SARS-CoV-2 Vaccine Responses in Individuals with Antibody Deficiency: Findings from the COV-AD Study

Background

Vaccination prevents severe morbidity and mortality from COVID-19 in the general population. The immunogenicity and efficacy of SARS-CoV-2 vaccines in patients with antibody deficiency is poorly understood.

Objectives

COVID-19 in patients with antibody deficiency (COV-AD) is a multi-site UK study that aims to determine the immune response to SARS-CoV-2 infection and vaccination in patients with primary or secondary antibody deficiency, a population that suffers from severe and recurrent infection and does not respond well to vaccination.

Methods

Individuals on immunoglobulin replacement therapy or with an IgG less than 4 g/L receiving antibiotic prophylaxis were recruited from April 2021. Serological and cellular responses were determined using ELISA, live-virus neutralisation and interferon gamma release assays. SARS-CoV-2 infection and clearance were determined by PCR from serial nasopharyngeal swabs.

Results

A total of 5.6% (n?=?320) of the cohort reported prior SARS-CoV-2 infection, but only 0.3% remained PCR positive on study entry. Seropositivity, following two doses of SARS-CoV-2 vaccination, was 54.8% (n?=?168) compared with 100% of healthy controls (n?=?205). The magnitude of the antibody response and its neutralising capacity were both significantly reduced compared to controls. Participants vaccinated with the Pfizer/BioNTech vaccine were more likely to be seropositive (65.7% vs. 48.0%, p?=?0.03) and have higher antibody levels compared with the AstraZeneca vaccine (IgGAM ratio 3.73 vs. 2.39, p?=?0.0003). T cell responses post vaccination was demonstrable in 46.2% of participants and were associated with better antibody responses but there was no difference between the two vaccines. Eleven vaccine-breakthrough infections have occurred to date, 10 of them in recipients of the AstraZeneca vaccine.

Conclusion

SARS-CoV-2 vaccines demonstrate reduced immunogenicity in patients with antibody deficiency with evidence of vaccine breakthrough infection.

     

A cytological comparison between regeneration, hyperplasia and early neoplasia in the rat liver

Regeneration, hyperplasia and neoplasia are three differentresponses to injury in the rat liver. These phenomena were inducedin rat liver and the parameters of ploidy, nuclearity and DNAsynthesis were examined. Analysis of hepatocytes from animalsundergoing liver regeneration following two-thirds partial hepatectomyrevealed that there is an increase in the cycling of diploidhepatocytes and a large increase in the frequency of binucleatedtetraploid cells undergoing DNA synthesis and amnitotic cytokinesisto mononucleated tetraploid cells. This results in an overallincrease in the ratio of tetra ploid:diploid cells but no changein the proportion of binucleated cells. The liver appears, temporarily,to undergo an increased rate of maturation. In both hyperplasiainduc ed by oral administration of 25 mg/kg methylclofenapateor diethylhexylphthalate (1 g/kg for 4 weeks) and neoplasiain duced by the hepatocarcinogens 3'-methyl-4-dimnethylamino-azobenzene(3'M), 6-p-dimethylaminophenylazobenzthiazole (6BT), 5-phenylazoindazole(5I), diethylnitrosamine (DEN) and thioacetamide (TA) the binucleatedcell is sensitive to the action of the chemicals, although itsresponse is different. Both types of carcinogen induce a reductionin the frequency of binucleated cells but the mononucleateddiploid cells produced by cytokinesis without a preceding Sphase as a result of the action of genotoxic carcinogens appearto be incapable of poly-ploidization and give rise to a liverwith a pennanently depressed tetraploid:diploid hepatocyte ratio.The non-genotoxic carcinogens methyiclofenapate and DEHP causean initial hyperplastic response due to the rapid conversionof binucleated cells to mononucleated tetraploids by amitoticcytokinesis following S phase. Over a longer period of ex posurethere is an increase in the tetraploid:diploid ratio due tothe continued conversion of newly formed binucleates to tetraploidmononucleates.     

Efficient differentiation of hepatocytes from human embryonic stem cells exhibiting markers recapitulating liver development in vivo

The potential to differentiate human embryonic stem cells (hESCs) in vitro to provide an unlimited source of human hepatocytes for use in biomedical research, drug discovery, and the treatment of liver diseases holds great promise. Here we describe a three-stage process for the efficient and reproducible differentiation of hESCs to hepatocytes by priming hESCs towards definitive endoderm with activin A and sodium butyrate prior to further differentiation to hepatocytes with dimethyl sulfoxide, followed by maturation with hepatocyte growth factor and oncostatin M. We have demonstrated that differentiation of hESCs in this process recapitulates liver development in vivo: following initial differentiation, hESCs transiently express characteristic markers of the primitive streak mesendoderm before turning to the markers of the definitive endoderm; with further differentiation, expression of hepatocyte progenitor cell markers and mature hepatocyte markers emerged sequentially. Furthermore, we have provided evidence that the hESC-derived hepatocytes are able to carry out a range of hepatocyte functions: storage of glycogen, and generation and secretion of plasma proteins. More importantly, the hESC-derived hepatocytes express several members of cytochrome P450 isozymes, and these P450 isozymes are capable of converting the substrates to metabolites and respond to the chemical stimulation. Our results have provided evidence that hESCs can be differentiated efficiently in vitro to functional hepatocytes, which may be useful as an in vitro system for toxicity screening in drug discovery.     

Identification of the proximate peroxisome proliferator(s) derived from di (2-ethylhexyl) adipate and species differences in response.

Identification of the proximate peroxisome proliferator(s) derived from di (2-ethylhexyl) adipate (DEHA) has been achieved using primary hepatocyte cultures derived from different species and cyanide-insensitive fatty acyl CoA oxidase (PCO) as a marker enzyme for peroxisome proliferation. In rat and mouse hepatocytes, the parent compound (DEHA) had no effect on peroxisomal beta-oxidation, but primary metabolites of DEHA, mono (2-ethylhexyl) adipate (MEHA) and 2-ethylhexanol (EH), were approximately equipotent in PCO induction (5-fold at 0.5 mM final concentration). The secondary metabolite of DEHA, 2-ethylhexanoic acid (EHA), was in both species the most potent peroxisome proliferator (25- and 9-fold induction in mice and rats, respectively, at 1 mM final concentration). At 2 mM final concentration a tertiary metabolite of DEHA, 2-ethyl-5-hydroxyhexan-1-oic acid, was less effective in mouse and rat hepatocytes at inducing PCO (15- and 5-fold, respectively). 2-Ethyl-5-oxohexan-1-oic acid and 2-ethylhexan-1,6-dioic acid had little effect (2-3-fold in both rat and mouse hepatocytes). Thus, EHA was identified as the proximate peroxisome proliferator of DEHA and mouse hepatocytes were approximately twice as sensitive as rat hepatocytes to peroxisome proliferation due to MEHA, EH and EHA. We investigated further species differences in response to peroxisome proliferators by using guinea pig and marmoset primary hepatocyte culture. None of the chemicals studied stimulated peroxisomal beta-oxidation in these species up to a final concentration of 2 mM. Higher concentrations lead to cytotoxicity. This lack of sensitivity of guinea pig and marmoset hepatocytes is in agreement with previous studies with di (2-ethylhexyl) phthalate metabolites, suggesting the absence of a threat of hepatocarcinogenic damage to these species and confirming that primary hepatocytes cultures are useful models for investigating the phenomenon of peroxisome proliferation.     

Alternatives to Aroclor 1254-induced S9 in in vitro genotoxicity assays.

A working party was set up by the UK Environmental Mutagen Society to consider alternatives to Aroclor 1254 (Aroclor)-induced S9 in in vitro genotoxicity assays, with the aims of considering whether a replacement for Aroclor in its role in general screening assays could be readily identified. The working party concluded that there was sufficient support in the literature to justify the use of an appropriate phenobarbital/beta-naphthoflavone regime as an acceptable alternative to Aroclor.     

Significance of induction phenomena.

A number of foreign compounds induce the proliferation of the hepatic smooth endoplasmatic reticulum and thereby increase the activity of monooxygenases that metabolize drugs and other foreign compound. With reference to the safety of food additives some antioxidants have been examined by various authors for their inducing capacity, in doses well above those ingested with treated food and above the stipulated accepted daily intake (ADI). Thus feeding of rats with the very high dose of 500 mg/kg body weight of butylated hydroxtoluene (BHT) resulted in an increase in its own oxidative metabolism. Also in monkeys BHT produces an inductive increase of microsomal enzyme activity, cytochrome p 450, and a proliferation of smooth endoplasmatic reticulum. The closely related antioxidant butylated hydroxyanisole (BHA) leads to similar effects in mice. Recent studies with ethoxyquin (EQ) have shown that this antioxidant stimulates the formation of a form of cytochrome P 450 which resembles the phenobarbital-inducible type. EQ itself is also an inhibitor of mixed function monooxygenase. The naturally occuring flavouring agents safrole and isosafrole in high doses also induce hepatic monooxygenations. Spectroscopic examination has revealed, however, that another type of cytochrome with characteristic binding of a suspected safrole metabolite is produced. This complex shows absorption maxima at 427 and 455 nm and can be dissociated by adding a number of different lipophilic agents, including safrole itself. In a time dependent displacement reaction the characteristic spectrum decreases and is replaced by a classical binding spectrum of the displacer itself. By these examples it can be shown that food additives exert a number of effects on hepatic drug inactivating systems. However, no risk of hepatic changes can be seen from the small amounts of antioxidants ingested in balanced human nutrition.     

Prediction of rodent nongenotoxic carcinogenesis: evaluation of biochemical and tissue changes in rodents following exposure to nine nongenotoxic NTP carcinogens

We studied nine presumed nongenotoxic rodent carcinogens, as defined by the U.S. National Toxicology Program (NTP), to determine their ability to induce acute or subacute biochemical and tissue changes that may act as useful predictors of nongenotoxic rodent carcinogenesis. The chemicals selected included six liver carcinogens (two of which are peroxisome proliferators), three thyroid gland carcinogens, and four kidney carcinogens. We administered the chemicals (diethylhexyl phthalate, cinnamyl anthranilate, chlorendic acid, 1,4-dichlorobenzene, monuron, ethylene thiourea, diethyl thiourea, trimethyl thiourea, and d-limonene to the same strains of mice and rats used in the original NTP bioassays (nine chemicals to rats and seven to mice). Selected tissues (liver, thyroid gland, and kidney) were collected from groups of animals at 7, 28, and 90 days for evaluation. Tissue changes selected for study were monitored for all of the test groups, irrespective of the specificity of the carcinogenic responses observed in those tissues. This allowed us to assess both the carcinogen specificity and the carcinogen sensitivity of the events being monitored. We studied relative weight, cell labeling indices, and pathologic changes such as hypertrophy in all tissues; a range of cytochrome P450 enzymes and palmitoyl coenzyme A oxidase in the liver; changes in the levels of plasma total triiodothyronine, total thyroxine, and thyroid-stimulating hormone (TSH) as markers of thyroid gland function; and hyaline droplet formation, tubular basophilia, and the formation of granular casts in the kidney. There were no single measurements that alerted specifically to the carcinogenicity of the agents to the rodent liver, thyroid gland, or kidney. However, in the majority of cases, the chemical induction of cancer in a tissue was preceded by a range of biochemical/morphologic changes, most of which were moderately specific for a carcinogenic outcome, and some of which were highly specific for it (e.g., increases in TSH in the thyroid gland and increases in relative liver weight in the mouse). The only measurements that failed to correlate usefully with carcinogenicity were the induction of liver enzymes (with the exception of the enzymes associated with peroxisome proliferation). Most of the useful markers were evident at the early times studied (7 days and 28 days), but no overall best time for the measurement of all markers was identified. The judicious choice of markers and evaluation times can aid the detection of potential nongenotoxic rodent carcinogens.