Development and application of a high-throughput screening assay for identification of small molecule inhibitors of the P. falciparum reticulocyte binding-like homologue 5 protein

Article abstract of DOI:10.1016/j.ijpddr.2020.10.008

The P. falciparum parasite, responsible for the disease in humans known as malaria, must invade erythrocytes to provide an environment for self-replication and survival. For invasion to occur, the parasite must engage several ligands on the host erythrocyte surface to enable adhesion, tight junction formation and entry. Critical interactions include binding of erythrocyte binding-like ligands and reticulocyte binding-like homologues (Rhs) to the surface of the host erythrocyte. The reticulocyte binding-like homologue 5 (Rh5) is the only member of this family that is essential for invasion and it binds to the basigin host receptor. The essential nature of Rh5 makes it an important vaccine target, however to date, Rh5 has not been targeted by small molecule intervention. Here, we describe the development of a high-throughput screening assay to identify small molecules which interfere with the Rh5-basigin interaction. To validate the utility of this assay we screened a known drug library and the Medicines for Malaria Box and demonstrated the reproducibility and robustness of the assay for high-throughput screening purposes. The screen of the known drug library identified the known leukotriene antagonist, pranlukast. We used pranlukast as a model inhibitor in a post screening evaluation cascade. We procured and synthesised analogues of pranlukast to assist in the hit confirmation process and show which structural moieties of pranlukast attenuate the Rh5 – basigin interaction. Evaluation of pranlukast analogues against P. falciparum in a viability assay and a schizont rupture assay show the parasite activity was not consistent with the biochemical inhibition of Rh5, questioning the developability of pranlukast as an antimalarial. The high-throughput assay developed from this work has the capacity to screen large collections of small molecules to discover inhibitors of P. falciparum Rh5 for future development of invasion inhibitory antimalarials.

Full text of DOI:10.1016/j.ijpddr.2020.10.008

2139
JACC March 21, 2017
Volume 69, Issue 11
FIT Clinical Decision Making
PARADOXICAL HYPERTENSIVE ADVERSE REACTION TO NITROGLYCERINE DURING TILT TABLE
TEST UNMASKING UNDIAGNOSED PHEOCHROMOCYTOMA: A CASE REPORT
Moderated Poster Contributions
Spotlight on Special Categories Moderated Poster Theater, Poster Hall, Hall C
Sunday, March 19, 2017, 1:30 p.m.-1:40 p.m.
Session Title: FIT Clinical Decision‐Making: Arrhythmias and Clinical Electrophysiology
Abstract Category: Prevention
Presentation Number: 1321M-11
Authors: Amit Arbune, Sharad Malavade, Timothy Edmiston, Zulfiqar Baloch, Saurabh Chokshi, MetroHealth Medical Center, Case
Western University, Cleveland, OH, USA, Brandon Regional Hospital, Brandon, FL, USA
Background: Tilt table test (TTT) is commonly used for dizziness work-up, that includes use of nitroglycerine(NTG) during provocation
phase. Paradoxical hypertensive reaction to NTG is extremely rare. Pheochromocytoma rarely presents as dizziness and hypertensive
emergency can rarely occur during TTT with NTG provocation in these patients.
Case: A 51year-old woman with controlled hypertension underwent TTT for dizziness with sublingual NTG for provocation phase. Within
14 minutes, she developed hypertensive emergency, pulmonary edema and cardio-pulmonary instability with room air SpO2 71%, heart
rate 103 bpm and BP 273/158mmHg, cold, clammy and greyish skin. Possible anaphylaxis to NTG with hypertensive emergency was
suspected and treated with hydralazine, metoprolol, intravenous methylprednisolone sodium succinate and diphenhydramine. Initially,
epinephrine was withheld due to severe hypertension. Despite treatment, patient decompensated with further respiratory distress and
developed hypotension. Epinephrine was given with respiratory support to stabilize her. She recovered uneventfully and was discharged
home.
Decision‐Making: Possible anaphylaxis to NTG was suspected given the rapidity of the patient’s adverse reaction. Skin test with NTG
was recommended, but posed risk of life threatening adverse reaction. Hyperadrenergic surge from undiagnosed pheochromocytoma was
considered as alternate diagnosis. Laboratory studies showed elevated 24-hour urine metanephrine 4266 mcg/24hr (normal range, 140
to 785) and 24-hour urine normetanephrine 4175 mcg/24hr (normal range, 75 to 375). MRI of abdomen revealed right suprarenal mass
consistent with diagnosis of pheochromocytoma.
Conclusions: NTG, an antihypertensive, with paradoxical hypertensive response should evoke suspicion for pheochromocytoma. Focused
laboratory and imaging studies prevented a potentially life threatening skin test with nitroglycerine in this patient.