Methods for treating HPV-related diseases

JP7905173B2Active Publication Date: 2026-08-14SQZ BIOTECHNOLOGIES CO
View PDF 6 Cites 0 Cited by

Patent Information

Authority / Receiving Office
JP · JP
Patent Type
Patents
Current Assignee / Owner
Filing Date
2019-03-11
Publication Date
2026-08-14

AI Technical Summary

Benefits of technology

【0064】 一部の態様では、本発明は、個体におけるHPV関連疾患を処置または予防するための方法であって、HPV抗原と会合した改変免疫細胞を個体に投与するステップを含み、改変免疫細胞は、a)HPV抗原がインプット細胞と会合するのを可能にするのに十分な時間、インプット細胞とHPV抗原および/またはアジュバントとをインキュベートし、それによって、抗原と会合した改変免疫細胞を作出するステップを含むプロセスによって調製される、方法を提供する。一部の実施形態では、HPV抗原は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含む。一部の実施形態では、HPV抗原は配列番号23のアミノ酸配列を含む。一部の実施形態では、アジュバントはCpG ODNである。一部の実施形態では、CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である。 特定の実施形態では、例えば、以下が提供される: (項目1) 個体におけるヒトパピローマウイルス(HPV)関連疾患を処置するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されている、方法。 (項目2) 個体におけるHPV関連疾患を予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されている、方法。 (項目3) HPV関連疾患を有する個体における免疫応答をモジュレートするための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されている、方法。 (項目4) 個体におけるHPV関連疾患を処置するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されており、 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記抗原および前記アジュバントが通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;ならびに b)前記HPV抗原および前記アジュバントが前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原および前記アジュバントとをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目5) 個体におけるHPV関連疾患を予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されており、 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原および前記アジュバントが通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;ならびに b)前記HPV抗原および前記アジュバントが前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原および前記アジュバントとをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目6) HPV関連疾患を有する個体における免疫応答をモジュレートするための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されており、 前記改変免疫細胞は、 a)HPV抗原を含むインプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原および前記アジュバントが通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;ならびに b)前記HPV抗原および前記アジュバントが前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記抗原および前記アジュバントとをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目7) 前記狭窄の前記直径は、前記細胞の前記直径未満である、項目4~6のいずれか一項に記載の方法。 (項目8) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~99%である、項目4~7のいずれか一項に記載の方法。 (項目9) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約60%未満である、項目4~8のいずれか一項に記載の方法。 (項目10) 前記狭窄は、チャネル中にある、項目4~9のいずれか一項に記載の方法。 (項目11) 前記インプット細胞に、それが前記狭窄を通過するときに変形力が印加される、項目4~10のいずれか一項に記載の方法。 (項目12) 前記HPV抗原および/または前記アジュバントは、細胞質基質および/またはエンドソームに存在する、項目1~11のいずれかに記載の方法。 (項目13) 前記抗原および/またはアジュバントは、前記細胞の多数の区画に存在する、項目1~12のいずれか一項に記載の方法。 (項目14) 前記改変免疫細胞は、前記細胞の外側に、HPV抗原および/またはアジュバントをさらに含む、項目1~13のいずれか一項に記載の方法。 (項目15) 前記摂動インプット細胞とインキュベートされるアジュバントの濃度は、約0.1μM~約1mMである、項目1~14のいずれか一項に記載の方法。 (項目16) 前記摂動インプット細胞とインキュベートされるHPV抗原の濃度は、約0.1μM~約1mMである、項目1~15のいずれか一項に記載の方法。 (項目17) 前記摂動インプット細胞とインキュベートされるHPV抗原のアジュバントに対する比は、約10000:1~約1:10000である、項目4~16のいずれか一項に記載の方法。 (項目18) 前記免疫応答は増強される、項目3または6に記載の方法。 (項目19) 前記HPV抗原に対する前記免疫応答は増強される、項目18に記載の方法。 (項目20) 前記アジュバントは、CpG ODN、IFN-α、STINGアゴニスト、RIG-Iアゴニスト、またはポリI:Cである、項目1~19のいずれか一項に記載の方法。 (項目21) 前記アジュバントはCpG ODNである、項目20に記載の方法。 (項目22) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目21に記載の方法。 (項目23) 前記改変免疫細胞は1つよりも多いアジュバントを含む、項目1~22のいずれか一項に記載の方法。 (項目24) 前記HPV抗原は、同じおよび/または異なるHPV抗原に対する応答を誘発する多数のポリペプチドのプールである、項目1~23のいずれか一項に記載の方法。 (項目25) 多数の抗原の前記プール中の抗原は、多数の抗原の前記プール中の他の抗原に向けられた前記免疫応答を減少させない、項目24に記載の方法。 (項目26) 前記HPV抗原は、抗原性HPVエピトープおよび1つまたは複数の異種ペプチド配列を含むポリペプチドである、項目1~25のいずれか一項に記載の方法。 (項目27) 前記HPV抗原は、それ自体と、他の抗原と、または前記アジュバントと複合体を形成する、項目1~26のいずれか一項に記載の方法。 (項目28) 前記HPVは細胞溶解物に由来する抗原である、項目1~27のいずれか一項に記載の方法。 (項目29) 前記HPV抗原は、HPV-16またはHPV-18抗原である、項目1~28のいずれか一項に記載の方法。 (項目30) 前記HPV抗原は、HLA-A2特異的エピトープで構成される、項目29に記載の方法。 (項目31) 前記HPV抗原は、HPV E6抗原またはHPV E7抗原である、項目1~30のいずれか一項に記載の方法。 (項目32) 前記改変免疫細胞は、HPV E6抗原およびHPV E7抗原を含む、項目1~31のいずれか一項に記載の方法。 (項目33) 前記HPV抗原は、1つまたは複数の異種ペプチド配列がそのN末端および/またはそのC末端で隣接している抗原性エピトープを含むポリペプチドである、項目1~32のいずれか一項に記載の方法。 (項目34) 前記HPV抗原は、配列番号18~26のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含む、項目33に記載の方法。 (項目35) 前記HPV抗原は、配列番号23と少なくとも90%の類似性を有するアミノ酸配列を含む、項目34に記載の方法。 (項目36) 前記HPV抗原は、MHCクラスI拘束性ペプチドにプロセシングされ得る、項目1~35のいずれか一項に記載の方法。 (項目37) 前記HPV抗原は、MHCクラスII拘束性ペプチドにプロセシングされ得る、項目1~36のいずれか一項に記載の方法。 (項目38) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記アジュバントを含む、項目1~37のいずれか一項に記載の方法。 (項目39) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記HPV抗原を含む、項目1~38のいずれか一項に記載の方法。 (項目40) 前記HPV抗原の前記アジュバントに対する比は、約10000:1~約1:10000である、項目1~39のいずれか一項に記載の方法。 (項目41) 前記改変免疫細胞は、作用物質をさらに含み、前記作用物質は、前記作用物質を含まない対応する改変免疫細胞と比較して前記改変免疫細胞の生存能力および/または機能を増強する、項目1~40のいずれか一項に記載の方法。 (項目42) 前記作用物質は、エンドサイトーシスを増強する化合物、安定剤、または補因子である、項目41に記載の方法。 (項目43) 前記作用物質はアルブミンである、項目41に記載の方法。 (項目44) 前記アルブミンは、マウス、ウシ、またはヒトアルブミンである、項目43に記載の方法。 (項目45) 前記作用物質は、二価金属カチオン、グルコース、ATP、カリウム、グリセロール、トレハロース、D-スクロース、PEG1500、L-アルギニン、L-グルタミン、またはEDTAである、項目41に記載の方法。 (項目46) 前記作用物質はマウス血清アルブミン(MSA)を含む、項目41に記載の方法。 (項目47) 前記改変免疫細胞は、共刺激分子のうちの1つまたは複数の発現を増加させるようにさらに改変されている、項目1~46のいずれか一項に記載の方法。 (項目48) 前記共刺激分子は、B7-H2、B7-1、B7-2、CD70、LIGHT、HVEM、CD40、4-1BBL、OX40L、TL1A、GITRL、CD30L、TIM4、SLAM、CD48、CD58、CD155、またはCD112である、項目47に記載の方法。 (項目49) 前記細胞は、前記1つまたは複数の共刺激分子の増加した発現をもたらす核酸を含む、項目47または48に記載の方法。 (項目50) 前記免疫細胞は、T細胞、樹状細胞、単球、マクロファージ、骨髄性細胞、顆粒球、好中球、マスト細胞、ナチュラルキラー細胞、自然リンパ球、好塩基球、または造血系前駆細胞である、項目1~49のいずれか一項に記載の方法。 (項目51) 前記免疫細胞はB細胞ではない、項目1~50のいずれか一項に記載の方法。 (項目52) 前記免疫細胞はB細胞である、項目1~50のいずれか一項に記載の方法。 (項目53) 前記免疫細胞はT細胞である、項目1~51のいずれか一項に記載の方法。 (項目54) 前記免疫細胞は混合細胞集団である、項目1~49のいずれか一項に記載の方法。 (項目55) 前記免疫細胞は複数のPBMCである、項目54に記載の方法。 (項目56) 前記T細胞は、MHCクラスI発現をモジュレートするさらなる改変を含む、項目53に記載の方法。 (項目57) 前記T細胞は、MHCクラスII発現をモジュレートするさらなる改変を含む、項目53に記載の方法。 (項目58) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を低下させるさらなる改変を含む、項目56または57に記載の方法。 (項目59) 前記さらなる改変は、siRNA、shRNA、CRISPR/Cas9、ZFN、TALEN、Creリコンビナーゼ、またはメガヌクレアーゼを使用して、MHCクラスIおよび/またはMHCクラスII発現を低下させることを含む、項目56または57に記載の方法。 (項目60) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を増加させるさらなる改変を含む、項目56または57に記載の方法。 (項目61) 前記さらなる改変は、RNAまたはプラスミドDNAを使用して、MHCクラスIおよび/またはMHCクラスII発現を増加させることを含む、項目56または57に記載の方法。 (項目62) 前記さらなる改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答は、前記さらなる改変を含まない対応する改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答と比較して低下している、項目53および56~59のいずれか一項に記載の方法。 (項目63) 前記さらなる改変T細胞の、それらが投与された個体における循環半減期は、前記さらなる改変を含まない対応する改変T細胞の、それらが投与された個体における循環半減期と比較してモジュレートされる、項目53および56~59のいずれか一項に記載の方法。 (項目64) 前記T細胞は、ヘルパーT細胞、細胞傷害性T細胞、メモリーT細胞、CIK細胞、およびナチュラルキラーT細胞のうちの1つまたは複数を含む、項目53および56~63のいずれか一項に記載の方法。 (項目65) 前記T細胞は、CD3+ T細胞、CD4+ T細胞、CD8+ T細胞、CD45RA+ T細胞、CD45RO+ T細胞、およびγδ-T細胞のうちの1つまたは複数を含む、項目53および56~63のいずれか一項に記載の方法。 (項目66) 前記改変細胞は前記個体に対して同種異系である、項目1~65のいずれか一項に記載の方法。 (項目67) 前記改変細胞は前記個体に対して自家である、項目1~65のいずれか一項に記載の方法。 (項目68) 前記個体は、モジュレートされた炎症および/またはモジュレートされた免疫応答を有するよう事前調整されている、項目1~67のいずれか一項に記載の方法。 (項目69) アジュバントを前記個体に投与するステップをさらに含む、項目1~68のいずれか一項に記載の方法。 (項目70) 前記アジュバントは、IFNαまたはCpG ODNである、項目69に記載の方法。 (項目71) 前記改変免疫細胞を含む前記組成物および前記アジュバントは同時に投与される、項目69または70に記載の方法。 (項目72) 前記改変免疫細胞を含む前記組成物および前記アジュバントは逐次的に投与される、項目69または70に記載の方法。 (項目73) 前記改変免疫細胞を含む前記組成物は、前記アジュバントを投与する前に投与される、項目72に記載の方法。 (項目74) 前記改変免疫細胞を含む前記組成物は、前記アジュバントの投与後に投与される、項目72に記載の方法。 (項目75) 前記改変免疫細胞を含む前記組成物は、免疫チェックポイント阻害剤の投与と組み合わせて投与される、項目1~74のいずれか一項に記載の方法。 (項目76) 前記改変免疫細胞を含む前記組成物および前記免疫チェックポイント阻害剤は同時に投与される、項目75に記載の方法。 (項目77) 前記改変免疫細胞を含む前記組成物および前記免疫チェックポイント阻害剤は逐次的に投与される、項目75に記載の方法。 (項目78) 前記改変免疫細胞を含む前記組成物は、前記免疫チェックポイント阻害剤を投与する前に投与される、項目77に記載の方法。 (項目79) 前記改変免疫細胞を含む前記組成物は、前記免疫チェックポイント阻害剤の投与後に投与される、項目77に記載の方法。 (項目80) 前記免疫チェックポイント阻害剤は、PD-1、PD-L1、CTLA-4、LAG3、TIM-3、TIGIT、VISTA、TIM1、B7-H4(VTCN1)、またはBTLAのうちの1つまたは複数を標的にする、項目75~79のいずれか一項に記載の方法。 (項目81) 前記改変免疫細胞を含む前記組成物は、化学療法の投与と組み合わせて投与される、項目1~80のいずれか一項に記載の方法。 (項目82) 前記改変免疫細胞を含む前記組成物および前記化学療法は同時に投与される、項目81に記載の方法。 (項目83) 前記改変免疫細胞を含む前記組成物および前記化学療法は逐次的に投与される、項目81に記載の方法。 (項目84) 前記改変免疫細胞を含む前記組成物は、前記化学療法を投与する前に投与される、項目83に記載の方法。 (項目85) 前記改変免疫細胞を含む前記組成物は、前記化学療法の投与後に投与される、項目83に記載の方法。 (項目86) 前記化学療法は白金系作用物質を含む、項目81~85のいずれか一項に記載の方法。 (項目87) 前記化学療法はシスプラチンを含む、項目81~86のいずれか一項に記載の方法。 (項目88) 前記個体への前記改変免疫細胞を含む前記組成物の投与は、前記HPV抗原に特異的な細胞傷害性Tリンパ球(CTL)の活性化および/または増殖をもたらす、項目1~87のいずれか一項に記載の方法。 (項目89) 前記個体への前記改変免疫細胞を含む前記組成物の投与は、前記抗原に特異的なヘルパーT(Th)細胞の活性化および/または増殖をもたらす、項目1~87のいずれか一項に記載の方法。 (項目90) 前記有効量の前記組成物は、約1×106~約1×1012個の改変免疫細胞を含む、項目1~89のいずれか一項に記載の方法。 (項目91) 前記改変免疫細胞を含む前記組成物の複数回投与を含む、項目1~90のいずれか一項に記載の方法。 (項目92) 前記改変免疫細胞を含む前記組成物の第1の投与、続いて、前記改変免疫細胞を含む前記組成物の第2の投与を含む、項目91に記載の方法。 (項目93) 前記第2の投与は、前記第1の投与の約1か月後である、項目92に記載の方法。 (項目94) 前記HPV関連疾患はHPV関連がんである、項目1~93のいずれか一項に記載の方法。 (項目95) 前記HPV関連がんは、子宮頸がん、肛門がん、中咽頭がん、膣がん、外陰がん、陰茎がん、皮膚がん、または頭頸部がんである、項目94に記載の方法。 (項目96) 前記HPV関連疾患は、HPV関連感染性疾患である、項目1~95のいずれか一項に記載の方法。 (項目97) 個体におけるヒトパピローマウイルス(HPV)関係疾患を処置するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸を含むHPV抗原を含む、方法。 (項目98) 個体におけるHPV関連疾患を予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含むHPV抗原を含む、方法。 (項目99) HPV関連疾患を有する個体における免疫応答をモジュレートするための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含むHPV抗原を含む、方法。 (項目100) 個体におけるHPV関連疾患を処置するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含むHPV抗原を含み、 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記抗原が通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;および b)前記HPV抗原が前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原とをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目101) 個体におけるHPV関連疾患を予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原を含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含むHPV抗原を含み、 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原が通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;および b)前記HPV抗原が前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原とをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目102) HPV関連疾患を有する個体における免疫応答をモジュレートするための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含むHPV抗原を含み、 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原が通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;および b)前記HPV抗原が前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原とをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目103) 前記狭窄の前記直径は、前記細胞の前記直径未満である、項目100~102のいずれか一項に記載の方法。 (項目104) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~99%である、項目100~103のいずれか一項に記載の方法。 (項目105) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約60%未満である、項目100~104のいずれか一項に記載の方法。 (項目106) 前記狭窄は、チャネル中にある、項目100~105のいずれか一項に記載の方法。 (項目107) 前記インプット細胞に、それが前記狭窄を通過するときに変形力が印加される、項目100~106のいずれか一項に記載の方法。 (項目108) アジュバントを前記個体に投与するステップをさらに含む、項目86~107のいずれか一項に記載の方法。 (項目109) 前記アジュバントは、IFNαまたはCpG ODNである、項目108に記載の方法。 (項目110) 前記改変免疫細胞を含む前記組成物および前記アジュバントは同時に投与される、項目108または109に記載の方法。 (項目111) 前記改変免疫細胞を含む前記組成物および前記アジュバントは逐次的に投与される、項目108または109に記載の方法。 (項目112) 前記改変免疫細胞を含む前記組成物は、前記アジュバントを投与する前に投与される、項目111に記載の方法。 (項目113) 前記改変免疫細胞を含む前記組成物は、前記アジュバントの投与後に投与される、項目111に記載の方法。 (項目114) 前記改変免疫細胞はアジュバントをさらに含む、項目97~113のいずれか一項に記載の方法。 (項目115) ステップbの前記摂動免疫細胞は、前記HPV抗原およびアジュバントとインキュベートされる、項目100~113のいずれか一項に記載の方法。 (項目116) 前記HPV抗原および/または前記アジュバントは、細胞質基質および/またはエンドソームに存在する、項目114または115に記載の方法。 (項目117) 前記抗原および/またはアジュバントは、前記細胞の多数の区画に存在する、項目114~116のいずれか一項に記載の方法。 (項目118) 前記改変免疫細胞は、前記細胞の外側に、HPV抗原および/またはアジュバントをさらに含む、項目114~117のいずれか一項に記載の方法。 (項目119) 前記摂動インプット細胞とインキュベートされるアジュバントの濃度は、約0.1μM~約1mMである、項目115~118のいずれか一項に記載の方法。 (項目120) 前記摂動インプット細胞とインキュベートされるHPV抗原の濃度は、約0.1μM~約1mMである、項目115~119のいずれか一項に記載の方法。 (項目121) 前記摂動インプット細胞とインキュベートされるHPV抗原のアジュバントに対する比は、約10000:1~約1:10000である、項目115~120のいずれか一項に記載の方法。 (項目122) 前記免疫応答は増強される、項目99または102に記載の方法。 (項目123) 前記HPV抗原に対する前記免疫応答は増強される、項目122に記載の方法。 (項目124) 前記アジュバントは、CpG ODN、IFN-α、STINGアゴニスト、RIG-Iアゴニスト、またはポリI:Cである、項目114~123のいずれか一項に記載の方法。 (項目125) 前記アジュバントはCpG ODNである、項目124に記載の方法。 (項目126) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目125に記載の方法。 (項目127) 前記改変免疫細胞は1つよりも多いアジュバントを含む、項目114~126のいずれか一項に記載の方法。 (項目128) 前記HPV抗原は、同じおよび/または異なるHPV抗原に対する応答を誘発する多数のポリペプチドのプールである、項目97~127のいずれか一項に記載の方法。 (項目129) 多数の抗原の前記プール中の抗原は、多数の抗原の前記プール中の他の抗原に向けられた前記免疫応答を減少させない、項目128に記載の方法。 (項目130) 前記HPV抗原は、抗原性HPVエピトープおよび1つまたは複数の異種ペプチド配列を含むポリペプチドである、項目97~129のいずれか一項に記載の方法。 (項目131) 前記HPV抗原は、それ自体と、他の抗原と、または前記アジュバントと複合体を形成する、項目97~130のいずれか一項に記載の方法。 (項目132) 前記HPV抗原は、HLA-A2特異的エピトープで構成される、項目97~131のいずれか一項に記載の方法。 (項目133) 前記HPV抗原は、MHCクラスI拘束性ペプチドにプロセシングされ得る、項目97~132のいずれか一項に記載の方法。 (項目134) 前記HPV抗原は、MHCクラスII拘束性ペプチドにプロセシングされ得る、項目97~133のいずれか一項に記載の方法。 (項目135) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記アジュバントを含む、項目114~134のいずれか一項に記載の方法。 (項目136) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記HPV抗原を含む、項目97~135のいずれか一項に記載の方法。 (項目137) 前記HPV抗原の前記アジュバントに対する比は、約10000:1~約1:10000である、項目114~136のいずれか一項に記載の方法。 (項目138) 前記改変免疫細胞は、作用物質をさらに含み、前記作用物質は、前記作用物質を含まない対応する改変免疫細胞と比較して前記改変免疫細胞の生存能力および/または機能を増強する、項目97~137のいずれか一項に記載の方法。 (項目139) 前記作用物質は、エンドサイトーシスを増強する化合物、安定剤、または補因子である、項目138に記載の方法。 (項目140) 前記作用物質はアルブミンである、項目138に記載の方法。 (項目141) 前記アルブミンは、マウス、ウシ、またはヒトアルブミンである、項目140に記載の方法。 (項目142) 前記作用物質は、二価金属カチオン、グルコース、ATP、カリウム、グリセロール、トレハロース、D-スクロース、PEG1500、L-アルギニン、L-グルタミン、またはEDTAである、項目138に記載の方法。 (項目143) 前記作用物質はMSAを含む、項目138に記載の方法。 (項目144) 共刺激分子のうちの1つまたは複数の発現を増加させるようにさらに改変されている、項目97~143のいずれか一項に記載の改変T細胞。 (項目145) 前記共刺激分子は、B7-H2(ICOSL)、B7-1(CD80)、B7-2(CD86)、CD70、LIGHT、HVEM、CD40、4-1BBL、OX40L、TL1A、GITRL、CD30L、TIM4、SLAM、CD48、CD58、CD155、またはCD112である、項目144に記載の改変T細胞。 (項目146) 前記1つまたは複数の共刺激分子の増加した発現をもたらす核酸を含む、項目144または145に記載の改変T細胞。 (項目147) 前記免疫細胞は、T細胞、樹状細胞、単球、マクロファージ、骨髄性細胞、顆粒球、好中球、マスト細胞、ナチュラルキラー細胞、自然リンパ球、好塩基球、または造血系前駆細胞である、項目97~146のいずれか一項に記載の方法。 (項目148) 前記免疫細胞はB細胞ではない、項目97~147のいずれか一項に記載の方法。 (項目149) 前記免疫細胞はB細胞である、項目97~148のいずれか一項に記載の方法。 (項目150) 前記免疫細胞はT細胞である、項目97~148のいずれか一項に記載の方法。 (項目151) 前記免疫細胞は混合細胞集団である、項目97~148のいずれか一項に記載の方法。 (項目152) 前記免疫細胞は複数のPBMCである、項目151に記載の方法。 (項目153) 前記T細胞は、MHCクラスI発現をモジュレートするさらなる改変を含む、項目150に記載の方法。 (項目154) 前記T細胞は、MHCクラスII発現をモジュレートするさらなる改変を含む、項目150に記載の方法。 (項目155) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を低下させるさらなる改変を含む、項目153または154に記載の方法。 (項目156) 前記さらなる改変は、siRNA、shRNA、CRISPR/Cas9、ZFN、TALEN、Creリコンビナーゼ、またはメガヌクレアーゼを使用して、MHCクラスIおよび/またはMHCクラスII発現を低下させることを含む、項目153または154に記載の方法。 (項目157) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を増加させるさらなる改変を含む、項目153または154に記載の方法。 (項目158) 前記さらなる改変は、RNAまたはプラスミドDNAを使用して、MHCクラスIおよび/またはMHCクラスII発現を増加させることを含む、項目153または154に記載の方法。 (項目159) 前記さらなる改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答は、前記さらなる改変を含まない対応する改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答と比較して低下している、項目150および153~156のいずれか一項に記載の方法。 (項目160) 前記さらなる改変T細胞の、それらが投与された個体における循環半減期は、前記さらなる改変を含まない対応する改変T細胞の、それらが投与された個体における循環半減期と比較してモジュレートされる、項目150および153~156のいずれか一項に記載の方法。 (項目161) 前記T細胞は、ヘルパーT細胞、細胞傷害性T細胞、メモリーT細胞、CIK細胞、およびナチュラルキラーT細胞のうちの1つまたは複数を含む、項目150および153~160のいずれか一項に記載の方法。 (項目162) 前記T細胞は、CD3+ T細胞、CD4+ T細胞、CD8+ T細胞、CD45RA+ T細胞、CD45RO+ T細胞、およびγδ-T細胞のうちの1つまたは複数を含む、項目150および153~160のいずれか一項に記載の方法。 (項目163) 前記改変細胞は前記個体に対して同種異系である、項目97~162のいずれか一項に記載の方法。 (項目164) 前記改変細胞は前記個体に対して自家である、項目97~162のいずれか一項に記載の方法。 (項目165) 前記個体は、モジュレートされた炎症および/またはモジュレートされた免疫応答を有するよう事前調整されている、項目97~164のいずれか一項に記載の方法。 (項目166) 前記改変免疫細胞を含む前記組成物は、免疫チェックポイント阻害剤の投与と組み合わせて投与される、項目97~165のいずれか一項に記載の方法。 (項目167) 前記改変免疫細胞を含む前記組成物および前記免疫チェックポイント阻害剤は同時に投与される、項目166に記載の方法。 (項目168) 前記改変免疫細胞を含む前記組成物および前記免疫チェックポイント阻害剤は逐次的に投与される、項目166に記載の方法。 (項目169) 前記改変免疫細胞を含む前記組成物は、前記免疫チェックポイント阻害剤を投与する前に投与される、項目168に記載の方法。 (項目170) 前記改変免疫細胞を含む前記組成物は、前記免疫チェックポイント阻害剤の投与後に投与される、項目168に記載の方法。 (項目171) 前記免疫チェックポイント阻害剤は、PD-1、PD-L1、CTLA-4、LAG3、TIM-3、TIGIT、VISTA、TIM1、B7-H4(VTCN1)、またはBTLAのうちの1つまたは複数を標的にする、項目152~156のいずれか一項に記載の方法。 (項目172) 前記改変免疫細胞を含む前記組成物は、化学療法の投与と組み合わせて投与される、項目97~171のいずれか一項に記載の方法。 (項目173) 前記改変免疫細胞を含む前記組成物および前記化学療法は同時に投与される、項目172に記載の方法。 (項目174) 前記改変免疫細胞を含む前記組成物および前記化学療法は逐次的に投与される、項目172に記載の方法。 (項目175) 前記改変免疫細胞を含む前記組成物は、前記化学療法を投与する前に投与される、項目174に記載の方法。 (項目176) 前記改変免疫細胞を含む前記組成物は、前記化学療法の投与後に投与される、項目174に記載の方法。 (項目177) 前記化学療法はシスプラチンを含む、項目172~176のいずれか一項に記載の方法。 (項目178) 前記個体への前記改変免疫細胞を含む前記組成物の投与は、前記HPV抗原に特異的な細胞傷害性Tリンパ球(CTL)の活性化および/または増殖をもたらす、項目97~177のいずれか一項に記載の方法。 (項目179) 前記個体への前記改変免疫細胞を含む前記組成物の投与は、前記HPV抗原に特異的なヘルパーT(Th)細胞の活性化および/または増殖をもたらす、項目97~177のいずれか一項に記載の方法。 (項目180) 前記有効量の前記組成物は、約1×106~約1×1012個の改変免疫細胞を含む、項目97~179のいずれか一項に記載の方法。 (項目181) 前記改変免疫細胞を含む前記組成物の複数回投与を含む、項目97~180のいずれか一項に記載の方法。 (項目182) 前記改変免疫細胞を含む前記組成物の第1の投与、続いて、前記改変免疫細胞を含む前記組成物の第2の投与を含む、項目181に記載の方法。 (項目183) 前記第2の投与は、前記第1の投与の約1か月後である、項目182に記載の方法。 (項目184) 前記HPV関連疾患はHPV関連がんである、項目97~183のいずれか一項に記載の方法。 (項目185) 前記HPV関連がんは、子宮頸がん、肛門がん、中咽頭がん、膣がん、外陰がん、陰茎がん、皮膚がん、または頭頸部がんである、項目184に記載の方法。 (項目186) 改変免疫細胞を含む組成物であって、前記改変免疫細胞は、CpG ODN、および配列番号18~25のいずれか1つと少なくとも90%の類似性を有するHPV抗原を細胞内に含む、組成物。 (項目187) 前記HPV抗原は、配列番号23と少なくとも90%の類似性を有するアミノ酸配列を含む、項目166に記載の組成物。 (項目188) 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原および前記CpG ODNが通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;ならびに b)前記HPV抗原および前記CpG ODNが前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原および前記CpG ODNとをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、項目186または187に記載の組成物。 (項目189) 前記狭窄の前記直径は、前記細胞の前記直径未満である、項目188に記載の組成物。 (項目190) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約99%である、項目188または189に記載の組成物。 (項目191) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約60%未満である、項目188~190のいずれか一項に記載の組成物。 (項目192) 前記狭窄は、チャネル中にある、項目188~191のいずれか一項に記載の組成物。 (項目193) 前記インプット細胞に、それが前記狭窄を通過するときに変形力が印加される、項目188~192のいずれか一項に記載の組成物。 (項目194) アジュバントをさらに含む、項目186~193のいずれか一項に記載の組成物。 (項目195) 前記HPV抗原および/または前記CpG ODNは、細胞質基質および/またはエンドソームに存在する、項目186~194のいずれかに記載の組成物。 (項目196) 前記抗原および/または前記CpG ODNは、前記細胞の多数の区画に存在する、項目186~195のいずれか一項に記載の組成物。 (項目197) 前記改変免疫細胞は、前記細胞の表面に、HPV抗原および/またはCpG ODNをさらに含む、項目186~196のいずれか一項に記載の組成物。 (項目198) 前記摂動インプット細胞とインキュベートされるCpG ODNの濃度は、約0.1μM~約1mMである、項目188~197のいずれか一項に記載の組成物。 (項目199) 前記摂動インプット細胞とインキュベートされるHPV抗原の濃度は、約0.1μM~約1mMである、項目188~198のいずれか一項に記載の組成物。 (項目200) 前記摂動インプット細胞とインキュベートされるHPV抗原のCpG ODNに対する比は、約10000:1~約1:10000である、項目188~199のいずれか一項に記載の組成物。 (項目201) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目186~200のいずれか一項に記載の組成物。 (項目202) 前記改変免疫細胞は1つよりも多いアジュバントを含む、項目186~201のいずれか一項に記載の組成物。 (項目203) 前記アジュバントは、CpG ODN、IFN-α、STINGアゴニスト、RIG-Iアゴニスト、またはポリI:Cを含む、項目202に記載の組成物。 (項目204) 前記HPV抗原は、同じおよび/または異なるHPV抗原に対する応答を誘発する多数のポリペプチドのプールである、項目186~203のいずれか一項に記載の組成物。 (項目205) 多数の抗原の前記プール中の抗原は、多数の抗原の前記プール中の他の抗原に向けられた前記免疫応答を減少させない、項目204に記載の組成物。 (項目206) 前記HPV抗原は、抗原性HPVエピトープおよび1つまたは複数の異種ペプチド配列を含むポリペプチドである、項目186~205のいずれか一項に記載の組成物。 (項目207) 前記HPV抗原は、それ自体と、他の抗原と、アジュバントと、または前記CpG ODNと複合体を形成する、項目186~206のいずれか一項に記載の組成物。 (項目208) 前記HPV抗原は、HLA-A2特異的エピトープで構成される、項目186~207に記載の組成物。 (項目209) 前記HPV抗原は、1つまたは複数の異種ペプチド配列がそのN末端および/またはそのC末端で隣接している抗原性エピトープを含むポリペプチドである、項目186~208のいずれか一項に記載の組成物。 (項目210) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記CpG ODNを含む、項目186~209のいずれか一項に記載の組成物。 (項目211) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記HPV抗原を含む、項目186~210のいずれか一項に記載の組成物。 (項目212) 前記HPV抗原の前記CpG ODNに対する比は、約10000:1~約1:10000である、項目186~211のいずれか一項に記載の組成物。 (項目213) 改変免疫細胞を含む組成物であって、前記改変免疫細胞は、HPV抗原を含み、前記HPV抗原は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含む、組成物。 (項目214) 前記HPV抗原は、配列番号23と少なくとも90%の類似性を有するアミノ酸配列を含む、項目213に記載の組成物。 (項目215) 前記改変免疫細胞は、 a)インプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原が通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;および b)前記HPV抗原が前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原とをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、項目213または214に記載の組成物。 (項目216) 前記狭窄の前記直径は、前記細胞の前記直径未満である、項目215に記載の組成物。 (項目217) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約99%である、項目215~216のいずれか一項に記載の組成物。 (項目218) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約60%未満である、項目215~217のいずれか一項に記載の組成物。 (項目219) 前記狭窄は、チャネル中にある、項目215~218のいずれか一項に記載の組成物。 (項目220) 前記インプット細胞に、それが前記狭窄を通過するときに変形力が印加される、項目215~219のいずれか一項に記載の組成物。 (項目221) アジュバントをさらに含む、項目213~220のいずれか一項に記載の組成物。 (項目222) 前記HPV抗原および/または前記アジュバントは、細胞質基質および/またはエンドソームに存在する、項目213~221のいずれかに記載の組成物。 (項目223) 前記抗原および/またはアジュバントは、前記細胞の多数の区画に存在する、項目213~222のいずれか一項に記載の組成物。 (項目224) 前記改変免疫細胞は、前記細胞の表面に、HPV抗原および/またはアジュバントをさらに含む、項目213~223のいずれか一項に記載の組成物。 (項目225) 前記摂動インプット細胞とインキュベートされるアジュバントの濃度は、約0.1μM~約1mMである、項目215~224のいずれか一項に記載の組成物。 (項目226) 前記摂動インプット細胞とインキュベートされるHPV抗原の濃度は、約0.1μM~約1mMである、項目215~225のいずれか一項に記載の組成物。 (項目227) 前記摂動インプット細胞とインキュベートされるHPV抗原のアジュバントに対する比は、約10000:1~約1:10000である、項目215~226のいずれか一項に記載の組成物。 (項目228) 前記アジュバントは、CpG ODN、IFN-α、STINGアゴニスト、RIG-Iアゴニスト、またはポリI:Cである、項目213~227のいずれか一項に記載の組成物。 (項目229) 前記アジュバントはCpG ODNである、項目228に記載の組成物。 (項目230) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目229に記載の組成物。 (項目231) 前記改変免疫細胞は1つよりも多いアジュバントを含む、項目213~230のいずれか一項に記載の組成物。 (項目232) 前記HPV抗原は、同じおよび/または異なるHPV抗原に対する応答を誘発する多数のポリペプチドのプールである、項目213~231のいずれか一項に記載の組成物。 (項目233) 多数の抗原の前記プール中の抗原は、多数の抗原の前記プール中の他の抗原に向けられた前記免疫応答を減少させない、項目232に記載の組成物。 (項目234) 前記HPV抗原は、抗原性HPVエピトープおよび1つまたは複数の異種ペプチド配列を含むポリペプチドである、項目213~233のいずれか一項に記載の組成物。 (項目235) 前記HPV抗原は、それ自体と、他の抗原と、または前記アジュバントと複合体を形成する、項目213~234のいずれか一項に記載の組成物。 (項目236) 前記HPV抗原は、HLA-A2特異的エピトープで構成される、項目213~235に記載の組成物。 (項目237) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記アジュバントを含む、項目213~236のいずれか一項に記載の組成物。 (項目238) 前記改変免疫細胞は、約0.1μM~約1mMの濃度で前記HPV抗原を含む、項目213~237のいずれか一項に記載の組成物。 (項目239) 前記HPV抗原の前記アジュバントに対する比は、約10000:1~約1:10000である、項目213~238のいずれか一項に記載の組成物。 (項目240) 前記HPV抗原は、MHCクラスI拘束性ペプチドにプロセシングされ得る、項目186~239のいずれか一項に記載の組成物。 (項目241) 前記HPV抗原は、MHCクラスII拘束性ペプチドにプロセシングされ得る、項目186~240のいずれか一項に記載の組成物。 (項目242) 前記改変免疫細胞は、作用物質をさらに含み、前記作用物質は、前記作用物質を含まない対応する改変免疫細胞と比較して前記改変免疫細胞の生存能力および/または機能を増強する、項目186~241のいずれか一項に記載の組成物。 (項目243) 前記作用物質は、エンドサイトーシスを増強する化合物、安定剤、または補因子である、項目242に記載の組成物。 (項目244) 前記作用物質はアルブミンである、項目242に記載の組成物。 (項目245) 前記アルブミンは、マウス、ウシ、またはヒトアルブミンである、項目244に記載の組成物。 (項目246) 前記作用物質は、二価金属カチオン、グルコース、ATP、カリウム、グリセロール、トレハロース、D-スクロース、PEG1500、L-アルギニン、L-グルタミン、またはEDTAである、項目242に記載の組成物。 (項目247) 前記作用物質はMSAを含む、項目242に記載の組成物。 (項目248) 前記細胞は、共刺激分子のうちの1つまたは複数の発現を増加させるようにさらに改変されている、項目186~247のいずれか一項に記載の組成物。 (項目249) 前記共刺激分子は、B7-H2(ICOSL)、B7-1(CD80)、B7-2(CD86)、CD70、LIGHT、HVEM、CD40、4-1BBL、OX40L、TL1A、GITRL、CD30L、TIM4、SLAM、CD48、CD58、CD155、またはCD112である、項目248に記載の組成物。 (項目250) 前記細胞は、前記1つまたは複数の共刺激分子の増加した発現をもたらす核酸を含む、項目248または249に記載の組成物。 (項目251) 前記免疫細胞は、T細胞、樹状細胞、単球、マクロファージ、骨髄性細胞、顆粒球、好中球、マスト細胞、ナチュラルキラー細胞、自然リンパ球、好塩基球、または造血系前駆細胞である、項目186~250のいずれか一項に記載の組成物。 (項目252) 前記免疫細胞はB細胞ではない、項目186~251のいずれか一項に記載の組成物。 (項目253) 前記免疫細胞はT細胞である、項目186~252のいずれか一項に記載の組成物。 (項目254) 前記T細胞は、MHCクラスI発現をモジュレートするさらなる改変を含む、項目253に記載の組成物。 (項目255) 前記T細胞は、MHCクラスII発現をモジュレートするさらなる改変を含む、項目253に記載の組成物。 (項目256) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を低下させるさらなる改変を含む、項目254または255に記載の組成物。 (項目257) 前記さらなる改変は、siRNA、shRNA、CRISPR/Cas9、ZFN、TALEN、Creリコンビナーゼ、またはメガヌクレアーゼを使用して、MHCクラスIおよび/またはMHCクラスII発現を低下させることを含む、項目254または255に記載の組成物。 (項目258) 前記T細胞は、MHCクラスIおよび/またはMHCクラスII発現を増加させるさらなる改変を含む、項目254または255に記載の組成物。 (項目259) 前記さらなる改変は、RNAまたはプラスミドDNAを使用して、MHCクラスIおよび/またはMHCクラスII発現を増加させることを含む、項目254または255に記載の組成物。 (項目260) 前記さらなる改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答は、前記さらなる改変を含まない対応する改変T細胞の同種異系の状況における投与に応答した個体において開始された自然免疫応答と比較して低下している、項目253~257のいずれか一項に記載の組成物。 (項目261) 前記さらなる改変T細胞の、それらが投与された個体における循環半減期は、前記さらなる改変を含まない対応する改変T細胞の、それらが投与された個体における循環半減期と比較してモジュレートされる、項目253~257のいずれか一項に記載の組成物。 (項目262) 前記T細胞は、ヘルパーT細胞、細胞傷害性T細胞、メモリーT細胞、CIK細胞、およびナチュラルキラーT細胞のうちの1つまたは複数を含む、項目253~261のいずれか一項に記載の組成物。 (項目263) 前記T細胞は、CD3+ T細胞、CD4+ T細胞、CD8+ T細胞、CD45RA+ T細胞、CD45RO+ T細胞、およびγδ-T細胞のうちの1つまたは複数を含む、項目253~261のいずれか一項に記載の組成物。 (項目264) 前記改変細胞は個体に対して同種異系である、項目186~263のいずれか一項に記載の組成物。 (項目265) 前記改変細胞は個体に対して自家である、項目186~263のいずれか一項に記載の組成物。 (項目266) 個体は、モジュレートされた炎症および/またはモジュレートされた免疫応答を有するよう事前調整されている、項目186~265のいずれか一項に記載の組成物。 (項目267) 免疫チェックポイント阻害剤をさらに含む、項目186~266のいずれか一項に記載の組成物。 (項目268) 前記免疫チェックポイント阻害剤は、PD-1、PD-L1、CTLA-4、LAG3、TIM-3、TIGIT、VISTA、TIM1、B7-H4(VTCN1)、またはBTLAのうちの1つまたは複数を標的にする、項目267に記載の組成物。 (項目269) 個体への前記改変免疫細胞を含む前記組成物の投与は、前記HPV抗原に特異的な細胞傷害性Tリンパ球(CTL)の活性化および/または増殖をもたらす、項目186~268のいずれか一項に記載の組成物。 (項目270) 個体への前記改変免疫細胞を含む前記組成物の投与は、前記抗原に特異的なヘルパーT(Th)細胞の活性化および/または増殖をもたらす、項目186~268のいずれか一項に記載の組成物。 (項目271) 前記有効量の前記組成物は、約1×106~約1×1012個の改変免疫細胞を含む、項目186~270のいずれか一項に記載の組成物。 (項目272) 抗原を含む組成物であって、前記抗原は、配列番号23と少なくとも90%の類似性を有するアミノ酸配列を含む、組成物。 (項目273) 前記抗原は配列番号23のアミノ酸配列を含む、項目272に記載の組成物。 (項目274) 個体におけるHPV関連疾患を処置または予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されており、 前記改変免疫細胞は、 a)HPV抗原を含むインプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記抗原および前記アジュバントが通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;ならびに b)前記アジュバントが前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記アジュバントとをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目275) 個体におけるHPV関連疾患を処置または予防するための方法であって、前記方法は、改変免疫細胞を含む有効量の組成物を前記個体に投与するステップを含み、前記改変免疫細胞は、HPV抗原およびアジュバントを含み、前記アジュバントは細胞内に供給されており、 前記改変免疫細胞は、 a)前記アジュバントを含むインプット細胞を含む細胞懸濁液を細胞変形狭窄に通すステップであって、前記狭窄の直径は、前記懸濁液中の前記インプット細胞の直径の関数であり、それによって、前記HPV抗原が通過するのに十分に大きな前記インプット細胞の摂動を引き起こして摂動インプット細胞を形成するステップ;および b)前記HPV抗原が前記摂動インプット細胞に入るのを可能にするのに十分な時間、前記摂動インプット細胞と前記HPV抗原とをインキュベートし、それによって、前記改変免疫細胞を作出するステップ によって調製される、方法。 (項目276) 前記狭窄の前記直径は、前記細胞の前記直径未満である、項目274または275に記載の方法。 (項目277) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~99%である、項目274~276のいずれか一項に記載の方法。 (項目278) 前記狭窄の前記直径は、前記細胞の前記直径の約20%~約60%未満である、項目274~277のいずれか一項に記載の方法。 (項目279) 前記狭窄は、チャネル中にある、項目274~278のいずれか一項に記載の方法。 (項目280) 前記インプット細胞に、それが前記狭窄を通過するときに変形力が印加される、項目274~279のいずれか一項に記載の方法。 (項目281) 前記HPV抗原および/または前記アジュバントは、細胞質基質および/またはエンドソームに存在する、項目274~280のいずれかに記載の方法。 (項目282) 前記抗原および/またはアジュバントは、前記細胞の多数の区画に存在する、項目274~281のいずれか一項に記載の方法。 (項目283) 前記摂動インプット細胞とインキュベートされるアジュバントの濃度は、約0.1μM~約1mMである、項目274に記載の方法。 (項目284) 前記摂動インプット細胞とインキュベートされるHPV抗原の濃度は、約0.1μM~約1mMである、項目275に記載の方法。 (項目285) 前記アジュバントは、CpG ODN、IFN-α、STINGアゴニスト、RIG-Iアゴニスト、またはポリI:Cである、項目274~285のいずれか一項に記載の方法。 (項目286) 前記アジュバントはCpG ODNである、項目285に記載の方法。 (項目287) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目286に記載の方法。 (項目288) 前記HPV抗原は細胞溶解物に由来する、項目274~287のいずれか一項に記載の方法。 (項目289) 前記HPV抗原は、HPV-16またはHPV-18抗原である、項目274~288のいずれか一項に記載の方法。 (項目290) 前記HPV抗原は、HPV E6抗原またはHPV E7抗原である、項目274~289のいずれか一項に記載の方法。 (項目291) 前記HPV抗原は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含む、項目290に記載の方法。 (項目292) 前記HPV抗原は、配列番号18~25のいずれか1つのアミノ酸配列を含む、項目289に記載の方法。 (項目293) 前記HPV抗原は、配列番号23と少なくとも90%の類似性を有するアミノ酸配列を含む、項目290に記載の方法。 (項目294) 前記HPV抗原は配列番号23のアミノ酸配列を含む、項目290に記載の方法。 (項目295) 個体におけるHPV関連疾患を処置または予防するための方法であって、HPV抗原と会合した改変免疫細胞を前記個体に投与するステップを含み、前記改変免疫細胞は、 a)前記HPV抗原がインプット細胞と会合するのを可能にするのに十分な時間、前記インプット細胞と前記HPV抗原および/またはアジュバントとをインキュベートし、それによって、前記抗原と会合した前記改変免疫細胞を作出するステップ を含むプロセスによって調製される、方法。 (項目296) 前記HPV抗原は、配列番号18~25のいずれか1つと少なくとも90%の類似性を有するアミノ酸配列を含む、項目295に記載の方法。 (項目297) 前記HPV抗原は配列番号23のアミノ酸配列を含む、項目296に記載の方法。 (項目298) 前記アジュバントはCpG ODNである、項目295~297のいずれか一項に記載の方法。 (項目299) 前記CpG ODNは、CpG ODN1018、CpG ODN1826、またはCpG ODN2006である、項目298に記載の方法。 (項目300) 医薬としての使用のための、項目186~273のいずれか一項に記載の改変免疫細胞を含む組成物。 (項目301) 手術、治療、または診断による、ヒトまたは動物身体の処置の方法における使用のための、項目186~273のいずれか一項に記載の改変免疫細胞を含む組成物。 (項目302) がん、感染性疾患、またはウイルス関連疾患の処置における使用のための、項目186~273のいずれか一項に記載の改変免疫細胞を含む組成物。 (項目303) 前記がんは、頭頸部がん、子宮頸がん、外陰がん、膣がん、陰茎がん、肛門がん、肛門周囲がん、肛門性器がん、口腔がん、または唾液腺がんである、項目186~273のいずれか一項に記載の改変免疫細胞を含む組成物。 (項目304) 前記改変PBMCは、免疫チェックポイント阻害剤の投与の前、それと並行して、または後に投与される、項目300~303のいずれか一項に記載の改変免疫細胞を含む組成物。 (項目305) 前記免疫チェックポイント阻害剤は、PD-1、PD-L1、CTLA-4、LAG3、VISTA、およびTIM-3のいずれか1つを標的にする、項目304に記載の組成物。 (項目306) 前記免疫チェックポイント阻害剤はPD-1を標的にする、項目305に記載の組成物。 (項目307) 前記免疫チェックポイント阻害剤はPD-L1を標的にする、項目305に記載の組成物。 (項目308) 前記改変PBMCは、治療剤の投与の前、それと並行して、または後に投与される、項目300~307のいずれか一項に記載の組成物。 (項目309) 前記治療剤は化学療法剤である、項目308に記載の組成物。 (項目310) 前記感染性疾患は、HIV、HPV、EBV、MCV、HBV、またはHCVと関連する、項目309に記載の組成物。

✦ Generated by Eureka AI based on patent content.

Smart Images

  • Figure 0007905173000019
    Figure 0007905173000019
  • Figure 0007905173000020
    Figure 0007905173000020
  • Figure 0007905173000021
    Figure 0007905173000021
Patent Text Reader

Abstract

The present application provides immune cells comprising HPV antigens and adjuvants, methods of making such modified immune cells, and methods of using such modified immune cells to treat HPV-associated disease, prevent HPV-associated disease, and / or modulate immune responses in individuals with HPV-associated disease. In some aspects, the present invention provides a method for treating human papillomavirus (HPV)-associated disease in an individual, the method comprising administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise HPV antigens and an adjuvant, and the adjuvant is provided intracellularly.
Need to check novelty before this filing date? Find Prior Art

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to U.S. Provisional Patent Application No. 62 / 641,988 filed on 12 March 2018, U.S. Provisional Patent Application No. 62 / 794,517 filed on 18 January 2019, and U.S. Provisional Patent Application No. 62 / 812,225 filed on 28 February 2019. This application also claims benefit to European Patent Application No. EP19161964.2 filed on 11 March 2019, and these in whole are thus incorporated herein by reference.

[0002] Submission of sequence listings in ASCII text files The contents of the following submission in ASCII text file are incorporated herein by reference in their entirety: Sequence listing in computer-readable format (CRF) (filename: 750322001640SEQLIST.TXT, date recorded: March 11, 2019, size: 14KB).

[0003] Field of Invention This disclosure generally relates to immune cells comprising antigens and adjuvants, methods for producing such modified immune cells, and methods for using such modified immune cells to treat HPV-related diseases, to prevent HPV-related diseases, and to modulate the immune response in individuals having HPV-related diseases. [Background technology]

[0004] Background of the Invention Papillomaviruses are small, non-enveloped DNA viruses with a virion size of approximately 55 nm in diameter. More than 100 HPV genotypes have been fully characterized, and it is estimated that many more exist. HPV is a known cause of cervical cancer, as well as some vulvar, vaginal, penile, oropharyngeal, anal, and rectal cancers. Most HPV infections are asymptomatic and resolve spontaneously, but persistent infection with one of the tumorigenic HPV types can progress to a precancerous condition or cancer. Other HPV-related diseases include verruca vulgaris, plantar warts, flat warts, anogenital warts, anal lesions, epidermal dysplasia, focal epithelial thickening, oral papillomas, verrucous cysts, laryngeal papillomatosis, squamous intraepithelial lesions (SILs), cervical intraepithelial neoplasia (CIN), vulvar intraepithelial neoplasia (VIN), and vaginal intraepithelial neoplasia (VAIN).

[0005] Many known human papillomavirus (HPV) types cause benign lesions due to a subset that is tumorigenic. Based on epidemiological and phylogenetic relationships, HPV types are classified into 15 "high-risk types" (HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82) and 3 "probably high-risk types" (HPV26, 53, and 66), which together are known to manifest as low- and high-grade cervical changes and cancers, as well as other anogential cancers such as vulvar, vaginal, penile, anal and perianal cancers, and head and neck cancers. Recently, associations between high-risk types HPV16 and 18 and breast cancer have also been described. Eleven HPV types classified as "low-risk" (HPV6, 11, 40, 42, 43, 44, 54, 61, 70, 72, and 81) are known to manifest as benign, low-grade cervical changes, genital warts, and recurrent respiratory papillomatosis. Cutaneous HPV types 5, 8, and 92 are associated with skin cancer. In some HPV-associated cancers, the immune system is suppressed, and consequently, the antitumor response is significantly impaired. See Suresh and Burtness Am J Hematol Oncol 13(6):20-27 (2017).

[0006] Immunotherapy can be divided into two main types of interventions: passive and active. Passive protocols include the administration of pre-activated and / or engineered cells (e.g., CAR T cells), disease-specific therapeutic antibodies, and / or cytokines. Active immunotherapy strategies target stimulating immune system effector function in vivo. Several current active protocols include vaccination strategies with disease-related peptides, lysates, or whole allogeneic cells, infusion of autologous DCs as vehicles for tumor antigen delivery, and infusion of immune checkpoint modulators. See Papaioannou, Nikos E., et al. Annals of Translational Medicine 4.14 (2016). Adoptive immunotherapy can be used to achieve the goals of modulating the immune response, enhancing antitumor activity, and treating or preventing HPV-related cancers. CD8 stimulated by disease-associated antigens + Cytotoxic T lymphocytes (CTLs) and CD4 + Helper T (Th) cells have the potential to target and destroy infected cells. The method described herein is used to produce modified immune cells de novo in a high-throughput and efficient manner, thereby inducing a robust T cell response to HPV antigens. All references cited herein, including patent applications and publications, are incorporated by reference in their entirety. Patent publications WO2017041050 and WO2016070136 are thus expressly incorporated herein by reference in their entirety. [Prior art documents] [Patent Documents]

[0007] [Patent Document 1] International Publication No. 2017 / 041050 [Patent Document 2] International Publication No. 2016 / 070136 [Non-patent literature]

[0008] [Non-Patent Document 1] Suresh and Burtness Am J Hematol Oncol 13(6):20-27 (2017) [Non-Patent Document 2] Papaioannou, Nikos E., et al. Annals of translational medicine 4.14 (2016) [Overview of the project] [Means for solving the problem]

[0009] Brief summary of the invention In some embodiments, the present invention provides a method for treating human papillomavirus (HPV)-related disease in an individual, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen and an adjuvant, and the adjuvant is supplied intracellularly. In some embodiments, the present invention provides a method for preventing HPV-related disease in an individual, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen and an adjuvant, and the adjuvant is supplied intracellularly. In some embodiments, the present invention provides a method for modulating the immune response in an individual having an HPV-related disease, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen and an adjuvant, and the adjuvant is supplied intracellularly.

[0010] In some embodiments, the present invention provides a method for treating an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells through a cell-deforming constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the present invention provides a method for preventing HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells.In some embodiments, the present invention provides a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells comprising an HPV antigen through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the diameter of the constriction is less than the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to 99% of the diameter of the cells. In some embodiments, the diameter of the constriction is approximately 20% to less than 60% of the cell diameter. In some embodiments, the constriction is located within the channel. In some embodiments, a deformable force is applied to the input cell as it passes through the constriction.

[0011] In some embodiments, the HPV antigen and / or adjuvant is present in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or adjuvant is present in multiple compartments of the cell. In some embodiments, the modified immune cells further contain the HPV antigen and / or adjuvant outside the cell. In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is about 0.1 μM to about 1 mM. In some embodiments, the concentration of the HPV antigen incubated with the perturbed input cells is about 0.1 μM to about 1 mM. In some embodiments, the ratio of the HPV antigen to the adjuvant incubated with the perturbed input cells is about 10000:1 to about 1:10000.

[0012] In some embodiments, the immune response is enhanced. In some embodiments, the immune response to HPV antigens is enhanced.

[0013] In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. In some embodiments, the adjuvant is CpG ODN. In some embodiments, the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. In some embodiments, the modified immune cells contain more than one adjuvant.

[0014] In some embodiments, the HPV antigen is a pool of numerous polypeptides that elicit responses to the same and / or different HPV antigens.

[0015] In some embodiments, an antigen in a pool of multiple antigens does not reduce the immune response directed towards other antigens in the pool of multiple antigens. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, the HPV antigen forms complexes with itself, with other antigens, or with adjuvants. In some embodiments, the HPV antigen is an HPV-16 or HPV-18 antigen. In some embodiments, the HPV antigen consists of an HLA-A2 specific epitope. In some embodiments, the HPV antigen is an HPV E6 antigen or an HPV E7 antigen. In some embodiments, modified immune cells contain both HPV E6 and HPV E7 antigens. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic epitope flanked by one or more heterologous peptide sequences at its N-terminus and / or C-terminus. In some embodiments, the HLA-A2 restricted peptide comprises one of the amino acid sequences of SEQ ID NOs: 1-4. In some embodiments, the N-terminal facilitation polypeptide comprises one of the amino acid sequences of SEQ ID NOs. 5-10, and / or the C-terminal facilitation polypeptide comprises one of the amino acid sequences of SEQ ID NOs. 11-17. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to one of SEQ ID NOs. 18-26. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO. 23. In some embodiments, the HPV antigen may be processed into an MHC class I-restricted peptide. In some embodiments, the HPV antigen may be processed into an MHC class II-restricted peptide.

[0016] In some embodiments, the modified immune cells contain an adjuvant at a concentration of about 0.1 μM to about 1 mM. In some embodiments, the modified immune cells contain HPV antigen at a concentration of about 0.1 μM to about 1 mM. In some embodiments, the ratio of HPV antigen to adjuvant is about 10,000:1 to about 1:10,000.

[0017] In some embodiments, the modified immune cells further contain an active agent that enhances the viability and / or function of the modified immune cells compared to corresponding modified immune cells that do not contain this active agent. In some embodiments, the active agent is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the active agent is albumin. In some embodiments, the albumin is mouse, bovine, or human albumin. In some embodiments, the active agent is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the active agent contains mouse serum albumin (MSA). In some embodiments, the modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules. In some embodiments, the co-stimulatory molecules are B7-H2, B7-1, B7-2, CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. In some embodiments, the cells contain nucleic acids that result in increased expression of one or more co-stimulatory molecules.

[0018] In some embodiments, the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, the immune cells are not B cells.

[0019] In some embodiments, the immune cells are T cells. In some embodiments, the T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using RNA or plasmid DNA to increase MHC class I and / or MHC class II expression. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting. In some embodiments, the circulating half-life of further modified T cells in the administered individual is modulated compared to the circulating half-life of corresponding modified T cells without further modification in the administered individual. In some embodiments, the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. In some embodiments, the modified cells are allogeneic to the individual. In some embodiments, the modified cells are autologous to the individual. In some embodiments, the individual is pre-conditioned to have modulated inflammation and / or a modulated immune response.

[0020] In some embodiments, the method further includes the step of administering an adjuvant to an individual. In some embodiments, the adjuvant is IFNα or CpG ODN. In some embodiments, the composition containing modified immune cells and the adjuvant are administered simultaneously. In some embodiments, the composition containing modified immune cells and the adjuvant are administered sequentially. In some embodiments, the composition containing modified immune cells is administered before the administration of the adjuvant. In some embodiments, the composition containing modified immune cells is administered after the administration of the adjuvant.

[0021] In some embodiments, the composition containing modified immune cells is administered in combination with the administration of an immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered simultaneously. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered sequentially. In some embodiments, the composition containing modified immune cells is administered before the administration of the immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells is administered after the administration of the immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA.

[0022] In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of antigen-specific helper T(T) lymphocytes. h ) This leads to the activation and / or proliferation of cells.

[0023] In some embodiments, an effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12The method includes multiple doses of the composition containing the modified immune cells. In some embodiments, the method includes a first dose of the composition containing the modified immune cells, followed by a second dose of the composition containing the modified immune cells. In some embodiments, the second dose is approximately one month after the first dose.

[0024] In some embodiments, HPV-related disease is HPV-related cancer. In some embodiments, HPV-related cancer is cervical cancer, anal cancer, oropharyngeal cancer, vaginal cancer, vulvar cancer, penile cancer, skin cancer, or head and neck cancer. In some embodiments, HPV-related disease is HPV-related infectious disease.

[0025] In some embodiments, the present invention provides a method for treating human papillomavirus (HPV)-related disease in an individual, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. In some embodiments, the present invention provides a method for preventing HPV-related disease in an individual, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. In some embodiments, the present invention provides a method for modulating an immune response in an individual having HPV-related disease, the method comprising the step of administering an effective amount of a composition containing modified immune cells to the individual, wherein the modified immune cells contain an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. In some embodiments, the modified immune cells contain an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. In some embodiments, the modified immune cells contain an HPV antigen comprising the amino acid sequence of SEQ ID NO: 23.

[0026] In some embodiments, the present invention provides a method for treating an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-25, the modified immune cells being prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the present invention provides a method for preventing HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen, the modified immune cells comprising an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-25, and the modified immune cells are prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells.In some embodiments, the present invention provides a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen comprising an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-25, and the modified immune cells are prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the modified immune cells comprise an HPV antigen comprising an amino acid sequence comprising any one of SEQ ID NOs: 18-25. In some embodiments, the modified immune cells comprise an HPV antigen comprising the amino acid sequence of SEQ ID NO: 23. In some embodiments, the diameter of the constriction is less than the diameter of the cell. In some embodiments, the diameter of the constriction is about 20% to 99% of the cell diameter. In some embodiments, the diameter of the constriction is about 20% to less than 60% of the cell diameter. In some embodiments, the constriction is located within the channel. In some embodiments, a deformable force is applied to the input cell as it passes through the constriction.

[0027] In some embodiments, the method further includes the step of administering an adjuvant to an individual. In some embodiments, the adjuvant is IFNα or CpG ODN. In some embodiments, the composition containing modified immune cells and the adjuvant are administered simultaneously. In some embodiments, the composition containing modified immune cells and the adjuvant are administered sequentially. In some embodiments, the composition containing modified immune cells is administered before the administration of the adjuvant. In some embodiments, the composition containing modified immune cells is administered after the administration of the adjuvant. In some embodiments, the modified immune cells further comprise the adjuvant. In some embodiments, the perturbed immune cells of step b are incubated with the HPV antigen and adjuvant. In some embodiments, the HPV antigen and / or adjuvant reside in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or adjuvant reside in multiple compartments of the cell. In some embodiments, the modified immune cells further comprise the HPV antigen and / or adjuvant outside the cell.

[0028] In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. In some embodiments, the concentration of the HPV antigen incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. In some embodiments, the ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is approximately 10,000:1 to approximately 1:10,000.

[0029] In some embodiments, the immune response is enhanced. In some embodiments, the immune response to HPV antigens is enhanced.

[0030] In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. In some embodiments, the adjuvant is CpG ODN. In some embodiments, the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. In some embodiments, the modified immune cells contain more than one adjuvant.

[0031] In some embodiments, the HPV antigen is a pool of multiple polypeptides that elicit responses to the same and / or different HPV antigens. In some embodiments, antigens in the pool of multiple antigens do not diminish the immune response directed to other antigens in the pool of multiple antigens. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, the HPV antigen forms complexes with itself, with other antigens, or with adjuvants. In some embodiments, the HPV antigen is the HPV-16 or HPV-18 antigen. In some embodiments, the HPV antigen consists of an HLA-A2 specific epitope. In some embodiments, the HPV antigen is the HPV E6 antigen or the HPV E7 antigen. In some embodiments, modified immune cells contain both the HPV E6 and HPV E7 antigens. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic epitope flanked by one or more heterologous peptide sequences at its N-terminus and / or C-terminus. In some embodiments, the HPV antigen can be processed into an MHC class I-restricted peptide. In some embodiments, the HPV antigen can be processed into an MHC class II-restricted peptide.

[0032] In some embodiments, the modified immune cells contain an adjuvant at a concentration of approximately 0.1 μM to approximately 1 mM. In some embodiments, the modified immune cells contain HPV antigen at a concentration of approximately 0.1 μM to approximately 1 mM. In some embodiments, the ratio of HPV antigen to adjuvant is approximately 10,000:1 to approximately 1:10,000.

[0033] In some embodiments, the modified immune cells further contain an active agent that enhances the viability and / or function of the modified immune cells compared to corresponding modified immune cells that do not contain this active agent. In some embodiments, the active agent is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the active agent is albumin. In some embodiments, the albumin is mouse, bovine, or human albumin. In some embodiments, the active agent is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the active agent contains mouse serum albumin (MSA). In some embodiments, the modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules. In some embodiments, the co-stimulatory molecules are B7-H2, B7-1, B7-2, CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. In some embodiments, the cells contain nucleic acids that result in increased expression of one or more co-stimulatory molecules.

[0034] In some embodiments, the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, the immune cells are not B cells.

[0035] In some embodiments, the immune cells are T cells. In some embodiments, the T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using RNA or plasmid DNA to increase MHC class I and / or MHC class II expression. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting. In some embodiments, the circulating half-life of further modified T cells in the administered individual is modulated compared to the circulating half-life of corresponding modified T cells without further modification in the administered individual. In some embodiments, the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. In some embodiments, the modified cells are allogeneic to the individual. In some embodiments, the modified cells are autologous to the individual. In some embodiments, the individual is pre-conditioned to have modulated inflammation and / or a modulated immune response.

[0036] In some embodiments, the method further includes the step of administering an adjuvant to an individual. In some embodiments, the adjuvant is IFNα or CpG ODN. In some embodiments, the composition containing modified immune cells and the adjuvant are administered simultaneously. In some embodiments, the composition containing modified immune cells and the adjuvant are administered sequentially. In some embodiments, the composition containing modified immune cells is administered before the administration of the adjuvant. In some embodiments, the composition containing modified immune cells is administered after the administration of the adjuvant.

[0037] In some embodiments, the composition containing modified immune cells is administered in combination with the administration of an immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered simultaneously. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered sequentially. In some embodiments, the composition containing modified immune cells is administered before the administration of the immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells is administered after the administration of the immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA.

[0038] In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of antigen-specific helper T(T) lymphocytes. h ) This leads to the activation and / or proliferation of cells.

[0039] In some embodiments, an effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12The method includes multiple doses of the composition containing the modified immune cells. In some embodiments, the method includes a first dose of the composition containing the modified immune cells, followed by a second dose of the composition containing the modified immune cells. In some embodiments, the second dose is approximately one month after the first dose.

[0040] In some embodiments, HPV-related disease is HPV-related cancer. In some embodiments, HPV-related cancer is cervical cancer, anal cancer, oropharyngeal cancer, vaginal cancer, vulvar cancer, penile cancer, skin cancer, or head and neck cancer. In some embodiments, HPV-related disease is HPV-related infectious disease.

[0041] In some embodiments, the present invention provides a composition comprising modified immune cells, wherein the modified immune cells contain within the cells a CpG ODN and an HPV antigen having at least 90% similarity to one of SEQ ID NOs. 18-25. In some embodiments, the HPV antigen contains an amino acid sequence having at least 90% similarity to SEQ ID NOs. 23. In some embodiments, the modified immune cells contain both a CpG ODN and an HPV antigen within the cells, and the HPV antigen contains an amino acid sequence from one of SEQ ID NOs. 18-25. In some embodiments, the HPV antigen contains the amino acid sequence of SEQ ID NOs. 23.

[0042] In some embodiments, modified immune cells are prepared by a) passing a cell suspension containing input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells large enough for the HPV antigen and CpG ODN to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and CpG ODN for a time sufficient to allow the HPV antigen and CpG ODN to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the diameter of the constriction is less than the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to about 99% of the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to less than about 60% of the diameter of the cells. In some embodiments, the constriction is located within a channel. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0043] In some embodiments, the composition further comprises an adjuvant. In some embodiments, the HPV antigen and / or CpG ODN are present in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or CpG ODN are present in multiple compartments of the cell. In some embodiments, the modified immune cells further comprise the HPV antigen and / or CpG ODN on the cell surface. In some embodiments, the concentration of CpG ODN incubated with the perturbation input cells is about 0.1 μM to about 1 mM. In some embodiments, the concentration of HPV antigen incubated with the perturbation input cells is about 0.1 μM to about 1 mM. In some embodiments, the ratio of HPV antigen to CpG ODN incubated with the perturbation input cells is about 10000:1 to about 1:10000. In some embodiments, the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. In some embodiments, the modified immune cells contain more than one adjuvant. In some embodiments, the adjuvant contains CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly(I:C).

[0044] In some embodiments, an HPV antigen is a pool of multiple polypeptides that elicit responses to the same and / or different HPV antigens. In some embodiments, antigens in the pool of multiple antigens do not diminish the immune response directed to other antigens in the pool of multiple antigens. In some embodiments, an HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, an HPV antigen forms complexes with itself, with other antigens, with adjuvants, or with CpG ODNs. In some embodiments, an HPV antigen consists of an HLA-A2 specific epitope. In some embodiments, an HPV antigen is a polypeptide comprising an antigenic epitope flanked by one or more heterologous peptide sequences at its N-terminus and / or C-terminus.

[0045] In some embodiments, the modified immune cells contain CpG ODN at a concentration of about 0.1 μM to about 1 mM. In some embodiments, the modified immune cells contain HPV antigen at a concentration of about 0.1 μM to about 1 mM. In some embodiments, the ratio of HPV antigen to CpG ODN is about 10000:1 to about 1:10000.

[0046] In some embodiments, the present invention comprises a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen, and the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18 to 25. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NOs. 23. In some embodiments, the HPV antigen comprises an amino acid sequence of any one of SEQ ID NOs. 18 to 25. In some embodiments, the HPV antigen comprises the amino acid sequence of SEQ ID NOs. 23.

[0047] In some embodiments, modified immune cells are prepared by a) passing a cell suspension containing input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells large enough for the HPV antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the diameter of the constriction is less than the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to about 99% of the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to less than 60% of the diameter of the cells. In some embodiments, the constriction is located within a channel. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0048] In some embodiments, the composition further comprises an adjuvant. In some embodiments, the HPV antigen and / or adjuvant is present in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or adjuvant is present in multiple compartments of the cell. In some embodiments, the modified immune cells further comprise the HPV antigen and / or adjuvant on the cell surface. In some embodiments, the concentration of the adjuvant incubated with the perturbation input cells is about 0.1 μM to about 1 mM. In some embodiments, the concentration of the HPV antigen incubated with the perturbation input cells is about 0.1 μM to about 1 mM. In some embodiments, the ratio of the HPV antigen incubated with the perturbation input cells to the adjuvant is about 10000:1 to about 1:10000. In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. In some embodiments, the adjuvant is CpG ODN. In some embodiments, the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. In some embodiments, the modified immune cells contain more than one adjuvant.

[0049] In some embodiments, the HPV antigen is a pool of multiple polypeptides that elicit responses to the same and / or different HPV antigens. In some embodiments, antigens in the pool of multiple antigens do not diminish the immune response directed to other antigens in the pool of multiple antigens. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, the HPV antigen forms complexes with itself, with other antigens, or with an adjuvant. In some embodiments, the HPV antigen consists of an HLA-A2 specific epitope. In some embodiments, modified immune cells contain the adjuvant at a concentration of about 0.1 μM to about 1 mM. In some embodiments, modified immune cells contain the HPV antigen at a concentration of about 0.1 μM to about 1 mM. In some embodiments, the ratio of HPV antigen to adjuvant is about 10000:1 to about 1:10000. In some embodiments, the HPV antigen can be processed into an MHC class I-restricted peptide. In some embodiments, the HPV antigen can be processed into an MHC class II-restricted peptide.

[0050] In some embodiments, the modified immune cells further contain an active agent that enhances the viability and / or function of the modified immune cells compared to corresponding modified immune cells that do not contain this active agent. In some embodiments, the active agent is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the active agent is albumin. In some embodiments, the albumin is mouse, bovine, or human albumin. In some embodiments, the active agent is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the active agent contains MSA. In some embodiments, the cells are further modified to increase the expression of one or more of the costimulatory molecules. In some embodiments, the co-stimulatory molecules are B7-H2(ICOSL), B7-1(CD80), B7-2(CD86), CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. In some embodiments, the cells contain nucleic acids that result in increased expression of one or more co-stimulatory molecules.

[0051] In some embodiments, the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, the immune cells are not B cells. In some embodiments, the immune cells are T cells. In some embodiments, the T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In some embodiments, further modifications include increasing MHC class I and / or MHC class II expression using RNA or plasmid DNA. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting. In some embodiments, the circulating half-life of further modified T cells in individuals to which they are administered is modulated compared to the circulating half-life of corresponding modified T cells without further modifications in individuals to which they are administered.

[0052] In some embodiments, the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. In some embodiments, the modified cells are allogeneic to the individual. In some embodiments, the modified cells are autologous to the individual. In some embodiments, the individual is pre-conditioned to have a modulated inflammation and / or a modulated immune response.

[0053] In some embodiments, the composition further comprises an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, LAG3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. In some embodiments, administration of the composition comprising modified immune cells to an individual results in activation and / or proliferation of cytotoxic T lymphocytes (CTLs) specific for the HPV antigen. In some embodiments, administration of the composition comprising modified immune cells to an individual results in activation and / or proliferation of helper T (T h ) cells specific for the antigen.

[0054] In some embodiments, an effective amount of the composition comprises from about 1×10 6 to about 1×10 12 modified immune cells. In some embodiments, the antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. In some embodiments, the antigen comprises the amino acid sequence of SEQ ID NO: 23.

[0055] In some embodiments, the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, the immune cells are not B cells. In some embodiments, the immune cells are T cells. In some embodiments, the T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In some embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In some embodiments, further modifications include increasing MHC class I and / or MHC class II expression using RNA or plasmid DNA. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting. In some embodiments, the circulating half-life of further modified T cells in individuals to which they are administered is modulated compared to the circulating half-life of corresponding modified T cells without further modifications in individuals to which they are administered.

[0056] In some embodiments, the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. In some embodiments, the modified cells are allogeneic to the organism. In some embodiments, the modified cells are autologous to the organism. In some embodiments, the organism is pre-conditioned to have modulated inflammation and / or a modulated immune response.

[0057] In some embodiments, the composition further comprises an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, administration of the composition containing modified immune cells to an individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). In some embodiments, administration of the composition containing modified immune cells to an individual results in the activation and / or proliferation of antigen-specific helper T(T) h ) This leads to the activation and / or proliferation of cells.

[0058] In some embodiments, an effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12 It includes modified immune cells. In some embodiments, the antigen includes an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. In some embodiments, the antigen includes the amino acid sequence of SEQ ID NO: 23.

[0059] In some embodiments, the present invention provides a method for treating or preventing an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells comprising the HPV antigen through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells and the adjuvant for a time sufficient to allow the adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the present invention provides a method for treating or preventing an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells comprising the adjuvant through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells.

[0060] In some embodiments, the diameter of the constriction is less than the diameter of the cell. In some embodiments, the diameter of the constriction is about 20% to about 99% of the cell diameter. In some embodiments, the diameter of the constriction is about 20% to less than 60% of the cell diameter. In some embodiments, the constriction is located within the channel. In some embodiments, a deformable force is applied to the input cell as it passes through the constriction.

[0061] In some embodiments, the HPV antigen and / or adjuvant are present in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or adjuvant are present in multiple compartments of the cell. In some embodiments, the concentration of the adjuvant incubated with the perturbation input cells is about 0.1 μM to about 1 mM. In some embodiments, the concentration of the HPV antigen incubated with the perturbation input cells is about 0.1 μM to about 1 mM.

[0062] In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. In some embodiments, the adjuvant is CpG ODN. In some embodiments, CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006.

[0063] In some embodiments, the HPV antigen is HPV-16 or HPV-18 antigen. In some embodiments, the HPV antigen is HPV E6 antigen or HPV E7 antigen. In some embodiments, the HPV antigen includes an amino acid sequence having at least 90% similarity to one of SEQ ID NOs. 18-25. In some embodiments, the HPV antigen includes an amino acid sequence having at least 90% similarity to SEQ ID NOs. 23. In some embodiments, the HPV antigen includes the amino acid sequence of SEQ ID NOs. 23.

[0064] In some embodiments, the present invention provides a method for treating or preventing an HPV-related disease in an individual, comprising the step of administering modified immune cells associated with an HPV antigen to the individual, wherein the modified immune cells are prepared by a process comprising the step of a) incubating input cells with the HPV antigen and / or adjuvant for a time sufficient to allow the HPV antigen to associate with the input cells, thereby producing modified immune cells associated with the antigen. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. In some embodiments, the HPV antigen comprises the amino acid sequence of SEQ ID NO. 23. In some embodiments, the adjuvant is CpG ODN. In some embodiments, the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. In certain embodiments, for example, the following are provided: (Item 1) A method for treating a human papillomavirus (HPV)-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. (Item 2) A method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, and the adjuvant is supplied intracellularly. (Item 3) A method for modulating an immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, and the adjuvant is supplied intracellularly. (Item 4) A method for treating an HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the antigen and the adjuvant to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells with the HPV antigen and the adjuvant for a sufficient time to allow the HPV antigen and the adjuvant to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 5) A method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and the adjuvant to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells with the HPV antigen and the adjuvant for a sufficient time to allow the HPV antigen and the adjuvant to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 6) A method for modulating the immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. The modified immune cells described above are a) a step of passing a cell suspension containing input cells containing HPV antigen through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and the adjuvant to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells with the HPV antigen and the adjuvant for a sufficient time to allow the HPV antigen and the adjuvant to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 7) The method according to any one of items 4 to 6, wherein the diameter of the stenosis is less than the diameter of the cell. (Item 8) The method according to any one of items 4 to 7, wherein the diameter of the stenosis is approximately 20% to 99% of the diameter of the cell. (Item 9) The method according to any one of items 4 to 8, wherein the diameter of the stenosis is about 20% to less than about 60% of the diameter of the cell. (Item 10) The aforementioned stenosis is in the channel as described in any one of items 4 to 9. (Item 11) The method according to any one of items 4 to 10, wherein a deformable force is applied to the input cell as it passes through the stenosis. (Item 12) The HPV antigen and / or the adjuvant present in the cytoplasm and / or endosomes, as described in any of items 1 to 11. (Item 13) The method according to any one of items 1 to 12, wherein the antigen and / or adjuvant is present in multiple compartments of the cell. (Item 14) The modified immune cells further comprise an HPV antigen and / or adjuvant outside the cells, according to any one of items 1 to 13. (Item 15) The method according to any one of items 1 to 14, wherein the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 16) The method according to any one of items 1 to 15, wherein the concentration of HPV antigen incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 17) The method according to any one of items 4 to 16, wherein the ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is approximately 10,000:1 to approximately 1:10,000. (Item 18) The immune response is enhanced, as described in item 3 or 6. (Item 19) The immune response to the HPV antigen is enhanced, as described in item 18. (Item 20) The adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C, as described in any one of items 1 to 19. (Item 21) The adjuvant is CpG ODN, as described in item 20. (Item 22) The method according to item 21, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 23) The modified immune cells contain more than one adjuvant, according to the method described in any one of items 1 to 22. (Item 24) The method according to any one of items 1 to 23, wherein the HPV antigen is a pool of numerous polypeptides that induce a response to the same and / or different HPV antigens. (Item 25) The method according to item 24, wherein the antigens in the pool of multiple antigens do not reduce the immune response directed towards other antigens in the pool of multiple antigens. (Item 26) The method according to any one of items 1 to 25, wherein the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. (Item 27) The method according to any one of items 1 to 26, wherein the HPV antigen forms a complex with itself, with other antigens, or with the adjuvant. (Item 28) The method according to any one of items 1 to 27, wherein the HPV is an antigen derived from cell lysates. (Item 29) The method according to any one of items 1 to 28, wherein the HPV antigen is HPV-16 or HPV-18 antigen. (Item 30) The method described in item 29, wherein the HPV antigen is composed of an HLA-A2 specific epitope. (Item 31) The method according to any one of items 1 to 30, wherein the HPV antigen is HPV E6 antigen or HPV E7 antigen. (Item 32) The modified immune cells are those comprising HPV E6 antigen and HPV E7 antigen, as described in any one of items 1 to 31. (Item 33) The method according to any one of items 1 to 32, wherein the HPV antigen is a polypeptide comprising an antigenic epitope in which one or more heterologous peptide sequences are flanked at its N-terminus and / or C-terminus. (Item 34) The method according to item 33, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of sequence numbers 18-26. (Item 35) The method according to item 34, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. (Item 36) The method according to any one of items 1 to 35, wherein the HPV antigen can be processed into an MHC class I-restricted peptide. (Item 37) The method according to any one of items 1 to 36, wherein the HPV antigen can be processed into an MHC class II-restricted peptide. (Item 38) The modified immune cells contain the adjuvant at a concentration of approximately 0.1 μM to approximately 1 mM, according to the method of any one of items 1 to 37. (Item 39) The modified immune cells contain the HPV antigen at a concentration of approximately 0.1 μM to approximately 1 mM, according to the method described in any one of items 1 to 38. (Item 40) The method according to any one of items 1 to 39, wherein the ratio of the HPV antigen to the adjuvant is approximately 10,000:1 to approximately 1:10,000. (Item 41) The method according to any one of items 1 to 40, wherein the modified immune cells further contain an active substance, the active substance enhances the viability and / or function of the modified immune cells compared to a corresponding modified immune cell that does not contain the active substance. (Item 42) The method according to item 41, wherein the active substance is a compound, stabilizer, or cofactor that enhances endocytosis. (Item 43) The method according to item 41, wherein the active substance is albumin. (Item 44) The method according to item 43, wherein the albumin is mouse, bovine, or human albumin. (Item 45) The method according to item 41, wherein the active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. (Item 46) The method according to item 41, wherein the active substance includes mouse serum albumin (MSA). (Item 47) The method according to any one of items 1 to 46, wherein the modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules. (Item 48) The method according to item 47, wherein the aforementioned co-stimulatory molecule is B7-H2, B7-1, B7-2, CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. (Item 49) The method according to item 47 or 48, wherein the cells contain nucleic acids that result in increased expression of one or more costimulatory molecules. (Item 50) The method according to any one of items 1 to 49, wherein the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. (Item 51) The immune cells described herein are not B cells, and the method described in any one of items 1 to 50. (Item 52) The method according to any one of items 1 to 50, wherein the immune cells are B cells. (Item 53) The method according to any one of items 1 to 51, wherein the immune cells are T cells. (Item 54) The method according to any one of items 1 to 49, wherein the immune cells are a mixed cell population. (Item 55) The immune cells are multiple PBMCs, as described in item 54. (Item 56) The T cells are further modified to modulate MHC class I expression, as described in item 53. (Item 57) The T cells are further modified to modulate MHC class II expression, as described in item 53. (Item 58) The method according to item 56 or 57, wherein the T cells include further modifications that reduce MHC class I and / or MHC class II expression. (Item 59) The further modifications described herein include reducing MHC class I and / or MHC class II expression using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease, as described in item 56 or 57. (Item 60) The method according to item 56 or 57, wherein the T cells include further modifications that increase MHC class I and / or MHC class II expression. (Item 61) The further modifications described herein include increasing MHC class I and / or MHC class II expression using RNA or plasmid DNA, as described in item 56 or 57. (Item 62) The method according to any one of items 53 and 56-59, wherein the innate immune response initiated in individuals in response to administration of the further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of the corresponding modified T cells without the further modification in an allogeneic setting. (Item 63) The method according to any one of items 53 and 56-59, wherein the circulating half-life of the further modified T cells in the administered individual is modulated compared to the circulating half-life of the corresponding modified T cells without the further modification in the administered individual. (Item 64) The method according to any one of items 53 and 56-63, wherein the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. (Item 65) The method according to any one of items 53 and 56-63, wherein the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. (Item 66) The modified cells are allogeneic with respect to the individual, according to any one of items 1 to 65. (Item 67) The method according to any one of items 1 to 65, wherein the modified cells are autologous to the individual. (Item 68) The method according to any one of items 1 to 67, wherein the individual is pre-conditioned to have a modulated inflammation and / or a modulated immune response. (Item 69) The method according to any one of items 1 to 68, further comprising the step of administering an adjuvant to the individual. (Item 70) The method according to item 69, wherein the adjuvant is IFNα or CpG ODN. (Item 71) The method according to item 69 or 70, wherein the composition containing the modified immune cells and the adjuvant are administered simultaneously. (Item 72) The method according to item 69 or 70, wherein the composition containing the modified immune cells and the adjuvant are administered sequentially. (Item 73) The composition comprising the modified immune cells is administered before administering the adjuvant, according to the method of item 72. (Item 74) The composition comprising the modified immune cells is administered after the administration of the adjuvant, according to item 72. (Item 75) The method according to any one of items 1 to 74, wherein the composition comprising the modified immune cells is administered in combination with the administration of an immune checkpoint inhibitor. (Item 76) The method according to item 75, wherein the composition containing the modified immune cells and the immune checkpoint inhibitor are administered simultaneously. (Item 77) The method according to item 75, wherein the composition containing the modified immune cells and the immune checkpoint inhibitor are administered sequentially. (Item 78) The composition comprising the modified immune cells is administered before administering the immune checkpoint inhibitor, according to item 77. (Item 79) The method according to item 77, wherein the composition comprising the modified immune cells is administered after the administration of the immune checkpoint inhibitor. (Item 80) The immune checkpoint inhibitor is one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA, as described in any one of items 75-79. (Item 81) The composition comprising the modified immune cells is administered in combination with the administration of chemotherapy, according to any one of items 1 to 80. (Item 82) The method according to item 81, wherein the composition containing the modified immune cells and the chemotherapy are administered simultaneously. (Item 83) The method according to item 81, wherein the composition containing the modified immune cells and the chemotherapy are administered sequentially. (Item 84) The composition comprising the modified immune cells is administered before administering the chemotherapy according to item 83. (Item 85) The composition comprising the modified immune cells is administered after the administration of the chemotherapy according to item 83. (Item 86) The chemotherapy described above is the method described in any one of items 81 to 85, comprising a platinum-based active agent. (Item 87) The chemotherapy described above includes cisplatin, as described in any one of items 81 to 86. (Item 88) The method according to any one of items 1 to 87, wherein administration of the composition comprising the modified immune cells to the individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). (Item 89) Administration of the composition containing the modified immune cells to the individual results in the antigen-specific helper T(T) h ) The method described in any one of items 1 to 87, which results in the activation and / or proliferation of cells. (Item 90) The effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12 The method described in any one of items 1 to 89, comprising individual modified immune cells. (Item 91) The method according to any one of items 1 to 90, comprising multiple administrations of the composition containing the modified immune cells. (Item 92) The method according to item 91, comprising a first administration of the composition containing the modified immune cells, followed by a second administration of the composition containing the modified immune cells. (Item 93) The method according to item 92, wherein the second dose is administered approximately one month after the first dose. (Item 94) The HPV-related disease is an HPV-related cancer, as described in any one of items 1 to 93. (Item 95) The method according to item 94, wherein the HPV-related cancer is cervical cancer, anal cancer, oropharyngeal cancer, vaginal cancer, vulvar cancer, penile cancer, skin cancer, or head and neck cancer. (Item 96) The HPV-related disease is an HPV-related infectious disease, as described in any one of items 1 to 95. (Item 97) A method for treating a human papillomavirus (HPV)-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having at least 90% similarity to any one of SEQ ID NOs. 18-25. (Item 98) A method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. (Item 99) A method for modulating an immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. (Item 100) A method for treating an HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the antigen to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the HPV antigen for a sufficient time to allow the HPV antigen to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 101) A method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen, and the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the HPV antigen for a sufficient time to allow the HPV antigen to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 102) A method for modulating an immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-25. The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the HPV antigen for a sufficient time to allow the HPV antigen to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 103) The method according to any one of items 100 to 102, wherein the diameter of the stenosis is less than the diameter of the cell. (Item 104) The method according to any one of items 100 to 103, wherein the diameter of the stenosis is approximately 20% to 99% of the diameter of the cell. (Item 105) The method according to any one of items 100 to 104, wherein the diameter of the stenosis is about 20% to less than about 60% of the diameter of the cell. (Item 106) The aforementioned stenosis is the method described in any one of items 100 to 105, which is located in the channel. (Item 107) The method according to any one of items 100 to 106, wherein a deformable force is applied to the input cell as it passes through the stenosis. (Item 108) The method according to any one of items 86 to 107, further comprising the step of administering an adjuvant to the individual. (Item 109) The method according to item 108, wherein the adjuvant is IFNα or CpG ODN. (Item 110) The method according to item 108 or 109, wherein the composition containing the modified immune cells and the adjuvant are administered simultaneously. (Item 111) The method according to item 108 or 109, wherein the composition containing the modified immune cells and the adjuvant are administered sequentially. (Item 112) The method according to item 111, wherein the composition comprising the modified immune cells is administered before administering the adjuvant. (Item 113) The composition comprising the modified immune cells is administered after the administration of the adjuvant, according to item 111. (Item 114) The modified immune cells further comprise an adjuvant, according to the method described in any one of items 97 to 113. (Item 115) The method according to any one of items 100 to 113, wherein the perturbed immune cells of step b are incubated with the HPV antigen and adjuvant. (Item 116) The method according to item 114 or 115, wherein the HPV antigen and / or the adjuvant are present in the cytoplasm and / or endosomes. (Item 117) The antigen and / or adjuvant present in multiple compartments of the cell, as described in any one of items 114 to 116. (Item 118) The modified immune cells further comprise an HPV antigen and / or adjuvant outside the cells, according to any one of items 114 to 117. (Item 119) The method according to any one of items 115 to 118, wherein the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 120) The method according to any one of items 115 to 119, wherein the concentration of HPV antigen incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 121) The method according to any one of items 115 to 120, wherein the ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is approximately 10,000:1 to approximately 1:10,000. (Item 122) The aforementioned immune response is enhanced by the method described in item 99 or 102. (Item 123) The method according to item 122, wherein the immune response to the HPV antigen is enhanced. (Item 124) The adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C, as described in any one of items 114 to 123. (Item 125) The adjuvant is CpG ODN, as described in item 124. (Item 126) The method according to item 125, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 127) The modified immune cells contain more than one adjuvant, as described in any one of items 114-126. (Item 128) The method according to any one of items 97 to 127, wherein the HPV antigen is a pool of numerous polypeptides that induce a response to the same and / or different HPV antigens. (Item 129) The method according to item 128, wherein the antigens in the pool of multiple antigens do not reduce the immune response directed towards other antigens in the pool of multiple antigens. (Item 130) The method according to any one of items 97 to 129, wherein the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. (Item 131) The method according to any one of items 97 to 130, wherein the HPV antigen forms a complex with itself, with other antigens, or with the adjuvant. (Item 132) The HPV antigen is comprised of an HLA-A2 specific epitope, as described in any one of items 97 to 131. (Item 133) The method according to any one of items 97 to 132, wherein the HPV antigen can be processed into an MHC class I-restricted peptide. (Item 134) The method according to any one of items 97 to 133, wherein the HPV antigen can be processed into an MHC class II-restricted peptide. (Item 135) The modified immune cells contain the adjuvant at a concentration of approximately 0.1 μM to approximately 1 mM, according to the method of any one of items 114 to 134. (Item 136) The modified immune cells contain the HPV antigen at a concentration of approximately 0.1 μM to approximately 1 mM, according to the method of any one of items 97 to 135. (Item 137) The method according to any one of items 114 to 136, wherein the ratio of the HPV antigen to the adjuvant is approximately 10,000:1 to approximately 1:10,000. (Item 138) The method according to any one of items 97 to 137, wherein the modified immune cells further comprise an active substance, the active substance enhances the viability and / or function of the modified immune cells compared to a corresponding modified immune cell that does not contain the active substance. (Item 139) The method according to item 138, wherein the active substance is a compound, stabilizer, or cofactor that enhances endocytosis. (Item 140) The method described in item 138, wherein the active substance is albumin. (Item 141) The method according to item 140, wherein the albumin is mouse, bovine, or human albumin. (Item 142) The method according to item 138, wherein the active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. (Item 143) The method described in item 138, wherein the active substance includes MSA. (Item 144) Modified T cells as described in any one of items 97-143, further modified to increase the expression of one or more of the costimulatory molecules. (Item 145) The aforementioned co-stimulatory molecules are B7-H2(ICOSL), B7-1(CD80), B7-2(CD86), CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112, as described in item 144, modified T cells. (Item 146) Modified T cells according to item 144 or 145, comprising nucleic acids that result in increased expression of one or more of the aforementioned costimulatory molecules. (Item 147) The method according to any one of items 97 to 146, wherein the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. (Item 148) The immune cells described herein are not B cells, and the method described in any one of items 97 to 147. (Item 149) The immune cells are B cells, as described in any one of items 97 to 148. (Item 150) The immune cells are T cells, as described in any one of items 97 to 148. (Item 151) The immune cells are a mixed cell population, as described in any one of items 97 to 148. (Item 152) The immune cells are multiple PBMCs, as described in item 151. (Item 153) The T cells are further modified to modulate MHC class I expression, as described in item 150. (Item 154) The T cells are further modified to modulate MHC class II expression, according to the method of item 150. (Item 155) The T cells according to item 153 or 154, further modifications including reducing MHC class I and / or MHC class II expression. (Item 156) The further modifications described herein include reducing MHC class I and / or MHC class II expression using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease, as described in item 153 or 154. (Item 157) The method according to item 153 or 154, wherein the T cells include further modifications that increase MHC class I and / or MHC class II expression. (Item 158) The further modifications described herein include increasing MHC class I and / or MHC class II expression using RNA or plasmid DNA, as described in item 153 or 154. (Item 159) The method according to any one of items 150 and 153-156, wherein the innate immune response initiated in individuals in response to administration of the further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of the corresponding modified T cells in an allogeneic setting without the further modification. (Item 160) The method according to any one of items 150 and 153-156, wherein the circulating half-life of the further modified T cells in the administered individual is modulated compared to the circulating half-life of the corresponding modified T cells without the further modification in the administered individual. (Item 161) The method according to any one of items 150 and 153-160, wherein the T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. (Item 162) The method according to any one of items 150 and 153-160, wherein the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. (Item 163) The method according to any one of items 97 to 162, wherein the modified cells are allogeneic with respect to the individual. (Item 164) The method according to any one of items 97 to 162, wherein the modified cells are autologous to the individual. (Item 165) The method according to any one of items 97 to 164, wherein the individual is pre-conditioned to have a modulated inflammation and / or a modulated immune response. (Item 166) The composition comprising the modified immune cells is administered in combination with the administration of an immune checkpoint inhibitor, according to any one of items 97 to 165. (Item 167) The method according to item 166, wherein the composition containing the modified immune cells and the immune checkpoint inhibitor are administered simultaneously. (Item 168) The method according to item 166, wherein the composition comprising the modified immune cells and the immune checkpoint inhibitor are administered sequentially. (Item 169) The composition comprising the modified immune cells is administered before administering the immune checkpoint inhibitor, according to the method of item 168. (Item 170) The composition comprising the modified immune cells is administered after the administration of the immune checkpoint inhibitor, according to the method of item 168. (Item 171) The immune checkpoint inhibitor is one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA, as described in any one of items 152 to 156. (Item 172) The composition comprising the modified immune cells is administered in combination with the administration of chemotherapy, according to any one of items 97 to 171. (Item 173) The method according to item 172, wherein the composition containing the modified immune cells and the chemotherapy are administered simultaneously. (Item 174) The method according to item 172, wherein the composition containing the modified immune cells and the chemotherapy are administered sequentially. (Item 175) The composition comprising the modified immune cells is administered before administering the chemotherapy according to item 174. (Item 176) The composition comprising the modified immune cells is administered after the administration of the chemotherapy according to item 174. (Item 177) The chemotherapy described herein includes cisplatin and is the method described in any one of items 172 to 176. (Item 178) The method according to any one of items 97 to 177, wherein administration of the composition comprising the modified immune cells to the individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). (Item 179) Administration of the composition containing the modified immune cells to the individual results in the HPV antigen-specific helper T(T) h ) The method described in any one of items 97 to 177, which results in the activation and / or proliferation of cells. (Item 180) The effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12 The method described in any one of items 97 to 179, comprising individual modified immune cells. (Item 181) The method according to any one of items 97 to 180, comprising multiple administrations of the composition containing the modified immune cells. (Item 182) The method according to item 181, comprising a first dose of the composition containing the modified immune cells, followed by a second dose of the composition containing the modified immune cells. (Item 183) The method according to item 182, wherein the second dose is administered approximately one month after the first dose. (Item 184) The HPV-related disease is an HPV-related cancer, as described in any one of items 97 to 183. (Item 185) The method according to item 184, wherein the HPV-related cancer is cervical cancer, anal cancer, oropharyngeal cancer, vaginal cancer, vulvar cancer, penile cancer, skin cancer, or head and neck cancer. (Item 186) A composition comprising modified immune cells, wherein the modified immune cells contain within the cells an HPV antigen having at least 90% similarity to CpG ODN and any one of SEQ ID NOs. 18-25. (Item 187) The composition according to item 166, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. (Item 188) The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and the CpG ODN to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells with the HPV antigen and the CpG ODN for a sufficient time to allow the HPV antigen and the CpG ODN to enter the perturbation input cells, thereby producing the modified immune cells. The composition described in item 186 or 187, prepared by [the specified method]. (Item 189) The composition according to item 188, wherein the diameter of the stenosis is less than the diameter of the cell. (Item 190) The composition according to item 188 or 189, wherein the diameter of the stenosis is about 20% to about 99% of the diameter of the cell. (Item 191) The composition according to any one of items 188 to 190, wherein the diameter of the stenosis is about 20% to less than about 60% of the diameter of the cell. (Item 192) The aforementioned stenosis is in the channel, and is a composition according to any one of items 188 to 191. (Item 193) The composition according to any one of items 188 to 192, wherein a deformable force is applied to the input cells as they pass through the stenosis. (Item 194) A composition according to any one of items 186 to 193, further comprising an adjuvant. (Item 195) The composition according to any one of items 186-194, wherein the HPV antigen and / or the CpG ODN are present in the cytoplasm and / or endosomes. (Item 196) The antigen and / or the CpG ODN is present in a number of compartments of the cell, as described in any one of items 186 to 195. (Item 197) The modified immune cells further comprise the HPV antigen and / or CpG ODN on the surface of the cells, according to any one of items 186 to 196. (Item 198) The composition according to any one of items 188 to 197, wherein the concentration of CpG ODN incubated with the perturbed input cells is about 0.1 μM to about 1 mM. (Item 199) The composition according to any one of items 188 to 198, wherein the concentration of HPV antigen incubated with the perturbed input cells is about 0.1 μM to about 1 mM. (Item 200) The composition according to any one of items 188 to 199, wherein the ratio of HPV antigen incubated with the perturbed input cells to CpG ODN is about 10,000:1 to about 1:10,000. (Item 201) The composition according to any one of items 186 to 200, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 202) The modified immune cells comprise one or more adjuvants, according to any one of items 186 to 201. (Item 203) The adjuvant is a composition according to item 202, comprising CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. (Item 204) The composition according to any one of items 186 to 203, wherein the HPV antigen is a pool of numerous polypeptides that induce a response to the same and / or different HPV antigens. (Item 205) The composition according to item 204, wherein the antigens in the pool of multiple antigens do not reduce the immune response directed towards other antigens in the pool of multiple antigens. (Item 206) The composition according to any one of items 186 to 205, wherein the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. (Item 207) The composition according to any one of items 186 to 206, wherein the HPV antigen forms a complex with itself, with other antigens, with an adjuvant, or with the CpG ODN. (Item 208) The HPV antigen is a composition described in items 186-207, comprising an HLA-A2 specific epitope. (Item 209) The composition according to any one of items 186 to 208, wherein the HPV antigen is a polypeptide comprising an antigenic epitope in which one or more heterologous peptide sequences are adjacent at its N-terminus and / or C-terminus. (Item 210) The modified immune cells are a composition according to any one of items 186 to 209, comprising the CpG ODN at a concentration of about 0.1 μM to about 1 mM. (Item 211) The modified immune cells are a composition according to any one of items 186 to 210, comprising the HPV antigen at a concentration of about 0.1 μM to about 1 mM. (Item 212) The composition according to any one of items 186 to 211, wherein the ratio of the HPV antigen to the CpG ODN is about 10,000:1 to about 1:10,000. (Item 213) A composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen, and the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18 to 25. (Item 214) The composition according to item 213, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. (Item 215) The modified immune cells described above are a) a step of passing a cell suspension containing input cells through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the HPV antigen for a sufficient time to allow the HPV antigen to enter the perturbation input cells, thereby producing the modified immune cells. The composition described in item 213 or 214, prepared by [the specified method]. (Item 216) The composition according to item 215, wherein the diameter of the stenosis is less than the diameter of the cell. (Item 217) The composition according to any one of items 215 to 216, wherein the diameter of the stenosis is about 20% to about 99% of the diameter of the cell. (Item 218) The composition according to any one of items 215 to 217, wherein the diameter of the stenosis is about 20% to less than about 60% of the diameter of the cell. (Item 219) The aforementioned stenosis is located in the channel and is a composition according to any one of items 215 to 218. (Item 220) The composition according to any one of items 215 to 219, wherein a deformable force is applied to the input cells as they pass through the stenosis. (Item 221) A composition according to any one of items 213 to 220, further comprising an adjuvant. (Item 222) The composition according to any one of items 213 to 221, wherein the HPV antigen and / or the adjuvant is present in the cytoplasm and / or endosomes. (Item 223) The composition according to any one of items 213 to 222, wherein the antigen and / or adjuvant is present in numerous compartments of the cell. (Item 224) The modified immune cells further comprise the HPV antigen and / or adjuvant on the surface of the cells, according to any one of items 213 to 223. (Item 225) The composition according to any one of items 215 to 224, wherein the concentration of the adjuvant incubated with the perturbed input cells is about 0.1 μM to about 1 mM. (Item 226) The composition according to any one of items 215 to 225, wherein the concentration of HPV antigen incubated with the perturbed input cells is about 0.1 μM to about 1 mM. (Item 227) The composition according to any one of items 215 to 226, wherein the ratio of HPV antigen incubated with the perturbed input cells to the adjuvant is about 10,000:1 to about 1:10,000. (Item 228) The composition according to any one of items 213 to 227, wherein the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C. (Item 229) The composition according to item 228, wherein the adjuvant is CpG ODN. (Item 230) The composition according to item 229, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 231) The modified immune cells are a composition according to any one of items 213 to 230, comprising one or more adjuvants. (Item 232) The composition according to any one of items 213 to 231, wherein the HPV antigen is a pool of numerous polypeptides that induce a response to the same and / or different HPV antigens. (Item 233) The composition according to item 232, wherein the antigens in the pool of multiple antigens do not reduce the immune response directed towards other antigens in the pool of multiple antigens. (Item 234) The composition according to any one of items 213 to 233, wherein the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. (Item 235) The composition according to any one of items 213 to 234, wherein the HPV antigen forms a complex with itself, with other antigens, or with the adjuvant. (Item 236) The HPV antigen is a composition described in items 213 to 235, comprising an HLA-A2 specific epitope. (Item 237) The modified immune cells are a composition according to any one of items 213 to 236, comprising the adjuvant at a concentration of about 0.1 μM to about 1 mM. (Item 238) The modified immune cells are a composition according to any one of items 213 to 237, comprising the HPV antigen at a concentration of about 0.1 μM to about 1 mM. (Item 239) The composition according to any one of items 213 to 238, wherein the ratio of the HPV antigen to the adjuvant is about 10,000:1 to about 1:10,000. (Item 240) The composition according to any one of items 186 to 239, wherein the HPV antigen can be processed into an MHC class I-restricted peptide. (Item 241) The composition according to any one of items 186 to 240, wherein the HPV antigen can be processed into an MHC class II-restricted peptide. (Item 242) The composition according to any one of items 186 to 241, wherein the modified immune cells further comprise an active substance, the active substance enhances the viability and / or function of the modified immune cells compared to a corresponding modified immune cell that does not contain the active substance. (Item 243) The composition according to item 242, wherein the active substance is a compound, stabilizer, or cofactor that enhances endocytosis. (Item 244) The composition described in item 242, wherein the active substance is albumin. (Item 245) The composition according to item 244, wherein the albumin is mouse, bovine, or human albumin. (Item 246) The composition according to item 242, wherein the active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. (Item 247) The composition according to item 242, comprising the active substance MSA. (Item 248) The composition according to any one of items 186 to 247, wherein the cells are further modified to increase the expression of one or more of the costimulatory molecules. (Item 249) The composition according to item 248, wherein the aforementioned co-stimulatory molecule is B7-H2(ICOSL), B7-1(CD80), B7-2(CD86), CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. (Item 250) The composition according to item 248 or 249, wherein the cells contain nucleic acids that result in increased expression of one or more costimulatory molecules. (Item 251) The composition according to any one of items 186 to 250, wherein the immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. (Item 252) The composition according to any one of items 186 to 251, wherein the immune cells are not B cells. (Item 253) The composition according to any one of items 186 to 252, wherein the immune cells are T cells. (Item 254) The T cells are further modified to modulate MHC class I expression, as described in item 253. (Item 255) The T cells are part of the composition according to item 253, further modified to modulate MHC class II expression. (Item 256) The T cells are further modified to reduce MHC class I and / or MHC class II expression, as described in item 254 or 255. (Item 257) The further modifications described above include reducing MHC class I and / or MHC class II expression using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease, as described in item 254 or 255. (Item 258) The composition according to item 254 or 255, wherein the T cells include further modifications that increase MHC class I and / or MHC class II expression. (Item 259) The composition according to item 254 or 255, wherein the further modification includes increasing MHC class I and / or MHC class II expression using RNA or plasmid DNA. (Item 260) The composition according to any one of items 253 to 257, wherein the innate immune response initiated in individuals in response to administration of the further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of the corresponding modified T cells without the further modification in an allogeneic setting. (Item 261) The composition according to any one of items 253 to 257, wherein the circulating half-life of the further modified T cells in the administered individual is modulated compared to the circulating half-life of the corresponding modified T cells without the further modification in the administered individual. (Item 262) The composition according to any one of items 253 to 261, wherein the T cells comprise one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells. (Item 263) The composition according to any one of items 253 to 261, wherein the T cells comprise one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. (Item 264) The modified cells are allogeneic with respect to the individual, according to any one of items 186 to 263. (Item 265) The modified cells are autologous to the individual, according to any one of items 186 to 263. (Item 266) The composition according to any one of items 186 to 265, wherein the individual is pre-conditioned to have a modulated inflammation and / or a modulated immune response. (Item 267) A composition according to any one of items 186 to 266, further comprising an immune checkpoint inhibitor. (Item 268) The composition described in item 267, wherein the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. (Item 269) The composition according to any one of items 186 to 268, wherein administration of the composition comprising the modified immune cells to an individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). (Item 270) Administration of the composition containing the modified immune cells to an individual is performed by the antigen-specific helper T(T) h ) A composition according to any one of items 186 to 268 that results in the activation and / or proliferation of cells. (Item 271) The effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12 A composition according to any one of items 186 to 270, comprising individual modified immune cells. (Item 272) A composition comprising an antigen, wherein the antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. (Item 273) The composition according to item 272, comprising the amino acid sequence of Sequence ID No. 23. (Item 274) A method for treating or preventing an HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. The modified immune cells described above are a) a step of passing a cell suspension containing input cells containing HPV antigen through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells large enough for the antigen and the adjuvant to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the adjuvant for a sufficient time to allow the adjuvant to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 275) A method for treating or preventing an HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly. The modified immune cells described above are a) a step of passing a cell suspension containing input cells containing the adjuvant through a cell deformation constriction, wherein the diameter of the constriction is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) Incubating the perturbation input cells and the HPV antigen for a sufficient time to allow the HPV antigen to enter the perturbation input cells, thereby producing the modified immune cells. A method prepared by (Item 276) The method according to item 274 or 275, wherein the diameter of the stenosis is less than the diameter of the cell. (Item 277) The method according to any one of items 274 to 276, wherein the diameter of the stenosis is about 20% to 99% of the diameter of the cell. (Item 278) The method according to any one of items 274 to 277, wherein the diameter of the stenosis is about 20% to less than about 60% of the diameter of the cell. (Item 279) The aforementioned stenosis is in the channel, as described in any one of items 274 to 278. (Item 280) The method according to any one of items 274 to 279, wherein a deformable force is applied to the input cell as it passes through the stenosis. (Item 281) The HPV antigen and / or the adjuvant present in the cytoplasm and / or endosomes, as described in any of items 274-280. (Item 282) The method according to any one of items 274-281, wherein the antigen and / or adjuvant is present in numerous compartments of the cell. (Item 283) The method according to item 274, wherein the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 284) The method according to item 275, wherein the concentration of HPV antigen incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM. (Item 285) The adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C, as described in any one of items 274-285. (Item 286) The adjuvant is CpG ODN, as described in item 285. (Item 287) The method according to item 286, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 288) The method according to any one of items 274 to 287, wherein the HPV antigen is derived from cell lysates. (Item 289) The method according to any one of items 274 to 288, wherein the HPV antigen is HPV-16 or HPV-18 antigen. (Item 290) The method according to any one of items 274 to 289, wherein the HPV antigen is HPV E6 antigen or HPV E7 antigen. (Item 291) The method according to item 290, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of sequence numbers 18-25. (Item 292) The HPV antigen is the method described in item 289, comprising one amino acid sequence from sequence numbers 18 to 25. (Item 293) The method according to item 290, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NO: 23. (Item 294) The HPV antigen is the method described in item 290, comprising the amino acid sequence of SEQ ID NO: 23. (Item 295) A method for treating or preventing an HPV-related disease in an individual, comprising the step of administering modified immune cells associated with an HPV antigen to the individual, wherein the modified immune cells are a) Incubating the input cells with the HPV antigen and / or adjuvant for a sufficient time to allow the HPV antigen to associate with the input cells, thereby producing the modified immune cells associated with the antigen. A method prepared by a process that includes the following. (Item 296) The method according to item 295, wherein the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of sequence numbers 18-25. (Item 297) The method described in item 296, wherein the HPV antigen comprises the amino acid sequence of SEQ ID NO: 23. (Item 298) The adjuvant is CpG ODN, as described in any one of items 295-297. (Item 299) The method according to item 298, wherein the CpG ODN is CpG ODN1018, CpG ODN1826, or CpG ODN2006. (Item 300) A composition comprising modified immune cells as described in any one of items 186 to 273, for use as a pharmaceutical. (Item 301) A composition comprising modified immune cells as described in any one of items 186 to 273, for use in methods of treating human or animal bodies by surgery, therapy, or diagnosis. (Item 302) A composition comprising modified immune cells as described in any one of items 186 to 273, for use in the treatment of cancer, infectious diseases, or virus-related diseases. (Item 303) A composition comprising modified immune cells as described in any one of items 186 to 273, wherein the cancer is head and neck cancer, cervical cancer, vulvar cancer, vaginal cancer, penile cancer, anal cancer, perianal cancer, anogenital cancer, oral cancer, or salivary gland cancer. (Item 304) The modified PBMC is a composition comprising modified immune cells as described in any one of items 300 to 303, which is administered before, concurrently with, or after the administration of an immune checkpoint inhibitor. (Item 305) The immune checkpoint inhibitor is a composition according to item 304, which targets one of PD-1, PD-L1, CTLA-4, LAG3, VISTA, and TIM-3. (Item 306) The composition described in item 305, wherein the immune checkpoint inhibitor targets PD-1. (Item 307) The composition described in item 305, wherein the immune checkpoint inhibitor targets PD-L1. (Item 308) The modified PBMC is a composition according to any one of items 300 to 307, administered before, concurrently with, or after the administration of a therapeutic agent. (Item 309) The aforementioned therapeutic agent is a chemotherapeutic agent, as described in item 308. (Item 310) The composition described in item 309, wherein the infectious disease is associated with HIV, HPV, EBV, MCV, HBV, or HCV. [Brief explanation of the drawing]

[0065] [Figure 1A] Figure 1A shows a typical schematic diagram of the treatment groups and schedule. [Figure 1B]Figure 1B shows tumor growth, measured by the formula ((length × width 2) / 2), compared between mice from the untreated group (no T cell adoptive transplantation) and mice from treatment groups B-E outlined in Figure 1A.

[0066] [Figure 2A] Figure 2A shows a typical schematic diagram for evaluating the E7 antigen. [Figure 2B] Figure 2B shows that the effect of SLP sequences on IFN-γ-producing CD8+ T cells occurred in response to TAPC vaccination.

[0067] [Figure 3] Figure 3 is a graph showing the ability of E6 SLPs to induce antigen-specific immune responses in E6 responder T cells in an in vitro human model.

[0068] [Figure 4] Figure 4 shows the ability of E7 SLP to induce antigen-specific immune responses in E711-20 responder T cells, and the effect of SLP sequences on SQZ T cell APC (Tapc) activation in an in vitro human model.

[0069] [Figure 5] Figure 5 shows the results of a study to evaluate the antigen dose for SQZ T cell APCs in an in vitro human model.

[0070] [Figure 6-1] Figure 6 shows the results of a study to determine donor variability for SQZ T cell APCs in an in vitro human model. [Figure 6-2] Figure 6 shows the results of a study to determine donor variability for SQZ T cell APCs in an in vitro human model.

[0071] [Figure 7A]Figure 7A is a schematic diagram of an experiment to compare the robustness of the immune response using different adjuvants. [Figure 7B] Figure 7B shows the results of experiments to compare the robustness of the immune response using poly(I:C) and CpG ODN.

[0072] [Figure 8A] Figure 8A is a schematic diagram of an experiment evaluating the effect of CpG ODN concentration on the immune response. [Figure 8B] Figure 8B shows the results of an experiment evaluating the effect of CpG ODN concentration on the immune response.

[0073] [Figure 9A] Figure 9A is a schematic diagram of an experiment evaluating the administration schedule of CpG ODNs in the immune response. [Figure 9B] Figure 9B shows the results of an experiment evaluating the administration schedule of CpG ODNs in the immune response.

[0074] [Figure 10A] Figure 10A is a schematic diagram of an experiment to evaluate the intracellular and systemic adjuvant administration combinations for TAPC antitumor function. [Figure 10B] Figure 10B shows the T cell response for each experimental group. [Figure 10C] Figure 10C shows tumor growth for each experimental group. [Figure 10D] Figure 10D shows tumor growth after rechallenge in animals treated with SQZ(E7+CpG) compared to untreated animals.

[0075] [Figure 11A] Figure 11A is a schematic diagram of an experiment to evaluate the effect of combining multiple HPV antigens on the antitumor function of TAPC. [Figure 11B] Figure 11B shows the T cell response for each experimental group. [Figure 11C] Figure 11C shows tumor growth for each experimental group.

[0076] [Figure 12A] Figure 12A shows the results of an experiment evaluating the importance of the administration route of CpG adjuvant for the E7-specific antitumor effect of TAPC. An administration schedule is provided. [Figure 12B] Figure 12B shows the tumor volume over time for individual mice within each treatment group.

[0077] [Figure 13] Figure 13 shows a schematic diagram of an experiment to assess the ability of co-administered adjuvants to induce E7-specific T-cell tumor invasion. The T-cell response is shown in the lower panel.

[0078] [Figure 14A] Figure 14A is a schematic diagram of an experiment to determine the vaccination schedule for both prime and booster doses of TA PC loaded with E7 synthetic long-chain peptide (SLP) + CpG. [Figure 14B] Figure 14B shows tumor growth for each experimental group.

[0079] [Figure 15] Figure 15 shows the results of an experiment demonstrating that SQZ-modified TAPCs can be directly presented to antigens.

[0080] [Figure 16] Figure 16 shows that SQZ delivery of the adjuvant does not significantly alter T cell cytokine levels in vitro.

[0081] [Figure 17] Figure 17 shows that SQZ delivery of antigen+ / - adjuvants does not significantly alter serum cytokine levels in vivo.

[0082] [Figure 18]Figure 18 shows that SQZ delivery of HPV-E7-containing cell lysates to dendritic cells (as APCs), followed by co-culture of SQZ-modified dendritic cells with CD8 T cell responders, results in a more robust T cell response compared to delivery of the same lysates to dendritic cells via endocytosis.

[0083] [Figure 19] Figure 19A shows a typical schematic diagram of an experiment to evaluate the ability of B cells as APCs to induce an endogenous response. Figure 19B shows the level of IFN-γ-positive CD8+ T cells induced by B9-23 challenge in response to OVA-loaded BAPC vaccination. Figure 19C shows the level of IFN-γ-positive CD8+ T cells induced by E7 challenge in response to E7-loaded BAPC vaccination.

[0084] [Figure 20] Figure 20A shows tumor volume over time in an experiment to determine the ability of SQZ-loaded B cells to function as APCs for prophylactic treatment of HPV-associated tumors. Figure 20B shows corresponding survival data over time from prophylactic treatment of B cell APCs for HPV-associated tumors.

[0085] [Figure 21] Figure 21A shows tumor volume over time in an experiment to determine the ability of SQZ-loaded B cells to function as APCs for therapeutic treatment of HPV-associated tumors. Figure 21B shows corresponding survival data over time from therapeutic treatment of B cell APCs for HPV-associated tumors.

[0086] [Figure 22] Figure 22A shows tumor volume over time in an experiment to determine the ability of SQZ-loaded B cells to function as APCs for therapeutic treatment of HPV-associated tumors. Figure 22B shows the phenotypic profiles and percentages of tumor-infiltrating cells recruited to the tumor.

[0087] [Figure 23] Figure 23 shows IFN-γ secretion by the E7 responder as an in vitro antigen-specific response to SQZ-loaded BAPCs induced by HPV16 E7 SLP.

[0088] [Figure 24] Figure 24 shows the relative amounts of tumor-infiltrating lymphocytes (TILs) mobilized to tumors by SQZ-loaded TAPC with HPV16 E7 SLP, with or without co-administration of adjuvant.

[0089] [Figure 25] Figure 25 shows tumor volume over time in experiments to determine the ability of SQZ-loaded T cells to function as APCs for prophylactic treatment against HPV-related tumors, for both shorter-term (right flank tumor, injected on day 0) and longer-term protection (left flank tumor, injected on day 60).

[0090] [Figure 26] Figure 26 shows tumor volume over time in experiments to determine the effects of T cell dose, adjuvant co-administration, and number of doses (prime vs. prime / boost) on the ability of SQZ-loaded T cells to function as APCs for therapeutic treatment of HPV-associated tumors. In Figure 26, "P" indicates prime and "B" indicates boost.

[0091] [Figure 27] Figure 27A shows tumor volume over time in experiments to determine the ability of SQZ-loaded B cells to function as APCs for therapeutic treatment of HPV-related tumors, compared to electroporated B cells and high-dose peptide vaccines (SC SLP). Figure 27B shows corresponding survival data over time from therapeutic treatment of B cell APCs for HPV-related tumors, compared to electroporated B cells and high-dose peptide vaccines.

[0092] [Figure 28] Figure 28A shows a typical schematic diagram of an experiment to evaluate the ability of splenocytes as APCs to induce an endogenous response. Figure 28B shows the level of IFN-γ-positive CD8+ T cells induced by B9-23 challenge in response to OVA-loaded splenocyte APC vaccination. Figure 28C shows the level of IFN-γ-positive CD8+ T cells induced by E7 challenge in response to E7-loaded splenocyte APC vaccination.

[0093] [Figure 29] Figure 29A shows tumor volume over time in an experiment to determine the ability of SQZ-loaded splenocytes to function as APCs for therapeutic treatment of HPV-related tumors. Figure 29B shows corresponding survival data over time from therapeutic treatment of splenocyte APCs for HPV-related tumors.

[0094] [Figure 30] Figure 30 shows IFN-γ secretion by the E7 responder as an in vitro antigen-specific response to SQZ-loaded PBMCAPC induced by HPV16 E7 SLP. [Modes for carrying out the invention]

[0095] Detailed description of the invention In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating an immune response in an individual having an HPV-related disease, comprising the step of administering to the individual a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cell. In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating an immune response in an individual having an HPV-related disease, the method comprising the steps of administering to an individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, the modified immune cells being prepared by first passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and then incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. Certain aspects of the present disclosure relate to a method for producing a composition comprising modified immune cells, wherein the immune cells are passed through a constriction, the constriction causing a perturbation of the cells so that the HPV antigen and / or adjuvant enters the modified immune cells.

[0096] In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating an immune response in an individual having an HPV-related disease, comprising the step of administering to an individual a composition comprising modified immune cells, wherein the modified immune cells contain HPV antigens within the cells. In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating an immune response in an individual having an HPV-related disease, comprising the step of administering to an individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells contain HPV antigens within the cells, and the modified immune cells are prepared by first passing a cell suspension containing input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen to pass through and forming perturbed input cells; and then incubating the perturbed input cells and the HPV antigens for a time sufficient to allow the HPV antigens to enter the perturbed input cells, thereby producing modified immune cells. Certain embodiments of this disclosure relate to a method for producing a composition comprising modified immune cells, wherein the immune cells are passed through a constriction, the constriction deforming the cells and thereby causing a perturbation of the cells so that the HPV antigen enters the modified immune cells. In some further embodiments, a method for treating and preventing HPV-related diseases, and / or modulating an immune response in an individual having an HPV-related disease, further comprises the step of administering an adjuvant to the individual. In some embodiments, the composition of modified immune cells further comprises an adjuvant (e.g., CpG oligonucleotide (CpG ODN) or IFNα). In some embodiments, the modified immune cells further contain an adjuvant such as CpG ODN within the cells.

[0097] general technique The techniques and procedures described or referenced herein are generally well understood, for example, Molecular Cloning: A Laboratory Manual (Sambrook et al., 4th ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 2012);Current Protocols in Molecular Biology (F.M. Ausubel, et al. eds., 2003);the series Methods in Enzymology (Academic Press, Inc.);PCR 2: A Practical Approach (M.J. MacPherson, B.D. Hames and G.R. Taylor eds., 1995);Antibodies, A Laboratory Manual (Harlow and Lane, eds., 1988);Culture of Animal Cells: A Manual of Basic Technique and Specialized Applications (R.I. Freshney, 6 thed., J. Wiley and Sons, 2010);Oligonucleotide Synthesis (M.J. Gait, ed., 1984);Methods in Molecular Biology, Humana Press; Cell Biology: A Laboratory Notebook (J.E. Cellis, ed., Academic Press, 1998);Introduction to Cell and Tissue Culture (J.P. Mather and P.E. Roberts, Plenum Press, 1998);Cell and Tissue Culture: Laboratory Procedures (A. Doyle, J.B. Griffiths, and D.G. Newell, eds., J. Wiley and Sons, 1993-8);Handbook of Experimental Immunology (D.M. Weir and C.C. Blackwell, eds., 1996);Gene Transfer Vectors for Mammalian Cells (J.M. Miller and M.P. Calos, eds., 1987);PCR: The Polymerase Chain Reaction, (Mullis et al., eds., 1994);Current Protocols in Immunology (J.E. Coligan et al., eds., 1991);Short Protocols in Molecular Biology (Ausubel et al., eds., J. Wiley and Sons, 2002);Immunobiology (C.A. Janeway et al., 2004);Antibodies (P. Finch, 1997);Antibodies: A Practical Approach (D. Catty., ed., IRL Press, 1988-1989);Monoclonal Antibodies: A Practical Approach (P. Shepherd and C.Conventional methodologies, such as those widely used by those skilled in the art, are employed, including those described in *Dean, eds., Oxford University Press, 2000*; *Using Antibodies: A Laboratory Manual* (E. Harlow and D. Lane, Cold Spring Harbor Laboratory Press, 1999); *The Antibodies* (M. Zanetti and JD Capra, eds., Harwood Academic Publishers, 1995); and *Cancer: Principles and Practice of Oncology* (VT DeVita et al., eds., JB Lippincott Company, 2011).

[0098] definition For the purpose of interpreting this Spec., the following definitions apply, and wherever appropriate, a singular term also includes the plural, and vice versa. In the event of any conflict between any definition described below and any document incorporated herein by reference, the definition described below shall prevail.

[0099] As used herein, the singular forms "a," "an," and "the" include plural references unless otherwise indicated.

[0100] The aspects and embodiments of the present invention described herein are understood to include aspects and embodiments that "comprising," "consisting," and "consisting essentially of."

[0101] As used herein, the term “approximately” refers to the normal margin of error for each value, which is readily known to those skilled in the art. References to “approximately” values ​​or parameters herein include (and are described) embodiments that apply to the value or parameter itself.

[0102] As used herein, “treatment” is an approach to obtain a beneficial or desired clinical outcome. As used herein, “treatment” encompasses any therapeutic administration or application for a disease in mammals, including humans. For the purposes of the present invention, beneficial or desired clinical outcomes include, but are not limited to, relief of one or more symptoms, reduction of disease severity, prevention or delay of disease progression (e.g., metastasis, e.g., to the lungs or lymph nodes), prevention or delay of disease recurrence, delay or slowing of disease progression, improvement of disease status, inhibition of the disease or its progression, inhibition or slowing of the disease or its progression, prevention of its occurrence, and remission (whether partial or complete). Reduction of the pathological outcomes of proliferative disorders is also encompassed by “treatment.” The methods of the present invention aim to achieve one or more of these forms of treatment.

[0103] In the context of cancer, the term “treating” includes any or all of the following: inhibiting the growth of cancer cells, inhibiting the replication of cancer cells, reducing the overall tumor burden, and improving one or more symptoms associated with the disease.

[0104] As used herein, the term “pore” refers to an opening, including but not limited to holes, cracks, cavities, openings, fissures, gaps, or perforations within a material. In some examples, (where indicated) the term refers to a pore within the surface of the present disclosure. In other examples, (where indicated) the term may refer to a pore in a cell membrane.

[0105] As used herein, the term “membrane” refers to a selective barrier or sheet containing pores. The term includes flexible, sheet-like structures that function as boundaries or backings. In some examples, the term refers to a surface or filter containing pores. This term is distinct from the term “cell membrane.”

[0106] As used herein, the term “filter” refers to a porous article that allows the selective passage of pores. In some examples, the term refers to a surface or film containing pores.

[0107] As used herein, the term “heterogeneous” refers to something that is mixed or not uniform in structure or composition. In some examples, the term refers to pores having varying sizes, shapes or distributions within a given surface.

[0108] As used herein, the term “homogeneous” refers to something that is consistent or uniform in structure or composition throughout. In some examples, the term refers to pores that have a consistent size, shape or distribution within a given surface.

[0109] As used herein, the term “homogeneous” refers to molecules originating from the same organism. In some instances, the term refers to nucleic acids or proteins that are commonly found or expressed within a given organism.

[0110] The term “heterogeneous,” when relating to nucleic acid sequences such as coding and regulatory sequences, typically refers to sequences that are not conjugated to each other and / or are not typically associated with a particular cell. Therefore, the “heterogeneous” region of a nucleic acid construct or vector is a segment of nucleic acid within, or bound to, another nucleic acid molecule that is not originally found in association with other molecules. For example, the heterogeneous region of a nucleic acid construct may include a coding sequence adjacent to a sequence that is not originally found in association with the coding sequence. Another example of a heterogeneous coding sequence is a coding sequence that is itself a construct that is not originally found (e.g., a synthetic sequence with codons different from the native gene). Similarly, a cell transformed with a construct that is not normally present in the cell should be considered heterogeneous for the purposes of this invention. Allelic variants or naturally occurring mutational events do not produce heterogeneous DNA as used herein.

[0111] The term "heterogeneous," when relating to amino acid sequences such as peptide and polypeptide sequences, typically refers to sequences that are not conjugated to each other and / or are not typically associated with a particular cell. Therefore, the "heterogeneous" region of a peptide sequence is a segment of amino acids within, or bound to, another amino acid molecule that is not originally found in association with other molecules. For example, the heterogeneous region of a peptide construct may include an amino acid sequence of the peptide adjacent to a sequence not originally found in association with the peptide's amino acid sequence. Another example of a heterogeneous peptide sequence is a peptide sequence that is itself a construct not originally found (e.g., a synthetic sequence having different amino acids encoded by a naturally occurring gene). Similarly, cells transformed with a vector expressing an amino acid construct not normally present in the cell should be considered heterogeneous for the purposes of this invention. Allelic mutations or naturally occurring mutational events do not produce heterogeneous peptides as used herein.

[0112] As used herein, the term “inhibit” may mean an action that blocks, reduces, eliminates, or otherwise antagonizes the presence or activity of a particular target. Inhibition may mean partial or complete inhibition. For example, inhibiting an immune response may mean any action that results in blocking, reducing, eliminating, or any other antagonism of the immune response. Other examples of inhibition of nucleic acid expression include, but are not limited to, a decrease in nucleic acid transcription, a decrease in mRNA abundance (e.g., silencing mRNA transcription), mRNA degradation, and inhibition of mRNA translation.

[0113] As used herein, the term “suppress” may mean an action that reduces, diminishes, interferes with, limits, mitigates, or otherwise diminishes the presence or activity of a particular target. Suppression may refer to partial or complete suppression. For example, suppressing an immune response may mean any action that results in a reduction, diminishing, interfering with, limiting, mitigating, or otherwise diminishing an immune response. Other examples of suppression of nucleic acid expression include, but are not limited to, a decrease in nucleic acid transcription, a decrease in mRNA abundance (e.g., silencing mRNA transcription), mRNA degradation, and inhibition of mRNA translation.

[0114] As used herein, the term “enhance” may mean an action that improves, boosts, enhances, or otherwise increases the presence or activity of a particular target. For example, enhancing an immune response may mean any action that results in improving, boosting, enhancing, or otherwise increasing an immune response. In one exemplary example, enhancing an immune response may mean using an antigen and / or adjuvant to improve, boost, enhance, or otherwise increase an immune response. Other examples of enhancing nucleic acid expression may include, but are not limited to, an increase in nucleic acid transcription, an increase in mRNA abundance (e.g., increasing mRNA transcription), a decrease in mRNA degradation, or an increase in mRNA translation.

[0115] As used herein, the term “modulate” may mean an action that alters, changes, fluctuates, or otherwise modifies the presence or activity of a particular target. For example, modulating an immune response may mean any action that results in altering, changing, fluctuating, or otherwise modifying an immune response. In some cases, “modulate” means enhancing the presence or activity of a particular target. In some cases, “modulate” means suppressing the presence or activity of a particular target. In other cases, modulating nucleic acid expression may include, but are not limited to, changes in nucleic acid transcription, changes in mRNA abundance (e.g., increasing mRNA transcription), corresponding changes in mRNA degradation, and changes in mRNA translation.

[0116] As used herein, the term “induce” may mean an action that initiates, prompts, stimulates, establishes, or otherwise produces a result. For example, inducing an immune response may mean any action that results in the initiation, prompting, stimulating, establishing, or otherwise producing a desired immune response. Other examples of inducing nucleic acid expression include, but are not limited to, the initiation of nucleic acid transcription or the initiation of mRNA translation.

[0117] As used herein, “peripheral blood mononuclear cells” or “PBMCs” refers to a heterogeneous population of blood cells having a round nucleus. Examples of cells that may be found in a population of PBMCs include lymphocytes such as T cells, B cells, and NK cells (including NKT cells and CIK cells), as well as monocytes such as macrophages and dendritic cells. As used herein, “multiple PBMCs” refers to a preparation of PBMCs containing at least two types of blood cells. In some embodiments, multiple PBMCs contain two or more of T cells, B cells, NK cells, macrophages, or dendritic cells. In some embodiments, multiple PBMCs contain three or more of T cells, B cells, NK cells, macrophages, or dendritic cells. In some embodiments, multiple PBMCs contain four or more of T cells, B cells, NK cells, macrophages, or dendritic cells. In some embodiments, multiple PBMCs contain T cells, B cells, NK cells, macrophages, and dendritic cells.

[0118] PBMCs can be isolated by means known in the art. For example, PBMCs can be obtained from the peripheral blood of an individual based on their density compared to other blood cells. In some embodiments, PBMCs are obtained from the peripheral blood of an individual using Ficoll (e.g., a ficoll gradient). In some embodiments, PBMCs are obtained from the peripheral blood of an individual using the ELUTRA® cell isolation system.

[0119] In some embodiments, the population of PBMCs is isolated from an individual. In some embodiments, the multiple PBMCs are an on-population of PBMCs, where the population originates from a specific individual, is manipulated by one of the methods described herein, and is returned to the specific individual. In some embodiments, the multiple PBMCs are an allogeneic population of PBMCs, where the population originates from one individual, is manipulated by one of the methods described herein, and is administered to a second individual.

[0120] In some embodiments, multiple PBMCs are reconstituted preparations of PBMCs. In some embodiments, multiple PBMCs can be produced by mixing cells typically found in a population of PBMCs, for example, by mixing two or more populations of T cells, B cells, NK cells, or monocytes.

[0121] As used herein, the terms “polynucleotide” or “nucleic acid” refer to nucleotides in polymeric form of either ribonucleotides or deoxyribonucleotides of any length. Therefore, the term includes, but is not limited to, single-stranded, double-stranded or multi-stranded DNA or RNA, genomic DNA, cDNA, DNA-RNA hybrids, or polymers containing purine and pyrimidine bases, or other natural, chemically or biochemically modified, unnatural, or derivatized nucleotide bases. The backbone of a polynucleotide may contain sugars and phosphate groups (as typically found in RNA or DNA), or modified or substituted sugars or phosphate groups. Alternatively, the backbone of a polynucleotide may contain polymers of synthetic subunits such as phosphoramides and phosphorothioates, and thus may be oligodeoxynucleoside phosphoramides (P-NH2), mixed phosphorothioate-phosphodiester oligomers, or mixed phosphoramides-phosphodiester oligomers. In addition, double-stranded polynucleotides can be obtained from chemically synthesized single-stranded polynucleotide products by either synthesizing a complementary strand and annealing the strand under appropriate conditions, or by synthesizing the complementary strand de novo using DNA polymerase with appropriate primers.

[0122] The terms “polypeptide” and “protein” are used interchangeably to refer to polymers of amino acid residues and are not limited to the minimum length. Such polymers of amino acid residues may, but are not limited to, contain native or non-native amino acid residues, and include peptides, oligopeptides, dimers, trimers, and polymers of amino acid residues. Full-length proteins and their fragments are both encompassed by definition. The term also includes post-expression modifications of polypeptides, such as glycosylation, sialylation, acetylation, and phosphorylation. Furthermore, for the purposes of this invention, “polypeptide” refers to a protein that includes modifications such as deletions, additions, and substitutions (generally, inherently conserved) of the native sequence, as long as the protein maintains the desired activity. These modifications may be planned, such as by site-directed mutagenesis, or accidental, such as due to mutations in the host producing the protein or errors resulting from PCR amplification.

[0123] As used herein, the term “adjuvant” refers to a substance that modulates and / or induces an immune response. Generally, adjuvants are administered in conjunction with an antigen to enhance the immune response to that antigen compared to the antigen alone. Various adjuvants are described herein.

[0124] The terms “CpG oligodeoxynucleotide” and “CpG ODN” refer, as herein, to DNA molecules of 10 to 30 nucleotides in length containing cytosine and guanine dinucleotides (also referred herein as “CpG” dinucleotide or “CpG”) that are separated by a phosphate. The CpG ODNs of this disclosure contain at least one unmethylated CpG dinucleotide; that is, the cytosine in the CpG dinucleotide is not methylated (i.e., not 5-methylcytosine). The CpG ODNs may have a partial or complete phosphorothioate (PS) backbone.

[0125] As used herein, “pharmaceutically acceptable” or “pharmaceutically acceptable” means a material that is not biologically or otherwise undesirable, for example, a material that can be incorporated into a pharmaceutical composition administered to a patient without causing any significant undesirable biological effect or interacting in an adverse manner with any other component of the composition in which it is contained. A pharmaceutically acceptable carrier or excipient is preferably one that meets the necessary standards of toxicological and manufacturing testing and / or is included in the Inactive Ingredient Guide made by the U.S. Food and Drug Administration.

[0126] Methods for determining any structural and functional features described herein are known in the art. Microfluidic systems and their components Microfluidic channels to provide cell deformation constriction In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating the immune response in an individual having an HPV-related disease, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, the modified immune cells being prepared by first passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and then incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, the constriction is contained within a microfluidic channel. In some embodiments, multiple constrictions can be arranged in parallel and / or in series within a microfluidic channel. In some embodiments, a deformable force is applied to the input cells as they pass through the constriction. An exemplary microfluidic channel containing a cellular deformable constriction for use in the methods disclosed herein is described in WO2013059343. An exemplary surface having a pore for use in the methods disclosed herein is described in WO2017041050.

[0127] In some embodiments, the microfluidic channel includes a lumen and is configured to allow immune cells suspended in a buffer to pass through, and the microfluidic channel includes a constriction. The microfluidic channel can be made from any one of several materials, including silicon, metal (e.g., stainless steel), plastic (e.g., polystyrene), ceramic, glass, crystalline substrate, amorphous substrate, or polymer (e.g., polymethyl methacrylate (PMMA), PDMS, cyclic olefin copolymer (COC), etc.). The microfluidic channel can be fabricated by any method known in the art, including dry etching, wet etching, photolithography, injection molding, laser ablation, or SU-8 masking.

[0128] In some embodiments, the constriction within the microfluidic channel includes an inlet portion, a center point, and an outlet portion. In some embodiments, the length, depth, and width of the constriction within the microfluidic channel can be varied. In some embodiments, the diameter of the constriction within the microfluidic channel is a function of the diameter of the immune cell. In some embodiments, the diameter of the constriction within the microfluidic channel is about 20% to about 99% of the diameter of the immune cell. In some embodiments, the size of the constriction is about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 99% of the diameter of the immune cell. In some embodiments, the size of the constriction is about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 99% of the minimum cross-sectional distance of the immune cell. In some embodiments, the channel includes a constriction width of about 2 μm to about 10 μm, or any width or range of widths between those. For example, the width of the constriction may be one of approximately 2 μm, 3 μm, 4 μm, 5 μm, 6 μm, or 7 μm. In some embodiments, the channel includes a constriction length of approximately 10 μm and a constriction width of approximately 4 μm. The cross-section, inlet, center point, and outlet of the channel can also be varied. For example, the cross-section may be circular, elliptical, elongated slit, square, hexagonal, or triangular in shape. The inlet defines the angle of the constriction, which is optimized to reduce channel clogging and for enhanced delivery of compounds to immune cells. The angle of the outlet can similarly be varied. For example, the angle of the outlet is configured to reduce the possibility of turbulence, which can result in non-laminar flow. In some embodiments, the walls of the inlet and / or outlet are straight. In other embodiments, the walls of the inlet and / or outlet are curved. The flow velocity through the channel can also be adjusted. In some embodiments, the flow velocity through the channel is approximately 0.001 mL / cm². 2 / sec ~ approx. 100L / cm 2 This is a speed or range of speeds between 1 / second and 1 / second.

[0129] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the diameter of the constriction is a function of the diameter of the PBMCs, e.g., the average diameter of the multiple PBMCs, or the average diameter of a subpopulation within the multiple PBMCs. In some embodiments, the diameter of the cells is measured by the minimum cross-sectional distance of the cells (e.g., cells within the multiple PBMCs).

[0130] In some embodiments of any one of the methods or compositions described herein, where the immune cells are a plurality of PBMCs, the diameter of the constriction is about 10% to about 99% of the average diameter of the plurality of input PBMCs. In some embodiments, the diameter of the constriction is one of about 10% to about 90%, about 10% to about 80%, about 10% to about 70%, about 20% to about 60%, about 40% to about 60%, or about 30% to about 45% of the average diameter of the plurality of input PBMCs. In some embodiments, the diameter of the constriction is one of about 10% to about 20%, about 20% to about 30%, about 30% to about 40%, about 40% to about 50%, about 50% to about 60%, about 60% to about 70%, about 70% to about 80%, about 80% to about 90%, or about 90% to about 99% of the average diameter of the plurality of input PBMCs. In some embodiments, the diameter of the stenosis is one of approximately 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% of the average diameter of the multiple input PBMCs.

[0131] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the diameter of the constriction is about 10% to about 99% of the average diameter of the subpopulation of cells having the minimum diameter in the multiple input PBMCs. In some embodiments, the diameter of the constriction is any one of about 10% to about 90%, about 10% to about 80%, about 10% to about 70%, about 20% to about 60%, about 40% to about 60%, about 30% to about 45%, about 50% to about 99%, about 50% to about 90%, about 50% to about 80%, about 50% to about 70%, about 60% to about 90%, about 60% to about 80%, or about 60% to about 70% of the average diameter of the subpopulation of cells having the minimum diameter in the multiple input PBMCs. In some embodiments, the diameter of the stenosis is one of approximately 10% to 20%, 20% to 30%, 30% to 40%, 40% to 50%, 50% to 60%, 60% to 70%, 70% to 80%, 80% to 90%, or 90% to 99% of the average diameter of the subpopulation of cells having the smallest diameter in multiple input PBMCs. In some embodiments, the subpopulation of cells having the minimum mean diameter within a plurality of input PBMCs is a population of lymphocytes, with a diameter of approximately 6 μm to approximately 10 μm. In some embodiments, the average diameter of the lymphocyte population is approximately 7 μm. In some embodiments, the lymphocyte population is a population of T cells. In some embodiments, the lymphocytes are T cells. In some embodiments, the subpopulation of cells having the minimum mean diameter within a plurality of input PBMCs is T cells.

[0132] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the diameter of the constriction is about 10% to about 99% of the average diameter of the subpopulation of cells having the largest diameter in the multiple input PBMCs. In some embodiments, the diameter of the constriction is any one of about 10% to about 90%, about 10% to about 80%, about 10% to about 70%, about 20% to about 60%, about 40% to about 60%, about 30% to about 45%, about 15% to about 30%, about 15% to about 20%, about 20% to about 25%, about 25% to about 30%, about 20% to about 30%, about 30% to about 70%, or about 30% to about 60% of the average diameter of the subpopulation of cells having the largest diameter in the multiple input PBMCs. In some embodiments, the diameter of the stenosis is one of approximately 5% to 10%, 10% to 20%, 20% to 30%, 30% to 40%, 40% to 50%, 50% to 60%, 60% to 70%, 70% to 80%, 80% to 90%, or 90% to 99% of the average diameter of the subpopulation of cells having the largest diameter in multiple input PBMCs. In some embodiments, the subpopulation of cells with the largest average diameter within a plurality of input PBMCs is a population of monocytes, with a diameter of approximately 15 μm to approximately 25 μm. In some embodiments, the average diameter of the monocyte population is approximately 20 μm. In some embodiments, the subpopulation of cells with the largest average diameter within a plurality of input PBMCs is monocytes.

[0133] In some embodiments of any one of the methods or compositions described herein, the diameter of the constriction is about 3 μm to about 15 μm. In some embodiments, the diameter of the constriction is about 3 μm to about 10 μm. In some embodiments, the diameter of the constriction is about 4 μm to about 10 μm. In some embodiments, the diameter of the constriction is about 4.2 μm to about 6 μm. In some embodiments, the diameter of the constriction is about 4.2 μm to about 4.8 μm. In some embodiments, the diameter of the constriction is one of the following: approximately 2 μm to approximately 14 μm, approximately 4 μm to approximately 12 μm, approximately 6 μm to approximately 9 μm, approximately 4 μm to approximately 6 μm, approximately 4 μm to approximately 5 μm, approximately 3.5 μm to approximately 7 μm, approximately 3.5 μm to approximately 6.3 μm, approximately 3.5 μm to approximately 5.6 μm, approximately 3.5 μm to approximately 4.9 μm, approximately 4.2 μm to approximately 6.3 μm, approximately 4.2 μm to approximately 5.6 μm, or approximately 4.2 μm to approximately 4.9 μm. In some embodiments, the diameter of the constriction is one of approximately 2 μm, 2.5 μm, 3 μm, 3.5 μm, 4 μm, 4.5 μm, 5 μm, 5.5 μm, 6 μm, 6.5 μm, 7 μm, 7.5 μm, 8 μm, 8.5 μm, 9 μm, 9.5 μm, 10 μm, 10.5 μm, 11 μm, 11.5 μm, 12 μm, 12.5 μm, 13 μm, 13.5 μm, 14 μm, 14.5 μm, or 15 μm. In some embodiments, the diameter of the constriction is one of approximately 4.0 μm, 4.1 μm, 4.2 μm, 4.3 μm, 4.4 μm, 4.5 μm, 4.6 μm, 4.7 μm, 4.8 μm, 4.9 μm, or 5.0 μm. In some embodiments, the diameter of the constriction is approximately 4.5 μm.

[0134] In some embodiments of any one of the methods or compositions described herein, the input immune cells pass through the constriction at a flow rate of about 0.001 mL / min to about 200 mL / min, or any speed or speed range in between. In some embodiments, the flow rate is approximately 0.001 mL / min to approximately 175 mL / min, approximately 0.001 mL / min to approximately 150 mL / min, approximately 0.001 mL / min to approximately 125 mL / min, approximately 0.001 mL / min to approximately 100 mL / min, approximately 0.001 mL / min to approximately 50 mL / min, approximately 0.001 mL / min to approximately 25 mL / min, approximately 0.001 mL / min to approximately 10 mL / min, approximately 0.001 mL / min to approximately 7.5 mL / min, approximately 0.001 mL / min to approximately 5.0 mL / min, approximately 0.001 mL / min to approximately 2.5 mL / min, approximately 0.001 mL / min to approximately 1 mL / min, approximately 0.001 mL / min to approximately 0.1 mL / min, or approximately 0.001 mL / min to approximately 0.01 mL / min. In some embodiments, the flow rate is approximately 0.001 mL / min to approximately 200 mL / min, approximately 0.01 mL / min to approximately 200 mL / min, approximately 0.1 mL / min to approximately 200 mL / min, approximately 1 mL / min to approximately 200 mL / min, approximately 10 mL / min to approximately 200 mL / min, approximately 50 mL / min to approximately 200 mL / min, approximately 75 mL / min to approximately 200 mL / min, approximately 100 mL / min to approximately 200 mL / min, and approximately 15 The flow rates are 0 mL / min to approximately 200 mL / min, approximately 0.5 mL / min to approximately 200 mL / min, approximately 1 mL / min to approximately 200 mL / min, approximately 2.5 mL / min to approximately 200 mL / min, approximately 5 mL / min to approximately 200 mL / min, approximately 7.5 mL / min to approximately 200 mL / min, approximately 10 mL / min to approximately 200 mL / min, approximately 25 mL / min to approximately 200 mL / min, or approximately 175 mL / min to approximately 200 mL / min. In some embodiments, input immune cells pass through the constriction at a flow rate of approximately 10 mL / min to approximately 200 mL / min. In some embodiments, input immune cells pass through the constriction at a flow rate of approximately 100 mL / min.

[0135] In some embodiments of any one of the methods or compositions described herein, the constriction may have any shape known in the art, for example, a three-dimensional shape or a two-dimensional shape. The two-dimensional shape, such as the cross-sectional shape of the constriction, may be, without limitation, circular, elliptical, round, square, star-shaped, triangular, polygonal, pentagonal, hexagonal, heptagonal, or octagonal. The three-dimensional shape of the constriction may be, without limitation, cylindrical, conical, or cubic. In some embodiments, the cross-sectional shape of the constriction is rectangular. In some embodiments, the cross-sectional shape of the constriction is a slit. In some embodiments, the cross-sectional shape of the constriction is a slit having a width of about 4 μm to about 10 μm and / or a depth of about 1 μm to about 200 μm. In some embodiments, the cross-sectional shape of the constriction is a slit having a width of about 3 μm to about 6 μm and / or a depth of about 20 μm to about 120 μm. In some embodiments, the constriction cross-sectional shape is a slit with a width of approximately 4.2 μm to approximately 6 μm and / or a depth of approximately 20 μm to approximately 120 μm. In some embodiments, the constriction cross-sectional shape is a slit with a width of approximately 4.2 μm to approximately 6 μm and / or a depth of approximately 40 μm to approximately 120 μm. In some embodiments, the constriction cross-sectional shape is a slit with a width of approximately 4.2 μm to approximately 6 μm and / or a depth of approximately 20 μm to approximately 80 μm. In some embodiments, the constriction cross-sectional shape is a slit with a width of approximately 4.5 μm and / or a depth of approximately 80 μm. In some embodiments, the slit has a length of approximately 5 μm to approximately 50 μm. In some embodiments, the slit has a length of approximately 10 μm to approximately 30 μm. In some embodiments, the slit has a length of approximately 2 μm to approximately 50 μm. In some embodiments, the slit includes one of the following lengths: approximately 2 μm to approximately 5 μm, approximately 5 μm to approximately 10 μm, approximately 10 μm to approximately 15 μm, approximately 15 μm to approximately 20 μm, approximately 20 μm to approximately 25 μm, approximately 25 μm to approximately 30 μm, approximately 30 μm to approximately 35 μm, approximately 35 μm to approximately 40 μm, approximately 40 μm to approximately 45 μm, or approximately 45 μm to approximately 50 μm. In some embodiments, the slit includes a length of approximately 10 μm.

[0136] Surface having pores to provide cell deformation constriction In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or enhancing the immune response in an individual having an HPV-related disease, the method comprising the steps of administering to an individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, the modified immune cells being prepared by first passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and then incubating the perturbed input cells and the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction. In some embodiments, the constriction is a pore or contained within a pore. In some embodiments, the pores are contained in the surface. Exemplary surfaces having pores for use in the methods disclosed herein are described in WO2017041050.

[0137] The surfaces disclosed herein are made of any one of several materials and can take any one of several forms. In some embodiments, the surface is a filter. In some embodiments, the surface is a membrane. In some embodiments, the filter is a tangential flow filter. In some embodiments, the surface is a sponge or sponge-like matrix. In some embodiments, the surface is a matrix.

[0138] In some embodiments, the surface is a meandering path surface. In some embodiments, the meandering path surface contains cellulose acetate. In some embodiments, the surface contains, without limitation, a material selected from synthetic or natural polymers, polycarbonate, silicon, glass, metal, alloy, cellulose nitrate, silver, cellulose acetate, nylon, polyester, polyethersulfone, polyacrylonitrile (PAN), polypropylene, PVDF, polytetrafluoroethylene, mixed cellulose esters, porcelain, and ceramics.

[0139] The surfaces disclosed herein may have any shape known in the art, for example, a three-dimensional shape. The two-dimensional shape of the surface may be, without limitation, circular, elliptical, round, square, star-shaped, triangular, polygonal, pentagonal, hexagonal, heptagonal, or octagonal. In some embodiments, the surface is round. In some embodiments, the three-dimensional shape of the surface is cylindrical, conical, or cubic.

[0140] The surface can have various cross-sectional widths and thicknesses. In some embodiments, the cross-sectional width of the surface is in the range of about 1 mm to about 1 m, or any cross-sectional width in between. In some embodiments, the surface has a defined thickness. In some embodiments, the surface thickness is uniform. In some embodiments, the surface thickness is variable. For example, in some embodiments, parts of the surface are thicker or thinner than other parts of the surface. In some embodiments, the surface thickness varies in the range of about 1% to about 90%, or any percentage in between. In some embodiments, the surface is in the range of about 0.01 μm to about 5 mm in thickness, or any thickness in between.

[0141] In some embodiments, the constriction is either a pore or contained within a pore. The width of the cross-sectional area of ​​the pore is related to the type of immune cells being treated. In some embodiments, the size of the pore is a function of the diameter of the immune cells or clusters of immune cells being treated. In some embodiments, the size of the pore is such that the immune cells are perturbed as they pass through the pore. In some embodiments, the size of the pore is less than the diameter of the immune cells. In some embodiments, the size of the pore is about 10% to about 99% of the diameter of the immune cells. In some embodiments, the size of the pore is about 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 99% of the diameter of the immune cells. The optimal pore size, or the width of the cross-sectional area of ​​the pore, can be varied based on the application and / or the type of immune cells. In some embodiments, the size of the pore is about 2 μm to about 14 μm. In some embodiments, the pore size is approximately 2 μm, 3 μm, 4 μm, 5 μm, 8 μm, 10 μm, 12 μm, or 14 μm. In some embodiments, the cross-sectional width is approximately 2 μm to 14 μm. In some embodiments, the cross-sectional dimensions of the pore are approximately 2 μm, 3 μm, 4 μm, 5 μm, 8 μm, 10 μm, 12 μm, or 14 μm.

[0142] The entrance and exit of a pore passage can have various angles. The pore angle can be selected to minimize pore clogging while immune cells pass through. For example, the angle of the entrance or exit portion may be about 0 to about 90 degrees. In some embodiments, the entrance or exit portion may be greater than 90 degrees. In some embodiments, pores have the same entrance and exit angles. In some embodiments, pores have different entrance and exit angles. In some embodiments, the pore edge is smooth, for example, rounded or curved. A smooth pore edge has a continuous, flat and smooth surface without protrusions, ridges, and uneven parts. In some embodiments, the pore edge is sharp. A sharp pore edge has a thin edge that is pointed or acute at the tip. In some embodiments, the pore passage is straight. A straight pore passage does not contain curves, bends, angles, and other irregularities. In some embodiments, the pore passage is curved. The passage of a curved pore is either bent or deviates from a straight line. In some embodiments, the passage of the pore has multiple curves, e.g., about 2, 3, 4, 5, 6, 7, 8, 9, 10 or more curves. The flow velocity through the pore can also be adjusted. In some embodiments, the flow velocity through the pore is about 0.001 mL / cm². 2 / sec ~ approx. 100L / cm 2 This is a speed or range of speeds between 1 / second and 1 / second.

[0143] Pores can have any shape known in the art, including two-dimensional or three-dimensional shapes. The shape of a pore (e.g., cross-sectional shape) can be, without limitation, circular, elliptical, round, square, star-shaped, triangular, polygonal, pentagonal, hexagonal, heptagonal, and octagonal. In some embodiments, the cross-section of a pore is round. In some embodiments, the three-dimensional shape of a pore is cylindrical or conical. In some embodiments, a pore has an inlet and outlet shape with grooves. In some embodiments, the shape of a pore is homogeneous (i.e., consistent or regular) among pores on a given surface. In some embodiments, the shape of a pore is heterogeneous (i.e., mixed or varied) among pores on a given surface.

[0144] The surfaces described herein may have a range of total pore numbers. In some embodiments, the pores comprise about 10% to about 80% of the total surface area. In some embodiments, the surface comprises about 1.0 × 10 5 ~Approx. 1.0×10 30 It contains a total of 10 pores, or any number or range between them. In some embodiments, the surface is approximately 10 to approximately 1.0 × 10 15 Pore / mm 2 This includes the surface area.

[0145] Pores can be distributed in a great many ways within a given surface. In some embodiments, pores are distributed parallel to each other within a given surface. In such an example, pores are distributed in the same direction and at the same distance from each other within a given surface. In some embodiments, the distribution of pores is regular or homogeneous. In such an example, pores are distributed in a regular, systematic pattern or at the same distance from each other within a given surface. In some embodiments, the distribution of pores is random or heterogeneous. In such an example, pores are distributed in an irregular, disordered pattern or at different distances from each other within a given surface. In some embodiments, a number of surfaces are distributed in series. The number of surfaces can be homogeneous or heterogeneous in size, shape, and / or roughness. The number of surfaces may further contain pores having homogeneous or heterogeneous pore sizes, shapes, and / or numbers, thereby enabling the simultaneous delivery of a range of compounds to different immunocytotypes.

[0146] In some embodiments, individual pores have a uniform width dimension (i.e., a constant width along the length of the pore passage). In some embodiments, individual pores have a variable width (i.e., a width that increases or decreases along the length of the pore passage). In some embodiments, pores in a given surface have the same individual pore depth. In some embodiments, pores in a given surface have different individual pore depths. In some embodiments, pores are directly adjacent to one another. In some embodiments, pores are separated from one another by a certain distance. In some embodiments, pores are separated from one another by a distance of about 0.001 μm to about 30 mm, or any distance or range between them.

[0147] In some embodiments, the surface is coated with a material. The material can be selected from any material known in the art, without limitation, including Teflon®, adhesive coatings, surfactants, proteins, adhesive molecules, antibodies, anticoagulants, factors that modulate cellular functions, nucleic acids, lipids, carbohydrates, or transmembrane proteins. In some embodiments, the surface is coated with polyvinylpyrrolidone (PVP). In some embodiments, the material is covalently attached to the surface. In some embodiments, the material is noncovalently attached to or adsorbed to the surface. In some embodiments, surface molecules are released when immune cells pass through the pore.

[0148] In some embodiments, the surface has modified chemical properties. In some embodiments, the surface is polar. In some embodiments, the surface is hydrophilic. In some embodiments, the surface is nonpolar. In some embodiments, the surface is hydrophobic. In some embodiments, the surface is charged. In some embodiments, the surface is positively and / or negatively charged. In some embodiments, the surface may be positively charged in some areas and negatively charged in other areas. In some embodiments, the surface has a positive charge as a whole, or a negative charge as a whole. In some embodiments, the surface may be smooth, electropolished, rough, or plasma-treated. In some embodiments, the surface contains zwitterions or bipolar compounds. In some embodiments, the surface is plasma-treated.

[0149] In some embodiments, the surface is contained within a larger module. In some embodiments, the surface is contained within a syringe, such as a plastic or glass syringe. In some embodiments, the surface is contained within a plastic filter holder. In some embodiments, the surface is contained within a pipette tip. Cell perturbation In some embodiments, the present invention provides a method for modulating an immune response by passing a cell suspension containing immune cells through a constriction, thereby causing a perturbation of the immune cells so that an antigen and / or adjuvant can enter the immune cells, wherein the perturbation in the immune cells is a cleavage (e.g., a hole, crack, cavity, opening, pore, fissure, gap, perforation) in the immune cells that allows material to move from the outside of the immune cells into the immune cells. In some embodiments, a deformable force is applied to the input cells as they pass through the constriction. The deformation can be caused, for example, by mechanical strain and / or shear force. In some embodiments, the perturbation is a perturbation within the immune cell membrane. In some embodiments, the perturbation is transient. In some embodiments, the immune cell perturbation is approximately 1.0 × 10⁻⁶ -9It lasts for a few seconds to about 2 hours, or any time or time range in between. In some embodiments, the immune cell perturbation is about 1.0 × 10⁻⁶ -9 It lasts for a few seconds to about 1 second, about 1 second to about 1 minute, or about 1 minute to about 1 hour. In some embodiments, immune cell perturbation is about 1.0 × 10⁻⁶ -9 ~Approx. 1.0×10 -1 , about 1.0×10 -9 ~Approx. 1.0×10 -2 , about 1.0×10 -9 ~Approx. 1.0×10 -3 , about 1.0×10 -9 ~Approx. 1.0×10 -4 , about 1.0×10 -9 ~Approx. 1.0×10 -5 , about 1.0×10 -9 ~Approx. 1.0×10 -6 , about 1.0×10 -9 ~Approx. 1.0×10 -7 , or approximately 1.0 × 10 -9 ~Approx. 1.0×10 -8 It lasts for any one of the following seconds. In some embodiments, the immune cell perturbation is approximately 1.0 × 10⁻⁶ -8 ~Approx. 1.0×10 -1 , about 1.0×10 -7 ~Approx. 1.0×10 -1 , about 1.0×10 -6 ~Approx. 1.0×10 -1 , about 1.0×10 -5 ~Approx. 1.0×10 -1 , about 1.0×10 -4 ~Approx. 1.0×10 -1 , about 1.0×10 -3 ~Approx. 1.0×10 -1 , or approximately 1.0 × 10 -2 ~Approx. 1.0×10 -1 It lasts for any one of the seconds. The immune cell perturbations (e.g., pores or holes) produced by the methods described herein are not formed as a result of the assembly of protein subunits to form a multimeric pore structure, such as those produced by complement or bacterial hemolysin.

[0150] When an immune cell passes through the constriction, the constriction temporarily damages the immune cell membrane, which enables passive diffusion of materials due to perturbation. In some embodiments, the immune cell is deformed for only a short period of approximately 100 μs to minimize the opportunity for the apoptosis pathway to be activated by the cell signaling mechanism, although other durations are possible (e.g., in the range of a few nanoseconds to several hours). In some embodiments, the immune cell is deformed for about 1.0×10 -9 seconds to about 2 hours, or any time or range of times therebetween. In some embodiments, the immune cell is deformed for about 1.0×10 -9 seconds to about 1 second, about 1 second to about 1 minute, or about 1 minute to about 1 hour. In some embodiments, the immune cell is deformed for about 1.0×10 -9 to about 1.0×10 -1 , about 1.0×10 -9 to about 1.0×10 -2 , about 1.0×10 -9 to about 1.0×10​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​The deformed cells are deformed for any one of the following seconds. In some embodiments, deforming immune cells includes, without limitation, deforming them for a time in the range of about 1 μs to at least about 750 μs, for example, at least about 1 μs, 10 μs, 50 μs, 100 μs, 500 μs, or 750 μs.

[0151] In some embodiments, the passage of antigens and / or adjuvants to immune cells occurs simultaneously with the passage of immune cells through the constriction and / or with the perturbation of immune cells. In some embodiments, the passage of compounds to immune cells occurs after the immune cells have passed through the constriction. In some embodiments, the passage of compounds to immune cells occurs approximately a few minutes after the immune cells have passed through the constriction. In some embodiments, the passage of compounds to immune cells occurs approximately 1.0 × 10⁻⁶ after the immune cells have passed through the constriction. -2 This occurs after a few seconds to at least about 30 minutes. For example, the passage of the compound into immune cells occurs approximately 1.0 × 10⁻¹⁶ times the time it takes for the immune cells to pass through the narrowing. -2 This occurs after a few seconds to about 1 second, about 1 second to about 1 minute, or about 1 minute to about 30 minutes. In some embodiments, the passage of the compound into immune cells occurs approximately 1.0 × 10⁻¹⁶ times the time it takes for the immune cells to pass through the constriction. -2 seconds to approximately 10 minutes, approximately 1.0×10 -2 seconds to approximately 5 minutes, approximately 1.0×10 -2 seconds to approximately 1 minute, approximately 1.0×10 -2 seconds ~ approx. 50 seconds, approx. 1.0×10 -2 seconds ~ approx. 30 seconds, approx. 1.0 x 10 -2 seconds ~ approx. 10 seconds, approx. 1.0×10 -2 seconds to approximately 1 second, or approximately 1.0 × 10 -2 This occurs after approximately 0.1 seconds. In some embodiments, the passage of the compound into immune cells occurs approximately 1.0 × 10⁻¹⁶ seconds after the immune cells have passed through the constriction. -1 This occurs after a few seconds to about 10 minutes, about 1 second to about 10 minutes, about 10 seconds to about 10 minutes, about 50 seconds to about 10 minutes, about 1 minute to about 10 minutes, or about 5 minutes to about 10 minutes. In some embodiments, perturbations in immune cells after passing through the constriction are corrected within approximately 5 minutes after the immune cells have passed through the constriction.

[0152] In some embodiments, the cell viability after passing through the constriction is approximately 5% to approximately 100%. In some embodiments, the cell viability after passing through the constriction is at least approximately 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%. In some embodiments, the cell viability is approximately 1.0 × 10⁻⁶ after immune cells have passed through the constriction. -2 It is measured from seconds to at least about 10 days later. For example, cell viability is about 1.0 × 10⁻⁶ when immune cells have passed through the stenosis. -2 Measurements are taken between seconds and approximately 1 second, between approximately 1 second and approximately 1 minute, between approximately 1 minute and approximately 30 minutes, or between approximately 30 minutes and approximately 2 hours. In some embodiments, cell viability is measured at approximately 1.0 × 10⁻⁶ times the immune cells have passed through the constriction. -2 Seconds to approximately 2 hours, approximately 1.0 × 10 -2 Seconds to approximately 1 hour, approximately 1.0 × 10 -2 seconds to approx. 30 minutes, approx. 1.0×10 -2 seconds to approximately 1 minute, approximately 1.0×10 -2 seconds ~ approx. 30 seconds, approx. 1.0 x 10 -2 seconds to approximately 1 second, or approximately 1.0 × 10 -2 Cell viability is measured approximately 0.1 seconds after the immune cells have passed through the constriction. In some embodiments, cell viability is measured approximately 1.5 hours to 2 hours, 1 hour to 2 hours, 30 minutes to 2 hours, 15 minutes to 2 hours, 1 minute to 2 hours, 30 seconds to 2 hours, or 1 second to 2 hours after the immune cells have passed through the constriction. In some embodiments, cell viability is measured approximately 2 hours to 5 hours, 5 hours to 12 hours, 12 hours to 24 hours, or 24 hours to 10 days after the immune cells have passed through the constriction.

[0153] Delivery parameters Several parameters may affect the delivery of compounds to immune cells for modulating the immune response by the method described herein. In some embodiments, the cell suspension comes into contact with the compound before, in parallel with, or after passing through the constriction. Immune cells may be suspended in a solution containing the compound for delivery and pass through the constriction, or the compound may be added to the cell suspension after the immune cells have passed through the constriction. In some embodiments, the compound to be delivered is coated onto the constriction.

[0154] Examples of parameters that may affect the delivery of compounds to immune cells include, but are not limited to, the dimensions of the constriction, the angle of the constriction entrance, the properties of the constriction surface (e.g., roughness, chemical modification, hydrophilicity, hydrophobicity, etc.), the operating flux (e.g., the time it takes for cells to pass through the constriction), the concentration of immune cells, the concentration of the compound in the cell suspension, and the amount of time allowed to recover or incubate the immune cells after passing through the constriction, all of which may affect the passage of compounds delivered to immune cells. Additional parameters that may affect the delivery of compounds to immune cells include the rate of immune cells in the constriction, the shear rate in the constriction, the viscosity of the cell suspension, the velocity component perpendicular to the flow rate, and the time in the constriction. Such parameters can be designed to control the delivery of compounds. In some embodiments, the concentration of immune cells is about 10 to at least about 10 12 The concentration ranges from individual cells / mL, or any concentration or range between those values. In some embodiments, the concentration of the delivery compound may range from about 10 ng / mL to about 1 g / mL, or any concentration or range between those values. In some embodiments, the concentration of the delivery compound may range from about 1 pM to at least about 2 M, or any concentration or range between those values.

[0155] The temperature used in the methods of this disclosure can be adjusted to affect compound delivery and cell viability. In some embodiments, the methods are carried out at a temperature of about -5°C to about 45°C. For example, the methods can be carried out at room temperature (e.g., about 20°C), physiological temperature (e.g., about 37°C), a temperature higher than physiological temperature (e.g., higher than about 37°C to 45°C, or higher), or lower (e.g., about -5°C to about 4°C), or at temperatures in between these exemplary temperatures.

[0156] Immune cells can be driven through a constriction using various methods. For example, pressure can be applied by an inlet pump (e.g., a compressor), vacuum can be applied by an outlet vacuum pump, capillary action can be applied by tubing, and / or gravity can be supplied to the system. Displacement-based fluid systems can also be used (e.g., syringe pumps, peristaltic pumps, manual syringes or pipettes, pistons, etc.). In some embodiments, immune cells pass through the constriction by positive or negative pressure. In some embodiments, immune cells pass through the constriction by constant or variable pressure. In some embodiments, pressure is applied using a syringe. In some embodiments, pressure is applied using a gas cylinder. In some embodiments, pressure is applied using a gas cylinder positive pressure method. In some embodiments, pressure is applied using a pump. In some embodiments, the pump is a peristaltic pump or a diaphragm pump. In some embodiments, pressure is applied using a vacuum. In some embodiments, immune cells pass through the constriction by g-force. In some embodiments, immune cells pass through the constriction by centrifugal force. In some embodiments, immune cells pass through the narrowing due to capillary pressure.

[0157] In some embodiments, the fluid flow directs immune cells through the constriction. In some embodiments, the fluid flow is turbulent before the immune cells pass through the constriction. Turbulent flow is the flow of a fluid in which the velocity at a given point fluctuates irregularly in magnitude and direction. In some embodiments, the fluid flow through the constriction is laminar flow. Laminar flow includes uninterrupted flow in a fluid near a solid boundary, where the direction of flow remains constant at all points. In some embodiments, the fluid flow is turbulent after the immune cells have passed through the constriction. The velocity at which immune cells pass through the constriction can be varied. In some embodiments, immune cells pass through the constriction at a uniform cell speed. In some embodiments, immune cells pass through the constriction at a fluctuating cell speed.

[0158] In other embodiments, the combination treatment is used to modulate an immune response by passing a cell suspension containing immune cells through a constriction, which deforms the immune cells, thereby causing perturbation of the immune cells so that antigens and / or adjuvants enter the immune cells, for example, in the method described herein, and then exposing them to an electric field downstream of the constriction. In some embodiments, after passing through the constriction, the immune cells pass through an electric field generated by at least one electrode. In some embodiments, the electric field assists in the delivery of compounds to a second location inside the immune cell, such as the nucleus of the immune cell. For example, the combination of cell deformation constriction and electric field delivers an antibody-encoding plasmid to an immune cell (e.g., the cell nucleus), resulting in de novo production of the antibody. In some embodiments, one or more electrodes are located close to the cell deformation constriction to generate the electric field. In some embodiments, the electric field is in the range of about 0.1 kV / m to about 100 MV / m, or any number or number in between. In some embodiments, an integrated circuit is used to provide an electrical signal to drive the electrodes. In some embodiments, immune cells are exposed to an electric field for a pulse width of about 1 ns to about 1 s and for a period of about 100 ns to about 10 s, or any time or range in between. Cell suspension for delivery to immune cells

[0159] A cell suspension can be a population of immune cells that are mixed or purified. In some embodiments, the cell suspension is a mixed population of cells such as whole blood or PBMCs. In further embodiments, the mixed cell population is a mixture of prescribed or purified populations. In some embodiments, the cell suspension is a purified cell population, such as a purified population of immune cells.

[0160] The composition of the cell suspension (e.g., volumetric osmolality, salt concentration, serum content, cell concentration, pH, etc.) can affect the delivery of compounds to modulate the immune response. In some embodiments, the suspension contains whole blood. Alternatively, the cell suspension is a mixture of cells in a physiological saline solution or physiological medium other than blood. In some embodiments, the cell suspension contains an aqueous solution. In some embodiments, the aqueous solution contains cell culture medium, (phosphate-buffered saline) PBS, salts, metal ions, sugars, growth factors, animal-derived products, fillers, surfactants, lubricants, lipids, vitamins, amino acids, proteins, cell cycle inhibitors, and / or active ingredients that affect actin polymerization. In some embodiments, the cell culture medium is X-VIVO® 10, X-VIVO® 15, DMEM, Opti-MEM®, IMDM, or RPMI. In addition, the solution buffer may contain one or more lubricants (Pluronic® or other surfactants) that can be designed, for example, to reduce or eliminate stenosis clogging and improve cell viability. Examples of surfactants include, but are not limited to, poloxamers; polysorbates; sugars or sugar alcohols such as mannitol and sorbitol; animal serum; and albumin proteins.

[0161] In some configurations involving certain types of immune cells, the immune cells can be incubated in one or more solutions to facilitate the delivery of compounds into the immune cells. In some embodiments, the aqueous solution contains an active agent that affects actin polymerization. In some embodiments, the active agent that affects actin polymerization is lantrunculin A, cytochalasin, and / or colchicine. For example, immune cells can depolymerize their actin cytoskeleton by incubation for 1 hour prior to delivery in a depolymerization solution such as lantrunculin A (0.1 μg / mL). As an additional example, immune cells can depolymerize their microtubule network by incubation for 2 hours prior to delivery in 10 μM colchicine (Sigma).

[0162] In some embodiments, the cell population is enriched before use in the disclosed method. For example, cells are obtained from a body fluid, e.g., peripheral blood, and are enriched or purified to concentrate immune cells, as necessary. The cells may be enriched by any method known in the Art, without limitation, including magnetic cell separation, fluorescence-activated cell sorting (FACS), or density gradient centrifugation.

[0163] The viscosity of the cell suspension may also affect the methods disclosed herein. In some embodiments, the viscosity of the cell suspension ranges from about 8.9 × 10⁻⁴ Pa·s to about 4.0 × 10⁻³ Pa·s, or any value or range between them. In some embodiments, the viscosity is about 8.9 × 10⁻⁴ Pa·s. -4 Pa·s~approx. 4.0×10 -3 The viscosity ranges between Pa·s, or any value between them or any range of values. In some embodiments, the viscosity is approximately 8.9 × 10⁻⁶. -4 Pa·s~approx. 4.0×10 -3 Pa·s, approx. 8.9×10 -4 Pa·s~approx. 3.0×10 -3 Pa·s, approx. 8.9×10 -4 Pa·s~approx. 2.0×10 -3 Pa·s, or approximately 8.9 × 10 -3 Pa·s~approx. 1.0×10 -3The viscosity ranges between any one of the following: Pa·s. In some embodiments, the viscosity ranges between any one of the following: about 0.89 cP to about 4.0 cP, about 0.89 cP to about 3.0 cP, about 0.89 cP to about 2.0 cP, or about 0.89 cP to about 1.0 cP. In some embodiments, a shear-thinning effect is observed, in which the viscosity of the cell suspension decreases under shear strain conditions. Viscosity can be measured by any method known in the art, including, but not limited to, a viscometer such as a glass capillary viscometer, or a rheometer. A viscometer measures viscosity under one flow condition, while a rheometer is used to measure viscosity that fluctuates with flow conditions. In some embodiments, viscosity is measured for shear-thinning solutions such as blood. In some embodiments, viscosity is measured between about -5°C and about 45°C. For example, viscosity is measured at room temperature (e.g., about 20°C), physiological temperature (e.g., about 37°C), above physiological temperature (e.g., above about 37°C to 45°C, or higher), below temperature (e.g., about -5°C to about 4°C), or between these exemplary temperatures.

[0164] Antigens and adjuvants to enhance the immune response Certain aspects of this disclosure relate to a method for treating a patient by introducing immune cells modified by the method described herein into the patient. In some embodiments, the immune cells are for use in immunotherapy. In some embodiments, this disclosure relates to a method for treating a human papillomavirus (HPV)-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells contain HPV antigens and adjuvants within the cells. In some embodiments, the present disclosure relates to a method for treating an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cells, and the modified immune cells are prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0165] In some embodiments, the present disclosure relates to a method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells contain HPV antigens and adjuvants within the cells. In some embodiments, the present disclosure relates to a method for preventing HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0166] In some embodiments, the present disclosure relates to a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells contain an HPV antigen and an adjuvant within the cells. In some embodiments, the present disclosure relates to a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the steps of: administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, and the modified immune cells being prepared by: a) passing a cell suspension comprising input cells comprising an HPV antigen through a microfluidic channel comprising a cellular deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction. In some embodiments, the immune response is enhanced. In some embodiments, the immune response to the HPV antigen is enhanced.

[0167] Some aspects of the present invention provide delivery of an antigen to an individual having an HPV-related disease for enhancing the immune response to an antigen by administering immune cells containing an intracellular antigen, wherein the antigen is delivered to the cells by any of the methods described herein. In some embodiments, the antigen is a single antigen. In some embodiments, the antigen is a mixture of antigens. An antigen is a substance that stimulates a specific immune response, such as a cell- or antibody-mediated immune response. An antigen binds to a receptor expressed by an immune cell, such as a T cell receptor (TCR), which is specific to a particular antigen. Antigen-receptor binding then triggers an intracellular signaling pathway that results in downstream immune effector pathways, such as cell activation, cytokine production, cell migration, secretion of cytotoxic factors, and antibody production.

[0168] In some embodiments, modified immune cells are prepared by a) passing a cell suspension containing input cells containing HPV antigen through a constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells. In further embodiments, the diameter of the constriction is less than the diameter of the cells. In some embodiments, the diameter of the constriction is smaller than the diameter of the immune cells. In some embodiments, the diameter of the constriction is about 20% to about 99% of the diameter of the cells. In some embodiments, the diameter of the constriction is about 20% to less than about 60% of the diameter of the cells. In some embodiments, the diameter of the constriction is one of approximately 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the cell diameter. In some embodiments, the diameter of the constriction is one of approximately 20%–30%, 30%–40%, 40%–50%, 50%–60%, 60%–70%, 70%–80%, 80%–90%, 90%–95%, or 95%–99% of the cell diameter. In some embodiments, a deformable force is applied to the input cell as it passes through the constriction. In some embodiments, the constriction is located within a channel. In some embodiments, the constriction is contained within a microfluidic channel. In some embodiments, the constriction is contained within a filter. In other embodiments, the constriction is a pore on the filter.

[0169] In some embodiments, the modified immune cells contain the HPV antigen and adjuvant intracellularly. In some embodiments, the HPV antigen and / or adjuvant are located in the cytoplasm and / or endosomes. In some embodiments, the antigen and / or adjuvant are located in multiple compartments of the cell. In further embodiments, the antigen and / or adjuvant are located in a compartment of the cell containing one or more of the endoplasmic rectum (ER), Golgi apparatus, lysosomes, or exosomes. In some embodiments, the antigen and adjuvant are located in the same compartment. In some embodiments, the antigen and adjuvant are located in different compartments. For example, in some embodiments, the antigen is located in the cytoplasm, while the adjuvant is located in endosomes. In some embodiments, the modified immune cells further contain the HPV antigen and / or adjuvant extracellularly.

[0170] In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.01 μM to approximately 10 mM. For example, in some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is one of approximately 0.01 μM, approximately 0.1 μM, approximately 1 μM, approximately 10 μM, approximately 100 μM, approximately 1 mM, or less than approximately 10 mM. In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is higher than approximately 10 mM. In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is one of approximately 0.01 μM to approximately 0.1 μM, approximately 0.1 μM to approximately 1 μM, approximately 1 μM to approximately 10 μM, approximately 10 μM to approximately 100 μM, approximately 100 μM to approximately 1 mM, or 1 mM to approximately 10 mM. In some embodiments, the concentration of the adjuvant incubated with the perturbed input cells is approximately 0.1 μM to approximately 1 mM.

[0171] In some embodiments, the concentration of HPV antigen incubated with perturbation input cells is approximately 0.01 μM to approximately 10 mM. For example, in some embodiments, the concentration of HPV antigen incubated with perturbation input cells is one of approximately 0.01 μM, approximately 0.1 μM, approximately 1 μM, approximately 10 μM, approximately 100 μM, approximately 1 mM, or less than approximately 10 mM. In some embodiments, the concentration of HPV antigen incubated with perturbation input cells is higher than approximately 10 mM. In some embodiments, the concentration of HPV antigen incubated with perturbation input cells is one of approximately 0.01 μM to approximately 0.1 μM, approximately 0.1 μM to approximately 1 μM, approximately 1 μM to approximately 10 μM, approximately 10 μM to approximately 100 μM, approximately 100 μM to approximately 1 mM, or 1 mM to approximately 10 mM. In some embodiments, the concentration of HPV antigen incubated with perturbed input cells is approximately 0.1 μM to approximately 1 mM.

[0172] In some embodiments, the molar ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is one of approximately 10,000:1 to approximately 1:10,000. For example, in some embodiments, the molar ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is one of approximately 10,000:1, approximately 1,000:1, approximately 100:1, approximately 10:1, approximately 1:1, approximately 1:10, approximately 1:100, approximately 1:1000, or approximately 1:10000. In some embodiments, the molar ratio of the HPV antigen incubated with the perturbed input cells to the adjuvant is one of the following: approximately 10000:1 to approximately 1000:1, approximately 1000:1 to approximately 100:1, approximately 100:1 to approximately 10:1, approximately 10:1 to approximately 1:1, approximately 1:1 to approximately 1:10, approximately 1:10 to approximately 1:100, approximately 1:100 to approximately 1:1000, or approximately 1:1000 to approximately 1:10000.

[0173] In some embodiments, the modified immune cells contain the adjuvant at a concentration of about 0.01 μM to about 10 mM. For example, in some embodiments, the immune cells contain the adjuvant at a concentration of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the immune cells contain the adjuvant at a concentration higher than about 10 mM. In some embodiments, the immune cells contain the adjuvant at a concentration of about 0.1 μM to about 1 μM, about 1 μM to about 10 μM, about 10 μM to about 100 μM, about 100 μM to about 1 mM, or 1 mM to about 10 mM. In some embodiments, the modified immune cells contain the adjuvant at a concentration of about 0.1 μM to about 1 mM.

[0174] In some embodiments, the concentration of HPV antigen in modified immune cells is approximately 0.01 μM to approximately 10 mM. For example, in some embodiments, the concentration of HPV antigen in modified immune cells is one of the following: approximately 0.01 μM, approximately 0.1 μM, approximately 1 μM, approximately 10 μM, approximately 100 μM, approximately 1 mM, or less than approximately 10 mM. In some embodiments, the concentration of HPV antigen in modified immune cells is higher than approximately 10 mM. In some embodiments, the concentration of HPV antigen in modified immune cells is one of the following: approximately 0.1 μM to approximately 1 μM, approximately 1 μM to approximately 10 μM, approximately 10 μM to approximately 100 μM, approximately 100 μM to approximately 1 mM, or 1 mM to approximately 10 mM. In some embodiments, the concentration of HPV antigen in modified immune cells is approximately 0.1 μM to approximately 1 mM.

[0175] In some embodiments, the molar ratio of HPV antigen to adjuvant in modified immune cells is one of approximately 10,000:1 to approximately 1:10,000. For example, in some embodiments, the molar ratio of HPV antigen to adjuvant in modified immune cells is one of approximately 10,000:1, approximately 1,000:1, approximately 100:1, approximately 10:1, approximately 1:1, approximately 1:10, approximately 1:100, approximately 1:1000, or approximately 1:10000. In some embodiments, the molar ratio of HPV antigen to adjuvant in modified immune cells is one of the following: approximately 10000:1 to approximately 1000:1, approximately 1000:1 to approximately 100:1, approximately 100:1 to approximately 10:1, approximately 10:1 to approximately 1:1, approximately 1:1 to approximately 1:10, approximately 1:10 to approximately 1:100, approximately 1:100 to approximately 1:1000, or approximately 1:1000 to approximately 1:10000.

[0176] In some embodiments, the antigen is a polypeptide antigen. In some embodiments, the antigen is modified with lipids. In some embodiments, the modified antigen is modified with a polysaccharide or carbohydrate moiety. In some embodiments, the antigen is associated with a virus. In some embodiments, the antigen is a viral antigen. An exemplary viral antigen is the HPV antigen. In further embodiments, the antigen is the HPV antigen. In some embodiments, the HPV antigen consists of a selection from the group HPV-16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73, and 82. HPV-16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82 are high-risk types that cause cancer, while HPV-26, 53, and 66 are "probably high-risk types" that cause cancer. In some embodiments, the HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, the antigen is the HPV-16 antigen or the HPV-18 antigen. In some embodiments, the HPV antigen consists of an HLA-A2 specific epitope. The HPV E6 and E7 genes are viral oncogenes, and the expression of these genes is required for malignant transformation. The E6 and E7 proteins target several negative regulators of the cell cycle, mainly p105Rb and p53, respectively, and thus interfere with cell cycle regulation. In further embodiments, the HPV antigen is the HPV E6 antigen or the HPV E7 antigen. In some embodiments, the modified immune cells contain the HPV E6 antigen and the HPV E7 antigen. In some embodiments, the HPV antigen is a polypeptide comprising an immunogenic epitope flanked by one or more heterologous peptide sequences at its N-terminus and / or C-terminus. In some embodiments, the HPV antigen is an HPV E7 epitope (E7.6) adjacent to a sequence derived from the HPV E6 polypeptide. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to one of sequence numbers 18-26. In some embodiments, the HPV antigen comprises the amino acid sequence of sequence number 23.In some embodiments, the HPV antigen includes one of the amino acid sequences of SEQ ID NOs. 18-26. In some embodiments, the HPV antigen includes the amino acid sequence of SEQ ID NO. 23.

[0177] In some embodiments, the antigen is derived from a cell lysate, such as a lysate of infected cells. In some embodiments, the antigen is present in the cell lysate. In some embodiments, the antigen is derived from a tumor lysate. In some embodiments, the antigen is derived from a lysate of HPV-related cancer cells. In some embodiments, the HPV-related cancer is one of the following: head and neck cancer, cervical cancer, vulvar cancer, vaginal cancer, penile cancer, anal cancer, perianal cancer, anogenital cancer, oral cancer, or salivary gland cancer.

[0178] In some embodiments, the present disclosure relates to a method for treating human papillomavirus (HPV)-related cancer in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-26. In some embodiments, the present disclosure relates to a method for treating an HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-26, and the modified immune cells are prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0179] In some embodiments, the present disclosure relates to a method for preventing HPV-related disease in an individual, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-26. In some embodiments, the present disclosure relates to a method for preventing HPV-related disease in an individual, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen, the modified immune cells comprising an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-26, and the modified immune cells are prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformation force is applied to the input cells as they pass through the constriction.

[0180] In some embodiments, the present disclosure relates to a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the step of administering to the individual an effective amount of a composition comprising modified immune cells, wherein the modified immune cells comprise an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-26. In some embodiments, the present disclosure relates to a method for modulating an immune response in an individual having an HPV-related disease, the method comprising the steps of administering to the individual an effective amount of a composition comprising modified immune cells, the modified immune cells comprising an HPV antigen having an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs: 18-26, and the modified immune cells being prepared by a) passing a cell suspension comprising input cells through a cell deformable constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells to be large enough for the HPV antigen to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In some embodiments, a deformable force is applied to the input cells as they pass through the constriction. In some embodiments, the immune response is enhanced. In some embodiments, the immune response to the HPV antigen is enhanced.

[0181] In some embodiments, the HPV antigen is a pool of numerous polypeptides that elicit responses to the same and / or different HPV antigens. In some embodiments, antigens in a pool of multiple antigens do not diminish the immune response directed to other antigens. For example, when using a pool of HPV E6 and E7 antigens, the respective immune responses directed to HPV E6 and E7 antigens should be comparable to those directed to HPV E6 alone or HPV E7 alone, respectively.

[0182] In some embodiments, the HPV antigen is a polypeptide comprising an immunogenic HPV epitope and one or more heterologous peptide sequences. In some embodiments, one or more HPV antigens form complexes with themselves, with other antigens, or with adjuvants.

[0183] As used herein, the term “adjuvant” refers to a substance that modulates and / or generates an immune response, directly or indirectly. Generally, adjuvants are administered in conjunction with an antigen to enhance the immune response to the antigen compared to the antigen alone. Thus, adjuvants can be used to boost the induction of an immune cell response (e.g., a T cell response) to an antigen. In some embodiments, the present invention provides immune cells modified to contain an HPV antigen and an adjuvant within the cell. In some embodiments, the perturbed immune cells as described herein are incubated with both the HPV antigen and the adjuvant. Exemplary intracellular adjuvants include, but are not limited to, CpG ODN, interferon-α (IFN-α), interferon gene-stimulating factor (STING) agonists, and retinoic acid-inducible gene I (RIG-I) agonists, as well as polyinosinic acid:polycytidylic acid (Poly-I:C). In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or polyI:C. In certain embodiments, the adjuvant is CpG ODN polynucleotide. In some embodiments, the CpG ODN adjuvant includes those selected from the group CpG ODN1018, CpG ODN1585, CpG ODN2216, CpG ODN2336, CpG ODN1668, CpG ODN1826, CpG ODN2006, CpG ODN2007, CpG ODN BW006, CpG ODN D-SL01, CpG ODN2395, CpG ODN M362, and CpG ODN D-SL03. In some embodiments, the CpG ODN adjuvant is CpG ODN1826 (TCCATGACGTTCCTGACGTT; SEQ ID NO: 30) or CpG ODN2006 (also known as CpG ODN7909) (TCGTCGTTTTGTCGTTTTGTCGTT; SEQ ID NO: 31) oligonucleotide. In some embodiments, the RIG-I agonist contains polyinosinic acid:polycytidylic acid (Poly-I:C). Multiple adjuvants may also be used in combination with antigens to enhance the induction of an immune response.In some embodiments, the modified immune cells contain more than one adjuvant. Multiple adjuvants can also be used in combination with antigens to enhance the induction of an immune response. In some embodiments, the modified immune cells contain more than one adjuvant. In some embodiments, the modified immune cells contain any combination of adjuvants such as CpG ODN, IFN-α, STING agonist, RIG-I agonist, or polyI:C.

[0184] Examples of adjuvants include, but are not limited to, CpG ODN, interferon-α (IFN-α), polyinosinic acid:polycytidylic acid (Poly-I:C), imiquimod (R837), reciquimod (R848), or lipopolysaccharide (LPS). In some embodiments, the adjuvant is CpG ODN, LPS, IFN-α, STING agonist, RIG-I agonist, Poly-I:C, R837, R848, TLR3 agonist, TLR4 agonist, or TLR9 agonist. In certain embodiments, the adjuvant is CpG ODN. In some embodiments, the adjuvant is CpG ODN. In some embodiments, CpG ODN is class A CpG ODN, class B CpG ODN, or class C CpG ODN. In some embodiments, the CpG ODN adjuvant includes those selected from the group consisting of CpG ODN1018, CpG ODN1585, CpG ODN2216, CpG ODN2336, CpG ODN1668, CpG ODN1826, CpG ODN2006, CpG ODN2007, CpG ODN BW006, CpG ODN D-SL01, CpG ODN2395, CpG ODN M362, and CpG ODN D-SL03. In some embodiments, the CpG ODN adjuvant is the CpG ODN1826 (TCCATGACGTTCCTGACGTT; SEQ ID NO: 30) or the CpG ODN2006 (also known as CpG ODN7909) (TCGTCGTTTTGTCGTTTTGTCGTT; SEQ ID NO: 31) oligonucleotide. In some embodiments, the adjuvant is CpG ODN7909. In some embodiments, the RIG-I agonist contains polyinosinic acid:polycytidylic acid (Poly-I:C). Multiple adjuvants may also be used in combination with antigens to enhance the induction of an immune response. In some embodiments, the modified immune cells contain more than one adjuvant. Multiple adjuvants may also be used in combination with antigens to enhance the induction of an immune response. In some embodiments, the modified immune cells contain more than one adjuvant.In some embodiments, the modified immune cells include any combination of adjuvants such as CpG ODN, LPS, IFN-α, STING agonist, RIG-I agonist, polyI:C, R837, R848, TLR3 agonist, TLR4 agonist, or TLR9 agonist.

[0185] In any of the embodiments described herein, unless otherwise indicated, adjuvant may refer to (a) an adjuvant incubated with and passing through perturbed input cells, (b) an adjuvant incubated with PMBCs to modulate PBMCs, or (c) an adjuvant co-administered to an individual with modified immune cells.

[0186] In some embodiments, the modified immune cells further contain an active agent that enhances the viability and / or function of the modified immune cells compared to the corresponding modified immune cells that do not contain this active agent. In some embodiments, the active agent is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the stabilizer forms a complex with the HPV antigen and / or adjuvant. In some embodiments, the stabilizer increases the solubility and / or solution half-life of the HPV antigen and / or adjuvant. In some embodiments, multiple modified immune cells have higher viability than the corresponding modified immune cells that do not contain the stabilizer. In some embodiments, the active agent is albumin. In further embodiments, the albumin is mouse, bovine, or human albumin. In further embodiments, the active agent is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the divalent metal cation is Mg 2+ Zn 2+ , or Ca 2+ It is one or more of the following. In some embodiments, the active ingredient includes MSA.

[0187] In some embodiments of any one of the methods or compositions described herein, the modified immune cells further contain an active substance that enhances the viability and / or function of the modified immune cells compared to a group of corresponding modified immune cells that do not contain this active substance. In some embodiments, the modified immune cells further contain an active substance that enhances the viability and / or function of the modified immune cells during freeze-thaw cycles compared to corresponding modified immune cells that do not contain this active substance. In some embodiments, the active substance is a cryopreservative and / or frost preservative. In some embodiments, neither the cryopreservative nor the frost preservative causes more than 10% or 20% cell death in modified immune cells containing the active substance compared to corresponding modified immune cells that do not contain the active substance before any freeze-thaw cycle. In some embodiments, at least about 70%, about 80%, or about 90% of the modified immune cells survive up to 1, 2, 3, 4, or 5 freeze-thaw cycles. In some embodiments, the active substance is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the active substance is albumin. In some embodiments, albumin is mouse, bovine, or human albumin. In some embodiments, the active ingredient is human albumin. In some embodiments, the active ingredient is one or more of the following: divalent metal cations, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the divalent metal cation is Mg 2+ Zn 2+ or Ca 2+One or more of the following. In some embodiments, the active substance is one or more of the following: sodium pyruvate, adenine, trehalose, dextrose, mannose, sucrose, human serum albumin (HSA), DMSO, HEPES, glycerol, glutathione, inosine, dibasic sodium phosphate, monobasic sodium phosphate, sodium metal ions, potassium metal ions, magnesium metal ions, chloride, acetate, gluconate, sucrose, potassium hydroxide, or sodium hydroxide. In some embodiments, the active ingredient is one or more of the following: sodium pyruvate, adenine, Rejuvesol®, trehalose, dextrose, mannose, sucrose, human serum albumin (HSA), PlasmaLyte®, DMSO, Cryostor® CS2, Cryostor® CS5, Cryostor® CS10, Cryostor® CS15, HEPES, glycerol, glutathione, and HypoThermosol®.

[0188] In some embodiments, the modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules. In further embodiments, the costimulatory molecules are B7-H2(ICOSL), B7-1(CD80), B7-2(CD86), CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. In some embodiments, the cells contain nucleic acids that result in increased expression of one or more costimulatory molecules.

[0189] In some embodiments, immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, immune cells are not B cells. In some embodiments, immune cells are T cells. In some embodiments, immune cells are other than B cells. In some embodiments, modified T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, or natural killer T cells. In some embodiments, T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. MHC expression in allogeneic T cells can result in an innate immune response initiated in an individual in response to their administration, leading to a shortening of the half-life of such T cells. In some embodiments, T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In certain embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In certain embodiments, the further modifications include using mRNA, plasmid DNA, or cDNA to increase MHC class I and / or MHC class II expression. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting.In some embodiments, the circulating half-life of further modified T cells in the administered individual is increased compared to the circulating half-life of the corresponding modified T cells without further modification in the administered individual. In some embodiments, the modified T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, or natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, or γδ-T cells.

[0190] Immune cells and other cells can be used as a source of autologous or allogeneic cells. In some embodiments, the modified immune cells are allogeneic to the organism. In other embodiments, the modified immune cells are autologous to the organism. In some embodiments, the organism being treated is pre-conditioned to modulate inflammation.

[0191] Adjuvants can be used to further enhance the immune response to HPV antigens. In some embodiments, the method for treatment further includes the step of administering an adjuvant to an individual. Exemplary adjuvants include, but are not limited to, IFN-α, CpG ODN, STING agonists, RIG-I agonists, and poly-I:C. In some embodiments, the adjuvant is IFN-α or CpG ODN. In some embodiments, the adjuvant is IFN-α, CpG ODN, STING agonist, RIG-I agonist, or poly-I:C. In some embodiments, the adjuvant includes any combination of IFN-α, CpG ODN, STING agonist, RIG-I agonist, or poly-I:C.

[0192] In some embodiments, the method includes multiple administrations of modified immune cells. In some embodiments, the method includes about 3 to about 9 administrations of modified immune cells. In some embodiments, the method includes one of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 administrations of modified immune cells. In some embodiments, the method includes, if necessary, consecutive administrations of modified immune cells. In some embodiments, the time interval between two consecutive administrations of modified immune cells is about 1 to about 30 days. In some embodiments, the time interval between two consecutive administrations of modified immune cells is about 21 days. In some embodiments, the time interval between two consecutive doses of modified immune cells is one of approximately 1, 2, 3, 4, 5, 6, 7, 8, 10, 12, 14, 16, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, or 150 days. In some embodiments, the modified immune cells are multiple modified PBMCs. In some embodiments, the modified immune cells are multiple modified PBMCs that have been prepared. Methods for preparing PBMCs are provided in U.S. Provisional Patent Application No. 62 / 812,225 and European Patent Application No. EP19161964.2, which are incorporated herein by reference in their entirety.

[0193] In some embodiments, the composition containing modified immune cells and the adjuvant are administered simultaneously. In some embodiments, the composition containing modified immune cells and the adjuvant are administered sequentially.

[0194] In some embodiments, the composition containing modified immune cells is administered before the administration of the adjuvant. For example, the composition containing modified immune cells is administered about 1 hour to about 1 week before the administration of the adjuvant. For example, in some embodiments, the composition containing modified immune cells is administered about 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days before the administration of the adjuvant. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days before the administration of the adjuvant.

[0195] In some embodiments, the composition containing modified immune cells is administered after the administration of the adjuvant. For example, the composition containing modified immune cells is administered approximately 1 hour to 1 week after the administration of the adjuvant. For example, in some embodiments, the composition containing modified immune cells is administered approximately 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days after the administration of the adjuvant. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days after the administration of the adjuvant.

[0196] Immune checkpoints are regulatory factors of the immune system that maintain the immune response during checkpoints. Immune checkpoint inhibitors can be used to enhance the immune response. In some embodiments, a composition containing modified immune cells is administered in combination with the administration of an immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered simultaneously. In some embodiments, the composition containing modified immune cells and the immune checkpoint inhibitor are administered sequentially.

[0197] In some embodiments, the composition containing modified immune cells is administered before the administration of an immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells is administered after the administration of an immune checkpoint inhibitor. For example, the composition containing modified immune cells is administered about 1 hour to about 1 week before the administration of an immune checkpoint inhibitor. For example, in some embodiments, the composition containing modified immune cells is administered about 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days before the administration of an immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days before administration of an immune checkpoint inhibitor.

[0198] In some embodiments, the composition containing modified immune cells is administered after the administration of an immune checkpoint inhibitor. For example, the composition containing modified immune cells is administered approximately 1 hour to 1 week after the administration of the immune checkpoint inhibitor. For example, in some embodiments, the composition containing modified immune cells is administered approximately 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days after the administration of the immune checkpoint inhibitor. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days after administration of the immune checkpoint inhibitor.

[0199] In some embodiments, the method includes multiple doses of a composition containing modified immune cells and / or multiple doses of a checkpoint inhibitor. For example, in some embodiments, the method includes 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 doses of the composition containing modified immune cells and / or the checkpoint inhibitor. For example, in some embodiments, the method includes fewer than 5, 10, 15, 20, 25, 30, 50, 75, 100, or 200 doses of the composition containing modified immune cells and / or the checkpoint inhibitor.

[0200] Exemplary immune checkpoint inhibitors target, without limitation, PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, the immune checkpoint inhibitor is one or more of antibodies that bind to PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In further embodiments, the antibody may be a full-length antibody or any variant, for example, an antibody fragment, a single-stranded variable fragment (ScFv), or an antigen-binding fragment (Fab), without limitation. In further embodiments, the antibody may be bispecific, tripspecific, or multispecific. In some embodiments, the immune checkpoint inhibitor is one or more chemical compounds that bind to and / or inhibit one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3. In some embodiments, the immune checkpoint inhibitor is one or more peptides that bind to and / or inhibit one or more of PD-1, PD-L1, CTLA-4, LAG3, or TIM-3.

[0201] Other exemplary immune checkpoint inhibitors target, without limitation, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. In some embodiments, the immune checkpoint inhibitor targets one or more of TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. In some embodiments, the immune checkpoint inhibitor is one or more of antibodies that bind to TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. In further embodiments, the antibody may be a full-length antibody or any variant, for example, an antibody fragment, a single-stranded variable fragment (ScFv), or an antigen-binding fragment (Fab), without limitation. In further embodiments, the antibody may be bispecific, tripspecific, or multispecific. In some embodiments, the immune checkpoint inhibitor is one or more chemical compounds that bind to and / or inhibit one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA. In some embodiments, the immune checkpoint inhibitor is one or more peptides that bind to and / or inhibit one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA.

[0202] Chemotherapy or radiotherapy can be used in combination with any one of the modified immune cells described herein to achieve additive or synergistic effects against cancer, such as HPV-related cancer. In some embodiments, the composition containing the modified immune cells is administered in combination with the administration of chemotherapy. In some embodiments, the composition containing the modified immune cells and the chemotherapy are administered simultaneously. In some embodiments, the composition containing the modified immune cells and the chemotherapy are administered sequentially.

[0203] In some embodiments, the composition containing modified immune cells is administered before the administration of chemotherapy. In some embodiments, the composition containing modified immune cells is administered after the administration of chemotherapy. For example, the composition containing modified immune cells is administered about 1 hour to about 1 week before the administration of chemotherapy. For example, in some embodiments, the composition containing modified immune cells is administered about 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days before the administration of chemotherapy. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days before chemotherapy is administered.

[0204] In some embodiments, the composition containing modified immune cells is administered after the administration of chemotherapy. For example, the composition containing modified immune cells is administered approximately 1 hour to 1 week after the administration of chemotherapy. For example, in some embodiments, the composition containing modified immune cells is administered approximately 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days after the administration of chemotherapy. In some embodiments, the composition containing modified immune cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days after the administration of chemotherapy.

[0205] In some embodiments, the method includes multiple doses of a composition containing modified immune cells and / or multiple doses of chemotherapy. For example, in some embodiments, the method includes 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 doses of the composition containing modified immune cells and / or chemotherapy. For example, in some embodiments, the method includes fewer than 5, 10, 15, 20, 25, 30, 50, 75, 100, or 200 doses of the composition containing modified immune cells and / or chemotherapy.

[0206] Exemplary chemotherapy may be cell cycle-dependent or cell cycle-independent. In some embodiments, chemotherapy comprises one or more chemotherapeutic agents. In some embodiments, the chemotherapeutic agents may target one or more of the following in cancer: cell division, DNA, or metabolism. In some embodiments, the chemotherapeutic agent is a platinum-based activator such as cisplatin, oxaliplatin, or carboplatin, but is not limited to these. In some embodiments, the chemotherapeutic agent is a taxane (such as docetaxel or paclitaxel). In some embodiments, the chemotherapeutic agent is 5-fluorouracil, doxorubicin, or irinotecan. In some embodiments, the chemotherapeutic agent is one or more of the following: alkylating agents, antimetabolites, antitumor antibiotics, topoisomerase inhibitors, or mitotic inhibitors. In some embodiments, chemotherapy comprises cisplatin. In some embodiments, one or more chemotherapy agents or immune checkpoint inhibitors may be combined with any of the modified immune cells described herein to treat or prevent HPV-related diseases.

[0207] Radiotherapy can be used in combination with any one of the modified T cells described herein to achieve additive or synergistic effects against cancer, such as HPV-related cancer. In some embodiments, the composition containing modified T cells is administered in combination with the administration of radiotherapy. In some embodiments, the composition containing modified T cells and radiotherapy are administered simultaneously. In some embodiments, the composition containing modified T cells and radiotherapy are administered sequentially. In some embodiments, the composition containing modified T cells is administered in combination with the administration of radiotherapy, in combination with chemotherapy, and / or in combination with an immune checkpoint inhibitor.

[0208] In some embodiments, the composition containing modified T cells is administered before the administration of radiotherapy. In some embodiments, the composition containing modified T cells is administered after the administration of radiotherapy. For example, the composition containing modified T cells is administered about 1 hour to about 1 week before the administration of radiotherapy. For example, in some embodiments, the composition containing modified T cells is administered about 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days before the administration of radiotherapy. In some embodiments, the composition containing modified T cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days before the administration of radiotherapy.

[0209] In some embodiments, the composition containing modified T cells is administered after the administration of radiotherapy. For example, the composition containing modified T cells is administered approximately 1 hour to 1 week after the administration of radiotherapy. For example, in some embodiments, the composition containing modified T cells is administered approximately 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 60 hours, 3 days, 4 days, 5 days, 6 days, or 7 days after the administration of radiotherapy. In some embodiments, the composition containing modified T cells is administered approximately 1 to 2 hours, 2 to 3 hours, 3 to 4 hours, 4 to 6 hours, 6 to 8 hours, 8 to 10 hours, 10 to 12 hours, 12 to 14 hours, 14 to 16 hours, 16 to 18 hours, 18 to 20 hours, 20 to 24 hours, 24 to 30 hours, 30 to 36 hours, 36 to 42 hours, 42 to 48 hours, 48 ​​to 60 hours, 60 hours to 3 days, 3 to 4 days, 4 to 5 days, 5 to 6 days, and 6 to 7 days after the administration of radiotherapy.

[0210] In some embodiments, the method includes multiple administrations of a composition containing modified T cells and / or multiple administrations of radiotherapy. For example, in some embodiments, the method includes 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 administrations of the composition containing modified T cells and / or radiotherapy. For example, in some embodiments, the method includes fewer than 5, 10, 15, 20, 25, 30, 50, 75, 100, or 200 administrations of the composition containing modified T cells and / or radiotherapy.

[0211] When HPV antigens are processed and presented to immune cells in the MHC, they can induce or enhance an immune response to the presented HPV epitope. In some embodiments, HPV antigens can be processed into MHC class I-restricted peptides. In some embodiments, HPV antigens can be processed into MHC class II-restricted peptides. In some embodiments, the immune response is enhanced. In further embodiments, the immune response to HPV antigens is enhanced. In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs). In some embodiments, administration of a composition containing modified immune cells to an individual results in the activation and / or proliferation of antigen-specific helper T(T) lymphocytes. h ) This leads to the activation and / or proliferation of cells.

[0212] In some embodiments, an effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 12 Contains modified immune cells. In some embodiments, an effective amount of the composition is about 1 × 10⁶ 6 pieces, about 1×10 7 pieces, about 1×10 8 pieces, about 1×10 9 pieces, about 1×10 10 pieces, about 1×10 11 pieces, or approximately 1 x 10 12 It contains one of the modified immune cells. In some embodiments, an effective amount of the composition is about 1 × 10 6 ~Approx. 1×10 7 pieces, about 1×10 7 ~Approx. 1×10 8 pieces, about 1×10 8 ~Approx. 1×10 9 pieces, about 1×10 9 ~Approx. 1×10 10 pieces, about 1×10 10 ~Approx. 1×10 11 pieces, or approximately 1 x 10 11 ~Approx. 1×10 12 Contains one of the individual modified immune cells.

[0213] In some embodiments, the method includes multiple doses of the composition containing modified immune cells. For example, in some embodiments, the method includes 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 doses of the composition containing modified immune cells. For example, in some embodiments, the method includes fewer than 5, 10, 15, 20, 25, 30, 50, 75, 100, or 200 doses of the composition containing modified immune cells. For example, in some embodiments, the method includes a first dose of the composition containing modified immune cells, followed by a second dose of the composition containing modified immune cells. The timing of the doses can also be modified to achieve the desired results. In some embodiments, the first dose of the composition to the individual occurs before the second dose of the composition. In some embodiments, the first dose is introduced to the individual more than one week, two weeks, three weeks, four weeks, one month, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, twelve months, eighteen months, or twenty-four months prior to the introduction of the second dose.

[0214] In some embodiments, the method includes multiple administrations of modified T cells. In some embodiments, the method includes any of approximately 2, 3, 4, 5, 6, 7, 8, 9, 10, or more than approximately 10 administrations. In some embodiments, the time interval between two consecutive administrations of modified T cells is approximately 1 day to approximately 1 month. In some embodiments, administrations are daily, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, weekly, every other week, or monthly. In some embodiments, consecutive administrations are given for up to 1 year or longer.

[0215] In certain embodiments, compositions comprising modified cells can be used to treat, prevent, and / or modulate the immune response in individuals having HPV-related diseases. In some embodiments, HPV-related diseases are HPV-related cancers. In some embodiments, HPV-related cancers are cervical diseases, anal diseases, oropharyngeal diseases, vaginal diseases, vulvar diseases, penile diseases, skin diseases, or head and neck diseases. In some embodiments, HPV-related diseases are HPV-related infectious diseases. Other HPV-related diseases include common warts, plantar warts, flat warts, anogenital warts, anal lesions, epidermal dysplasia, focal epithelial thickening, oral papillomas, verrucous cysts, and laryngeal papillomatosis.

[0216] In some aspects, this disclosure relates to the use of modified immune cells for treating HPV-related diseases, wherein the modified immune cells contain HPV antigens and adjuvants within the cells. In some embodiments, the Disclosure relates to the use of modified immune cells for treating HPV-related diseases, the method comprising the steps of administering an effective amount of a composition comprising modified immune cells to an individual, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cells, and the modified immune cells are prepared by a) passing a cell suspension comprising an input cell deformable constriction such that a deformable force is applied to the input cells as they pass through the constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells.

[0217] In some embodiments, the disclosure relates to a composition comprising modified immune cells for the manufacture of a pharmaceutical product used to treat an HPV-related disease, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cell. In some embodiments, the present disclosure relates to a composition comprising modified immune cells for the manufacture of a pharmaceutical used to treat HPV-related diseases, wherein the method comprises the steps of administering an effective amount of the composition comprising modified immune cells to an individual, the modified immune cells comprising an HPV antigen and an adjuvant within the cells, and the composition being prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction such that a deformation force is applied to the input cells as they pass through the constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells.

[0218] In some embodiments, the disclosure relates to a composition comprising modified immune cells for use in a method of medical treatment, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cell. In some embodiments, the present disclosure relates to a composition comprising modified immune cells for use in a method of medical treatment, the method comprising the steps of administering an effective amount of the composition comprising modified immune cells to an individual, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cells, and the composition is prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction such that a deformation force is applied to the input cells as they pass through the constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells.

[0219] In some embodiments, the disclosure relates to a composition comprising modified immune cells for use in methods of treating cancer, infectious diseases, or virus-related diseases, wherein the modified immune cells comprise HPV antigens and adjuvants within the cells. In some embodiments, the present disclosure relates to a composition comprising modified immune cells for use in the treatment of HPV-related diseases, the method comprising the steps of administering an effective amount of the composition comprising modified immune cells to an individual, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cells, and the composition is prepared by a) passing a cell suspension comprising input cells through a cell deformation constriction such that a deformation force is applied to the input cells as they pass through the constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells that is large enough for the HPV antigen and adjuvant to pass through, thereby forming perturbed input cells; and b) incubating the perturbed input cells with the HPV antigen and adjuvant for a time sufficient to allow the HPV antigen and adjuvant to enter the perturbed input cells, thereby producing modified immune cells.

[0220] In some embodiments, the present disclosure relates to a method for treating or preventing an HPV-related disease in an individual, comprising the step of administering modified immune cells associated with an HPV antigen to the individual, wherein the modified immune cells are prepared by a process comprising the step of a) incubating input cells with the HPV antigen and / or adjuvant for a time sufficient to allow the HPV antigen to associate with the cell surface of the input cells, thereby producing modified immune cells associated with the antigen.

[0221] In some embodiments, the modified immune cells of the present invention do not induce resistance in the individual. In some embodiments, the modified immune cells do not suppress the immune response in the individual. In some embodiments, the modified immune cells do not contain immunotolerogenic factors. In some embodiments, the modified immune cells are not administered in combination with immunotolerogenic factors. In some embodiments, the modified immune cells are not administered before, simultaneously with, or after the administration of immunotolerogenic factors.

[0222] composition In certain embodiments, the present invention provides a composition comprising modified immune cells, wherein the modified immune cells contain an HPV antigen and CpG ODN within the cells. In other embodiments, the present disclosure relates to a composition comprising modified immune cells, wherein the modified immune cells contain an HPV antigen within the cells, and the HPV antigen comprises an amino acid sequence having at least 90% similarity to any one of SEQ ID NOs. 18-26. In some embodiments, the HPV antigen comprises an amino acid sequence having at least 90% similarity to SEQ ID NOs. 23. In some embodiments, the HPV antigen comprises the amino acid sequence of SEQ ID NOs. 23. In some embodiments, modified immune cells are prepared by a) passing a cell suspension containing input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension, thereby causing a perturbation of the input cells large enough for the HPV antigen to pass through and forming perturbed input cells; and b) incubating the perturbed input cells and the HPV antigen for a time sufficient to allow the HPV antigen to enter the perturbed input cells, thereby producing modified immune cells. In further embodiments, a deformation force is applied to the input cells as they pass through the constriction. In some embodiments, the composition further comprises an adjuvant within the cells.

[0223] In some embodiments, the HPV antigen and / or adjuvant is located in the cytoplasm or endosomes. In some embodiments, the antigen and / or adjuvant is located in multiple compartments of the cell. In further embodiments, the antigen and / or adjuvant is located in compartments of the cell including the endoplasmic reticulum (ER), Golgi apparatus, lysosomes, exosomes, cell surface, or cell membrane. In some embodiments, the antigen and adjuvant are located in the same compartment. In some embodiments, the antigen and adjuvant are located in different compartments. For example, in some embodiments, the antigen is located in the cytoplasm, while the adjuvant is located in endosomes. In some embodiments, the modified immune cell further includes the HPV antigen and / or adjuvant outside the cell.

[0224] In some embodiments, the antigen is a polypeptide antigen. In some embodiments, the antigen is a modified antigen. For example, the antigen may be fused with a therapeutic agent or a targeted peptide. In some embodiments, the modified antigen is fused with a polypeptide. In some embodiments, the antigen is modified with a lipid. In some embodiments, the antigen is modified with a polysaccharide or carbohydrate moiety. In some embodiments, the antigen is associated with a virus. In some embodiments, the antigen is a viral antigen. An exemplary viral antigen is the HPV antigen. In further embodiments, the antigen is an HPV antigen. In some embodiments, the HPV antigen consists of a selection from the group HPV-16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73, and 82. HPV-16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82 are high-risk types that cause cancer, while HPV-26, 53, and 66 are "probably high-risk types" that cause cancer. In some embodiments, the antigen is the HPV-16 antigen or the HPV-18 antigen. In some embodiments, the HPV antigen consists of an HLA-A2 specific epitope. The HPV E6 and E7 genes are viral oncogenes, and the expression of these genes is required for malignant transformation. The E6 and E7 proteins target several negative regulators of the cell cycle, mainly p105Rb and p53, respectively, and thus interfere with cell cycle regulation. In further embodiments, the HPV antigen is the HPV E6 antigen or the HPV E7 antigen. In some embodiments, the modified immune cells contain the HPV E6 antigen and the HPV E7 antigen. In some embodiments, the HPV antigen is a polypeptide comprising an immunogenic epitope flanked by one or more heterologous peptide sequences at its N-terminus and / or C-terminus. In some embodiments, the HPV antigen is an HPV E7 epitope flanked by a sequence derived from the HPV E6 polypeptide. In some embodiments, the HPV antigen comprises amino acids having at least 90% similarity to any one of SEQ ID NOs: 18-26. In some embodiments, the HPV antigen comprises the amino acid sequence of SEQ ID NO: 23.

[0225] When adjuvants are added to immunogenic agents, they nonspecifically enhance or increase the immune response to the agent in the recipient host upon exposure to the mixture. Therefore, adjuvants can be used to boost the induction of immune cell responses (e.g., T cell responses) to antigens. In some embodiments, perturbed cells are incubated with both the HPV antigen and the adjuvant. Exemplary intracellular adjuvants include, but are not limited to, CpG ODN, interferon-α (IFN-α), interferon gene-stimulating factor (STING) agonists, retinoic acid-inducible gene I (RIG-I) agonists, and polyinosinic acid:polycytidylic acid (Poly-I:C). In some embodiments, the adjuvant is CpG ODN, IFN-α, STING agonist, RIG-I agonist, or Poly-I:C. In certain embodiments, the adjuvant is CpG ODN polynucleotide. In some embodiments, the CpG ODN adjuvant includes those selected from the group consisting of CpG ODN1585, CpG ODN2216, CpG ODN2336, CpG ODN1668, CpG ODN1826, CpG ODN2006, CpG ODN2007, CpG ODN BW006, CpG ODN D-SL01, CpG ODN2395, CpG ODN M362, and CpG ODN D-SL03 (InvivoGen). In some embodiments, the CpG ODN adjuvant is the CpG ODN1826 (TCCATGACGTTCCTGACGTT; SEQ ID NO: 30) or the CpG ODN2006 (also known as CpG ODN7909) (TCGTCGTTTTGTCGTTTTGTCGTT; SEQ ID NO: 31) oligonucleotide. Multiple adjuvants can also be used in conjunction with antigens to enhance the induction of an immune response. In some embodiments, the modified immune cells contain more than one adjuvant. In some embodiments, the modified immune cells contain any combination of adjuvants CpG ODN, IFN-α, STING agonist, RIG-I agonist, and poly(I:C).

[0226] In some embodiments, the modified immune cells contain the adjuvant at a concentration of about 0.01 μM to about 10 mM. For example, in some embodiments, the modified immune cells contain the adjuvant at a concentration of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the modified immune cells contain the adjuvant at a concentration higher than about 10 mM. In some embodiments, the modified immune cells contain the adjuvant at a concentration of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the modified immune cells contain the adjuvant at a concentration higher than about 10 mM. In some embodiments, the modified immune cells contain an adjuvant at a concentration of approximately 0.1 μM to 1 μM, 1 μM to 10 μM, 10 μM to 100 μM, 100 μM to 1 mM, or 1 mM to 10 mM.

[0227] In some embodiments, the modified immune cells contain HPV antigen at concentrations ranging from about 0.01 μM to about 10 mM. For example, in some embodiments, the modified immune cells contain adjuvants at concentrations of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the modified immune cells contain adjuvants at concentrations higher than about 10 mM. In some embodiments, the modified immune cells contain HPV antigen at concentrations of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the modified immune cells contain adjuvants at concentrations higher than about 10 mM. In some embodiments, the modified immune cells contain HPV antigen at one of the following concentrations: approximately 0.1 μM to approximately 1 μM, approximately 1 μM to approximately 10 μM, approximately 10 μM to approximately 100 μM, approximately 100 μM to approximately 1 mM, or 1 mM to approximately 10 mM.

[0228] In some embodiments, the ratio of HPV antigen to adjuvant is approximately 10,000:1 to approximately 1:10,000. For example, in some embodiments, the ratio of HPV antigen to adjuvant is one of approximately 10,000:1, approximately 1,000:1, approximately 200:1, approximately 100:1, approximately 10:1, approximately 1:1, approximately 1:10, approximately 1:100, approximately 1:1000, or approximately 1:10000. In some embodiments, the ratio of HPV antigen to adjuvant is approximately 10000:1 to 1000:1, approximately 1000:1 to 100:1, approximately 100:1 to 10:1, approximately 10:1 to 1:1, approximately 1:1 to 1:10, approximately 1:10 to 1:100, approximately 1:100 to 1:1000, and approximately 1:1000 to 1:10000.

[0229] In some embodiments, the modified immune cells further contain an active agent that enhances the viability and / or function of the modified immune cells compared to the corresponding modified immune cells that do not contain this active agent. In some embodiments, the active agent is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the stabilizer forms a complex with the HPV antigen and / or adjuvant. In some embodiments, the stabilizer increases the solubility and / or solution half-life of the HPV antigen and / or adjuvant. In some embodiments, multiple modified immune cells have higher viability than the corresponding modified immune cells that do not contain the stabilizer. In some embodiments, the active agent is albumin. In further embodiments, the albumin is mouse, bovine, or human albumin. In further embodiments, the active agent is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the divalent metal cation is Mg 2+ Zn 2+ , or Ca 2+ It is one or more of the following. In some embodiments, the active ingredient includes MSA.

[0230] In some embodiments of any one of the methods or compositions described herein, the modified immune cells further contain an active substance that enhances the viability and / or function of the modified immune cells compared to a group of corresponding modified immune cells that do not contain this active substance. In some embodiments, the modified immune cells further contain an active substance that enhances the viability and / or function of the modified immune cells during freeze-thaw cycles compared to corresponding modified immune cells that do not contain this active substance. In some embodiments, the active substance is a cryopreservative and / or frost preservative. In some embodiments, neither the cryopreservative nor the frost preservative causes more than 10% or 20% cell death in modified immune cells containing the active substance compared to corresponding modified immune cells that do not contain the active substance before any freeze-thaw cycle. In some embodiments, at least about 70%, about 80%, or about 90% of the modified immune cells survive up to 1, 2, 3, 4, or 5 freeze-thaw cycles. In some embodiments, the active substance is a compound, stabilizer, or cofactor that enhances endocytosis. In some embodiments, the active substance is albumin. In some embodiments, albumin is mouse, bovine, or human albumin. In some embodiments, the active ingredient is human albumin. In some embodiments, the active ingredient is one or more of the following: divalent metal cations, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. In some embodiments, the divalent metal cation is Mg 2+ Zn 2+ or Ca 2+One or more of the following. In some embodiments, the active substance is one or more of the following: sodium pyruvate, adenine, trehalose, dextrose, mannose, sucrose, human serum albumin (HSA), DMSO, HEPES, glycerol, glutathione, inosine, dibasic sodium phosphate, monobasic sodium phosphate, sodium metal ions, potassium metal ions, magnesium metal ions, chloride, acetate, gluconate, sucrose, potassium hydroxide, or sodium hydroxide. In some embodiments, the active ingredient is one or more of the following: sodium pyruvate, adenine, Rejuvesol®, trehalose, dextrose, mannose, sucrose, human serum albumin (HSA), PlasmaLyte®, DMSO, Cryostor® CS2, Cryostor® CS5, Cryostor® CS10, Cryostor® CS15, HEPES, glycerol, glutathione, and HypoThermosol®.

[0231] In some embodiments, the modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules. In further embodiments, the costimulatory molecules are B7-H2(ICOSL), B7-1(CD80), B7-2(CD86), CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112. In some embodiments, the cells contain nucleic acids that result in increased expression of one or more costimulatory molecules.

[0232] In some embodiments, immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphoid cells, basophils, or hematopoietic progenitor cells. In some embodiments, immune cells are not B cells. In some embodiments, immune cells are T cells. In some embodiments, immune cells are other than B cells. In some embodiments, modified T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, or natural killer T cells. In some embodiments, T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. MHC expression in allogeneic T cells can result in an innate immune response initiated in an individual in response to their administration, leading to a shortening of the half-life of such T cells. In some embodiments, T cells include further modifications that modulate MHC class I expression. In some embodiments, the T cells include further modifications that modulate MHC class II expression. In some embodiments, the T cells include further modifications that reduce MHC class I and / or MHC class II expression. In certain embodiments, the further modifications include using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease to reduce MHC class I and / or MHC class II expression. In some embodiments, the T cells include further modifications that increase MHC class I and / or MHC class II expression. In certain embodiments, the further modifications include using mRNA, plasmid DNA, or cDNA to increase MHC class I and / or MHC class II expression. In some embodiments, the innate immune response initiated in individuals in response to administration of further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to administration of corresponding modified T cells without further modifications in an allogeneic setting.In some embodiments, the circulating half-life of further modified T cells in the administered individual is increased compared to the circulating half-life of the corresponding modified T cells without further modification in the administered individual. In some embodiments, the modified T cells include one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, or natural killer T cells. In some embodiments, the T cells include one or more of CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, or γδ-T cells.

[0233] Immune cells and other cells can be used as a source of autologous or allogeneic cells. In some embodiments, the modified immune cells are allogeneic to the organism. In other embodiments, the modified immune cells are autologous to the organism. In some embodiments, the organism being treated is pre-conditioned to have reduced inflammation or a modulated inflammatory response.

[0234] PBMC composition As used herein, PBMCs can be isolated by leukocyte removal from whole blood obtained from an individual. PBMC compositions are also provided that are reconstituted by mixing different pools of PBMCs from the same or different individuals. In other examples, PBMCs may also be reconstituted by mixing different cell populations into a mixed cell composition having a production profile. In some embodiments, the cell population used to reconstitute the PBMCs is a mixed cell population (such as a mixture of one or more of T cells, B cells, NK cells, or monocytes). In some embodiments, the cell population used to reconstitute the PBMCs is a purified cell population (such as purified T cells, B cells, NK cells, or monocytes). In additional examples, the different cell populations used in the reconstitution of the PBMC composition may be isolated from the same individual (e.g., autologous) or from different individuals (e.g., allogeneic and / or heterogeneous).

[0235] Accordingly, in some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple input PBMCs include one or more T cells, B cells, NK cells, monocytes, dendritic cells, or NK-T cells. In some embodiments, the multiple input PBMCs include T cells, B cells, NK cells, monocytes, dendritic cells, or NK-T cells. In some embodiments, the multiple input PBMCs include one or more CD3+ T cells, CD20+ B cells, CD14+ monocytes, and CD56+ NK cells. In some embodiments, the multiple input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the multiple input PBMCs is essentially the same as the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in whole blood. In some embodiments, the input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the input PBMCs is essentially the same as the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the leukocyte removal product from whole blood. In some embodiments, the input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the input PBMCs differs from the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in whole blood by one of the following: 1%, 2%, 5%, 10%, 15%, 20%, 25%, 30%, 40%, or 50% or less. In some embodiments, the input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the input PBMCs differs by 10% or less from the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in whole blood.In some embodiments, the input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the input PBMCs differs from the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the leukocyte removal product from whole blood by one of the following: 1%, 2%, 5%, 10%, 15%, 20%, 25%, 30%, 40%, or 50% or less. In some embodiments, the input PBMCs include T cells, B cells, NK cells, and monocytes, and the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the input PBMCs differs from the ratio of T cells, B cells, NK cells, and monocytes to the total number of PBMCs in the leukocyte removal product from whole blood by one of the following: 10% or less.

[0236] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, about 25% to about 70% of the modified PBMCs are T cells. In some embodiments, about 2.5% to about 14% of the modified PBMCs are B cells. In some embodiments, about 3.5% to about 35% of the modified PBMCs are NK cells. In some embodiments, about 4% to about 25% of the modified PBMCs are NK cells. In some embodiments, where the immune cells are multiple PBMCs, at least about 90% to about 99% of the input PBMCs consist of T cells, B cells, NK cells, and monocytes. In some embodiments, at least one of about 80% to about 85%, about 85% to about 90%, about 90% to about 95%, or about 95% to about 99% of the input PBMCs consists of T cells, B cells, NK cells, and monocytes. In some embodiments, at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the input PBMCs consist of T cells, B cells, NK cells, and monocytes. In some embodiments, at least about 90% of the input PBMCs consist of T cells, B cells, NK cells, and monocytes. In some embodiments, the input PBMCs consist of T cells, B cells, NK cells, and monocytes.

[0237] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, at least about 90% to about 99% of the modified PBMCs consist of T cells, B cells, NK cells, and monocytes. In some embodiments, at least one of about 80% to about 85%, about 85% to about 90%, about 90% to about 95%, or about 95% to about 99% of the modified PBMCs consists of T cells, B cells, NK cells, and monocytes. In some embodiments, at least one of about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the modified PBMCs consists of T cells, B cells, NK cells, and monocytes. In some embodiments, at least about 90% of the modified PBMCs consist of T cells, B cells, NK cells, and monocytes.

[0238] In some embodiments of any one of the methods or compositions described herein, in which the immune cells are multiple PBMCs, at least about 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, or 75% of the input PBMCs are T cells. In some embodiments, at least about 25% of the input PBMCs are T cells. In some embodiments, at least about 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% of the input PBMCs are B cells. In some embodiments, at least about 2.5% of the input PBMCs are B cells. In some embodiments, at least about 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% of the input PBMCs are NK cells. In some embodiments, at least about 3.5% of the input PBMCs are NK cells. In some embodiments, at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 12%, 14%, 16%, 18%, 20%, 25%, 30%, 35%, or 40% of the input PBMCs are monocytes. In some embodiments, at least about 4% of the input PBMCs are monocytes. In some embodiments, at least about 25% of the input PBMCs are T cells; at least about 2.5% of the input PBMCs are B cells; at least about 3.5% of the input PBMCs are NK cells; and at least about 4% of the input PBMCs are monocytes.

[0239] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, at least about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, or 70% of the modified PBMCs are T cells. In some embodiments, at least about 20% of the modified PBMCs are T cells. In some embodiments, at least about 0.25%, 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% of the modified PBMCs are B cells. In some embodiments, at least about 2% of the modified PBMCs are B cells. In some embodiments, at least about 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% of the modified PBMCs are NK cells. In some embodiments, at least about 3% of the modified PBMCs are NK cells. In some embodiments, at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 12%, 14%, 16%, 18%, 20%, 25%, 30%, 35%, or 40% of the modified PBMCs are monocytes. In some embodiments, at least about 3% of the modified PBMCs are monocytes. In some embodiments, at least about 20% of the modified PBMCs are T cells; at least about 2% are B cells; at least about 3% are NK cells; and at least about 3% are monocytes.

[0240] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, one of about 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, or 90% or less of the input PBMCs are T cells. In some embodiments, about 70% or less of the input PBMCs are T cells. In some embodiments, one of about 5%, 10%, 12%, 14%, 16%, 18%, 20%, 22%, 25%, 30%, 35%, 40%, or 50% or less of the input PBMCs are B cells. In some embodiments, about 14% or less of the input PBMCs are B cells. In some embodiments, one of the following proportions of the input PBMCs is NK cells: approximately 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, or 60% or less. In some embodiments, one of the following proportions of the input PBMCs is NK cells: approximately 35% or less. In some embodiments, one of the following proportions of the input PBMCs is monocytes: approximately 5%, 10%, 12%, 14%, 16%, 18%, 20%, 22%, 25%, 30%, 35%, 40%, or 50% or less. In some embodiments, one of the following proportions of the input PBMCs is monocytes: approximately 4% or less. In some embodiments, one of the following proportions of the input PBMCs is T cells: approximately 25% or less; approximately 2.5% or less; approximately 3.5% or less; NK cells: approximately 4% or less.

[0241] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, one of about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, or 70% or less of the modified PBMCs are T cells. In some embodiments, about 20% or less of the modified PBMCs are T cells. In some embodiments, one of about 0.25%, 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% or less of the modified PBMCs are B cells. In some embodiments, about 2% or less of the modified PBMCs are B cells. In some embodiments, one of the following percentages of modified PBMCs is NK cells: approximately 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, 6%, 7%, 7.5%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, or 30% or less. In some embodiments, one of the following percentages of modified PBMCs is NK cells: approximately 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 12%, 14%, 16%, 18%, 20%, 25%, 30%, 35%, or 40% or less. In some embodiments, one of the following percentages of modified PBMCs is monocytes: approximately 3% or less. In some embodiments, less than 20% of the modified PBMCs are T cells; less than 2% are B cells; less than 3% are NK cells; and less than 3% are monocytes.

[0242] In some embodiments of any one of the methods or compositions described herein, in which the immune cells are multiple PBMCs, one of about 20%–25%, 25%–30%, 30%–35%, 35%–40%, 40%–45%, 45%–50%, 50%–55%, 55%–60%, 60%–65%, 65%–70%, or 70%–75% of the modified PBMCs is a T cell. In some embodiments, about 25%–70% of the modified PBMCs is a T cell. In some embodiments, one of about 1%–2.5%, 2.5%–4%, 4%–6%, 6%–8%, 8%–10%, 10%–12%, 12%–14%, 14%–16%, 16%–20%, or 20%–25% of the modified PBMCs is a B cell. In some embodiments, approximately 2.5% to 14% of the modified PBMCs are B cells. In some embodiments, one of the following percentages of the modified PBMCs is B cells: approximately 1% to 2%, 2% to 3.5%, 3.5% to 5%, 5% to 8%, 8% to 10%, 10% to 12%, 12% to 14%, 14% to 16%, 16% to 20%, or 20% to 25%. In some embodiments, approximately 3.5% to 35% of the modified PBMCs are NK cells. In some embodiments, one of the following percentages of the modified PBMCs is monocytes: approximately 2% to 4%, 4% to 6%, 6% to 8%, 8% to 10%, 10% to 12%, 12% to 14%, 14% to 16%, 16% to 20%, 20% to 25%, 25% to 30%, 30% to 35%, or 35% to 40%. In some embodiments, approximately 4% to 25% of the modified PBMCs are monocytes.

[0243] As used herein, PBMCs may also be produced after manipulating the composition of a mixed cell population of mononuclear blood cells (such as lymphocytes and monocytes). In some cases, input PBMCs are produced after depleting (e.g., depleting) a specific subpopulation (such as B cells) within a mixed cell population of mononuclear blood cells. The composition of a mixed cell population of mononuclear blood cells in an individual can be manipulated to produce a cell population that more closely resembles a leukocyte-removed product from whole blood in the same individual. In other cases, the composition of a mixed cell population of mononuclear blood cells (e.g., mouse splenocytes) can also be manipulated to produce a cell population that more closely resembles human PBMCs isolated from a leukocyte-removed product from human whole blood.

[0244] In some embodiments, construction-mediated delivery does not differentially modulate the viability of different subpopulations within the PBMC (such as B cells, T cells, NK cells, or monocytes) in a significant manner. In some embodiments, the conditioning process does not differentially modulate the viability of different subpopulations within the PBMC in a significant manner. In some embodiments, further addition of active agents (including, but not limited to, biopreservatives or active agents that enhance the function and / or viability of the PBMC) does not differentially modulate the viability of different subpopulations within the PBMC in a significant manner. Therefore, in some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the percentage of T cells in the multiple modified PBMCs and the percentage of T cells in the multiple input PBMCs differ by about 10% or less in number. In some embodiments, the percentage of T cells in multiple modified PBMCs and the percentage of T cells in multiple input PBMCs differ by one of the following numbers: approximately 5%, 8%, 10%, 12%, 14%, 16%, 18%, or 20% or less. In some embodiments, the percentage of B cells in multiple modified PBMCs and the percentage of B cells in multiple input PBMCs differ by one of the following numbers: approximately 10% or less. In some embodiments, the percentage of B cells in multiple modified PBMCs and the percentage of B cells in multiple input PBMCs differ by one of the following numbers: approximately 5%, 8%, 10%, 12%, 14%, 16%, 18%, or 20% or less. In some embodiments, the percentage of NK cells in multiple modified PBMCs and the percentage of NK cells in multiple input PBMCs differ by one of the following numbers: approximately 10% or less. In some embodiments, the percentage of NK cells in multiple modified PBMCs and the percentage of NK cells in multiple input PBMCs differ by one of the following numbers: approximately 5%, 8%, 10%, 12%, 14%, 16%, 18%, or 20% or less. In some embodiments, the percentage of monocytes in multiple modified PBMCs and the percentage of monocytes in multiple input PBMCs differ by approximately 10% or less.In some embodiments, the percentage of monocytes in multiple modified PBMCs and the percentage of monocytes in multiple input PBMCs differ by number by one of the following: approximately 5%, 8%, 10%, 12%, 14%, 16%, 18%, or 20% or less.

[0245] PBMC Conditioning In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple modified PBMCs are conditioned. In further embodiments, the multiple modified PBMCs are mature. In some embodiments, the multiple PBMCs are conditioned after stenosis-mediated delivery. Thus, in some embodiments, the process for preparing multiple modified PBMCs further includes the step of incubating the multiple modified PBMCs containing the antigen and / or adjuvant with a second adjuvant for a time sufficient to conditioned the modified PBMCs containing the antigen, thereby producing conditioned multiple modified PBMCs containing the antigen and / or adjuvant. In some embodiments, the process further includes the step of isolating the multiple modified PBMCs containing the antigen and / or adjuvant from the cell suspension prior to incubation with the adjuvant for conditioned the modified PBMCs.

[0246] In some embodiments, the concentration of the antigen incubated with the modified PBMC is about 0.01 μM to about 10 mM. For example, in some embodiments, the concentration of the antigen incubated with the modified PBMC is one of the following: about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the concentration of the antigen incubated with the modified PBMC is higher than about 10 mM. In some embodiments, the concentration of the antigen incubated with the modified PBMC is one of the following: about 0.01 μM to about 0.1 μM, about 0.1 μM to about 1 μM, about 1 μM to about 10 μM, about 10 μM to about 100 μM, about 100 μM to about 1 mM, or 1 mM to about 10 mM. In some embodiments, the concentration of the antigen incubated with the modified PBMC is about 0.1 μM to about 1 mM. In some embodiments, the concentration of the antigen incubated with the modified PBMC is approximately 0.1 μM to approximately 10 μM. In some embodiments, the concentration of the antigen incubated with the modified PBMC is 1 μM.

[0247] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple modified PBMCs are incubated with an adjuvant for about 1 to about 24 hours to prepare the modified PBMCs. In some embodiments, the multiple modified PBMCs are incubated with an adjuvant for about 2 to about 10 hours to prepare the modified PBMCs. In some embodiments, the multiple modified PBMCs are incubated with an adjuvant for about 3 to about 6 hours to prepare the modified PBMCs. In some embodiments, the multiple modified PBMCs are incubated with an adjuvant for any one of the following periods: about 1 hour, 2 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, 6 hours, 8 hours, 12 hours, 16 hours, 20 hours, or 24 hours to prepare the modified PBMCs. In some embodiments, the multiple modified PBMCs are incubated with an adjuvant for about 4 hours to prepare the modified PBMCs.

[0248] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple modified PBMCs are prepared before stenosis-mediated delivery. Thus, in some embodiments, the process of preparing the multiple modified PBMCs further includes the step of incubating the multiple input PBMCs and the adjuvant for a sufficient time to prepare the input PBMCs, thereby producing the prepared multiple input PBMCs. In some embodiments, a plurality of modified PBMCs containing an antigen are provided, prepared by a process comprising: a) incubating a plurality of input PBMCs and an adjuvant for a time sufficient to prepare the input PBMCs, thereby producing a plurality of prepared input PBMCs; b) passing a cell suspension containing the plurality of prepared input PBMCs through a cell deformation constriction, the diameter of which is a function of the diameter of the input PBMCs in the suspension, thereby causing a perturbation of the input PBMCs large enough for the antigen to pass through, thereby forming a plurality of prepared perturbed input PBMCs; and c) incubating the plurality of prepared perturbed input PBMCs and an antigen for a time sufficient to allow the antigen to enter the perturbed input PBMCs, thereby producing a plurality of prepared modified PBMCs containing the antigen. In some embodiments, the process further comprises isolating the plurality of prepared input PBMCs from the adjuvant to be prepared before passing the plurality of prepared input PBMCs through a cell deformation constriction.

[0249] In some embodiments, the concentration of the antigen incubated with the input PBMC is approximately 0.01 μM to approximately 10 mM. For example, in some embodiments, the concentration of the antigen incubated with the input PBMC is one of approximately 0.01 μM, approximately 0.1 μM, approximately 1 μM, approximately 10 μM, approximately 100 μM, approximately 1 mM, or less than approximately 10 mM. In some embodiments, the concentration of the antigen incubated with the input PBMC is higher than approximately 10 mM. In some embodiments, the concentration of the antigen incubated with the input PBMC is one of approximately 0.01 μM to approximately 0.1 μM, approximately 0.1 μM to approximately 1 μM, approximately 1 μM to approximately 10 μM, approximately 10 μM to approximately 100 μM, approximately 100 μM to approximately 1 mM, or 1 mM to approximately 10 mM. In some embodiments, the concentration of the antigen incubated with the input PBMC is approximately 0.1 μM to approximately 1 mM. In some embodiments, the concentration of the antigen incubated with the input PBMC is approximately 0.1 μM to approximately 10 μM. In some embodiments, the concentration of the antigen incubated with the input PBMC is 1 μM.

[0250] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple input PBMCs are incubated with the adjuvant for about 1 to about 24 hours to prepare the input PBMCs. In some embodiments, the multiple input PBMCs are incubated with the adjuvant for about 2 to about 10 hours to prepare the input PBMCs. In some embodiments, the multiple input PBMCs are incubated with the adjuvant for about 3 to about 6 hours to prepare the input PBMCs. In some embodiments, the multiple input PBMCs are incubated with the adjuvant for any one of the following periods: about 1 hour, 2 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, 6 hours, 8 hours, 12 hours, 16 hours, 20 hours, or 24 hours to prepare the input PBMCs. In some embodiments, the multiple input PBMCs are incubated with the adjuvant for about 4 hours to prepare the input PBMCs.

[0251] In some embodiments, a plurality of antigen-containing prepared PBMCs are provided, prepared by a step of incubating a plurality of PBMCs containing an antigen with an adjuvant for a sufficient time to prepare the PBMCs, thereby producing a plurality of antigen-containing prepared PBMCs. In some embodiments, a plurality of antigen-containing prepared PBMCs are provided, prepared by a step of incubating a plurality of PBMCs containing an adjuvant for a sufficient time to prepare the PBMCs before introducing the antigen into the PBMCs, thereby producing a plurality of antigen-containing prepared PBMCs.

[0252] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the concentration of the antigen incubated with the PBMCs is about 0.01 μM to about 10 mM. For example, in some embodiments, the concentration of the antigen incubated with the PBMCs is any of about 0.01 μM, about 0.1 μM, about 1 μM, about 10 μM, about 100 μM, about 1 mM, or less than about 10 mM. In some embodiments, the concentration of the antigen incubated with the PBMCs is higher than about 10 mM. In some embodiments, the concentration of the antigen incubated with the PBMCs is any of about 0.01 μM to about 0.1 μM, about 0.1 μM to about 1 μM, about 1 μM to about 10 μM, about 10 μM to about 100 μM, about 100 μM to about 1 mM, or 1 mM to about 10 mM. In some embodiments, the concentration of the antigen incubated with the PBMCs is about 0.1 μM to about 1 mM. In some embodiments, the concentration of the antigen incubated with the PBMC is approximately 0.1 μM to approximately 10 μM. In some embodiments, the concentration of the antigen incubated with the PBMC is 1 μM.

[0253] In some embodiments of any one of the methods or compositions described herein, where the immune cells are multiple PBMCs, the multiple PBMCs are incubated with an adjuvant for about 1 to about 24 hours to prepare the PBMCs. In some embodiments, the multiple PBMCs are incubated with an adjuvant for about 2 to about 10 hours to prepare the PBMCs. In some embodiments, the multiple PBMCs are incubated with an adjuvant for about 3 to about 6 hours to prepare the PBMCs. In some embodiments, the multiple PBMCs are incubated with an adjuvant for any one of the following periods: about 1 hour, 2 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, 5 hours, 5.5 hours, 6 hours, 8 hours, 12 hours, 16 hours, 20 hours, or 24 hours to prepare the PBMCs. In some embodiments, the multiple PBMCs are incubated with an adjuvant for about 4 hours to prepare the PBMCs.

[0254] In some embodiments, one or more co-stimulatory molecules are upregulated in a tuned modified PBMC compared to an untuned modified PBMC. In some embodiments, one or more co-stimulatory molecules are upregulated in a subpopulation of cells in a tuned modified PBMC compared to a subpopulation of cells in an untuned modified PBMC. In some embodiments, one or more co-stimulatory molecules are upregulated in B cells of a tuned modified PBMC compared to B cells in an untuned modified PBMC. In some embodiments, the co-stimulatory molecules are CD80 and / or CD86. In some embodiments, the co-stimulatory molecule is CD86. In some embodiments, CD80 and / or CD86 are upregulated in B cells of a tuned modified PBMC at levels higher than approximately 1.2 times, 1.5 times, 1.8 times, 2 times, 3 times, 4 times, 5 times, 8 times, or 10 times compared to B cells in an untuned modified PBMC. In some embodiments, CD80 and / or CD86 are upregulated in B cells of a combination of combinations In some embodiments, the expression of one or more of IFN-γ, IL-6, MCP-1, MIP-1β, IP-10, or TNF-α is increased in a subpopulation of cells in a modified PBMC compared to a subpopulation of cells in an unmodified PBMC.In some embodiments, the expression of one or more of IFN-γ, IL-6, MCP-1, MIP-1β, IP-10, or TNF-α is increased in the modified PBMCs by approximately 1.2-fold, 1.5-fold, 1.8-fold, 2-fold, 3-fold, 4-fold, 5-fold, 8-fold, or 10-fold compared to the unmodified modified PBMCs. In some embodiments, the expression of one or more of IFN-γ, IL-6, MCP-1, MIP-1β, IP-10, or TNF-α is increased in the modified modified PBMCs by approximately 1.2 to 1.5 times, 1.5 to 1.8 times, 1.8 to 2 times, 2 to 3 times, 3 to 4 times, 4 to 5 times, 5 to 8 times, 8 to 10 times, 10 to 20 times, 20 to 50 times, 50 to 100 times, 100 to 200 times, 200 to 500 times, or higher than 500 times, compared to the modified modified PBMCs.

[0255] Applicable In some embodiments, the present invention provides a method for treating and preventing HPV-related diseases and / or modulating an immune response in an individual having an HPV-related disease, comprising the step of administering a composition comprising modified immune cells to the individual, wherein the modified immune cells comprise an HPV antigen and an adjuvant within the cells. In some embodiments, the cells are isolated from a patient, modified by the disclosed method, and reintroduced into the patient. For example, a population of immune cells is isolated from a patient, passed through a stenosis to achieve delivery of the HPV antigen and adjuvant, and then reinjected into the patient to increase a therapeutic immune response to the HPV antigen. In some embodiments, the cells are isolated from an individual having an HPV-related disease, modified by the disclosed method, and reintroduced into ...

Claims

1. A composition comprising modified immune cells, wherein the modified immune cells contain within the cells the HPV antigen that has entered a perturbed input cell containing a perturbation large enough for the human papillomavirus (HPV) antigen to pass through, wherein the HPV antigen comprises one amino acid sequence of SEQ ID NOs. 18, 21, 23, or 25, and the modified immune cells further comprise an active substance, wherein the active substance enhances the viability and / or function of the modified immune cells compared to a corresponding modified immune cell that does not contain the active substance. The composition wherein the active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA.

2. The composition according to claim 1, wherein the modified immune cells further contain an adjuvant within the cells.

3. The composition according to claim 2, wherein the adjuvant supplied into the cell enters a perturbed input cell containing the HPV antigen, or the HPV antigen and the adjuvant, which contains a perturbation large enough for the adjuvant to pass through.

4. A composition comprising modified immune cells, wherein the modified immune cells contain the adjuvant and the HPV antigen within the cells, which have entered a perturbed input cell containing a perturbation large enough for the adjuvant and the HPV antigen to pass through, and the modified immune cells further contain an active substance, the active substance enhancing the viability and / or function of the modified immune cells compared to a corresponding modified immune cell that does not contain the active substance. The active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. A composition wherein the HPV antigen comprises one amino acid sequence of sequence numbers 18, 21, 23, or 25.

5. The composition is (a) Treatment or prevention of HPV-related diseases in individuals; and / or (b) Modulating the immune response in individuals with HPV-related disease. A composition according to any one of claims 1 to 4, for use in the following context.

6. A composition for use in treating or preventing HPV-related disease in an individual, or in modulating the immune response in an individual having HPV-related disease, wherein the composition comprises modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the adjuvant being supplied intracellularly and entering perturbation input cells comprising the HPV antigen and a perturbation large enough for the adjuvant to pass through, the modified immune cells further comprising an active substance, the active substance enhancing the viability and / or function of the modified immune cells compared to a corresponding modified immune cell without the active substance, The active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. A composition wherein the HPV antigen comprises one amino acid sequence of sequence numbers 18, 21, 23, or 25.

7. A composition for use in treating or preventing HPV-related disease in an individual, or in modulating the immune response in an individual having HPV-related disease, wherein the composition comprises modified immune cells, the modified immune cells comprising an HPV antigen and an adjuvant, the HPV antigen being supplied intracellularly and the adjuvant into a perturbation input cell comprising a perturbation large enough for the HPV antigen to pass through, the modified immune cells further comprising an active substance, the active substance enhancing the viability and / or function of the modified immune cells compared to a corresponding modified immune cell without the active substance, The active substance is a divalent metal cation, glucose, ATP, potassium, glycerol, trehalose, D-sucrose, PEG1500, L-arginine, L-glutamine, or EDTA. A composition wherein the HPV antigen comprises one amino acid sequence of sequence numbers 18, 21, 23, or 25.

8. The perturbation is caused by passing a cell suspension containing input cells through a cell deformation constriction, the diameter of which is a function of the diameter of the input cells in the suspension. (a) The diameter of the stenosis is less than the diameter of the cell; and / or (b) The stenosis is located in the channel; and / or (c) A deformable force is applied to the input cell as it passes through the constriction; and / or (d) The modified immune cells comprise an adjuvant, the concentration of the adjuvant incubated with the perturbation input cells being 0.1 μM to 1 mM; and / or (e) The concentration of the HPV antigen incubated with the perturbed input cells is 0.1 μM to 1 mM; and / or (f) The composition according to any one of claims 3 to 7, wherein the modified immune cells comprise an adjuvant, and the ratio of HPV antigen incubated with the perturbation input cells to the adjuvant is 10,000:1 to 1:10,000.

9. The composition according to claim 8, wherein the diameter of the stenosis is 20% to 99% of the diameter of the cell or 20% to less than 60% of the diameter of the cell.

10. (a) The HPV antigen and / or the adjuvant are present in the cytoplasm and / or endosomes; and / or (b) The antigen and / or adjuvant are present in numerous compartments of the cell; and / or (c) The modified immune cells further comprise an HPV antigen and / or adjuvant outside the cell; and / or (d) The modified immune cells contain an adjuvant, and the modified immune cells contain the adjuvant at a concentration of 0.1 μM to 1 mM; and / or (e) The modified immune cells contain the HPV antigen at a concentration of 0.1 μM to 1 mM; and / or (f) The composition according to any one of claims 1 to 9, wherein the modified immune cells comprise an adjuvant, and the ratio of the HPV antigen contained in the modified immune cells to the adjuvant is 10,000:1 to 1:10,000.

11. (a) The modified immune cells include an adjuvant, the adjuvant being CpG ODN, IFN-α, STING agonist, RIG-I agonist, or poly-I:C; and / or (b) The modified immune cells include an adjuvant, and the modified immune cells include more than one adjuvant; and / or (c) The HPV antigen is a pool of numerous polypeptides that elicit responses to the same and / or different HPV antigens; and / or (d) The HPV antigen is a polypeptide comprising an antigenic HPV epitope and one or more heterologous peptide sequences; and / or (e) The HPV antigen forms a complex with itself, with other antigens, or with the adjuvant; and / or (f) The HPV is an antigen derived from a cell lysate; and / or (g) The composition according to any one of claims 1 to 10, wherein the HPV antigen is HPV-16 or HPV-18 antigen.

12. (a) The modified immune cells include an adjuvant, the adjuvant being CpG ODN; and / or (b) The HPV antigen is composed of an HLA-A2 specific epitope; and / or (c) The HPV antigen is the HPV E6 antigen or the HPV E7 antigen; and / or (d) The modified immune cells contain HPV E6 antigen and HPV E7 antigen; and / or (e) The composition according to any one of claims 1 to 11, wherein the HPV antigen is a polypeptide comprising an antigenic epitope in which one or more heterologous peptide sequences are adjacent at its N-terminus and / or C-terminus.

13. (a) The modified immune cells comprise an adjuvant, the adjuvant being CpG ODN1018, CpG ODN1826, or CpG ODN2006; and / or (b) The HPV antigen is a pool of numerous polypeptides that elicit responses to the same and / or different HPV antigens, and the antigens in the pool of numerous antigens do not diminish the immune response directed to other antigens in the pool of numerous antigens; and / or (c) The composition according to claim 12, wherein the HPV antigen comprises the amino acid sequence of SEQ ID NO:

23.

14. (a) The HPV antigen may be processed into an MHC class I-restricted peptide; and / or (b) The composition according to any one of claims 1 to 13, wherein the HPV antigen can be processed into an MHC class II restrictive peptide.

15. (a) The modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules; and / or (b) The composition according to any one of claims 1 to 14, wherein the modified immune cells include T cells, the T cells comprising further modifications to modulate MHC class I expression and / or further modifications to modulate MHC class II expression.

16. (a) The modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules, the costimulatory molecules being B7-H2, B7-1, B7-2, CD70, LIGHT, HVEM, CD40, 4-1BBL, OX40L, TL1A, GITRL, CD30L, TIM4, SLAM, CD48, CD58, CD155, or CD112; and / or (b) The composition according to claim 15, wherein the modified immune cells include T cells, the T cells comprising modifications that reduce MHC class I and / or MHC class II expression, or the T cells comprising modifications that increase MHC class I and / or MHC class II expression.

17. (a) The modified immune cells are further modified to increase the expression of one or more of the costimulatory molecules, and the cells contain nucleic acids that result in the increased expression of the one or more costimulatory molecules; and / or (b) The composition according to claim 15 or 16, wherein the modified immune cells include T cells, the T cells comprising modifications that reduce MHC class I and / or MHC class II expression using siRNA, shRNA, CRISPR / Cas9, ZFN, TALEN, Cre recombinase, or meganuclease, or the T cells comprising modifications that increase MHC class I and / or MHC class II expression using RNA or plasmid DNA.

18. (a) The immune cells are T cells, dendritic cells, monocytes, macrophages, myeloid cells, granulocytes, neutrophils, mast cells, natural killer cells, innate lymphocytes, basophils, or hematopoietic progenitor cells; and / or (b) The composition according to any one of claims 1 to 17, wherein the immune cells are a mixed cell population.

19. (a) The immune cells are B cells or T cells; and / or (b) The composition according to any one of claims 1 to 18, wherein the immune cells are a plurality of PBMCs.

20. The composition is intended for use in treating or preventing human papillomavirus (HPV)-related diseases in individuals, or in modulating the immune response in individuals having HPV-related diseases, wherein the immune cells include T cells. (a) The innate immune response initiated in individuals in response to the administration of the further modified T cells in an allogeneic setting is reduced compared to the innate immune response initiated in individuals in response to the administration of the corresponding modified T cells in an allogeneic setting without the further modification; and / or (b) The composition according to any one of claims 1 to 19, wherein the circulating half-life of the further modified T cells in the administered individual is modulated compared to the circulating half-life of the corresponding modified T cells without the further modification in the administered individual.

21. The aforementioned immune cells include T cells, and the T cells are, (a) one or more of helper T cells, cytotoxic T cells, memory T cells, CIK cells, and natural killer T cells; or (b) One or more of the following: CD3+ T cells, CD4+ T cells, CD8+ T cells, CD45RA+ T cells, CD45RO+ T cells, and γδ-T cells. A composition according to any one of claims 1 to 20, comprising:

22. The composition is intended for use in the treatment or prevention of human papillomavirus (HPV)-related diseases in individuals, or in modulating the immune response in individuals with HPV-related diseases. (a) The composition is intended for use in modulating the immune response in an individual having an HPV-related disease, and the immune response is enhanced; and / or (b) The modified immune cells are allogeneic or autologous to the individual; and / or (c) The individual is pre-conditioned to have modulated inflammation and / or a modulated immune response; and / or (d) The modified immune cells are formulated for administration in combination with further adjuvants; and / or (e) The modified immune cells are formulated for administration in combination with the administration of immune checkpoint inhibitors; and / or (f) The modified immune cells are formulated for administration in combination with chemotherapy; and / or (g) The effective amount of the composition for use is 1 × 10 ６ ~1 x 10 １２ Including individual modified immune cells; and / or (h) The composition according to any one of claims 1 to 21, wherein the modified immune cells are formulated for multiple administrations.

23. The composition is intended for use in the treatment or prevention of human papillomavirus (HPV)-related diseases in individuals, or in modulating the immune response in individuals with HPV-related diseases. (a) The composition is intended for use in modulating the immune response in an individual having an HPV-related disease, and the immune response to the HPV antigen is enhanced; and / or (b) The modified immune cells are formulated for administration in combination with a further adjuvant, the further adjuvant being IFNα or CpG ODN; and / or (c) The modified immune cells are formulated for administration in combination with an immune checkpoint inhibitor, the immune checkpoint inhibitor targets one or more of PD-1, PD-L1, CTLA-4, LAG3, TIM-3, TIGIT, VISTA, TIM1, B7-H4 (VTCN1), or BTLA; and / or (d) The modified immune cells are formulated for administration in combination with chemotherapy, the chemotherapy comprising a platinum-based active agent; and / or (e) The composition according to claim 22, wherein the modified immune cells are formulated for multiple administrations, and the time interval between two consecutive administrations of the modified T cells is 1 to 150 days.

24. The composition is intended for use in the treatment or prevention of human papillomavirus (HPV)-related diseases in individuals, or in modulating the immune response in individuals with HPV-related diseases. (a) The modified immune cells are formulated for administration before, concurrently with, or after the administration of further adjuvants; and / or (b) The modified immune cells are formulated for administration before, concurrently with, or after administration of an immune checkpoint inhibitor; and / or (c) The modified immune cells are formulated for administration before, concurrently with, or after chemotherapy; and / or (d) The modified immune cells are formulated for administration in combination with chemotherapy, the chemotherapy comprising cisplatin; and / or (e) The composition according to claim 22 or 23, wherein the modified immune cells are formulated for multiple administrations, and the time interval between two consecutive administrations of the modified T cells is 1 to 30 days.

25. The composition is intended for use in the treatment or prevention of human papillomavirus (HPV)-related diseases in individuals, or in modulating the immune response in individuals with HPV-related diseases. (a) Administration of the composition containing the modified immune cells to the individual results in the activation and / or proliferation of HPV antigen-specific cytotoxic T lymphocytes (CTLs); and / or (b) Administration of the composition containing the modified immune cells to the individual is effective in activating the antigen-specific helper T(T) ｈ A composition according to any one of claims 1 to 24, which results in the activation and / or proliferation of cells.

26. The composition according to any one of claims 5 to 25, wherein the HPV-related disease is HPV-related cancer or HPV-related infectious disease.

27. The composition according to any one of claims 5 to 26, wherein the HPV-related disease is an HPV-related cancer, and the HPV-related cancer is cervical cancer, anal cancer, oropharyngeal cancer, vaginal cancer, vulvar cancer, penile cancer, skin cancer, or head and neck cancer.

Citation Information

Patent Citations

  • Intradermal HPV peptide vaccination

    JP2010530362A

  • Vaccine immunotherapy

    JP2012526853A

  • Delivery of biomolecules to immune cells

    JP2017533702A

  • Delivery of biomolecules to immune cells

    WO2016070136A1

  • Intracellular delivery of biomolecules mediated by a surface with pores

    WO2017041050A1