Expansion and genetic modification of human natural killer cells for adoptive immunotherapy of cancer
Abstract: A ?century ?after ?the ?initial? proposition ?that ?the ?immune? system ?has ?the? capacity ?to? fight? against ?tumors, ?evading ?destruction? by ?immune? cells? is ?now? well ?recognized? as? a? hallmark? of? cancer.? Recent? decades? have? witnessed? extraordinary? improvements ?in? the ?use? of? immunotherapy ?against ?malignancies ?and? adoptive? transfer? of? Natural? Killer? (NK)? cells? stands? among? promising? tools? in? the? fight? against? cancer.? Clinical? studies? have? demonstrated? the? anti?tumor? responses? generated ?by? NK ?cells ?both ?in? the? autologous ?and? allogeneic? settings? in? various? cancers.?Direct ?adoptive ?transfer,? ex? vivo? activation ?and/o r?expansion, ?as ?well? as? genetic? modification? of? NK? cells? aspire? novel? improvements? to? current? immunotherapy? strategies.? As? such? interventions? develop,? the? quest? for? better? preparation? of ?NK ?cell? based ?therapies ?continues. This ?thesis, ?primarily? investigates ?the? feasibility? and ?potential? of? ex? vivo? expanded? NK? cells? for? cancer ?immunotherapy. ?Our ?results?p roduced? a ?system ?that? has ?the? capacity ?to? expand? polyclonal ?and ?highly ?cytotoxic? NK ?cells showing ?selective? anti? tumor? activity.? Protocols? for? expansion? of? these? cells? from? healthy? donors? and? patients? with? Multiple? Myeloma? (MM)? using? current? Good? Manufacturing? Practice? (cGMP)?compliant? methods? have? been? optimized? in? conventional? cell? culture ?systems ?as? well? as?automated? bioreactors.?The? elevated? cytotoxic?activity? of ?expanded ?NK? cells ?against? autologous? tumor ?cells,? along? with? detailed? analysis? of? phenotypic? changes? during? the? expansion? process? has? subsequently? shifted? attention ?to? the? interaction ?between ?NK ?and? tumor ?cells. Both ?as? a ?basic ?method ?to? identify? these? interactions,? and ?as ?part ?of? further? plans? to? use? genetically? retargeted? NK? cells? in? cancer? immunotherapy,? we? have? investigated? methods? for ?efficient? lentiviral? genetic? modification ?of? NK ?cells.?This? study ?has ?resulted ?in? an? optimized ?stimulation ?and? genetic? modification ?process? for? NK? cells? that? greatly? enhances? viral? gene? delivery.? Along? with? NK? cell? stimulating? cytokines,? an? inhibitor? of? innate? immune? receptor? signaling? that? blocks? the? intracellular? detection? of? viral? RNA? introduced? by? the? vector? was? successfully? utilized ?to ?enhance ?gene? transfer ?efficiency,? also? constituting ?a? proof? of?concept ?for? various? other? gene? therapy? approaches. Taken? together,? the? work? presented? in? this? thesis? aims? to? bring? us? closer? to? optimal? ex ?vivo? manipulation ?of? NK ?cells? for? immunotherapy.? Clinical ?trials ?with? the ?long?term? expanded ?NK ?cells? as? well? as ?further ?preclinical ?development? of ?NK? cell? genetic ?modification ?processes ?are? warranted.
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