. In this study, we highlight the overlapping distribution of Amblyomma eburneum and Amblyomma variegatum found on African buffaloes (Syncerus caffer) at Coutada 11, Central Mozambique. In total, 1,039 Amblyomma ticks were collected and morphologically identified using taxonomic keys, and genomic DNA was extracted. They were subjected to reverse line blotting for pathogen identification followed by molecular analysis (COI sequencing) of both tick species. Pathogens such as Ehrlichia ruminantium, Anaplasma centrale, Theileria sp., Babesia sp. and Rickettsia africae were detected, of which R. africae is zoonotic. Ehrlichia ruminantium, R. africae, Theileria mutans and Theileria velifera are well-established pathogens transmitted by Amblyomma ticks; however, Anaplasma spp. and Babesia spp. are not, suggesting residual parasite DNA in the bloodmeal. Little is mentioned in the literature about A. eburneum, including its role as a vector and reservoir for pathogens. In Mozambique A. eburneum is currently restricted to wildlife but the spread of the tick may be observed given the climate change that is occurring. The infection rates for the pathogens in both Amblyomma tick species were lower than expected, but this may be due to the low host density in the forest niche and the innate immunity of these hosts. With the propensity of ticks of the Amblyomma genus to form parapatric distributions, the mechanisms that allows for the overlapping distribution of these two Amblyomma species while maintaining tick species identity is of great interest.

Ectoparasites associated with the Bushveld gerbil (Gerbilliscus leucogaster) and the role of the host and habitat in shaping ectoparasite diversity and infestations.

Rodents are known hosts for various ectoparasite taxa such as fleas, lice, ticks and mites. South Africa is recognized for its animal diversity, yet little is published about the parasite diversity associated with wild rodent species. By focusing on a wildlife-human/domestic animal interface, the study aims to record ectoparasite diversity and levels of infestations of the Bushveld gerbil, Gerbilliscus leucogaster, and to establish the relationship between ectoparasite infestation parameters and host- and habitat factors. Rodents (n = 127) were trapped in 2 habitat types (natural and agricultural) during 2014­2020. More than 6500 individuals of 32 epifaunistic species represented by 21 genera and belonging to 5 taxonomic groups (fleas, sucking lice, ticks, mesostigmatan mites and trombiculid mites) were collected. Mesostigmatan mites and lice were the most abundant and fleas and mesostigmatan mites the most prevalent groups. Flea and mesostigmatan mite numbers and mesostigmatan mite species richness was significantly higher on reproductively active male than female rodents. Only ticks were significantly associated with habitat type, with significantly higher tick numbers and more tick species on rodents in the natural compared to the agricultural habitat. We conclude that the level of infestation by ectoparasites closely associated with the host (fleas and mites) was affected by host-associated factors, while infestation by ectoparasite that spend most of their life in the external environment (ticks) was affected by habitat type.

Impact of host sex and age on the diversity of endoparasites and structure of individual-based host-parasite networks in nyalas (Tragelaphus angasii Angas) from three game reserves in KwaZulu-Natal province, South Africa.

In recent years, numerous studies have examined the effect of host sex and age on the structure of parasite communities in several host taxa under various environmental conditions and in different geographic regions. However, the influence of such factors on the structure of host-parasite networks has received less attention, and remarkably few studies have been carried out on large terrestrial mammals. In this study, we investigated the effects of host age and sex on the parasite infra- and component communities of nyalas (Tragelaphus angasii) and on the structure of individual-based nyala-endoparasite networks. We also aimed to evaluate to what extent these effects vary spatially and if they are mediated by conservation management. Based on a data set of internal macroparasites of 74 nyalas from three game reserves in KwaZulu-Natal province, we found that host age strongly influenced parasite community structure as well as the structure of parasite-nyala networks, whereas host sex played a minor role. However, the effects of both host sex and age were mediated by environmental conditions and thus led to different patterns at the three localities. Our findings highlight that host-parasite communities from different localities should not be pooled when conducting host-parasite network and community studies as this may bias results and mask patterns that are typical for a given locality.

First-in-human trial exploring safety, antitumor activity, and pharmacokinetics of Sym013, a recombinant pan-HER antibody mixture, in advanced epithelial malignancies.

PURPOSE: Sym013 contains six humanized monoclonal antibodies that bind to non-overlapping epitopes on three human epidermal growth factor receptors (HER1-3). Preclinical studies suggested Sym013 strongly suppresses growth of multiple epithelial tumors. This is a first-in-human study exploring safety and efficacy of Sym013 in patients with advanced epithelial malignancies. METHODS: Dose escalation used single-patient cohorts until the stopping rule was met, followed by 3 + 3 design. Dose levels planned were: 1, 2, 4, 6, 9, 12, 15, and 18 mg/kg. Treatment cycles were 28 days with imaging every eight weeks. Serum samples were collected at multiple time points for assessment of pharmacokinetics and development of anti-drug antibodies. RESULTS: Thirty-two patients were enrolled with multiple solid tumors, most common being colorectal cancer (CRC; 10/32, 31%). Due to mucositis, rash, and diarrhea at 4 mg/kg once-weekly, dosing was changed to biweekly (Q2W). Mandatory prophylaxis was added due to Grade 3 infusion-related reaction and oral mucositis at 9 mg/kg Q2W. The 15 mg/kg Q2W cohort was enrolling when the study was terminated for business reasons. Most common adverse events were skin (81%) and gastrointestinal (75%) disorders, including dermatitis/rash, stomatitis, and diarrhea. One patient with CRC achieved a partial response; 12 patients with varied malignancies had stable disease. CONCLUSION: During the conduct of the study, management of frequent infusion-related reactions, skin toxicities, and mucosal disorders, which are indicative of HER inhibition, necessitated multiple protocol amendments. The investigators, in concert with the Sponsor, agreed that achieving a tolerated regimen with acceptable target saturation was unlikely. TRIAL REGISTRY: www. CLINICALTRIALS: gov ; NCT02906670 (September 20, 2016).

Similarity in ixodid tick communities harboured by wildlife and livestock in the Albany Thicket Biome of South Africa.

South Africa has a diverse fauna of ixodid tick species, several of which are of medical or veterinary importance. Elucidation of which host or environmental characteristics determine the composition and abundance of tick assemblages is, therefore, highly relevant to disease management and wildlife conservation efforts. Here, we analysed the similarity in ixodid tick assemblages of three wildlife and three livestock species in a natural (Great Fish River Reserve) and anthropogenic (an adjacent farm) habitat in South Africa. We compared tick infracommunities of three wild host species between the reserve and the farm; of three wild host species within the reserve and of wild and livestock species on the farm (considering body size). Hosts examined in this study harboured the adults and immature stages of 11 tick species belonging to five ixodid genera. Notably, several tick species of South African wildlife have successfully made the switch to livestock and thus both wild hosts and livestock now contribute to the pool of maintenance hosts for these ticks as well as their associated pathogens. This is an important consideration when translocating wildlife or livestock as part of farming or conservation activities.

Paediatric Strategy Forum for medicinal product development of chimeric antigen receptor T-cells in children and adolescents with cancer: ACCELERATE in collaboration with the European Medicines Agency with participation of the Food and Drug Administration.

The seventh multi-stakeholder Paediatric Strategy Forum focused on chimeric antigen receptor (CAR) T-cells for children and adolescents with cancer. The development of CAR T-cells for patients with haematological malignancies, especially B-cell precursor acute lymphoblastic leukaemia (BCP-ALL), has been spectacular. However, currently, there are scientific, clinical and logistical challenges for use of CAR T-cells in BCP-ALL and other paediatric malignancies, particularly in acute myeloid leukaemia (AML), lymphomas and solid tumours. The aims of the Forum were to summarise the current landscape of CAR T-cell therapy development in paediatrics, too identify current challenges and future directions, with consideration of other immune effector modalities and ascertain the best strategies to accelerate their development and availability to children. Although the effect is of limited duration in about half of the patients, anti-CD19 CAR T-cells produce high response rates in relapsed/refractory BCP-ALL and this has highlighted previously unknown mechanisms of relapse. CAR T-cell treatment as first- or second-line therapy could also potentially benefit patients whose disease has high-risk features associated with relapse and failure of conventional therapies. Identifying patients with very early and early relapse in whom CAR T-cell therapy may replace haematopoietic stem cell transplantation and be definitive therapy versus those in whom it provides a more effective bridge to haematopoietic stem cell transplantation is a very high priority. Development of approaches to improve persistence, either by improving T cell fitness or using more humanised/fully humanised products and co-targeting of multiple antigens to prevent antigen escape, could potentially further optimise therapy. Many differences exist between paediatric B-cell non-Hodgkin lymphomas (B-NHL) and BCP-ALL. In view of the very small patient numbers with relapsed lymphoma, careful prioritisation is needed to evaluate CAR T-cells in children with Burkitt lymphoma, primary mediastinal B cell lymphoma and other NHL subtypes. Combination trials of alternative targets to CD19 (CD20 or CD22) should also be explored as a priority to improve efficacy in this population. Development of CD30 CAR T-cell immunotherapy strategies in patients with relapsed/refractory Hodgkin lymphoma will likely be most efficiently accomplished by joint paediatric and adult trials. CAR T-cell approaches are early in development for AML and T-ALL, given the unique challenges of successful immunotherapy actualisation in these diseases. At this time, CD33 and CD123 appear to be the most universal targets in AML and CD7 in T-ALL. The results of ongoing or planned first-in-human studies are required to facilitate further understanding. There are promising early results in solid tumours, particularly with GD2 targeting cell therapies in neuroblastoma and central nervous system gliomas that represent significant unmet clinical needs. Further understanding of biology is critical to success. The comparative benefits of autologous versus allogeneic CAR T-cells, T-cells engineered with T cell receptors T-cells engineered with T cell receptor fusion constructs, CAR Natural Killer (NK)-cell products, bispecific T-cell engager antibodies and antibody-drug conjugates require evaluation in paediatric malignancies. Early and proactive academia and multi-company engagement are mandatory to advance cellular immunotherapies in paediatric oncology. Regulatory advice should be sought very early in the design and preparation of clinical trials of innovative medicines, for which regulatory approval may ultimately be sought. Aligning strategic, scientific, regulatory, health technology and funding requirements from the inception of a clinical trial is especially important as these are very expensive therapies. The model for drug development for cell therapy in paediatric oncology could also involve a 'later stage handoff' to industry after early development in academic hands. Finally, and very importantly, strategies must evolve to ensure appropriate ease of access for children who need and could potentially benefit from these therapies.

Adaptive radiation and speciation in Rhipicephalus ticks: A medley of novel hosts, nested predator-prey food webs, off-host periods and dispersal along temperature variation gradients.

Rhipicephalus are a species-diverse genus of ticks, mainly distributed in the Afrotropics with some species in the Palearctic and Oriental regions. Current taxonomic consensus comprise nine informal species groups/lineages based on immature morphology. This work integrates biogeographic, ecological and molecular lines of evidence to better understand Rhipicephalus evolution. Phylogenetic analysis based on four genes (12S, 16S, 28S-D2 and COI) recovered five distinct clades with nine descendant clades that are generally congruent with current taxonomy, with some exceptions. Historical biogeography is inferred from molecular divergence times, ancestral distribution areas, host-use and climate niches of four phylogenetically significant bioclimatic variables (isothermality, annual, seasonal and diurnal temperature range). Novel hosts enabled host-linked dispersal events into new environments, and ticks exploited new hosts through nested predator-prey connections in food webs. Diversification was further induced by climate niche partitioning along gradients in temperature range during off-host periods. Ancestral climate niche estimates corroborated dispersal events by indicating hypothetical ancestors moved into environments with different annual and seasonal temperature ranges along latitudinal gradients. Host size for immature and adult life stages was important for dispersal and subsequent diversification rates. Clades that utilise large, mobile hosts (ungulates and carnivores) early in development have wider geographic ranges but slower diversification rates, and those utilising small, less mobile hosts (rodents, lagomorphs and afroinsectivores) early in development have smaller ranges but higher diversification rates. These findings suggest diversification is driven by a complex set of factors linked to both host-associations (host size, ranges and mobility) and climate niche partitioning along annual and seasonal temperature range gradients that vary with latitude. Moreover, competitive interactions can reinforce these processes and drive speciation. Off-host periods facilitate adaptive radiation by enabling host switches along nested predator-prey connections in food webs, but at the cost of environmental exposure that partitions niches among dispersing progenitors, disrupting geneflow and driving diversification. As such, the evolution and ecological niches of Rhipicephalus are characterised by trade-offs between on- and off-host periods, and these trade-offs interact with nested predator-prey connections in food webs, host-use at different life stages, as well as gradients in annual and seasonal temperature ranges to drive adaptive radiation and speciation.

Some gastrointestinal nematodes and ixodid ticks shared by several wildlife species in the Kruger National Park, South Africa.

Parasite surveys were conducted for 1­2 years in the Kruger National Park (KNP), South Africa on blue wildebeest, impalas, greater kudus, common warthogs and scrub hares. The host associations of some of the gastrointestinal nematode species infecting ≥60% of at least one of the five host species, were determined. These were Agriostomum gorgonis, Cooperia acutispiculum, Cooperia connochaeti, Cooperia hungi, Cooperia neitzi, Cooperioides hamiltoni, Gaigeria pachyscelis, Haemonchus bedfordi, Haemonchus krugeri, Haemonchus vegliai, Impalaia tuberculata, Longistrongylus sabie, Strongyloides papillosus, Trichostrongylus deflexus and Trichostrongylus thomasi. Although the prevalence of Trichostrongylus falculatus did not exceed 50% in any host species, it was present in all five hosts. Nematodes in the KNP range from those exhibiting strict host associations to generalists. Nematode-host associations may be determined by host feeding patterns and habitat use. Eight ixodid tick species were commonly collected from the same animals and in 2­3 year long surveys from plains zebras and helmeted guinea fowls: Amblyomma hebraeum, Amblyomma marmoreum, Hyalomma truncatum, Rhipicephalus appendiculatus, Rhipicephalus decoloratus, Rhipicephalus evertsi evertsi, Rhipicephalus simus and Rhipicephalus zambeziensis. Host specificity was less pronounced in ixodid tick species than in nematodes and the immature stages of five tick species infested all host species examined.

Deep neural networks identify signaling mechanisms of ErbB-family drug resistance from a continuous cell morphology space.

It is well known that the development of drug resistance in cancer cells can lead to changes in cell morphology. Here, we describe the use of deep neural networks to analyze this relationship, demonstrating that complex cell morphologies can encode states of signaling networks and unravel cellular mechanisms hidden to conventional approaches. We perform high-content screening of 17 cancer cell lines, generating more than 500 billion data points from ∼850 million cells. We analyze these data using a deep learning model, resulting in the identification of a continuous 27-dimension space describing all of the observed cell morphologies. From its morphology alone, we could thus predict whether a cell was resistant to ErbB-family drugs, with an accuracy of 74%, and predict the potential mechanism of resistance, subsequently validating the role of MET and insulin-like growth factor 1 receptor (IGF1R) as drivers of cetuximab resistance in in vitro models of lung and head/neck cancer.

Ticks and tick-borne pathogens infecting livestock and dogs in Tchicala-Tcholoanga, Huambo Province, Angola.

The diversity of ticks and tick-borne pathogens (TBPs) infesting domestic animals in Tchicala-Tcholoanga, Angola, in 2016 was investigated. Seventeen tick species were recorded, Amblyomma pomposum being the most abundant on cattle (40%), goats (38%) and sheep (35%); Rhipicephalus turanicus was the most abundant on dogs (46%). This study presents new records of Haemaphysalis paraleachi, R. compositus, R. kochi and R. sulcatus in Angola, the first georeferenced population of Ha. leachi in southern Africa and the second record of R. microplus in Angola. Using the reverse line blot (RLB) hybridisation assay, fifteen TBP species were detected in blood samples from cattle (n = 88), goats (n = 82), sheep (n = 85) and dogs (n = 85). F The most frequently detected species were Theileria velifera in cattle (78%), Theileria ovis in sheep (80%) and Babesia vogeli in dogs (35%). Species-specific quantitative PCR assays detected Babesia bigemina in 43% (35/80) of blood samples of cattle, while E. ruminantium was detected in 4% (3/70) of blood samples and in 7% of A. pomposum ticks. Anaplasma platys was detected from cattle (18%) and sheep (6%) during RLB analysis. These findings constitute pioneering research in Angola.

Colorectal cancer residual disease at maximal response to EGFR blockade displays a druggable Paneth cell-like phenotype.

Blockade of epidermal growth factor receptor (EGFR) causes tumor regression in some patients with metastatic colorectal cancer (mCRC). However, residual disease reservoirs typically remain even after maximal response to therapy, leading to relapse. Using patient-derived xenografts (PDXs), we observed that mCRC cells surviving EGFR inhibition exhibited gene expression patterns similar to those of a quiescent subpopulation of normal intestinal secretory precursors with Paneth cell characteristics. Compared with untreated tumors, these pseudodifferentiated tumor remnants had reduced expression of genes encoding EGFR-activating ligands, enhanced activity of human epidermal growth factor receptor 2 (HER2) and HER3, and persistent signaling along the phosphatidylinositol 3-kinase (PI3K) pathway. Clinically, properties of residual disease cells from the PDX models were detected in lingering tumors of responsive patients and in tumors of individuals who had experienced early recurrence. Mechanistically, residual tumor reprogramming after EGFR neutralization was mediated by inactivation of Yes-associated protein (YAP), a master regulator of intestinal epithelium recovery from injury. In preclinical trials, Pan-HER antibodies minimized residual disease, blunted PI3K signaling, and induced long-term tumor control after treatment discontinuation. We found that tolerance to EGFR inhibition is characterized by inactivation of an intrinsic lineage program that drives both regenerative signaling during intestinal repair and EGFR-dependent tumorigenesis. Thus, our results shed light on CRC lineage plasticity as an adaptive escape mechanism from EGFR-targeted therapy and suggest opportunities to preemptively target residual disease.

Simultaneous targeting of HER family pro-survival signaling with Pan-HER antibody mixture is highly effective in TNBC: a preclinical trial with PDXs.

BACKGROUND: The human epidermal growth factor receptor (HER) family, notably EGFR, is overexpressed in most triple-negative breast cancer (TNBC) cases and provides cancer cells with compensatory signals that greatly contribute to the survival and development of resistance in response to therapy. This study investigated the effects of Pan-HER (Symphogen, Ballerup, Denmark), a novel mixture of six monoclonal antibodies directed against members of the HER family EGFR, HER2, and HER3, in a preclinical trial of TNBC patient-derived xenografts (PDXs). METHODS: Fifteen low passage TNBC PDX tumor samples were transferred into the right mammary fat pad of mice for engraftment. When tumors reached an average size of 100-200 mm3, mice were randomized (n ≥ 6 per group) and treated following three 1-week cycles consisting of three times/week intraperitoneal (IP) injection of either formulation buffer (vehicle control) or Pan-HER (50 mg/kg). At the end of treatment, tumors were collected for Western blot, RNA, and immunohistochemistry analyses. RESULTS: All 15 TNBC PDXs were responsive to Pan-HER treatment, showing significant reductions in tumor growth consistent with Pan-HER-mediated tumor downmodulation of EGFR and HER3 protein levels and significantly decreased activation of associated HER family signaling pathways AKT and ERK. Tumor regression was observed in five of the models, which corresponded to those PDX tumor models with the highest level of HER family activation. CONCLUSIONS: The marked effect of Pan-HER in numerous HER family-dependent TNBC PDX models justifies further studies of Pan-HER in TNBC clinical trials as a potential therapeutic option.

Invasive cattle ticks in East Africa: morphological and molecular confirmation of the presence of Rhipicephalus microplus in south-eastern Uganda.

BACKGROUND: Rhipicephalus microplus, an invasive tick species of Asian origin and the main vector of Babesia species, is considered one of the most widespread ectoparasites of livestock. The tick has spread from its native habitats on translocated livestock to large parts of the tropical world, where it has replaced some of the local populations of Rhipicephalus decoloratus ticks. Although the tick was reported in Uganda 70 years ago, it has not been found in any subsequent surveys. This study was carried out to update the national tick species distribution on livestock in Uganda as a basis for tick and tick-borne disease control, with particular reference to R. microplus. METHODS: The study was carried out in Kadungulu, Serere district, south-eastern Uganda, which is dominated by small scale livestock producers. All the ticks collected from 240 cattle from six villages were identified microscopically. Five R. microplus specimens were further processed for phylogenetic analysis and species confirmation. RESULTS: The predominant tick species found on cattle was Rhipicephalus appendiculatus (86.9 %; n = 16,509). Other species found were Amblyomma variegatum (7.2 %; n = 1377), Rhipicephalus evertsi (2.3 %; n = 434) and R. microplus (3.6 %; n = 687). Phylogenetic analysis of the 12S rRNA, 16S rRNA and ITS2 gene sequences of R. microplus confirmed the morphological identification. CONCLUSIONS: It is concluded that R. microplus has replaced R. decoloratus in the sampled villages in Kadungulu sub-county, since the latter was not any longer found in this area. There is currently no livestock movement policy in force in Uganda, which could possibly limit the further spread of R. microplus ticks. Future surveys, but also retrospective surveys of museum specimens, will reveal the extent of distribution of R. microplus in Uganda and also for how long this tick has been present on livestock without being noticed.

CD4+ and CD8a+ PET imaging predicts response to novel PD-1 checkpoint inhibitor: studies of Sym021 in syngeneic mouse cancer models.

Predicting the outcome of immunotherapy is essential for efficient treatment. The recent clinical success of immunotherapy is increasingly changing the paradigm of cancer treatment. Accordingly, the development of immune-based agents is accelerating and the number of agents in the global immuno-oncology pipeline has grown 60-70% over the past year. However, despite remarkable clinical efficacy in some patients, only few achieve a lasting clinical response. Treatment failure can be attributed to poorly immunogenic tumors that do not attract tumor infiltrating lymphocytes (TILs). Therefore, we developed positron emission tomography (PET) radiotracers for non-invasive detection of CD4+ and CD8a+ TILs in syngeneic mouse tumor models for preclinical studies. Methods: Seven syngeneic mouse tumor models (B16F10, P815, CT26, MC38, Renca, 4T1, Sa1N) were quantified for CD4+ and CD8a+ TILs using flow cytometry and immunohistochemistry (IHC), as well as for tumor growth response to Sym021, a humanized PD-1 antibody cross-reactive with mouse PD-1. Radiotracers were generated from F(ab)'2 fragments of rat-anti-mouse CD4 and CD8a antibodies conjugated to the p-SCN-Bn-Desferrioxamine (SCN-Bn-DFO) chelator and radiolabeled with Zirconium-89 (89Zr-DFO-CD4/89Zr-DFO-CD8a). Tracers were optimized for in vivo PET/CT imaging in CT26 tumor-bearing mice and specificity was evaluated by depletion studies and isotype control imaging. 89Zr-DFO-CD4 and 89Zr-DFO-CD8a PET/CT imaging was conducted in the panel of syngeneic mouse models prior to immunotherapy with Sym021. Results: Syngeneic tumor models were characterized as "hot" or "cold" according to number of TILs determined by flow cytometry and IHC. 89Zr-DFO-CD4 and 89Zr-DFO-CD8a were successfully generated with a radiochemical purity >99% and immunoreactivity >85%. The optimal imaging time-point was 24 hours post-injection of ~1 MBq tracer with 30 µg non-labeled co-dose. Reduced tumor and spleen uptake of 89Zr-DFO-CD8a was observed in CD8a+ depleted mice and the uptake was comparable with that of isotype control (89Zr-DFO-IgG2b) confirming specificity. PET imaging in syngeneic tumor models revealed a varying maximum tumor-to-heart ratio of 89Zr-DFO-CD4 and 89Zr-DFO-CD8a across tumor types and in-between subjects that correlated with individual response to Sym021 at day 10 relative to start of therapy (p=0.0002 and p=0.0354, respectively). The maximum 89Zr-DFO-CD4 tumor-to-heart ratio could be used to stratify mice according to Sym021 therapy response and overall survival was improved in mice with a 89Zr-DFO-CD4 ratio >9 (p=0.0018). Conclusion: We developed 89Zr-DFO-CD4 and 89Zr-DFO-CD8a PET radiotracers for specific detection and whole-body assessment of CD4+ and CD8a+ status. These radiotracers can be used to phenotype preclinical syngeneic mouse tumor models and to predict response to an immune checkpoint inhibitor. We foresee development of such non-invasive in vivo biomarkers for prediction and evaluation of clinical efficacy of immunotherapeutic agents, such as Sym021.

Ixodid ticks (Acari: Ixodidae) collected from African savanna elephants (Loxodonta africana) and African forest elephants (Loxodonta cyclotis).

Eight ixodid tick species were collected from 173 African savanna elephants (Loxodonta africana) in Kenya, northern Mozambique and Zimbabwe, and two species were collected from six African forest elephants (Loxodonta cyclotis) in the Republic of Congo. A new host record is reported for Amblyomma eburneum. A list of ticks collected from elephants in various African countries, and stored in the United States National Tick Collection, is supplied as well as an annotated checklist of the 27 ixodid tick species that have been collected from African elephants. The geographic distributions and alternative hosts of the various tick species collected from elephants are briefly discussed.

Sym021, a promising anti-PD1 clinical candidate antibody derived from a new chicken antibody discovery platform.

Discovery of therapeutic antibodies is a field of intense development, where immunization of rodents remains a major source of antibody candidates. However, high orthologue protein sequence homology between human and rodent species disfavors generation of antibodies against functionally conserved binding epitopes. Chickens are phylogenetically distant from mammals. Since chickens generate antibodies from a restricted set of germline genes, the possibility of adapting the Symplex antibody discovery platform to chicken immunoglobulin genes and combining it with high-throughput humanization of antibody frameworks by "mass complementarity-determining region grafting" was explored. Hence, wild type chickens were immunized with an immune checkpoint inhibitor programmed cell death 1 (PD1) antigen, and a repertoire of 144 antibodies was generated. The PD1 antibody repertoire was successfully humanized, and we found that most humanized antibodies retained affinity largely similar to that of the parental chicken antibodies. The lead antibody Sym021 blocked PD-L1 and PD-L2 ligand binding, resulting in elevated T-cell cytokine production in vitro. Detailed epitope mapping showed that the epitope recognized by Sym021 was unique compared to the clinically approved PD1 antibodies pembrolizumab and nivolumab. Moreover, Sym021 bound human PD1 with a stronger affinity (30 pM) compared to nivolumab and pembrolizumab, while also cross-reacting with cynomolgus and mouse PD1. This enabled direct testing of Sym021 in the syngeneic mouse in vivo cancer models and evaluation of preclinical toxicology in cynomolgus monkeys. Preclinical in vivo evaluation in various murine and human tumor models demonstrated a pronounced anti-tumor effect of Sym021, supporting its current evaluation in a Phase 1 clinical trial. Abbreviations: ADCC, antibody-dependent cellular cytotoxicity; CD, cluster of differentiation; CDC, complement-dependent cytotoxicity; CDR, complementarity determining region; DC, dendritic cell; ELISA, enzyme-linked immunosorbent assay; FACS, fluorescence activated cell sorting; FR, framework region; GM-CSF, granulocyte-macrophage colony-stimulating factor; HRP, horseradish peroxidase; IgG, immunoglobulin G; IL, interleukin; IFN, interferon; mAb, monoclonal antibody; MLR, mixed lymphocyte reaction; NK, natural killer; PBMC, peripheral blood mono-nuclear cell; PD1, programmed cell death 1; PDL1, programmed cell death ligand 1; RT-PCR, reverse transcription polymerase chain reaction; SEB, Staphylococcus Enterotoxin B; SPR, surface Plasmon Resonance; VL, variable part of light chain; VH, variable part of heavy chain.

Identity of Haemaphysalis (Rhipistoma) muhsamae Santos Dias, 1954 (Acari: Ixodidae) and H. (R.) subterra Hoogstraal, El Kammah & Camicas, 1992, parasites of carnivores and rodents in eastern and southern Africa.

Haemaphysalis (Rhipistoma) muhsamae Santos Dias, 1954 (Acari: Ixodidae) and H. (R.) subterra Hoogstraal, El Kammah & Camicas, 1992, are redescribed based on males and females. Adults of H. muhsamae were mostly collected from various mongooses (Carnivora: Herpestidae) but also from the striped polecat, Ictonyx striatus (Perry) (Carnivora: Mustelidae), serval, Leptailurus serval (Schreber) (Carnivora: Felidae), red veld rat, Aethomys chrysophilus (de Winton) and Selinda veld rat, Aethomys silindensis Roberts (Rodentia: Muridae) in Botswana, Kenya, Mozambique, South Africa, Tanzania and Zimbabwe. Adults of H. subterra were mostly found on various species of African mole rats, Tachyoryctes spp. (Rodentia: Spalacidae) but also on striped polecat, I. striatus and slender mongoose, Galerella sanguinea (Rüppell) (Carnivora: Herpestidae) in Ethiopia and Kenya. Males and females of both species can be differentiated from each other and other H. spinulosa-like ticks by their size, pattern of punctations on conscutum/scutum, shape of genital structures, shape and size of posterodorsal and posteroventral spurs on palpal segment II, hypostome dentition, and shape and size of spurs on coxae. Taxonomic issues of both species and those related to the identity of H. (R.) spinulosa Neumann, 1906 are discussed and a neotype of H. muhsamae has been designated.