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Anti-Acinetobacter Baumannii single-chain variable fragments provide therapeutic efficacy in an immunocompromised mouse pneumonia model

Abstract

Background

The emergence of carbapenem-resistant and extensively drug-resistant (XDR) Acinetobacter baumannii as well as inadequate effective antibiotics calls for an urgent effort to find new antibacterial agents. The therapeutic efficacy of two human scFvs, EB211 and EB279, showing growth inhibitory activity against A. baumannii in vitro, was investigated in immunocompromised mice with A. baumannii pneumonia.

Results

The data revealed that infected mice treated with EB211, EB279, and a combination of the two scFvs showed better survival, reduced bacterial load in the lungs, and no marked pathological abnormalities in the kidneys, liver, and lungs when compared to the control groups receiving normal saline or an irrelevant scFv.

Conclusions

The results from this study suggest that the scFvs with direct growth inhibitory activity could offer promising results in the treatment of pneumonia caused by XDR A. baumannii.

Peer Review reports

Background

The carbapenem-resistant Acinetobacter baumannii (CRAB) has become one of the most clinically critical pathogens in medical care due to its severe threat to human health [1, 2]. Nosocomial infections, including pneumonia, bloodstream infection, urinary tract infection, and skin, soft tissue and bone infections caused by CRAB particularly threaten newborns, immunocompromised individuals, and patients hospitalized at intensive care unit (ICU) [3,4,5,6,7,8,9,10,11]. In particular, CRAB pneumonia, which has a high in-hospital mortality rate of up to 46%, has received a great deal of attention [1, 12]. Antimicrobial resistance (AMR), ranking as one of the top ten health threats in the world, has severely compromised the treatment of CRAB infections, particularly its tendency to be extensively drug-resistant (XDR), only susceptible to limited antibiotics (e.g. colistin and tigecycline), and pan-resistant isolates that are resistant to all antibiotics [11, 13]. Accordingly, the World Health Organization (WHO) has classified CRAB as a critical priority pathogen (Priority 1) for which functional antibiotics capable of combating CRAB infections are urgently needed [13].

The use of immunotherapy to combat respiratory infections caused by particular pathogens can significantly improve the effectiveness of treatment and minimize the side effects resulted from antibiotics [14,15,16]. Monoclonal antibody (mAb) is considered the pronounced arm of immunotherapy due to its great ability in recognition and binding to particular antigens leading to a range of simple (e.g. blockade of target) to multifaceted events (e.g. antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity) [16,17,18]. Of note, mAbs can enhance the efficacy of existing antibiotics (e.g. additive or synergistic effect) and minimize the adverse effects of toxic antibiotics (e.g. colistin) [19], or improve drug delivery to target sites (e.g. DSTA4637S comprised of a monoclonal THIOMAB™ IgG1 specific to S. aureus β-O-linked N-acetylglucosamine conjugated with a rifamycin analogue [dmDNA31]) [20, 21]. In this regard, several studies developed mAbs targeting different sites of A. baumannii (e.g. capsule, outer membrane protein A (OmpA), and Omp34) [22,23,24,25,26]. Compared to mAbs, antibody fragments have been less explored but include benefits such as better accessibility to hidden epitopes, development of bispecific antibodies, and cost-effective routes of production [27, 28]. Among the various antibody fragments, single-chain variable fragments (scFvs) stand out due to their particular construct containing only the variable regions of both the heavy and light chains [27,28,29,30]. The scFv antibody fragments benefit from their small size, suitable binding ability, and low immunogenicity [28, 31, 32]. In particular, the emergence of bactericidal scFvs as new antimicrobial biotherapeutics, exhibiting significant growth inhibitory activity against some pathogens in vitro and in vivo [30, 33,34,35,36,37,38,39], has opened up a new window of hope in combating against health-threatening pathogens. Targeting bacterial cell membranes and causing the disintegration of the lipid bilayer which leads to cytoplasmic leakage and bacterial death are introduced the major mechanism of actions of bactericidal scFvs [30, 34, 40]. Several bactericidal scFvs have been developed against pathogens such as A. baumannii [40], Pseudomonas aeruginosa [34, 35], relapsing fever Borrelia [41], and Staphylococcus aureus [30, 33], some of which could provide protection in the mouse model of infection.

In the previous report, we identified the two fully human scFvs, EB211 and EB279, which exerted bactericidal activity against A. baumannii [40] by displacing Mg2+ and interrupting the integrity of the outer membrane (data not shown). As part of the present study, we first determined histopathologically whether EB211, EB279, and a cocktail of the two scFvs have undesirable effects on healthy mice. Afterwards, the protective activity of the scFv was assessed in immunocompromised mice with pneumonia caused by an XDR A. baumannii strain, either alone or in a combination with another scFv twice a day (BID) for three days.

Results

The non-toxic potential of EB211 and EB279 in healthy mice

The in vivo assessment of toxicity of EB211 and EB279 demonstrated no marked histopathological changes in the kidneys and liver of mice injected intraperitoneally twice daily with EB211 (12 mg/kg) and EB279 (12 mg/kg), and the combination of the EB211 and EB279 scFvs (6 mg/kg of each) for 72 h, similar to mice injected intraperitoneally twice daily with colistimethate sodium (CMS; 30 mg/kg) or normal saline (NS) (Fig. 1).

Fig. 1
figure 1

EB211 and EB279 caused no toxicity to the kidneys and liver of mice. No noticeable histopathological changes were observed in the kidneys and liver of mice injected intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), or a cocktail of the two scFvs (CKT; 6 mg/kg of each) for 72 h, similar to mice injected intraperitoneally twice daily with colistimethate sodium (CMS; 30 mg/kg) or normal saline (NS). Green arrowheads: sinusoids, V: central venules, Yellow arrowheads: hepatocytes, Yellow arrows: renal tubule and renal corpuscle

Treatment effectiveness of EB211 and EB279 in immunocompromised mice with A. baumannii pneumonia

Twice daily treatment of immunocompromised mice, infected with A. baumannii A.b.56 at the 50% lethal dose (LD50; 4×107 CFU per mouse), with EB211 (12 mg/kg), EB279 (12 mg/kg), a combination of the two scFvs (6 mg/kg of each), or CMS (30 mg/kg) for 72 h, resulted in 100% survival rate after seven days, while a 50% survival rate was observed in mice treated twice daily with NS or MEH158 (an irrelevant scFv; 12 mg/kg) (Fig. 2A). Furthermore, once daily treatment of immunocompromised infected mice with EB211 for 72 h increased the survival rate to 75% after seven days (Fig. 2A).

Fig. 2
figure 2

Administration of EB211 and EB279 led to an increased survival rate and a reduced bacterial burden in the lungs of immunocompromised mice with A. baumannii pneumonia. Immunocompromised mice were inoculated intranasally with XDR A. baumannii A.b.56 at a 50% lethal dose (LD50; 4×107 CFU per mouse). Immunocompromised infected mice were administered intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), a cocktail of the two scFvs (CKT; 6 mg/kg of each), an irrelevant scFv (MEH158; an S. aureus-specific scFv) (12 mg/kg), colistimethate sodium (CMS; 30 mg/kg), or normal saline (NS), or once daily with EB211 (12 mg/kg) (Q24H) two hours after inoculation for 72 h. (A) The survival rate of mice (n = 8) monitored daily for up to seven days after infection. The log-rank test was used to compare Kaplan-Meier survival curves. *p < 0.05 for EB211, EB279, CKT, or CMS versus NS. The results represent three independent experiments. (B) Bacterial burden in the lungs of immunocompromised infected mice (n = 6) administered with the scFvs, CMS, or NS after 24, 48, and 72 h of infection. The results represent the mean ± SEM of three independent experiments. *p < 0.05 for EB211 (Q24H) versus NS; **p < 0.01 for EB211, EB211 (Q24H), EB279, CKT, or CMS versus NS.

The administration of EB211, EB279, a cocktail of the two scFvs, or CMS at two hours post-infection led to a significant reduction in A. baumannii burden in the lungs of mice at 24, 48, and 72 h of infection compared to mice treated with NS or MEH158 (Fig. 2B). Despite a high bacterial load in the lungs of NS-treated mice, twice daily administration of EB211, EB279, a cocktail of the two scFvs, or CMS resulted in the highest bacterial load reduction in the lungs of mice at 72 h of infection (Fig. 2B).

The histopathological examination of the kidneys, liver, and lungs from immunocompromised infected mice administered with EB211, EB279, or a cocktail of the two scFvs twice daily for 72 h demonstrated the significant protective effect of the scFvs against pneumonia caused by A. baumannii A.b.56. After 72 h of infection, tubular degeneration and necrosis, inflammatory cell infiltration (leukocytes) into the interstitial tissue surrounding the tubules in the cortex and medulla, and diminished and distorted glomeruli, and eosinophilic casts were observed in immunocompromised mice treated twice daily with NS, MEH158, or CMS, or once daily with EB211 (Fig. 3). In contrast, immunocompromised infected mice treated twice daily with EB211 and EB279 (alone and a cocktail of the two scFvs) showed no marked pathological symptoms, except cytoplasmic vacuolation and pale-staining plus diminished and distorted glomeruli, which might associate with cyclophosphamide (Fig. 3).

After 72 h, micro and macro vesicles were detected in the livers of immunocompromised uninfected and infected mice (Fig. 4). Bacterial communities and infiltration of inflammatory cells (neutrophils and mononuclear cells) along with spotty necrosis‌ were the marked pathological symptoms found in immunocompromised infected mice treated twice daily with NS or MEH158 (Fig. 4). Furthermore, mice treated twice daily with CMS or once daily with EB211 showed similar pathologic symptoms, except for bacterial foci. None of the immunocompromised infected mice treated twice daily with EB211, EB279, or a cocktail of the two scFvs showed severe pathological changes (Fig. 4).

Fig. 3
figure 3

Administration of EB211 and EB279 prevented severe tissue damage in the kidneys of immunocompromised mice with A. baumannii pneumonia. Immunocompromised mice were inoculated intranasally with XDR A. baumannii A.b.56 at a 50% lethal dose (LD50; 4×107 CFU per mouse). Immunocompromised infected mice were administered intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), a cocktail of the two scFvs (CKT; 6 mg/kg of each), an irrelevant scFv (MEH158; an S. aureus-specific scFv) (12 mg/kg), colistimethate sodium (CMS; 30 mg/kg), or normal saline (NS), or once daily with EB211 (12 mg/kg) (Q24H) two hours after inoculation for 72 h. Healthy (H), immunocompromised uninfected (IU), and immunocompromised infected (Iinf) mice were killed at 72 h of infection, and the kidneys were collected. Tissue sections were stained with hematoxylin-eosin and microscopically evaluated for histopathological alterations. Tubular degeneration and necrosis, and infiltration of inflammatory cells (leukocytes) into the interstitial tissue surrounding the tubules in the cortex and medulla were the marked pathological symptoms in Iinf mice treated twice daily with NS, MEH158, or CMS, or once daily with EB211 (Q24H). After 72 h of infection, no severe pathological signs were detected in the kidneys of Iinf mice treated twice daily with EB211, EB279, or CKT for 72 h. Black arrowheads: tubular necrosis, Green arrows: diminished and distorted glomeruli, White arrows: infiltration of inflammatory cells (leukocytes), White arrowheads: cytoplasmic vacuolation and pale-staining plus fragmented cytoplasm

Fig. 4
figure 4

Administration of EB211 and EB279 prevented severe tissue damage in the liver of immunocompromised mice with A. baumannii pneumonia. Immunocompromised mice were inoculated intranasally with XDR A. baumannii A.b.56 at a 50% lethal dose (LD50; 4×107 CFU per mouse). Immunocompromised infected mice were administered intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), a cocktail of the two scFvs (CKT; 6 mg/kg of each), an irrelevant scFv (MEH158; an S. aureus-specific scFv) (12 mg/kg), colistimethate sodium (CMS; 30 mg/kg), or normal saline (NS), or once daily with EB211 (12 mg/kg) (Q24H) two hours after inoculation for 72 h. Healthy (H), immunocompromised uninfected (IU), and immunocompromised infected (Iinf) mice were killed at 72 h of infection, and the liver were collected. Tissue sections were stained with hematoxylin-eosin and microscopically evaluated for histopathological alterations. Bacterial communities and infiltration of inflammatory cells (neutrophils and mononuclear cells) along with spotty necrosis‌ were observed in the liver of Iinf mice treated twice daily with NS or MEH158. After 72 h of infection, no severe pathological signs were detected in the liver of Iinf mice treated twice daily with EB211, EB279, or CKT for 72 h. Black arrows: bacterial foci, Black arrowheads: necrosis of hepatocytes, Red arrowheads: bi-nucleated cells, White arrows: infiltration of inflammatory cells (mononuclear cells) along with spotty necrosis, White arrowheads: micro and macro vesicles

After 72 h of infection, bacterial communities in perivascular areas, infiltration of inflammatory cells (neutrophils and mononuclear cells) in the perivascular and peribronchial areas, parenchymal necrosis, necrosis of bronchial epithelial cells, edema of perivascular spaces, and hemorrhage in the alveoli and interstitium were the significant pathological symptoms observed in the lungs of immunocompromised infected mice treated twice daily with NS or MEH158 (Fig. 5A and B). On the contrary, none of these pathological abnormalities were observed in the lungs of immunocompromised infected mice treated twice daily with EB211, EB279, or a cocktail of the two scFvs, or once daily with EB211 (Fig. 5A and B).

Fig. 5
figure 5

Administration of EB211 and EB279 prevented severe tissue damage in the lungs of immunocompromised mice with A. baumannii pneumonia. Immunocompromised mice were inoculated intranasally with XDR A. baumannii A.b.56 at a 50% lethal dose (4×107 CFU per mouse). Immunocompromised infected mice were administered intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), a cocktail of the two scFvs (CKT; 6 mg/kg of each), an irrelevant scFv (MEH158; an S. aureus-specific scFv) (12 mg/kg), colistimethate sodium (CMS; 30 mg/kg), or normal saline (NS), or once daily with EB211 (12 mg/kg) (Q24H) two hours after inoculation for 72 h. (A) and (B) Healthy (H), immunocompromised uninfected (IU), and immunocompromised infected (Iinf) mice were killed at 72 h of infection, and the lungs were collected. Tissue sections were stained with hematoxylin-eosin and microscopically evaluated for histopathological alterations. Bacterial communities in perivascular areas, infiltration of inflammatory cells (neutrophils and mononuclear cells) in the perivascular and peribronchial areas, parenchymal necrosis, necrosis of bronchial epithelial cells, edema of perivascular spaces, and hemorrhage in the alveoli and interstitium were detected in the lungs of Iinf mice treated twice daily with NS or MEH158. After 72 h of infection, no severe pathological signs were detected in the lungs of Iinf mice treated twice daily with EB211, EB279, or CKT for 72 h. Black arrows: bacterial foci, Black arrowheads: necrosis of bronchial epithelial cells, Black asterisks: edema of perivascular spaces, Red arrow: macrophage accumulation in the perivascular and peribronchial areas, White arrows: infiltration of inflammatory cells (neutrophils and mononuclear cells) in the perivascular and peribronchial areas, and parenchymal necrosis, White asterisks: areas of hemorrhage within alveoli and interstitium

Discussion

Up to now, several mAbs targeting A. baumannii have been developed, including anti-capsule mAbs [24, 42, 43], anti-OmpA and anti-Omp34 egg yolk antibodies (IgYs) [26], anti-OmpA mAbs [44], anti-siderophore-specific receptors (Iron Regulated Outer Membrane Proteins [IROMPs]) mAbs [45], and AR-401 (a fully human mAb; Aridis Pharmaceuticals, Inc.). There are controversial results about the efficacy of anti-capsule mAbs (13D6, C8, and 8E3 mAbs) against A. baumannii infection [24, 42, 43]. Two mAbs, 13D6 (an IgM specific to K1 capsular polysaccharide) [42] and C8 (an IgG1 specific to K4 capsular polysaccharide) [24], showed antibacterial activity in vitro and in vivo models of A. baumannii infections. In contrast, mAb 8E3, a mouse IgG3 specific to K2 capsular polysaccharide, exacerbated the disease in mice with pneumonia caused by A. baumannii strain AB899 [43]. Wang-Lin et al. indicated that antibody-dependent enhancement (ADE) of infection, previously reported for viruses such as Ebola virus [46], human immunodeficiency virus type 1 (HIV-1) [47, 48], dengue virus [49], and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [50], and “the sink effect” (binding of capsule-specific mAbs to shed capsule), led to the loss of therapeutic activity of mAb 8E3, followed by an increased bacterial burden in the blood, lungs, and spleen, and a low survival rate in infected mice [43]. Moreover, Jahangiri et al. reported a low survival rate in immunocompromised mice with A. baumannii pneumonia treated with a combination of anti-OmpA and Omp34 IgYs (particularly at a high dose) at four hours post-infection compared to immunocompromised infected mice receiving anti-OmpA IgY or anti-Omp34 IgY alone [26]. They hypothesized that some similarity between OmpA and Omp34 proteins with murine proteins and subsequent binding of anti-OmpA and anti-Omp34 IgYs to mouse proteins (as decoys) led to the ADE of A. baumannii infection in immunocompromised mice [26]. Of note, Goel et al. indicated that five mouse IgM mAbs targeting IROMPs, involved in the uptake of siderophore-iron complex, inhibited the growth of A. baumannii cells in vitro by two mechanisms, including the augmentation of opsonization and the interference with the biological activity of IROMPs (direct bactericidal activity) [45]. In any case, the ADE of bacterial infection might be observed with antibody-based therapy [21], exacerbating the disease in patients. In contrast, antibody fragments such as bactericidal scFvs [30, 35, 37, 38, 41], having remarkable features including target-specific binding, small size, great tissue penetration, phagocytes- and complement-independent antimicrobial activity (particularly in immunocompromised patients), and subsequently, no ADE effect (Fc-related unwanted events), can be substantial alternatives to conventional mAbs [28, 35, 51, 52]. Aiming to isolate bactericidal scFvs specific to A. baumannii, two scFvs, EB211 and EB279, with unique sequences and significant binding ability to A. baumannii, were identified in our previous study [40]. Of note, we demonstrated that EB211 and EB279 elicited an antibacterial effect against A. baumannii [40] possibly through the disruption in the outer membrane.

Off-target toxicity is one of the major causes impeding the evaluation of many therapeutics in clinical trials due to a group of various unwanted events in patients [53]. As abovementioned, Jahangiri et al. showed the similarity between peptides derived from OmpA and Omp34 proteins with some peptides of Mus musculus proteins (as decoys), resulting in the inefficacy of the combination anti-OmpA and anti-Omp34 IgYs in the treatment of mice with A. baumannii pneumonia [26]. In this regard, we investigated the off-target effect of both scFvs in vivo. The histopathological examination of the kidneys and liver of mice receiving EB211 (12 mg/kg), EB279 (12 mg/kg), or a cocktail of the two scFvs (6 mg/kg of each) twice daily for 72 h indicated no inflammation or tissue damage, confirming the safety of the scFvs in healthy mice.

In our previous study, EB211 (200 µg/ml), EB279 (200 µg/ml), and a cocktail of the two scFvs (100 µg/ml of each) showed significant growth inhibitory activity against an XDR A. baumannii strain in vitro (about 60%, 22%, and 60% reduction, respectively) [40]. Markedly, EB211 and EB279 alone did not exhibit a significant effect on the growth of A. baumannii at a concentration of 100 µg/ml; however, when these two scFvs were combined, acceptable results were obtained at the same concentration [40]. We hypothesized that combining two different antibodies at appropriate concentrations would be more advantageous than using a high concentration of one antibody. Since the latter may result in saturation of antibodies around the outer membranes of bacteria, leading to the presence of free antibodies that do not have the opportunity to bind and result in off-target binding in the host [54]. It is noteworthy that prior studies showed promising results when several antibodies were used in combination against pathogens such as S. aureus [30, 33]. In this regard, we investigated the therapeutic efficacy of a cocktail of the two scFvs (6 mg/kg of each) in addition to EB211 (12 mg/kg) alone and EB279 (12 mg/kg) alone in immunocompromised mice infected with an XDR A. baumannii strain. The administration dose of 12 mg/kg (for EB211 and EB279 alone) and 6 mg/kg (as a cocktail of two scFvs) was selected based on the concentration of the scFv which showed the highest activity against A. baumannii in vitro (alone, 200 µg/ml; as a cocktail, 100 µg/ml of each; [40]). EB211, EB279, and a cocktail of the two scFvs conferred complete protection against lethal pneumonia. Besides, immunocompromised infected mice administered twice daily with EB211, EB279, or a cocktail of the two scFvs for 72 h showed a lower bacterial load in the lungs and no marked histopathological abnormalities in the kidneys, liver, and lungs after 72 h of infection compared to immunocompromised infected mice administered twice daily with NS or an irrelevant scFv. A few studies assessed the protective efficacy of anti-A. baumannii mAbs in the murine model of pneumonia [24, 26, 43]. Nielsen et al. assessed the therapeutic efficacy of mAb C8, targeting the K4 capsular polysaccharide of A. baumannii, in a murine model of aspiration pneumonia [24]. Mice intratracheally infected with an XDR A. baumannii (strain HUMC1) received mAb C8 (5 µg/mouse/intravenously or 50 µg/mouse/intraperitoneally) immediately or four hours after infection [24]. At both doses and routes of administration, mAb C8 exhibited significant protective activity in infected mice after seven days. Moreover, the administration of mAb C8 (50 µg/mouse/intraperitoneally) led to a decreased bacterial burden in the blood and lungs and, no pathological symptoms in the lungs of infected mice compared to infected mice receiving an isotype control mAb (50 µg/mouse/intraperitoneally) at 24 h of infection [24]. Nonetheless, Wang-Lin et al. reported that the administration of mAb 8E3 (against the K2 capsular polysaccharide) (50 or 200 µg/g/intraperitoneally) immediately after intratracheal inoculation with A. baumannii strain AB899 caused increased disease severity (100% mortality at 48 h of infection) [43]. Additionally, a higher bacterial load in the blood, lungs, and spleen was observed in 8E3-treated mice than PBS-treated mice at 24 h of infection. Based on the findings, mAb 8E3 could not confer protection in infected mice [43]. In another study, Jahangiri et al. evaluated the therapeutic efficacy of a group of IgYs specific to OmpA, Omp34, and the whole inactivated cells of A. baumannii in immunocompromised mice intranasally inoculated with A. baumannii strain AbI101 [26]. All mice were intranasally administered with IgYs at four hours after inoculation. Based on the results, IgYs against OmpA and the whole inactivated cells had significant therapeutic activity and decreased the mortality by 25% in infected mice (75% survival) at eight days after inoculation [26]. In contrast, the combination of anti-OmpA and anti-Omp34 IgYs exacerbated the disease in mice due to the ADE effect, as previously mentioned [26].

Conclusions

Two fully human anti-A. baumannii scFvs, EB211 and EB279, demonstrated no toxic activity in mice after receiving them (alone or combined). Of note, they exhibited significant therapeutic efficacy in immunocompromised mice with pneumonia caused by XDR A. baumannii. These data suggest that EB211 and EB279 may be effective against A. baumannii pneumonia.

Methods

Investigation of the toxic potential of EB211 and EB279 in healthy mice

To investigate the possible cytotoxic effects of EB211 and EB279, female C57BL/6 mice (groups of six, seven to eight-week-old), obtained from the Animal Laboratory of the Pasteur Institute of Iran, were injected intraperitoneally twice daily with EB211 (12 mg/kg), EB279 (12 mg/kg), or a cocktail of the two scFvs (6 mg/kg of each) for 72 h. Mice injected intraperitoneally twice daily with 30 mg/kg of CMS (Colistin is administered as the prodrug colistimethate sodium [CMS] [55,56,57,58]; Sigma-Aldrich, Saint Louis, USA) or NS for 72 h were used as the controls. All mice were sacrificed by intraperitoneal injection of an overdose (5 times the anesthetic dose; [59]) of ketamine and xylazine (Alfasan, Woerden, The Netherlands) at 72 h of infection. The kidneys and liver were aseptically removed and then fixed in 10% formalin for 24 h, followed by embedding in paraffin. Then, thin sections of tissues stained with hematoxylin-eosin were microscopically analyzed for histopathological changes.

Evaluation of the protective efficacy of EB211 and EB279 in an immunocompromised mouse model of A. baumannii pneumonia

An immunocompromised mouse pneumonia model was established as described previously [60]. In brief, female C57BL/6 mice (groups of eight, seven to eight-week-old), acquired from the Animal Laboratory of the Pasteur Institute of Iran, were injected intraperitoneally with 150 mg/kg of cyclophosphamide (Sigma-Aldrich), four days and one day before intranasal challenge with A. baumannii [60]. On day zero, mice anesthetized by intraperitoneal injection of a combination of ketamine (100 mg/kg) and xylazine (10 mg/kg) were intranasally inoculated with XDR A. baumannii A.b.56 [40, 61] (at a 50% lethal dose [LD50]; 4×107 CFU per mouse) [62]. Two hours after inoculation [63], mice were given EB211(12 mg/kg), EB279 (12 mg/kg), a cocktail of the two scFvs (6 mg/kg of each), or an irrelevant scFv (MEH158; an S. aureus-specific scFv) (12 mg/kg) intraperitoneally twice daily for 72 h. Additionally, in a different group, immunocompromised infected mice were treated intraperitoneally once daily with 12 mg/kg of EB211 for 72 h to investigate the dosing frequency on the success of treatment of pneumonia. The control mice were administered intraperitoneally twice daily with 30 mg/kg of CMS or NS for 72 h. Mice were monitored for disease symptoms [64, 65] and body weight [64], and the number of surviving animals was recorded daily for seven days [64].

Moreover, immunocompromised infected mice (groups of six), treated twice daily with EB211, EB279, a cocktail of the two scFvs, MEH158, CMS, or NS, or once daily with EB211 for 72 h, were euthanized with an overdose of ketamine and xylazine [59] after 24, 48, and 72 h of infection, and the lungs were harvested. The A.baumannii burden in the lungs was determined by plating the diluted tissue samples on Luria-Bertani agar (Merck, Darmstadt, Germany) containing imipenem (Sigma-Aldrich), followed by the enumeration of colonies grown after 18 h of incubation at 37 °C [60, 64, 66, 67].

Healthy mice, immunocompromised uninfected mice, and immunocompromised infected mice treated with EB211, EB279, a cocktail of the two scFvs, MEH158, CMS, or NS (groups of six) were sacrificed with an overdose of ketamine and xylazine [59] at 72 h after of infection, and the kidneys, liver, and lungs were harvested [64]. The thin sections of paraffin-embedded tissues were stained with hematoxylin-eosin and then investigated for histopathological alterations [64, 66,67,68].

Statistical analyses

Statistical differences between the experimental groups were analyzed by Student’s t-test. The log-rank test was used to compare Kaplan-Meier survival curves. GraphPad Prism version 6 software (https://www.graphpad.com/) was used for all analyses, and differences were considered statistically significant at p values of < 0.05.

Data Availability

All data generated or analyzed during this study are included in the manuscript. The datasets generated and/or analysed during the current study are available in the GenBank repository under accession numbers OQ970166 (EB211) and OQ970167 (EB279).

References

  1. Liang CA, Lin YC, Lu PL, Chen HC, Chang HL, Sheu CC. Antibiotic strategies and clinical outcomes in critically ill patients with Pneumonia caused by carbapenem-resistant Acinetobacter baumannii. Clin Microbiol Infect. 2018;24(8):908–e1.

    Article  Google Scholar 

  2. Abdul-Mutakabbir JC, Griffith NC, Shields RK, Tverdek FP, Escobar ZK. Contemporary perspective on the treatment of Acinetobacter baumannii Infections: insights from the Society of Infectious Diseases pharmacists. Infect Dis Ther. 2021;10(4):2177–202.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Appaneal HJ, Lopes VV, LaPlante KL, Caffrey AR, Treatment. Clinical outcomes, and predictors of mortality among a National Cohort of admitted patients with Acinetobacter baumannii Infection. Antimicrob Agents Chemother. 2022;66(3):e0197521.

    Article  PubMed  Google Scholar 

  4. Medioli F, Bacca E, Faltoni M, Burastero GJ, Volpi S, Menozzi M, et al. Is it possible to Eradicate Carbapenem-Resistant Acinetobacter baumannii (CRAB) from endemic hospitals? Antibiotics. 2022;11(8):1015.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Hu H, Lou Y, Feng H, Tao J, Shi W, Ni S, et al. Molecular characterization of Carbapenem-Resistant Acinetobacter baumannii isolates among Intensive Care Unit patients and Environment. Infect Drug Resist. 2022;15:1821–29.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Kumar A, Randhawa VS, Nirupam N, Rai Y, Saili A. Risk factors for carbapenem-resistant Acinetobacter baumanii blood stream Infections in a neonatal intensive care unit, Delhi, India. J Infect Dev Ctries. 2014;8(8):1049–54.

    Article  PubMed  Google Scholar 

  7. Jung C-J, Liao Y-D, Hsu C-C, Huang T-Y, Chuang Y-C, Chen J-W, et al. Identification of potential therapeutic antimicrobial peptides against Acinetobacter baumannii in a mouse model of Pneumonia. Sci Rep. 2021;11(1):1–10.

    Google Scholar 

  8. Kyriakidis I, Vasileiou E, Pana ZD, Tragiannidis A. Acinetobacter baumannii Antibiotic Resistance Mechanisms Pathogens. 2021;10(3):373.

    CAS  PubMed  Google Scholar 

  9. Walsh BJC, Wang J, Edmonds KA, Palmer LD, Zhang Y, Trinidad JC, et al. The response of Acinetobacter baumannii to Hydrogen Sulfide reveals two Independent persulfide-sensing systems and a connection to Biofilm Regulation. mBio. 2020;11(3):e01254–20.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Zhen X, Stalsby Lundborg C, Sun X, Gu S, Dong H. Clinical and economic Burden of Carbapenem-resistant Infection or colonization caused by Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii: a Multicenter Study in China. Antibiotics. 2020;9(8):514.

    Article  PubMed  PubMed Central  Google Scholar 

  11. Magda M, Bettoni S, Laabei M, Fairley D, Russo TA, Riesbeck K, et al. Clinical isolates of Acinetobacter spp. are highly serum resistant despite efficient recognition by the complement system. Front Immunol. 2022;13:814193.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Zheng YL, Wan YF, Zhou LY, Ye ML, Liu S, Xu CQ, et al. Risk factors and mortality of patients with nosocomial carbapenem-resistant Acinetobacter baumannii Pneumonia. Am J Infect Control. 2013;41(7):e59–63.

    Article  PubMed  Google Scholar 

  13. Piperaki ET, Tzouvelekis LS, Miriagou V, Daikos GL. Carbapenem-resistant Acinetobacter baumannii: in pursuit of an effective treatment. Clin Microbiol Infect. 2019;25(8):951–7.

    Article  CAS  PubMed  Google Scholar 

  14. Desoubeaux G, Reichert JM, Sleeman M, Reckamp KL, Ryffel B, Adamczewski JP, et al. Therapeutic monoclonal antibodies for Respiratory Diseases: current challenges and perspectives, March 31 - April 1, 2016, tours, France. MAbs. 2016;8(6):999–1009.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Yang B, Fang D, Lv Q, Wang Z, Liu Y. Targeted therapeutic strategies in the battle against pathogenic Bacteria. Front Pharmacol. 2021;12:673239.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Ramamurthy D, Nundalall T, Cingo S, Mungra N, Karaan M, Naran K, et al. Recent advances in immunotherapies against infectious Diseases. Immunother Adv. 2021;1(1):ltaa007.

    Article  Google Scholar 

  17. Vacca F, Sala C, Rappuoli R. Monoclonal antibodies for bacterial pathogens: mechanisms of Action and Engineering approaches for enhanced effector functions. Biomedicines. 2022;10(9):2126.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Motley MP, Banerjee K, Fries BC. Monoclonal antibody-based therapies for bacterial Infections. Curr Opin Infect Dis. 2019;32(3):210–16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Motley MP, Fries BC. A New take on an old remedy: Generating antibodies against Multidrug-Resistant Gram-negative Bacteria in a Postantibiotic World. mSphere. 2017;2(5):10–28.

    Article  Google Scholar 

  20. Peck M, Rothenberg ME, Deng R, Lewin-Koh N, She G, Kamath AV, et al. A phase 1, Randomized, single-ascending-dose study to investigate the Safety, Tolerability, and pharmacokinetics of DSTA4637S, an Anti-Staphylococcus Aureus Thiomab antibody-antibiotic Conjugate, in healthy volunteers. Antimicrob Agents Chemother. 2019;63(6):e02588–02518.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Wang-Lin SX, Balthasar JP. Pharmacokinetic and pharmacodynamic considerations for the use of monoclonal antibodies in the treatment of bacterial Infections. Antibodies. 2018;7(1):5.

    Article  PubMed  PubMed Central  Google Scholar 

  22. Slarve M, Reyna Z, Burk E, Ruiz-Delgado J, Li R, Yan J, et al. Therapeutic, humanized monoclonal antibody exhibits broad binding and protective efficacy against Acinetobacter baumannii. Antimicrob Agents Chemother. 2023;67(3):e00086–00023.

    PubMed  PubMed Central  Google Scholar 

  23. Nielsen TB, Yan J, Slarve M, Li R, Junge JA, Luna BM, et al. Development of a bispecific antibody targeting clinical isolates of Acinetobacter baumannii. J Infect Dis. 2023;227:1042–49.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Nielsen TB, Pantapalangkoor P, Luna BM, Bruhn KW, Yan J, Dekitani K, et al. Monoclonal antibody protects against Acinetobacter baumannii Infection by enhancing bacterial clearance and evading sepsis. J Infect Dis. 2017;216(4):489–501.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Nielsen TB, Yan J, Slarve M, Lu P, Li R, Ruiz J, et al. Monoclonal antibody therapy against Acinetobacter baumannii. Infect Immun. 2021;89(10):e00162–00121.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Jahangiri A, Owlia P, Rasooli I, Salimian J, Derakhshanifar E, Aghajani Z, et al. Specific egg yolk immunoglobulin as a promising non-antibiotic biotherapeutic product against Acinetobacter baumannii Pneumonia Infection. Sci Rep. 2021;11(1):1–11.

    Article  Google Scholar 

  27. Knauf GA, Groover KE, O’Donnell AC, Davies BW. Generation of Synthetic Acinetobacter baumannii-Specific nanobodies. ACS Infect Dis. 2023;9:1190–95.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Ahamadi-Fesharaki R, Fateh A, Vaziri F, Solgi G, Siadat SD, Mahboudi F, et al. Single-chain variable fragment-based bispecific antibodies: hitting two targets with one sophisticated arrow. Mol Ther Oncolytics. 2019;14:38.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Di Mambro T, Vanzolini T, Bianchi M, Crinelli R, Canonico B, Tasini F, et al. Development and in vitro characterization of a humanized scFv against fungal Infections. PLoS ONE. 2022;17(10):e0276786.

    Article  PubMed  PubMed Central  Google Scholar 

  30. Soltanmohammadi B, Piri-Gavgani S, Basardeh E, Ghanei M, Azizi M, Khaksar Z, et al. Bactericidal fully human single-chain fragment variable antibodies protect mice against methicillin-resistant Staphylococcus aureus bacteraemia. Clin Transl Immunol. 2021;10(7):e1302.

    Article  CAS  Google Scholar 

  31. Weisser NE, Hall JC. Applications of single-chain variable fragment antibodies in therapeutics and diagnostics. Biotechnol Adv. 2009;27(4):502–20.

    Article  CAS  PubMed  Google Scholar 

  32. Holliger P, Hudson PJ. Engineered antibody fragments and the rise of single domains. Nat Biotechnol. 2005;23(9):1126–36.

    Article  CAS  PubMed  Google Scholar 

  33. Wang M, Zhang Y, Zhu J. Anti-Staphylococcus Aureus single-chain variable region fragments provide protection against mastitis in mice. Appl Microbiol Biotechnol. 2016;100(5):2153–62.

    Article  CAS  PubMed  Google Scholar 

  34. Richard G, MacKenzie CR, Henry KA, Vinogradov E, Hall JC, Hussack G. Antibody binding to the O-Specific Antigen of Pseudomonas aeruginosa O6 inhibits cell growth. Antimicrob Agents Chemother. 2020;64(4):e02168–02119.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Xie X, McLean MD, Hall JC. Antibody-dependent cell-mediated cytotoxicity-and complement-dependent cytotoxicity-independent bactericidal activity of an IgG against Pseudomonas aeruginosa O6ad. J Immunol. 2010;184(7):3725–33.

    Article  CAS  PubMed  Google Scholar 

  36. Oren Z, Shai Y. Mode of action of linear amphipathic α-helical antimicrobial peptides. Pept Sci. 1998;47(6):451–63.

    Article  CAS  Google Scholar 

  37. Richard G. Investigating the bactericidal mechanism of Anti-LPS antibodies against Pseudomonas aeruginosa Serotype O6. Ontario, Canada: University of Guelph; 2017.

    Google Scholar 

  38. LaRocca TJ, Katona LI, Thanassi DG, Benach JL. Bactericidal action of a complement-independent antibody against relapsing Fever Borrelia resides in its variable region. J Immunol. 2008;180(9):6222–28.

    Article  CAS  PubMed  Google Scholar 

  39. Nasirudeen A, Tan KS. Caspase-3-like protease influences but is not essential for DNA fragmentation in Blastocystis undergoing apoptosis. Eur J Cell Biol. 2004;83(9):477–82.

    Article  CAS  PubMed  Google Scholar 

  40. Basardeh E, Piri-Gavgani S, Soltanmohammadi B, Ghanei M, Omrani MD, Soezi M, et al. Anti-Acinetobacter Baumannii single-chain variable fragments show direct bactericidal activity. Iran J Basic Med Sci. 2022;25:1141–49.

    PubMed  PubMed Central  Google Scholar 

  41. LaRocca TJ, Holthausen DJ, Hsieh C, Renken C, Mannella CA, Benach JL. The bactericidal effect of a complement-independent antibody is osmolytic and specific to Borrelia. PNAS. 2009;106(26):10752–57.

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  42. Russo TA, Beanan JM, Olson R, MacDonald U, Cox AD, St. Michael F, et al. The K1 capsular polysaccharide from Acinetobacter baumannii is a potential therapeutic target via passive immunization. Infect Immun. 2013;81(3):915–22.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Wang-Lin SX, Olson R, Beanan JM, MacDonald U, Russo TA, Balthasar JP. Antibody dependent enhancement of Acinetobacter baumannii Infection in a mouse Pneumonia model. J Pharmacol Exp Ther. 2019;368(3):475–89.

    Article  CAS  PubMed  Google Scholar 

  44. Wang-Lin SX, Olson R, Beanan JM, MacDonald U, Balthasar JP, Russo TA. The capsular polysaccharide of Acinetobacter baumannii is an obstacle for therapeutic passive immunization strategies. Infect Immun. 2017;85(12):e00591–17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Goel VK, Kapil A. Monoclonal antibodies against the iron regulated outer membrane proteins of Acinetobacter baumannii are bactericidal. BMC Microbiol. 2001;1(1):16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Takada A, Watanabe S, Okazaki K, Kida H, Kawaoka Y. Infectivity-enhancing antibodies to Ebola virus glycoprotein. J Virol. 2001;75(5):2324–30.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Robinson JRWE, Montefiori D, Mitchell W. Antibody-dependent enhancement of human immunodeficiency virus type 1 Infection. Lancet. 1988;331(8589):790–94.

    Article  Google Scholar 

  48. Tirado SM, Yoon KJ. Antibody-dependent enhancement of virus Infection and Disease. Viral Immunol. 2003;16(1):69–86.

    Article  CAS  PubMed  Google Scholar 

  49. Dejnirattisai W, Jumnainsong A, Onsirisakul N, Fitton P, Vasanawathana S, Limpitikul W, et al. Cross-reacting antibodies enhance dengue virus Infection in humans. Science. 2010;328(5979):745–48.

    Article  ADS  CAS  PubMed  Google Scholar 

  50. Lee WS, Wheatley AK, Kent SJ, DeKosky BJ. Antibody-dependent enhancement and SARS-CoV-2 vaccines and therapies. Nat Microbiol. 2020;5(10):1185–91.

    Article  CAS  PubMed  Google Scholar 

  51. Monnier P, Vigouroux R, Tassew N. In vivo applications of single chain fv (variable domain)(scFv) fragments. Antibodies. 2013;2(2):193–208.

    Article  CAS  Google Scholar 

  52. Li Z, Krippendorff B-F, Sharma S, Walz AC, Lavé T, Shah DK. Influence of molecular size on tissue distribution of antibody fragments. MAbs. 2016;8:113–19.

    Article  CAS  PubMed  Google Scholar 

  53. Huang Y, Furuno M, Arakawa T, Takizawa S, de Hoon M, Suzuki H, et al. A framework for identification of on-and off-target transcriptional responses to drug treatment. Sci Rep. 2019;9(1):1–9.

    Article  Google Scholar 

  54. Jamnani FR, Rahbarizadeh F, Shokrgozar MA, Ahmadvand D, Mahboudi F, Sharifzadeh Z. Targeting high affinity and epitope-distinct oligoclonal nanobodies to HER2 over-expressing Tumor cells. Exp Cell Res. 2012;318(10):1112–24.

    Article  CAS  PubMed  Google Scholar 

  55. Bartal C, Rolston KVI, Nesher L. Carbapenem-resistant Acinetobacter baumannii: colonization, Infection and current treatment options. Infect Dis Ther. 2022;11(2):683–94.

    Article  PubMed  PubMed Central  Google Scholar 

  56. Nation RL, Rigatto MHP, Falci DR, Zavascki AP. Polymyxin Acute kidney Injury: Dosing and other strategies to reduce toxicity. Antibiotics. 2019;8(1):24.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Katip W, Uitrakul S, Oberdorfer P. Clinical efficacy and nephrotoxicity of the Loading Dose Colistin for the treatment of Carbapenem-Resistant Acinetobacter baumannii in critically Ill patients. Pharmaceutics. 2021;14(1):31.

    Article  PubMed  PubMed Central  Google Scholar 

  58. Ayoub Moubareck C. Polymyxins and bacterial membranes: a review of Antibacterial Activity and mechanisms of Resistance. Membranes. 2020;10(8):181.

    Article  PubMed  PubMed Central  Google Scholar 

  59. Underwood W, Anthony R. AVMA guidelines for the euthanasia of animals: 2020 edition. Retrieved on March. 2020;2013(30):2020–1.

    Google Scholar 

  60. Ma X-L, Guo Y-Z, Wu Y-M, Gong W-T, Sun J, Huang Z. In vivo bactericidal effect of colistin–linezolid combination in a murine model of MDR and XDR Acinetobacter baumannii Pneumonia. Sci Rep. 2020;10(1):1–8.

    Article  ADS  CAS  Google Scholar 

  61. Irani N, Basardeh E, Samiee F, Fateh A, Shooraj F, Rahimi A, et al. The inhibitory effect of the combination of two new peptides on biofilm formation by Acinetobacter baumannii. Microb Pathog. 2018;121:310–17.

    Article  CAS  PubMed  Google Scholar 

  62. DiGiandomenico A, Keller AE, Gao C, Rainey GJ, Warrener P, Camara MM, et al. A multifunctional bispecific antibody protects against Pseudomonas aeruginosa. Sci Transl Med. 2014;6(262):262ra155.

    Article  PubMed  Google Scholar 

  63. Chiang SR, Chuang YC, Tang HJ, Chen CC, Chen CH, Lee NY, et al. Intratracheal colistin sulfate for BALB/c mice with early Pneumonia caused by carbapenem-resistant Acinetobacter baumannii. Crit Care Med. 2009;37(9):2590–95.

    Article  PubMed  Google Scholar 

  64. Harris G, Lee RK, Lam CK, Kanzaki G, Patel GB, et al. A mouse model of Acinetobacter baumannii-associated Pneumonia using a clinically isolated hypervirulent strain. Antimicrob Agents Chemother. 2013;57(8):3601–13.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. van Faassen H, KuoLee R, Harris G, Zhao X, Conlan JW, Chen W. Neutrophils play an important role in host resistance to Respiratory Infection with Acinetobacter baumannii in mice. Infect Immun. 2007;75(12):5597–608.

    Article  PubMed  PubMed Central  Google Scholar 

  66. Ku NS, Lee S-H, Lim Y-s, Choi H, Ahn JY, Jeong SJ, et al. In vivo efficacy of combination of colistin with fosfomycin or minocycline in a mouse model of multidrug-resistant Acinetobacter baumannii Pneumonia. Sci Rep. 2019;9(1):1–7.

    Article  Google Scholar 

  67. Kang MJ, Jang AR, Park JY, Ahn JH, Lee TS, Kim DY, et al. IL-10 protects mice from the lung Infection of Acinetobacter baumannii and contributes to bacterial clearance by regulating STAT3-Mediated MARCO expression in macrophages. Front Immunol. 2020;11:270.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Dietert K, Gutbier B, Wienhold SM, Reppe K, Jiang X, Yao L, et al. Spectrum of pathogen- and model-specific histopathologies in mouse models of acute Pneumonia. PLoS ONE. 2017;12(11):e0188251.

    Article  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

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Funding

This project was granted by the Pasteur Institute of Iran (grant No.: 1537).

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FRJ supervised, directed, and managed the study, and wrote the manuscript. EB performed the experiments. SPG performed the experiments and was involved in the manuscript preparation. HRM performed histopathological analysis. MA helped interpret the data. PM and FN were involved in revised the manuscript. MGH and FM helped design the study and provided important critical review. All authors read and approved the final manuscript.

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Correspondence to Fatemeh Rahimi-Jamnani.

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Animal experiments were conducted in accordance with ARRIVE guidelines (https://arriveguidelines.org) and approved by the Animal Care and Use Committees of the Pasteur Institute of Iran (IR.PII.REC.1397.036). All methods were performed in accordance with relevant guidelines and regulations.

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Basardeh, E., Piri-Gavgani, S., Moradi, H.R. et al. Anti-Acinetobacter Baumannii single-chain variable fragments provide therapeutic efficacy in an immunocompromised mouse pneumonia model. BMC Microbiol 24, 55 (2024). https://doi.org/10.1186/s12866-023-03080-9

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