Sporozoite motility as a quantitative readout for anti-CSP antibody inhibition.

Antibodies can prevent malaria by neutralizing the infectious Plasmodium falciparum sporozoites (SPZ) before they establish an infection in the liver. Circumsporozoite protein (CSP), the most abundant surface protein of SPZ is the leading candidate for passive (and subunit) immunization approaches against malaria. Comprehensive assessment of the parasite-inhibitory capacity of anti-CSP monoclonal antibodies (mAbs) is an important step in advancing CSP-based immunization strategies. In this study, we employed a quantitative imaging-based motility assay to quantify the effect of anti-CSP mAbs on SPZ motility, both in vitro and in human skin.Our assay provided a quantitative measure of mAb parasite-inhibitory capacity through measurement of the half-maximal motility inhibitory concentration (IC50M) value for anti-CSP mAbs (IC50M 2A10: 24 nM, IC50M 3SP2: 71 nM). We found a sevenfold discrepancy between the IC50M and the binding saturation concentration measured by ELISA, possibly related to the observed shedding of CSP-mAb complexes during SPZ movement. In a subset of SPZ (5%), in vitro motility was unaffected by the presence of 2A10 while 3SP2 was able to completely block movement. In our ex vivo skin explant model, SPZ proved less susceptible to anti-CSP mAbs compared to SPZ in an in vitro environment. By quantitatively assessing motility, we created a valuable tool that can be used for comprehensive assessment of anti-CSP mAb potency. Insight that will help deepen our understanding of anti-CSP mAb potency and guide selection of the most promising anti-CSP mAbs for downstream clinical development.

Clustering and Erratic Movement Patterns of Syringe-Injected versus Mosquito-Inoculated Malaria Sporozoites Underlie Decreased Infectivity.

Malaria vaccine candidates based on live, attenuated sporozoites have led to high levels of protection. However, their efficacy critically depends on the sporozoites' ability to reach and infect the host liver. Administration via mosquito inoculation is by far the most potent method for inducing immunity but highly impractical. Here, we observed that intradermal syringe-injected Plasmodium berghei sporozoites (syrSPZ) were 3-fold less efficient in migrating to and infecting mouse liver than mosquito-inoculated sporozoites (msqSPZ). This was related to a clustered dermal distribution (2-fold-decreased median distance between syrSPZ and msqSPZ) and, more importantly, a 1.4-fold (significantly)-slower and more erratic movement pattern. These erratic movement patterns were likely caused by alteration of dermal tissue morphology (>15-µm intercellular gaps) due to injection of fluid and may critically decrease sporozoite infectivity. These results suggest that novel microvolume-based administration technologies hold promise for replicating the success of mosquito-inoculated live, attenuated sporozoite vaccines.IMPORTANCE Malaria still causes a major burden on global health and the economy. The efficacy of live, attenuated malaria sporozoites as vaccine candidates critically depends on their ability to migrate to and infect the host liver. This work sheds light on the effect of different administration routes on sporozoite migration. We show that the delivery of sporozoites via mosquito inoculation is more efficient than syringe injection; however, this route of administration is highly impractical for vaccine purposes. Using confocal microscopy and automated imaging software, we demonstrate that syringe-injected sporozoites do cluster, move more slowly, and display more erratic movement due to alterations in tissue morphology. These findings indicate that microneedle-based engineering solutions hold promise for replicating the success of mosquito-inoculated live, attenuated sporozoite vaccines.

Anatomical localization of radiocolloid tracer deposition affects outcome of sentinel node procedures in prostate cancer.

PURPOSE: Diagnostic imaging modalities have moderate sensitivity for the identification of lymph node (LN) metastases in prostate cancer (PCa) patients. Mapping the lymphatic drainage from the prostate can help to identify the LNs directly draining from the tumour (sentinel nodes (SNs)); the LNs stated to have the highest chance of containing metastatic cancer cells. Although the lymphatic drainage may differ between segments within the prostate, the location of the primary tumour is not routinely taken into account during peripheral zone-aimed tracer administration. This study evaluates whether linking the SN procedure to the primary cancer deposits increases the identification accuracy of lymphatic metastases. METHODS: Sixty-seven PCa patients, scheduled for robot-assisted laparoscopic prostatectomy (RALP) and extended lymph node dissection (ePLND) with subsequent SN biopsy, were included in this retrospective study. After injection of the hybrid tracer ICG-99mTc-nanocolloid in the prostate, SN mapping was performed based on lymphoscintigraphy and SPECT/CT. SNs were resected using a combination of radio- and fluorescence guidance. Pathology was used to determine the primary tumour location and metastatic spread. Fluorescence imaging of paraffin-embedded prostate tissue was used to determine the location of the tracer deposits in the prostate. This deposition was related to the primary tumour location, the lymphatic drainage pattern of the injected tracer, and the metastatic spread. RESULTS: In total 265 radioactive LNs (211 SNs and 54 higher-echelon nodes in 64 patients; 4.3 LNs per patient; IQR: 2-6) were identified. In three patients (4%) preoperative imaging did not allow identification of SNs. Tumour-positive SN visualization within the pelvis was shown to be influenced by intraprostatic location of tracer administration. This could be concluded from (1) a clear correlation between lymphatic drainage to the right or left side of the body and tracer deposition on the right or left side of the prostate, (2) visualization of a higher number of LNs after dorsal tracer deposition compared with ventral tracer deposition, (3) different drainage patterns observed for tracer deposition into the base or apex of the prostate, and (4) the indication that intratumoural tracer deposition increases the chance of visualizing nodal metastases compared with extratumoural tracer deposition. CONCLUSIONS: The correlation between the location of the tracer deposits, the location of the primary tumour, and the visualization of the (tumour-positive) SNs indicated that placement of tracer deposits is of influence on the visualized lymphatic drainage pattern. This suggests that tracer injection near or into the primary tumour site is beneficial for the identification of metastatic spread.