Animal model or human cells: efficacy evidence for approved monoclonal antibodies

“In a cynomolgus monkey model of collagen-induced arthritis (CIA), tocilizumab was shown to prevent both the local joint and the systemic inflammatory disease manifestations.”

“The cynomolgus monkey was chosen as the pharmacologically relevant species because tocilizumab cross-reacts with monkey IL-6R under in vitro and in vivo conditions.”

“Fremanezumab was tested in 2 rat models of headache (post-concussion headache and medication overuse headache) in which pain-related behaviors were measured after application of a presumed migraine stimulus.”

“No complete animal model of migraine is available. Restriction of blood flow via inhibition of CGRP after administration of fremanezumab was assessed in several animal models, including animal models of pain.”

“In vivo proof-of-concept studies were conducted in a cuticle bleeding model in New Zealand White (NZW) rabbits where hemophilia A-like conditions were simulated by administration of a neutralizing FVIII antibody.”

“Based on the pharmacology data and the lack of cross reactivity to rodent species, the rabbit was selected as the nonclinical efficacy species (hemophilia animal models).”

“In an angiotensin converting enzyme inhibitor (captopril)-induced edema murine model (heterozygous C1-INH-deficient (C1INH+/-) mice treated with 2.5 mg/kg captopril), garadacimab significantly inhibited the angiotensin-converting enzyme inhibitor-induced increase of vascular permeability.”

“Since garadacimab and the parental antibody 3F7 demonstrated cross-reactivity in mice, an in vivo efficacy was investigated in murine PSA and PCA models.”

“The efficacy of ANTHIM for treatment and prophylaxis of inhalational anthrax was studied in multiple studies in the cynomolgus macaque and NZW rabbit models of inhalational anthrax.”

“Because it is not feasible or ethical to conduct controlled clinical trials in humans with inhalational anthrax, the efficacy of ANTHIM for the treatment of inhalational anthrax is based on efficacy studies in NZW rabbits and cynomolgus macaques.”

“The potential therapeutic effects of ofatumumab in vivo were examined in severe combined immunodeficiency (SCID) mouse tumour xenograft models.”

“In contrast, ofatumumab did not bind to spleen sections obtained from mouse, rat, rabbit, pig and dog.”

“Mono-therapy with A4.6.1, bevacizumab, paclitaxel [76, 79], or doxorubicin [88] inhibited tumour growth and angiogenesis in tumours from human breast cancer [88], prostate cancer [79] and ovarian cancer [76].”

“The pharmacokinetics of bevacizumab was investigated in species where the antibody is expected to bind to and neutralise endogenous VEGF, i.e., cynomolgus monkeys [41] and rabbits (albeit with lower affinity than for human VEGF, see toxicology), and in mice and rats, where it is reported not to bind endogenous VEGF [42].”

“To assess the in vivo anti-tumor activity of avelumab, the Applicant utilized a syngeneic mouse model of colon cancer.”

“In vitro pharmacology studies demonstrated the ability of avelumab to bind to human, mouse, and cynomolgus monkey PD-L1 with comparable affinities.”

“In vivo studies showed a preventive action of brolucizumab on retinal vascular permeability, choroidal neovascularisation (both by IVT administration) and preretinal neovascularisation (IVT and IP administration).”

“In addition to binding to human VEGF-A, brolucizumab also binds to rodent VEGF-A allowing for evaluation in rodent efficacy models.”

“In Zhu et al, 2017, the antiviral activity of 1G7 was investigated in vivo by administering 1G7 i.m. at 8 different dose levels to cotton rats 1 day prior to challenging with RSV A2 or RSV B9320 strains at doses up to 3 mg/kg and 1.5 mg/kg, respectively.”

“The YTE modification was added to the mAb to prolong the terminal half-life of the antibody in humans, though the modification significantly decreases antibody exposure in rodents resulting in decreased serum antibody levels relative to the native human Fc. Therefore, the parental molecule, 1G7, was utilized for nonclinical pharmacology studies conducted in rodents.”

“The in vivo efficacy of caplacizumab was evaluated in a baboon model of TTP after preventive and therapeutic treatment. In this model mimicking the early onset of acquired TTP, the disease is induced by functional inhibition of ADAMTS13 through administration of an inhibitory monoclonal antibody (3H9).”

“The non-clinical pharmacokinetic properties of caplacizumab were analysed in guinea pigs and baboons as cross-reactive species, as well as in mice (not cross-reactive).”

“Certolizumab pegol was studied in two in vivo models where normal mice or rabbits were challenged with hTNFα. In these models, certolizumab pegol inhibited hTNFα induced effects. Effective doses ranged from 3-3000 µg/kg. certolizumab pegol also protected against development of arthritis in one transgenic mouse, serving as a disease model for polyarthritis, at 10 and 30 mg/kg”

“Certolizumab pegol cross-reacts poorly with TNF from rodents and rabbits, therefore in vivo efficacy was evaluated using animal models in which human TNFα was the physiologically active molecule.”

“Treatment with reslizumab decreased pulmonary eosinophilia after ovalbumin challenge in mice and guinea pigs, as well as in Ascaris-challenged monkeys. Reslizumab also reduced eosinophil influx into the skin of antigen challenged New Zealand rabbits.”

“Reslizumab binds with similar affinity to human, monkey, and mouse IL-5 (KD = 24, 20, and 31 pM; respectively, as determined by BIAcore biosensor assays).”

“Functionally in vivo, secukinumab inhibited IL-17A induced inflammatory reactions in two mouse models, the knee joint inflammatory model and the air pouch model where human IL-17A was produced locally.”

“Secukinumab does not recognize mouse and rat IL-17A, and therefore mouse surrogate antibody BZN035 was developed.”

“The mouse model used in the biodistribution study, athymic nude mice harboring subcutaneous human neuroblastoma xenografts, was the same model used for in vivo primary pharmacodynamics studies evaluating the efficacy of naxitamab in an animal model of disease.”

“Treatment with daratumumab effectively reduced growth of human lymphoma cells in SCID mice, both in the preventive and therapeutic settings.”

“Daratumumab binds to human and chimpanzee CD38, but not to CD38 of species typically used for nonclinical toxicology testing, ie, mouse, rat, rabbit, pig, and cynomolgus and rhesus monkey.”

“Although no preclinical models of atopic dermatitis were available a transgenic mouse expressing hIL-4 and hIL-4Rα was generated and used for proof of efficacy studies with dupilumab in models of type 2 immune responses.”

“Due to the lack of an in vivo model of atopic dermatitis the proof of principle experiments have been performed using classical models of type 2 immune response which have been shown to be mediated at least in part through signalling via IL-4 and IL-13.”

“The activity of ansuvimab-zykl was evaluated in three in vivo proof-of-concept studies in which rhesus monkeys were challenged with uniformly lethal doses of Kikwit 8U EBOV (1000 pfu). These studies demonstrated that a single dose of 50 mg/kg ansuvimab-zykl prevented EBOV-induced mortality in these animals as late as 5-days postchallenge relative to controls.”

“Given the challenges of conducting adequate and well-controlled trials for treatment of EBOV infection, the development program for ansuvimab-zykl was initially based on fulfilling the necessary criteria for potential approval under the Animal Rule pathway.”

“The anti-tumor activities of MuLuc63 and HuLuc63 were studied in xenograft mouse models with tumors grown from the human multiple myeloma cell lines L363 and OPM2.”

“Remarkably that this is true for mouse studies with human xenografts, indicating that the SLAMF7-mediated stimulation of NK-cell activity (not possible with mouse NK-cells) is not essential for the full activity of elotuzumab.”

“Clesrovimab and RB-1 LALA were administered at doses of 0.03, 0.09, 0.28, 0.83, or 2.5 mg/kg to cotton rats. Challenge with 1 x 105 pfu RSV strain A2 followed on the day after immunization.”

“The use of the parental antibody was justified by the fact that the YTE modification leads to increased binding to the rodent FcRn which, in turn, reduces the overall exposure to the Ab.”

“In immuno-deficient mice, GEO (human colon cancer) cell tumour growth was markedly reduced by cetuximab 1 mg twice weekly for 3-5 weeks and even more by a combination of cetuximab and VEGF antisense.”

“These cells do not express EGFR and the result supports the hypothesis that the antitumour effect of cetuximab is linked to blockade of EGFR.”

“In ovariectomized rats and monkeys (animal models of postmenopausal osteoporosis), romosozumab increased bone mass and bone strength without negatively affecting bone quality.”

“The Sprague Dawley rat and cynomolgus monkey were selected as the main rodent and non-rodent toxicology species as romosozumab binds to human, rat, and cynomolgus monkey sclerostin with similar affinity and is pharmacologically active in these species.”

“Evidence of pharmacodynamic activity of evinacumab was established following single and multiple dose (8 week) SC administration in normolipidemic (C57BL/6) and multiple hyperlipidemic (ApoE-/- , Ldlr-/- , db/db and high-fat diet fed) mouse models”

“In vitro, evinacumab binds the N-terminal domain of ANGPTL3 proteins containing LPL binding regions from humans, mice, rats, and monkeys with high affinity (sub to low-nanomolar KD values ranging from 0.3 to 1.3 nM).”

“In in vivo pharmacology studies, benralizumab, when given to naive or allergen-challenged cynomolgus monkeys, demonstrated rapid and pronounced reduction of eosinophils in blood, bone marrow and bronchoalveolar lavage fluid, demonstrating its pharmacological activity in vivo in this species.”

“Benralizumab binds to human and cynomolgus monkey IL-5Rα with KD values of 11 and 42 pM, respectively (as measured by BIAcore). Benralizumab did not cross-react with murine IL-5Rα.”

“IV Q7D obinutuzumab displayed superior anti-tumour efficacy in SCID mice carrying tumours derived from SUDHL-4 DLBCL, OCI-Ly-18 NHL, Z138 MCL, RL NHL and WSU-DLCL2 B cell lymphoma cell lines when compared to rituximab.”

“Obinutuzumab binds to Cynomolgus monkey (but not rodent) CD20 with an affinity and ADCC potency similar to that of human CD20; consequently, the Cynomolgus monkey was considered the only appropriate species in which to assess the safety of obinutuzumab.”

“The combination with paclitaxel produced the greatest tumour regression in vivo with the BT-474 cell line.”

“Since trastuzumab is a humanised MAb, significant species differences in pharmacokinetics are to be expected: rodent p185neu (corresponding receptor protein to human p185HER2) is not recognised, whereas in non-human primates trastuzumab recognises a receptor (as yet uncharacterised) in epithelial cells. However, unlike humans these primate species do not overexpress p185HER2 or produce shed antigen.”

“In vivo, adalimumab inhibited hTNFα-induced lethality in D-galactosamine sensitised mice and hTNFα-induced pyrexia in rabbits. Moreover, it prevented development of arthritis in two transgenic mouse models, the Tg197 (expressing a soluble human TNFα transgene) and Tg5453 (expressing a transgene that encodes human pro-TNFα), serving as disease models of polyarthritis.”

“Neutralisation of murine TNFα was considerably weaker (>2.0 x 10-7), and adalimumab did not bind rat TNFα.”

“The pharmacological activity of marstacimab was evaluated in vivo in mouse models of HA (FVIII-deficient) and HB (FIX-deficient). Marstacimab dose-dependently reduced blood loss in both HA (76%) and HB (65%) mouse models relative to controls.”

“Marstacimab showed similar binding affinity to the combined K1K2 domains (via the K2 domain) of TFPI from human, cynomolgus monkey, rabbit, mouse, and the Wistar Han rat.”

“Improvement of the disease pathology was demonstrated in the collagen antibody-induced arthritis (CAIA) Tg32 mouse model and increasing platelet levels was shown in the immune thrombocytopenia (ITP) Tg32 mouse model.”

“No proof-of-concept studies were conducted in gMG models; however, IV administration of nipocalimab to transgenic mice expressing human FcRn reduced autoantibody-mediated effects in models of collagen-induced arthritis and immune-mediated thrombocytopenia.”

“The Applicant used A375 (human melanoma) and HPAC (human pancreatic adenocarcinoma) cells enriched for CD4+ and CD8+ T cells. Treatment with durvalumab reduced the tumour volume in the HPAC xenograft model, resulting in a maximum tumour growth inhibition of 74%.”

“Durvalumab was found to only bind to human and cynomolgus monkey PD-L1, and the cynomolgus monkey was found to be the only relevant nonclinical species for the safety and toxicity studies.”

“In keeping with these findings, olaratumab exhibited in vivo anti-tumor activity in nude mice bearing human sarcoma xenografts.”

“The Applicant investigated the in vivo anti-tumor activity of REGN2810 in huPD-1 mice bearing syngeneic colorectal cancer xenografts”

“Since REGN2810 does not bind mouse PD-1, mice were genetically engineered to express a human/mouse PD-1 chimeric receptor (huPD-1 mice)”

“In vivo toripalimab was found to inhibit the growth of a human melanoma cell lines (624Mel) in a Xenograft Tumour NSG mouse model with adoptive CTL transfer and to inhibit the growth of murine colon MC38 cell line in a human PD-1 Knock-in Mouse.”

“Toripalimab did not bind to the ECD of mouse or rat PD-1.”

“The in vivo disease model used was the well established model of laser-induced CNV, in this case, in monkeys.”

“Identification of the sequence differences between monkey and humans (99% sequence homology, alignment predicting the protein products) resulted in a prediction of a 100% homology between the species on a protein level.”

“Margetuximab activities were demonstrated in in vivo efficacy studies applying different mouse xenograft models with HER2+ human breast cancer lines.”

“The modified Fc domain reduced binding to mFcγRIII (mCD16), mFcγRII (mCD32), and mFcγRI (mCD64).”

“The in vivo antitumor activity of tafasitamab monotherapy was evaluated using subcutaneous xenograft models in severe combined immunodeficiency (SCID) mice engrafted with human Raji and RAMOS lymphoma cells. Tafasitamab treatment resulted in reduction in tumor growth.”

“Finally, tafasitamab binds with substantially greater affinity to mouse FcγRs compared to the native human IgG1 antibody. Hence, human xenograft mouse tumour models allow for testing of the antitumour efficacy of tafasitamab in vivo.”

“In vivo, mepolizumab decreased peripheral eosinophil counts in Cynomolgus monkeys and reduced pulmonary eosinophilia in response to A. suum challenge in an acute model of asthma.”

“The species specificity of mepolizumab was restricted to human and nonhuman primate IL-5 only. Mepolizumab did not bind to mouse or rat IL-5.”

“reduced HO formation in two conditional knock-in mouse models of FOP, along with efficacy data from the phase 2 Trial R2477-FOP-1623 (Trial 1623, LUMINA-1).”

“Given that pempulimab does not bind mouse PD-1, the Applicant investigated the in vivo antitumor activity of pempulimab in a human PD-1 knock-in genetically engineered mouse model (HuGEMM) implanted subcutaneously with MC38 mouse colon adenocarcinoma cells.”

“Given that pempulimab does not bind mouse PD-1, the Applicant investigated the in vivo antitumor activity of pempulimab in a human PD-1 knock-in genetically engineered mouse model (HuGEMM)”

“Weekly IV/IP administrations of pertuzumab demonstrated anti-tumour efficacy in various tumour xenografts models based on breast cancer, prostate, non-small cell lung cancer and ovarian cell lines expressing either low or high levels of HER2.”

“Treatment of mice implanted with tumor cell lines derived from NSCLC, pancreatic, and colorectal tumors with necitumumab resulted in varying degrees of tumor growth inhibition, depending, in part, on EGFR status of cells.”

“The antitumor activity of mogamulizumab was evaluated in mouse xenograft models of CTCL and ATL. Male SCID mice (Fox CHASE C.B-17/Icr-scidJcl) were subcutaneously injected with the human CTCL cell line HH or the ATL cell line TL-Om1.”

“One mouse study not only evaluated the effects of alirocumab on lipid turnover, but also the effects of alirocumab to prevent cardiovascular adverse events. Alirocumab, atorvastatin, or alirocumab/atorvastatin combination therapy was administered to APOE*Leiden.CETP mice fed a western diet.”

“Administration of alirocumab to a hyperlipaemic ApoE-/- mouse model, which was also fed a modified diet, profoundly reduced serum LDL-C concentrations as a result of binding to murine PCSK-9.”

“The rat tail bleeding model represents a situation with limited blood loss. Also the intracranial haemorrhage in mice is associated with limited bleeding. In contrast, the pig liver trauma model is associated with severe blood loss, haemorrhagic shock and ultimately mortality.”

“As it is difficult to obtain controlled information from humans treated in emergency situations, these non-clinical PD studies were considered justified and the results were adequately reflected in section 5.1 of the SmPC.”

“The disease model in ovariectomized cynomolgus monkeys is considered a relevant model for the indication of postmenopausal osteoporosis.”

“Denosumab is a fully human IgG2 mAb and does not recognize rodent RANKL. Several studies have been conducted with a transgenic huRANKL knock-in mouse model expressing chimeric RANKL that can be recognized and neutralized by denosumab.”

“HGS submitted the BLA for Raxibacumab to the FDA on May 14, 2009, containing efficacy studies of raxibacumab alone and in combination with antibiotic treatment against inhalational anthrax in both rabbits and non-human primates.”

“Raxibacumab is the first monoclonal antibody antitoxin against anthrax to seek licensure under Animal Rule (21 CFR 601, Subpart H, 2002).”

“Studies in normal mice given rhTNFα and different transgenic mice models that expressed recombinant forms of soluble or transmembrane human TNFα showed that infliximab inhibited the pathological effects mediated by human TNFα.”

“It was also shown that infliximab could neutralise human and chimpanzee TNFα with similar potency, while TNFα from a number of other species (rhesus cynomolgous, pigtail macaque, cotton-top tamarin, baboon, marmoset, pig, rabbit, rat and mouse) was not affected by infliximab.”

“A xenograft model with a Non-Hodgkin Lymphoma (NHL) cell line in severe combined immunodeficient (SCID) mice (SU DHL-8 tumours; study IMP23­018) was used to provide in vivo POC for isatuximab.”

“From the pharmacokinetic point of view, the SCID mice with a xenograft consisting of cells from a CD38 expressing cancer cell line was the most relevant species for non-clinical efficacy testing.”

“Investigations were also performed in vivo in a mouse model of arthritis. A single dose of golimumab (1, 3, 10 or 30 mg/kg) significantly delayed onset of clinical symptoms compared to a vehicle control group.”

“No neutralization of rat or mouse TNFα by golimumab was observed.”

“In an in vivo mouse ear thickening model, risankizumab inhibits human IL-23-induced ear thickening as well as the induction of tissue IL-17 and IL-22.”

“Risankizumab inhibits cynomolgus monkey and human IL-23 functions, but not mouse or rat.”

“CNTO 1275 (STELARA, ustekinumab) was pharmacologically active in non-human primates in a marmoset experimental autoimmune encephalomyelitis model (EAE) and was effective in suppressing EAE incidence and severity in marmosets.”

“Direct evaluation of carcinogenic potential of CNTO 1275 (STELARA, ustekinumab) in carcinogenicity studies or rodent tumour immune surveillance models is precluded by the limited species reactivity of CNTO 1275 (STELARA, ustekinumab) which does not bind to or neutralise IL-12 or IL-23 from mice or rats.”

“In a study of mice bearing H460 human lung tumour xenografts, twice-weekly dosing with CNTO 328 (25 mg/kg) caused a reduction in tumour volume and a delay in time to tumour endpoint by 7.6 days compared to PBS control.”

“Siltuximab binds to and neutralizes human and non-human primate IL-6 but does not inhibit mouse, rat, dog, pig or guinea pig IL-6.”

“Palivizumab was able to reduce the virus titres in the lungs when applied prophylactically or as a treatment for an ongoing infection from approx. 105 to 102 Pfu/gram lung at concentrations of 5 mg/kg after either IM or IV administration.”

“For in vivo pharmacology studies, the Applicant generated murine/human chimeric antibodies of atezolizumab to minimize immunogenicity in mice and evaluated PD-L1 blockade in syngeneic mouse models of melanoma and colorectal cancer. Anti-PD-L1 antibody reduced tumor growth by ≥ 80%.”

“The Applicant generated mouse IgG2a/human chimeric antibodies for murine in vivo studies in order to reduce immunogenicity, which was observed in prior mouse studies.”

“In vivo studies using murine xenograft tumour models and human PBMCs showed tislelizumab efficacy against several tumour cell lines.”

“From the results shown, tislelizumab seems to specifically bind to PD-1 (and to cynomolgus PD-1, but not to murine PD-1) with KD in the order of 0.1-0.2 nM.”

“The humanized form of ibalizumab was also tested in SIV-infected rhesus monkeys. Anti-ibalizumab antibodies were produced in the treated monkeys, which neutralized ibalizumab, reduced exposure and inhibited efficacy. Nevertheless, reductions in SIV viral load were seen in the monkeys up to day 10 of treatment.”

“Since it was shown in vitro that ibalizumab is also active against SIV, this is considered an appropriate animal model.”

“The studies in the guinea pig EAE model on the efficacy of natalizumab were performed at 3 mg/kg dose (MAD) administered s.c. on day 7 and 14 days after EAE induction. Natalizumab induced beneficial effects on active EAE, as a model of MS.”

“The affinity of AN100226m and AN100226 (the humanized form of the antibody) for guinea pig lymphocytes was very similar to that for human lymphocytes (Kd = 0.3 nM).”

“In a separate study, administration of ch14.18 with PBMCs and IL-2 prevented the establishment and growth of GD2+ melanoma tumours, whereas treatment with ch14.18 alone or PBMCs and IL-2 alone had no effect on tumour growth.”

“Treatment with inebilizumab effectively depleted CD19+ B cells and plasma cells in EAE huCD19 Tg mice. This led to a global reduction of autoantibodies in blood and in the CNS and to significantly reduced disease scores (Chen et al, 2014 and Chen et al, 2016).”

“The in vivo pharmacological activity of inebilizumab was evaluated in huCD19 Tg mice as the relevant species since this mAb was specific for human CD19 and had poor cross-reactivity with CD19 on the surface of B cells from rat, rabbit, baboon, rhesus monkey, African green monkey, or cynomolgus monkey.”

“Panitumumab was also shown to inhibit cell growth in vitro and in vivo (xenotransplants in mice) in the same cell line.”

“In vivo anti-tumour activity of zolbetuximab was evaluated in mouse xenograft models of human gastric cancer.”

“the epitope recognised was identified in the 1st extracellular loop of CLDN18.2, a region that is highly conserved across species.”

“Except for skin test reactivity studies in Cynomolgus monkeys sensitised to ragweed, no animal model of disease was used.”

“At the time the studies were performed there were no well-characterised primate asthma models available. Rodent asthma models were not considered because of the species specificity of omalizumab”

“Key pharmacodynamic effects of OMS721 were evaluated in an ex vivo model of aHUS and an in vivo mouse model of TMA.”

“OMS721 displayed functional activity in mouse, rabbit, and cynomolgus monkey, with potencies 3.8, 2.7 and 10-fold less than human. No functional activity was observed in rats.”

“The MC-38 colon carcinoma tumour line was implanted into the human CTLA-4 transgenic mice to evaluate the anti-tumour activity of ipilimumab.”

“Ipilimumab did not show cross-reactivity with or binding to CTLA-4 from rats, mice or rabbits, but showed specific binding to cynomolgus monkey recombinant CTLA-4 and activated T-cells”

“Preclinical in vivo efficacy of actoxumab+bezlotoxumab was evaluated in the Golden Syrian hamster model of infection (at doses of 0.5 to 50 mg/Kg) and in the mouse CDI model (at doses of 2, 5, 10 and 50 mg/Kg).”

“The protective effects of bezlotoxumab have been shown in vitro in TcdB neutralization assays, and in nonclinical studies in mouse and hamster models of CDI (in combination with actoxumab, since bezlotoxumab alone has poor efficacy in rodent models).”

“The in vivo proof of principle experiments have been performed using models of type 2 immune responses mediated through signalling via IL-13. While some of the in vitro data suggest applicability of tralokinumab in treatment of AD, no relevant model of AD (e.g. ex vivo or in vitro human skin models) was presented in a proof of concept study.”

“Several of the models were performed in mice whereas previously discussed the endogenous mouse IL-13 is not bound by tralokinumab. For these studies, injection of human recombinant IL-13 was used since it can bind the mouse IL-13 receptor and mediate signalling in this manner, however, the activity of human IL-13 at the mouse receptor is less than that of mouse IL-13 at the same receptor.”

“In vivo activity was demonstrated in a cynomolgus model of capsaicin induced increase in dermal blood flow, an effect mediated at least partly by CGRP. There are no animal migraine models.”

“There are no animal migraine models. Therefore, all evidence for a therapeutic effect is derived from clinical data.”

“The in vivo studies in mice as well as in monkeys demonstrated that belimumab inhibits splenocyte proliferation and decreased the number of B-cells in spleen, lymph node and circulating B cells.”

“Although belimumab binds mouse BLyS, it does so with a 10-fold lower affinity for murine BLyS compared with belimumab binding to human or cynomolgus monkey BLyS. Moreover, multiple injections of belimumab in mice led to a strong anti-drug antibody response that altered the PK and in some cases led to death by an anaphylactoid–like response.”

“B cell depletion by ublituximab was also observed in in vivo mouse models of the clearance of cells derived from a patient suffering from B cell chronic lymphocytic leukaemia (B-CLL).”

“However, the feasibility of such a model due to ublituximab cross-reaction with rodent CD20 and its the strong immunogenicity observed in nonhuman primate studies, is questioned.”

“The primary pharmacodynamic effect evaluated in the in vivo pharmacology studies was lymphocyte cell depletion. The studies were conducted in the human CD52 (huCD52) transgenic mouse model and in the cynomolgus monkey.”

“Efficacy studies in animal models of MS were not conducted with alemtuzumab, because clinical efficacy data were already available from clinical experience with alemtuzumab for MS at the time huCD52 transgenic mice were developed and characterised.”

“In an in vivo C57BL/6 mouse airway inflammation model, lebrikizumab reduced human IL-13-induced inflammatory reactions;”

“Nevertheless, this issue is not pursued further as the AD and airway inflammation model share some similarities regarding the IL-13 / STAT6 signalling routes, and lack of binding to rodent target limits further the studies in rodents.”

“In vivo activity was demonstrated in male rat and in female cynomolgus models of capsaicin-induced increase of dermal blood flow (DBF).”

“Chronic administration of sutimlimab in monkeys led to nearly complete inhibition of classical complement activity (up to 98% reduced activity) at both dose levels.”

“The pharmacology of satralizumab and tocilizumab was studied in terms of inhibitory effects on IL-6-stimulated C-reactive protein (CRP) production (i.e. on classical signaling), free and total (both free IL-6R and receptor bound to satralizumab) soluble IL-6R concentration (i.e. on trans-signaling).”

“As satralizumab showed cross-reactivity in cynomolgus monkey but not in mice or rats, in vivo pharmacology of satralizumab was studied in cynomolgus monkeys. The efficacy of satralizumab in NMOSD models in vivo have not been investigated since no NMOSD model has been established in monkeys.”

“whereas the clearest effect was demonstrated following depemokimab 1 mg/kg treatment with a normalised EO ratio below 0.5 from day 2 to 155, showing a prolonged inhibition of eosinophil count.”

“No in vivo efficacy studies were conducted, what is agreed given the lack of cross reactivity with mouse, rat, rabbit and dog IL-5.”

“Since canakinumab does not react with rodent IL-1β, the in vivo activity of canakinumab was demonstrated in models of inflammation induced by human IL-1β in rodents (i.e. mouse joint inflammation, rat fever, neutrophil infiltration into mouse air pouch).”

“Due to the lack of crossreactivity to IL-1β from rodent species, the in vivo activity of Canakinumab could not be demonstrated in classical animal disease models.”

“Primary PD studies and studies to characterize the antibody and its mechanism of action were conducted in vitro in human and cynomolgus monkey monocytes, CHO cells expressing the CSF-1R extracellular domain, and human cancer cells. Additionally, PD endpoints were incorporated into single-and repeat-dose studies in cynomolgus monkeys.”

“The in vivo toxicology of axatilimab was assessed in the cynomolgus monkey, the only toxicology species in which axatilimab binds to CSF-1R and blocks CSF-1-mediated signaling of the receptor.”

“Because ocrelizumab binds only to human and non-human primate CD20, in vitro studies were conducted using human-derived cell lines and in vivo studies were conducted in cynomolgus monkey.”

“Ocrelizumab specifically binds CD20 in humans and nonhuman primates. It does not bind CD20 in rodents. Therefore, cynomolgus monkey was selected as the relevant species for nonclinical investigation.”

“A pharmacology study in an animal model of PNH was not conducted; however, the Applicant assessed complement inhibition in a pharmacokinetics/pharmacodynamics study and in repeat-dose toxicology studies in monkeys as well as in patients with PNH.”

“These in vitro results confirmed that rat, mice and rabbit were not pharmacologically relevant species and therefore were excluded to be selected as species in the toxicity studies.”

“Evolocumab elicited expected pharmacological responses in hamsters and monkeys; evolocumab lowered total cholesterol and LDL-cholesterol in the species tested and decreased HDL-cholesterol in hamsters.”

“The Applicant identified the hamster and monkey as pharmacologically relevant species for toxicology testing with evolocumab; both species express PCSK9, to which evolocumab binds with high affinity.”

“The B-cell depleting effect was demonstrated in Cynomolgus monkeys both in peripheral blood and lymphatic tissues.”

“Antibodies against rituximab may appear after 2 weeks of treatment. Therefore, no xenograft experiments are feasible. No animal model is available for evaluating the antineoplastic effect.”

“Lanadelumab pretreatment was found to inhibit carrageenan induced rat oedema in a dose-dependent manner with a maximum inhibition at 30 mg lanadelumab/kg by 73% following SC and 62% following IV infusion, respectively, suggesting an inhibition of pKal functional effects in vivo.”

“Due to the lack of established non-clinical animal models of HAE, the in vivo pharmacodynamics of lanadelumab was evaluated in the carrageenan-induced rat paw oedema model, in which bradykinin is a known proinflammatory mediator, and that can be used to access mechanistic aspects of pKal inhibition.”

“In nude mice bearing human tumours expressing IGF-1R, single dose of teprotumumab (6 mg/kg IP) lead to down-regulation of IGF-1R on the tumour cells as demonstrated ex vivo.”

“Guselkumab administration was shown to attenuate rhIL-23 induced increases in serum levels of cytokines IL- 1α and G-CSF without any significant change in TNFα, IP-10 or GM-CSF levels.”

“It is accepted that no nonclinical model of plaque psoriasis that recapitulates all facets of human psoriasis is available.”

“The findings from this study suggest that in the presence of human C5, ALXN1210 showed more than three-fold extension in half-life compared with eculizumab. In addition, the serum half-life of ALXN1210 relative to eculizumab resulted in increased PD, as evidenced by prolonged haemolytic inhibition.”

“Neither ALXN1210 nor eculizumab exhibit detectable pharmacological activity against any non‐human species tested, as was predicted by epitope mapping studies and sequence alignments of human and non‐human C5 molecules, and confirmed by in vitro binding and activity studies.”

“Following administration to C5 humanized mice and normal monkeys, pozelimab blocked classical and alternative pathway-initiated hemolysis in serum samples ex vivo.”

“Importantly, though a CD55 knockout mouse model exists, it does not display the CHAPLE disease phenotype, and thus is not considered an acceptable animal disease model in which one could demonstrate potential clinical effectiveness of pozelimab in subjects with CHAPLE disease.”

“In vivo activity of eptinezumab was demonstrated in a model of capsaicin-induced increase in dermal blood flow in rats and cynomolgus monkeys and a model of vasodilation induced intradermal injection of CRGP in rabbits.”

“There are no animal models for migraine; thus, the evidence for a therapeutic effect are derived from the clinical data.”

“These studies in monkeys included the evaluation of a pharmacodynamic endpoint – retifanlimab binding to peripheral T cells by flow cytometry”

“Retifanlimab cross reacts with PD-1 from cynomolgus and rhesus monkeys, but not with PD-1 from non￾primate species. Thus, in vivo studies were limited to cynomolgus monkeys with retifanlimab delivered by IV infusion.”

“In vivo studies in the Tg2576 mouse model were conducted using the murine analog, ch12F6A, a chimeric anti-Aβ monoclonal antibody.”

“However, the intended target of aducanumab is aggregated forms of Aβ peptide. Neither rat nor monkey is a particularly relevant species for general toxicity because of the lack of aggregated amyloid in rodent or non-aged monkey.”

“A murine anti-FGF23 antibody was used to characterize the PD effects and mode of action of inhibiting FGF23 in Hyp mice.”

“it was shown to bind with similar binding affinity (KD~10-11 M) to human, cynomolgus monkey and rabbit FGF23. KRN23 shows no binding to mouse, rat or dog FGF23.”

“DC101 alone significantly inhibited the in vivo tumor growth of both NCI-N87 and MKN-45 gastric carcinoma cells. In addition, DC101 administered as a monotherapy inhibited 10 of 17 human primary gastric tumor fragments implanted into athymic mice.”

“Since ramucirumab is not cross-reactive with mouse VEGFR2, its antitumor activity and anti-angiogenic mechanism of action could not be studied in the mouse.”

“In vivo, colitic cotton-top tamarins (these develop a type of colitis spontaneously) were used to determine the functional relevance of α4β7 in the pathogenesis of chronic inflammatory disease. Animals were given 2.0 mg/kg Act-1, the murine vedolizumab precursor, for 8 days.”

“In vivo, colitic cotton-top tamarins (these develop a type of colitis spontaneously) were used to determine the functional relevance of α4β7 in the pathogenesis of chronic inflammatory disease.”

“For the disease model, perforin-deficient (pfp-/-) mice were infected with lymphocytic choriomeningitis virus and received single injections of XMG1.2 at 3, 10, 30 or 100 mg/kg on day 6 post-infection.”

“Emapalumab neutralized human IFNγ in a dose dependent manner, with an IC50 of 0.9 μg/mL (equivalent to 6 nM), but was not active in mice or rats.”

“Two different models of colitis, a T-cell dependent and an anti-CD40 model, were used and where efficacy of SCH-900598 was assessed.”

“Due to the lack of cross-reactivity with mouse IL-23p19 and in order to test the potency efficacy in vivo a surrogate antibody, SCH 900598, was generated against the mouse IL-23p19.”

“A surrogate tremelimumab anti‐CTLA‐4 antibody prolonged survival in multiple subcutaneous mouse tumor models when administered alone and demonstrated increased response rates when administered in combination with an anti‐mouse PD‐L1 antibody.”

“Tremelimumab also bound CTLA‐4 expressed on human and cynomolgus T cells stimulated to express CTLA‐4, but did not bind stimulated mouse, rat, hamster, or rabbit T cells;”

“Due to lack of cross‐reactivity between dostarlimab and mouse PD‐1, a surrogate anti‐mouse PD‐1 antibody (RMP1‐14) was used to assess the potential for PD‐1 inhibition to suppress tumor growth in an MC38 syngeneic mouse colon tumor model.”

“Because dostarlimab did not bind to mouse PD‐1, the Applicant developed a mouse surrogate anti‐PD‐1 antibody (RMP1‐14) to investigate the effects of PD‐1 pathway inhibition on tumor growth.”

“Pertinent to the proposed indication of rheumatoid arthritis, the effect of blocking IL-6Rα was evaluated in a murine model of collagen-induced arthritis. In a prophylactic setting, REGN844 prevented development of joint inflammation and bone erosion.”

“Sarilumab binds human and Cynomolgus monkey IL-6Rα with an equilibrium dissociation constant (Kd) of 54 pM and 123 pM, respectively, but no binding of sarilumab to the murine IL-6Rα was observed.”

“The antitumor activity induced by blocking PD-1 was shown in vivo in mice in a syngeneic tumour model where MC38 colon adenocarcinoma was injected subcutaneously and mice were treated using a commercially available homologous anti-mouse PD-1 monoclonal antibody J43 at 2 mg/kg and 10 mg/kg doses every 3-4 weeks.”

“Given the lack of cross-reactivity of MK-3475 with murine PD-1, a commercially available hamster anti-murine PD-1 mAb, J43, was used as surrogate for in vivo pharmacology studies.”

“Donanemab binds to N3pE containing amyloid beta plaques, which were demonstrated to be reduced via typical antibody clearance mechanisms in aged PDAPP mice treated with murine surrogate mE8c in a dose-and effector-function dependent manner.”

“Donanemab is not pharmacologically active in normal rodent or nonrodent; therefore, toxicity studies were conducted in cynomolgus monkey (6-week IV donanemab) to assess for any off-target toxicity, and in the PDAPP transgenic mouse model (6-week and 6-month SC mE8c, a murine surrogate) to assess for relevant on-target effects”

“Therefore, a series of in vivo studies were performed to assess the ability of the murine analog, mAb158, to alter the levels of Aβ protofibrils in the brains of two mouse models of Alzheimer’s disease (Tg2576 and APPArcSwe).”

“In vivo studies in transgenic animal models to characterize the pharmacodynamic effects of lecanemab were not conducted because of immunogenicity.”

“In vivo pharmacology studies investigating the itch-inhibiting effect of nemolizumab in a cynomolgus monkey model and effects of an anti-mouse IL-31RA surrogate antibody in a mite antigeninduced AD mouse model were also provided.”

“Based on presented data nemolizumab appears to cross react only to IL-31RA in cynomolgus monkeys. No monkey AD model exists, however, in vivo studies were conducted to assess the effect of nemolizumab in the cynomolgus monkey in an IL-31-induced itch model.”

“Neutralization of IL-23 using a surrogate mouse antibody, LSN2479016, demonstrated efficacy in two murine models of inflammatory bowel disease.”

“Considering the absence of binding to rodent IL-23, a mouse surrogate antibody, LSN2479016, was also developed to enable its use in proof of concept studies.”

“The administration of anti-mouse PD-1 antibody 4H2 resulted in delayed tumor progression in the MC38 colon carcinoma, Sa1N fibrosarcoma, and J558”

“Because nivolumab was unable to bind to PD-1 from rodents, the Applicant developed a murine surrogate anti-PD-1 antibody to investigate the effects of PD-1 pathway inhibition on tumor growth.”

“The ability of a mouse surrogate (4470) of rozanolixizumab to increase clearance of IgG was evaluated in a mouse disease model of immune thrombocytopenia (ITP).”

“Affinity (KDs) for rozanolixizumab for mouse and rat FcRn was much weaker (nanomolar range) and binding to rabbit FcRn was not observed at all.”

“The in vivo proof-of-concept was investigated in a mouse disease model of lupus, using a mouse surrogate anti-IFNAR1 mAb.”

“Anifrolumab does not bind to murine IFNAR1 and does not inhibit the biological activity of murine IFN-α.”

“The proof of concept for brodalumab was demonstrated in a mouse model of skin inflammation by the use of surrogate antibodies against mouse IL-17RA.”

“No cross-reactivity was found with rat or mouse IL-17RA.”

“Due to the species specificity of eculizumab, potential pharmacological activities in vivo have been explored with murine antibodies targeting rodent C5. Since there are no murine models of PNH, studies targeting other therapeutic areas with potential involvement of C5 activation (arthritis, lupus, airway inflammation and cardiac allograft models) have been presented.”

“Due to the species specificity of eculizumab, potential pharmacological activities in vivo have been explored with murine antibodies targeting rodent C5. Since there are no murine models of PNH, studies targeting other therapeutic areas with potential involvement of C5 activation (arthritis, lupus, airway inflammation and cardiac allograft models) have been presented.”

“In vivo, BI 674304 reduced Imiquimod-induced skin inflammation in mice. BI 674304 also reduced IL-36-induced skin inflammation and IL-33 production in the skin of mice.”

“Spesolimab does not bind to IL36R in common nonclinical species.”

“Two surrogate antibodies that cross react with murine IL-17 were used in nonclinical collagen-induced arthritis (CIA) and experimental autoimmune encephalomyelitis (EAE) models.”

“Ixekizumab is a humanized IgG4 monoclonal antibody that binds to human and monkey IL-17A. Ixekizumab does not bind to mouse or rat IL-17A.”

“M702 (anti-mouse TSLP surrogate antibody) decreased airway hyperresponsiveness and total leukocytes (neutrophils and eosinophils) in the bronchoalveolar lavage fluid (BALF) of animals treated with M702 in a mouse model of OVA-induced asthma.”

“Because of the lack of binding of tezepelumab to mouse TSLP, M702, a mouse surrogate antibody was developed for in vivo studies.”

“In pharmacology studies in non-obese diabetic (NOD) mice, a model of spontaneous autoimmune development of diabetes, a single 5-day treatment cycle with the surrogate delayed the onset and reduced the incidence of diabetes by approximately 5 weeks and 30%, respectively.”

“CD3 is poorly conserved across species. Teplizumab cross-reacts only with human, gorilla, chimpanzee, and gibbon CD3+ T cells.”

“The impact of blocking APRIL interaction with its receptors was evaluated in a grouped ddY (gddY) IgA mouse nephropathy model using the mAb 4540 mouse surrogate and in healthy cynomolgus monkeys with sibeprenlimab.”

“Due to lack of binding to rodent APRIL, direct effects of sibeprenlimab were assessed in rodent models using a mouse surrogate antibody (mAb 4540).”

“The published in vitro data demonstrate that crizanlizumab blocks binding of human P-selectin to its ligand PSG-1, blocks neutrophil binding to P-selectin, blocks leukocyte rolling and adhesion to human vascular endothelium under shear stress, inhibits interaction of neutrophils with stimulated platelets, and blocks binding of human sickled red cells to human endothelium under flow.”

“Primary pharmacodynamics was only studied ex vivo with SelG1 and SEG101 because of occurrence of immunological reactions and species differences in target recognition of P-selectin, which hamper animal PD studies.”

“Primary and secondary pharmacodynamics studies with bimekizumab were conducted in vitro. Pharmacodynamic activity of bimekizumab was investigated in primary cultures of normal human dermal fibroblasts (NHEK) and epithelial keratinocytes (NHDF).”

“No in vivo pharmacodynamics assessment has been conducted given the low expression of IL- 17A and IL-17F in normal tissues, as no direct pharmacodynamics markers are available, and as no appropriate and relevant animal model of PSO is available.”

“Receptor blocking profiles of Simulect and its murine progenitor RFT5 were comparable; they demonstrate equal potency in their ability to inhibit the binding of radiolabelled IL-2 on the IL-2 receptor expressed on T-lymphocytes.”

“Simulect bound specifically to the α-chain of the primate IL-2-receptor, but not to the dog IL-2 receptor.”

“A significant increase in fold change IFN-γ production was observed for cosibelimab relative to the isotype control antibody (huIgG1).”

“cosibelimab selectively binds to monkey and human PD-L1 with high affinities, but not to murine PD-L1.”

“In vitro studies demonstrated that DAC-HYP prevents IL-2 induced proliferation and cytokine secretion in human and monkey primary lymphocytes.”

“DAC-HYP had no pharmacological effect on primary lymphocytes isolated from rats or mice.”