Acetylcysteine Treatment of Acetaminophen Overdose: Foundational and Clinical Development

Abstract

N-acetyl para-aminophenol was suggested as a safer alternative to other drugs on the market for pain and fever in 1948. It was given the generic name “acetaminophen” in 1951 and the trade name “Tylenol” when it was put on the market in the USA in 1955 as a prescription drug to treat pediatric fever. It also received the generic name “paracetamol” in the UK where it was initially marketed in 1956 under the name “Panadol.” Toxicity from overdose of acetaminophen was reported in 1966. Research at the US National Institutes of Health uncovered the mechanisms of toxicity and proposed a treatment in a foundational series of papers in 1973 and 1974. A nomogram was developed in 1973 and published in 1975 to guide estimation of patient risk of hepatic toxicity. Rapid development followed utilizing acetylcysteine given both orally and intravenously. Various protocols and methods of administration have been employed over time with the primary use today of acetylcysteine intravenously as the therapeutic method. The nomogram has been revised over time to the current version, published in 2023, which allows stratification of patients to a high-risk group over 300 mg/L at 4 h and standard risk above 150 mg/L at 4 h, except in the UK where the standard risk is defined very conservatively with a line above 100 mg/L at 4 h. Adjunct therapy with fomepizole in patients with massive ingestions, delay until arrival in a health care facility or renal injury has been proposed. The mortality rate with treatment has been substantially reduced and recovery from hepatic injury is achieved in almost all patients.

1. Clinical Application of Acetaminophen (Paracetamol) for Fever and Pain

It has been 77 years since n-acetyl-p-aminophenol was suggested for clinical use in a publication “…that it may have distinct advantages over acetanilide and related as an analgesic”. This publication by Bernard B. Brodie and Julius Axelrod in 1948 demonstrated in humans that when “…administered orally, it was not attended by the formation of methemoglobin” and should be considered as a less toxic alternative to acetophenetidin (phenacetin) [1].

Commercial development was considered a few years later. Robert Lincoln McNeil Jr. joined the family business, McNeil Laboratories, and decided he wanted to compete with aspirin (https://en.wikipedia.org/wiki/Robert_L._McNeil_Jr. (accessed on 21 April 2025)). He was aware of the work by Brodie and Axelrod and McNeil Laboratories began work to bring the analgesic and antipyretic to market in 1951. Robert McNeil coined the generic name “acetaminophen” (APAP) from the chemical name as a derivative N-acetyl-p-aminophenol and a marketing colleague at the company also derived the brand name “Tylenol” (N-acetyl-p-aminophenol) from the chemical name. Tylenol was approved by the FDA as a prescription drug starting in 1955 for pediatric use as “Children’s Tylenol Elixir” and became an over-the-counter drug, also marketed to adults, after the purchase of McNeil Laboratories in 1960 by Johnson and Johnson. It was introduced in the United Kingdom in 1956 by Frederick Stearns and Co, also initially as a prescription drug. They utilized the generic term “paracetamol” as a derivative from an alternative chemical name “paraacetyl-amino-phenol with the trade name “Panadol.”

2. Mechanisms of Toxicity of Acetaminophen (Paracetamol)

Within a few years following its introduction as an over-the-counter drug, cases of overdose were reported [2,3]. These reports were initially in the United Kingdom and then elsewhere. There were two key publications from Edinburgh in 1970 and 1971 describing the course of hepatic centrilobular necrosis from acetaminophen and the pharmacokinetics (Figure 1) without any known treatment [4,5].

During this time-period, Bernard Brodie at NIH was looking at carcinogenic compounds that were being activated to reactive species that were bound covalently to cellular macromolecules which produced toxicity. He postulated that this might occur with bromobenzene that also produced centrilobular necrosis in the liver. As part of this research, acetaminophen was used to block glucuronidation and sulfation of bromobenzene. His laboratory observed that acetaminophen control animals developed centrilobular necrosis without bromobenzene. Thus began an extraordinary research effort by Mitchell and colleagues at NIH focused on acetaminophen, which produced a landmark series of papers published in 1973 and 1974 [6,7,8,9,10,11,12]. These papers defined the mechanism of toxicity from acetaminophen and proposed a therapeutic approach. Further details from this time were recently published by one of the participants, David Jollow, in a comprehensive reminiscence [13].

Holtzman and colleagues had previously demonstrated that glutathione conjugated epoxides and further work by Calder had recently demonstrated such a metabolite from phenacetin [14,15]. The research in the Mitchell laboratory demonstrated that glutathione depletion and the administration of a surrogate could interfere with the toxic metabolite. They postulated a toxic intermediate which was then identified as N-acetyl-p-benzoquinone imine (NAPQI) as a highly electrophilic substance which could be conjugated by glutathione. Substantial additional work on NAPQI was carried out in the Nelson Laboratory at NIH and published in the 1980s [16,17]. A recent review discussed in detail that NAPQI covalently binds to different key mitochondrial enzymes, thus resulting in oxidative phosphorylation impairment and severe ATP depletion. As the number of overdoses producing hepatic toxicity increased and were reported clinically, there was substantial interest on the part of investigators in finding a treatment regimen which could be administered to these patients [18].

The author of this paper, Barry Rumack (BHR), had worked in the Holtzman laboratory at the National Institutes of Health from 1969 to 1971 on various aspects of hepatic metabolism. Toward the end of that time, there was discussion regarding the work being carried out in the Mitchell Laboratory on acetaminophen but none of the acetaminophen work was done in the Holtzman laboratory.

Barry Rumack was invited to do a clinical and research fellowship at the Royal Infirmary of Edinburgh Poisons Unit in 1973 with Henry Matthew who was Physician-in-Charge and Director of the Scottish Poisons Information Bureau. Two recently published papers provide historical information over 50 years of the clinical and research work, with a primary focus on the work carried out in Edinburgh and Denver [19,20].

The Royal Infirmary of Edinburgh Poisons Unit admitted patients with overdoses of acetaminophen during the time that Barry Rumack was in residence. The journal “Pediatrics” had asked Barry Rumack to write a paper regarding acetaminophen as there had been reports in the United States as well. As part of that paper, the journal requested a graphic representation of the risk from overdose rather than just the pharmacokinetic formulas. Rumack and Matthew wrote the paper together and utilized data from 30 cases from the paper published in 1971 and collected additional data from 34 cases and created a nomogram in 1973, which was published in 1975 [21]. This nomogram (Figure 2) permitted physicians to rapidly interpret levels of acetaminophen and provide a guide to the risk of hepatic toxicity. During that time, acetaminophen hepatic toxicity was defined with transaminase levels greater than 1000 IU/L. This was because the laboratory had to dilute samples and re-assay over this concentration in order to provide a transaminase measurement.

3. Development of n-Acetylcysteine (NAC) Treatment of Acetaminophen Overdose

Mitchell et al. suggested the administration of cysteamine (4-mercaptoethanol) or other nucleophiles might provide a basis for treatment in humans [10]. This 1974 publication had elegantly demonstrated the production of metabolites following radioactively labeled acetaminophen being administered to human volunteers. Two human volunteers were given radioactive 3H-APAP 1200 mg with 100 microcuries (µc) and urine was collected at 12 h intervals for 48 h. The results were measured with 2.1% free APAP, 52.1% sulfate, 42% glucuronide and 3.8% mercapturic acid conjugate. Mercapturic acid, which is derived from glutathione catabolism, had previously been identified in animal studies because of the conversion of acetaminophen to an electrophilic molecule. These animal studies also demonstrated that mercapturic acid is proportional to an administered dose of acetaminophen [6,7,8,9,10,11]. Of the 12 volunteers, the remaining ten were given non-radioactive APAP doses 900, 1200 and 1800 mg, and 10 were given desipramine or metyrapone followed by APAP while 7 subjects received phenobarbital and then amobarbital prior to APAP. Phenobarbital and amobarbital increased mercapturic acid in 6 of 7 and desipramine decreased mercapturic acid in 4 of 10. The mouse studies in this paper demonstrated that pretreatment with cysteamine reduced the mortality rate and hepatic necrosis if given within 2 to 3 h. Dimercaprol also reduced necrosis but desipramine did not. Timing in mice is faster than humans and is longer than the murine 2 to 3 h, perhaps 6 to 8 or 10 in humans. The authors concluded, “cysteamine protects mice from arylation of hepatic macromolecules and glutathione may serve a similar protective role in humans” [10].

Based on this groundbreaking work by Mitchell et al. suggesting cysteamine, Prescott and colleagues at the Royal Infirmary of Edinburgh were able to treat a series of patients with cysteamine in early 1974. Cysteamine showed clinical success although with some significant side effects [22]. In fact, Prescott had to be admitted to his own poisons unit when he and a registrar suffered substantial toxicity after administering cysteamine to themselves to determine dosing [20]. This toxicity was primarily gastrointestinal and dehydration. Cysteamine administered to seven patients with severe acetaminophen overdosage treated within 4 to 10 h post ingestion resulted in no hepatic necrosis in five and mild transient hepatic injury in two. Eleven untreated patients all developed severe hepatic necrosis and two died [22]. This study was crucial to the understanding that a sulfhydryl containing medication could intervene to prevent toxicity from the toxic metabolite N-acetyl-p-benzoquinone imine (NAPQI) from acetaminophen in humans.

The medical director of McNeil Laboratoires, Thomas Gates, MD was aware in early 1974 of the paper by Rumack and Matthew which had not yet been published. In addition to the nomogram, the paper included epidemiologic information on 156 reported ingestions and two fatalities recorded in Denver from acetaminophen. There were a series of meetings and discussions and McNeil Laboratories, manufacturer of Tylenol, (a subsidiary of Johnson and Johnson at the time and now part of Kenvue) agreed to provide funding for further investigation of acetaminophen toxicity and potential treatments.

Cysteamine had been previously investigated as a radio protective agent by the US Army in the 1950s and was found to be too toxic for use in circumstances involving radioactive agents, primarily the atomic bomb tests being carried out at the time. Soldiers administered cysteamine developed gastrointestinal distress, nausea and vomiting. Rumack was aware of this toxicity from cysteamine and presented this information at meetings with several scientists including those from McNeil Laboratories regarding the development of a treatment for the increasing number of cases of acetaminophen overdose being seen by US poison centers.

Two investigators at McNeil Laboratories, Elliott Piperno and D.A. Berssenbruegge began examining all medications on the market which contained sulfhydryl groups in 1974. Agents investigated included methionine, cysteine, Dimercaprol (British Anti-Lewisite-BAL), penicillamine and Mucomyst (acetylcysteine). Working with Beagle dogs they determined that the most effective treatment with the least toxicity was acetylcysteine. The results of their work which were known starting in 1974 were eventually published [23,24]. Methionine, which was being investigated clinically at Guy’s Hospital in London, was rejected for use in the United States as the survival rate in mice was decreased at higher dosages when administered at 1 or 4.5 h after acetaminophen [24]. Methionine continued to be utilized orally in some centers [25].

Rumack, Gates and colleagues then began working with the United States Food and Drug Administration (FDA) to develop an ANDA (Amended New Drug Application) to begin clinical trials in the United States of acetylcysteine (n-acetylcysteine, NAC, trade name Mucomyst).

The owner of the patent acetylcysteine (Mucomyst) was Mead Johnson (unrelated to Johnson and Johnson) and they declined to provide the work necessary to demonstrate that the orally administered sterile Mucomyst did not contain pyrogens so it could be administered intravenously. They did agree to allow access to the drug master file. During discussions with the FDA, it was made clear that the medication would only be approved for oral use. There was some consideration that oral acetylcysteine might be superior to intravenous acetylcysteine given its first-pass effect on the liver through the splanchnic circulation and absorption. It was clear from the primary use of acetylcysteine in patients with cystic fibrosis that oral administration would result in nausea and vomiting given the strong smell, despite being masked with various diluents [26].

Once the decision was made in early 1974 to use acetylcysteine, a dosing methodology was developed. A series of calculations were made and included in the submission to the FDA. The work of Mitchell et al. had shown that hepatic injury occurred in animals when glutathione was depleted by 70% and this was one of the bases for dosing calculations [10].

Calculation of acetaminophen ingestion that would produce hepatic toxicity or injury was based on 70% depletion of glutathione as demonstrated by experimental work [9,10].

The following assumptions were made:

(1)

Normal human liver has approximately 4 mmoles of glutathione (GSH) per liter [10].

(2)

Using the 1974 FDA standard 70 kg patient:

• The human liver in a 70 kg patient is about 1.5 L so ≅ 6 mmoles of GSH.
• Hepatic necrosis was reported in mice when there is 70% depletion of GSH.
• Seventy percent (0.70) × 6 mmoles = 4.2 mmoles depletion in a 70 kg patient to produce necrosis.

(3)

APAP is 151.2 g/mole (151.2 mg/mmol)

• NAPQI production is 4% of an APAP absorbed dose.
• The measurement of 4.2 mmoles GSH is ≅ to 4.2 mmoles of NAPQI so roughly equivalent
• Four percent (0.04) × APAP dose = the amount of NAPQI produced.
• Solving the toxic dose of APAP to produce a 70% depletion of GSH:
  • A measurement of 4.2 mmoles NAPQI/0.04 ≅ 105 mmols APAP.
  • A measurement of 105 mmoles APAP × 151.2 mg/mmol = 15,876 mg APAP.
• So, an absorbed dose of 15.9 g of APAP in a 70 kg human would be sufficient to deplete GSH by 70% and produce necrosis.
• Four percent of 15,876 mg of APAP would produce 635 mg of NAPQI.

Calculation of the initial acetylcysteine dosing regimen.

The assumption in 1974 was to match acetylcysteine administration with GSH depletion based on 4% of the 15.9 g dose producing 635 mg of NAPQI.

      a.

The 1.5 L liver contains ≅ 6 mmol of GSH.

      b.

Acetylcysteine is 163.2 g per mole or 163.2 mg/mmol and 1 g is 6.1 mmol.

      c.

Acetaminophen is 151.2 g per mole or 151.2 mg/mmol and 1 g is 6.6 mmol.

      d.

Acetylcysteine and acetaminophen are roughly equivalent on a molar basis.

      e.

GSH turnover was estimated to be 1.5 mmol/h and replacing 25% of the 4.2 mmol depletion was ≅1 mmol which resulted in a balance of ~2.5 mmol/h replacement per h.

      f.

Dividing 2.5 mmol/h by the 70 kg patient = 0.036 mmol/kg/hr.

      g.

A measurement of 0.036 mmol/kg × 163.2 mg/mmol acetylcysteine = 5.88 mg/kg/hr rounded to a 6 mg/kg/hr dose of acetylcysteine to replace the GSH turnover and binding to NAPQI.

Calculation of additional safety factors and accounting for 4 h dosing intervals.

(1)

Initial protocol from the calculations gives the following:

• Administering 6 mg/kg/hr of acetylcysteine on a 4 h schedule resulted in 24 mg/kg/4 h.
• An initial loading dose of twice the maintenance dose was 48 mg/kg.
• The initial protocol was a loading dose followed by 11 maintenance doses over 48 h. However, this was changed to 60 h and 14 doses when in a correction to the first submission to be consistent with a 12 h half-life of APAP.

(2)

We were fully aware that many patients consumed an overdose of greater than 15.9 g. Following several discussions with the FDA, the second resubmission of the protocol contained the following dosing methodology to account for higher doses and adding empirical safety factors, along with other changes:

• Loading dose of 140 mg/kg
• Seventeen maintenance doses every four hours of 70 mg/kg
• Acetylcysteine was administered for 72 h with a total dose of 1330 mg/kg.

4. Acetylcysteine Clinical Studies in Acetaminophen Overdose

A randomized controlled trial of acetylcysteine was an important part of the original submission to the FDA. The FDA was aware of the concerns that had been expressed about the ethics of a controlled trial in discussions held in 1974 and 1975. One concern was expressed at the conclusion of an article published in 1974 and the others were not published until 1976 [27,28,29]. Several letters were also received by the FDA questioning the ethics of a trial that would withhold acetylcysteine from the control group.

The FDA rejected the inclusion in our submission of a randomized controlled trial. Instead, the FDA required that it be re-written to treat all patients and use the historical patients from the Prescott et al. 1971 publication as the comparator [4]. This came as a great surprise to us as it was quite unusual for the FDA to require anything other than a randomized controlled trial. There has never been a placebo-controlled trial of acetylcysteine and no dose-response studies have ever been performed.

Further changes to our submission required that the nomogram developed in 1973 starting at 200 mg/L at 4 h have a new line with the same slope beginning at 150 mg/L at 4 h (the “treatment line”) so that there would be a 25% safety factor. This “study design nomogram” also required a line at 300 mg/L at 4 h to see if there were differences with high-risk patients [30] (Figure 3). We added study lines at 400 mg/L and 500 mg/L to see if other risk levels could be determined. Other changes included the requirement that the acetylcysteine dosage in the protocol would have a loading dose and 17 additional doses over 72 h. The protocol also included the requirement that all samples of blood adhering to the protocol were to be shipped to the University of Colorado School of Medicine Drug Assay Laboratory by Federal Express. Thus, the concentrations could be compared as they were all analyzed by the same laboratory.

We had proposed in our submission that the study would be a United States National Multicenter Study (USNMS) where any physician in the United States could call a toll-free number at the Rocky Mountain Poison and Drug Center in Denver and after agreeing to the full protocol could administer acetylcysteine. Acetylcysteine was widely available in hospitals as Mucomyst. Patients were entered into the study based only on the history obtained by their physician. Very few hospitals were able to measure acetaminophen. This meant that many patients were treated but when acetaminophen levels were later obtained, they were below the treatment line. These patients were considered “acetylcysteine safety” patients as they received all 18 doses of acetylcysteine. The intent of the USNMS was to rapidly collect cases to figure out if acetylcysteine was a safe and effective treatment for an acetaminophen overdose in a large population.

The FDA approved the protocol incorporating several changes as described. The United States National Multicenter Study officially began accepting patients on 1 September 1976. The first patient treated with acetylcysteine for an acetaminophen overdose was reported in JAMA in 1977 [31]. Details of the approved protocol were presented at a symposium in New York City in November of 1977 and published in the proceedings in 1978 [32].

During this time the group in Edinburgh were also determining a protocol for intravenous administration of acetylcysteine as it was less toxic than cysteamine. The Edinburgh protocol was empirical but based on sound pharmacologic principles [33]. The UK was willing to permit acetylcysteine administration by the intravenous route. An intravenous regimen over 20.25 h was utilized and was published in 1979 [34]. Intravenous acetylcysteine was given as an initial dose of 150 mg/kg in 200 mL 5% dextrose over 15 min followed by 50 mg/kg in 500 mL 5% dextrose over four hours and 100 mg/kg in one liter 5% dextrose over the next 16 h. The total dose of the intravenous protocol was 300 mg/kg over 20.25 h as compared to the oral protocol with a total dose of 1330 mg/kg over 72 h. Intravenous acetylcysteine produced an anaphylactoid reaction along with some flushing, nausea and vomiting in some patients. These side effects have decreased with changes in the protocol extending the length of time over which the highest concentration is administered and decreasing the length of acetylcysteine administration in some cases [35,36].

The Guy’s Hospital group of toxicologists in London investigated the use of methionine for acetaminophen overdose [37,38]. The publication from this group in 1981 reported that 132 cases of severe acetaminophen (paracetamol) poisoning were treated with oral methionine. Seven of 96 patients who received the antidote within ten hours of ingestion of the overdose had severe liver damage (aspartate transaminase level, greater than 1000 IU/L), but none of these patients died. Thirty-six patients received methionine between 10 and 24 h of ingestion; severe liver damage occurred in 47%, and two patients died. Despite some success in the treatment of acetaminophen overdose, methionine is no longer utilized.

The results of the first 100 patients treated with oral acetylcysteine in the USNMS were reported at a symposium held in New York City in November of 1977 [32]. Of the 49 patients treated within 10 h or less post overdose (by history), 17% (8) had transaminases of over 1000 IU/L and 45% (23) of those treated between 10 and 24 h developed transaminases over 1000 IU/L. For those patients who received the full acetylcysteine course but were later determined not to have acetaminophen levels above the nomogram line (“safety controls”), there were no abnormalities in any of the laboratory measurements. Acetylcysteine was therefore demonstrated to be effective when compared to the historical controls who, with supportive care, developed transaminases over 1000 IU/L in 55 to 71% of cases [39,40].

Prescott and colleagues published 100 cases of acetaminophen overdose treated with intravenous acetylcysteine in 1979 [34]. Of the 62 patients treated within 10 h or less post ingestion, only 2% (1) of patients developed a transaminase greater than 1000 IU/L. Of 38 patients treated between 10 and 24 h, 53% (20) of patients developed a transaminase greater than 1000 IU/L. There were 57 patients in this study that received supportive care only (controls) and 58% (33) developed a transaminase greater than 1000 IU/L. Further data were provided after stratifying patients into various risk groups. It was concluded that intravenous acetylcysteine was effective in protecting against hepatic toxicity when given within 8 to 10 h post ingestion.

The USNMS results were further detailed at a symposium in 1979 and published in 1981 [30]. The data utilized in this publication formed the basis for the submission to the FDA in support of the Abbreviated New Drug Application. The submission to the FDA was not required to provide historical controls as the data from the “safety” and untreated patients were considered sufficient. The FDA gave formal approval on 31 January 1985 for the oral use of acetylcysteine in the treatment of acetaminophen overdose. It carried the designation Mead Johnson 13–601 as that company owned the patent on Mucomyst.

The USNMS did not include children under the age of 13 but a parallel study was conducted at the same time. A paper looking at pharmacokinetics in children was published in 1978 and showed that based on half-life determinations, the metabolism is slower in the neonate but comparable to adults in both children and adolescents [41]. A previous pharmacokinetics paper after a dose of 10 mg/kg demonstrated that in children aged 9 and younger, sulfate conjugation was the primary route, but that at age 12 and above the “adult” glucuronide was predominant [42]. We were concerned that if sulfate conjugation was rate limiting, we might see higher levels of toxicity in children but in fact we saw lower levels of toxicity with similar acetaminophen concentrations. An initial publication in 1981 looking at 300 potential exposures reported that 17 had plasma levels measured with 5 above the 150 mg/L at the 4 h treatment line and 11 received acetylcysteine. The highest transaminase was 1660 IU/L and none of the remainder reached 1000 IU/L. There were 417 total cases of suspected exposure to an acetaminophen overdose collected in the parallel study to the USNMS and published in 1984 [43]. There were 55 patients with acetaminophen concentrations in the toxic range with the remaining either non-toxic or not interpretable. Of the 55 patients with potentially toxic levels, 43 were treated with acetylcysteine and 3 developed a transaminase of 1000 IU/L or greater. An additional 56 patients in the non-toxic group were treated with acetylcysteine without any laboratory abnormalities. The current recommendation is to treat children under the age of 13 in the same manner as adolescents and adults are treated, although the incidence of hepatic toxicity appears lower than that in patients aged 13 or greater.

The USNMS concluded in 1985 and a full analysis was published in 1988 in the New England Journal of Medicine [26]. There were 11,195 cases entered into the study, of which 2540 were treated with acetylcysteine and met inclusion criteria. There were 517 patients who received the full acetylcysteine treatment protocol but were later determined to be below the 150 mg/L line and were considered “acetylcysteine safety patients” as they did not meet inclusion criteria. The patients were stratified on a study nomogram utilizing lines beginning at 4 h and starting at 150 mg/L, 200 mg/L, 300 mg/L, 400 mg/L and 500 mg/L and further defined by delay to treatment. Those patients above the 300 mg/L line were characterized as “high risk”. The 400 mg/L and 500 mg/L nomogram lines were not used in the final analysis. A standard nomogram simplified from the study design nomogram for assessing risk was in use from 1985 to 2023 (Figure 4).

As previously shown in smaller studies, those patients treated within the first 8 to 10 h post ingestion of acetaminophen (probable risk) had the lowest incidence of hepatic toxicity (transaminase greater than 1000 IU/L), which was 6.1% (32/527) in this study. The subset of those treated within the first 10 h whose concentrations were in the high-risk range had a higher incidence of hepatic toxicity, at 8.3% (17/206). There was a stepwise increase in hepatic toxicity related to longer time from ingestion to treatment between 10 and 24 h, with probable risk patients at 26.4% (247/935) and high-risk patients at 34.4% (199/578). Those patients between 16 and 24 h were all high risk with 41% (116/283) showing hepatic toxicity. These reports from the USNMS indicated that oral and intravenous acetylcysteine had a similar therapeutic benefit.

Because most hospitals could not measure acetaminophen concentrations, there were 517 patients treated based on history who did not have levels in excess of the 150 mg/L nomogram line. These patients had an incidence of 1.08% to 3.16% of transaminases greater than 1000 IU/L showing a non-zero risk of hepatotoxicity, possibly due to the inaccurate history of the time of ingestion of the overdose [44] (Figure 5).

Intravenous acetylcysteine, following the protocol from Edinburgh, became the preferred method of administration. However, the UK regulatory authorities have required treatment based on a very conservative “100 mg/L at 4 h” post ingestion treatment line since 2012 [44].

Oral acetylcysteine remained the primary treatment for acetaminophen overdose in the United States until the intravenous version was approved by the FDA in January 2004. Intravenous acetylcysteine is now the most common method of treatment throughout the world. Oral acetylcysteine is still utilized occasionally although the smell, taste and resultant vomiting make it considerably less desirable.

A comparative study of intravenously treated patients compared to oral acetylcysteine-treated patients reported some may do better with the oral protocol [45]. This is not likely related to the route of administration but rather the total dose and duration of the acetylcysteine administered. The data comparison was between 1963 US patients treated orally and 2086 Canadian patients treated intravenously. The 20 h IV protocol was more effective when instituted early while the 72 h oral protocol was more effective when instituted in delayed patients. This is likely due to the 300 mg/kg total IV dose versus the 1330 mg/kg oral dose. Additional publications support this conclusion [26,33,45,46,47,48].

5. Acetylcysteine in Pregnancy

There were 113 patients who were pregnant in the USNMS and 60 cases had full pregnancy outcome data [49]. There were 24 patients with acetaminophen levels above the treatment line of 150 mg/L. Of the ten treated with acetylcysteine within 10 h post-ingestion, eight delivered normal infants and two had elective abortions. Of ten patients treated 10–16 h post ingestion, five delivered viable infants, two had elective abortions, and three had spontaneous abortions. Of four treated within 16–24 h post ingestion, there was one spontaneous abortion, one stillbirth, one elective abortion, one delivery of a normal infant and one mother died. There was a statistically significant increase in the incidence of spontaneous abortion or fetal death when treatment was begun late. Acetylcysteine crosses the placenta and can be measured in the neonate [50]. Mean level of acetylcysteine in cord blood was 9.4 mcg/mL which is between the peak 13.9 mcg/mL and trough 5 mcg/mL levels previously reported [51]. The recommendation based on this small series is to treat as early as possible with acetylcysteine following normal protocols.

6. Acetylcysteine Failure in Renal Toxicity

Renal toxicity, while associated with fulminant hepatic failure in acetaminophen overdose, may also occur in patients with less severe cases of overdose. Acetylcysteine is ineffective in the treatment of renal toxicity and experimental work has uncovered the mechanism and provided an explanation [52]. While formation of APAP-CYS protein adducts occurs in the mitochondria of hepatocytes, these adducts are formed in the endoplasmic reticulum (ER) of proximal tubular cells of the kidney. The reactive metabolites trigger an ER stress-mediated activation of caspase-12 which results in apoptotic cell deaths in this area of the kidney. This contrasts with necrotic cell death in the liver. Acetylcysteine does not have any effect on this process in the kidney but fomepizole does prevent injury by attenuating renal cell death from apoptotic death in primary human kidney cells. Thus, based on the experimental evidence, fomepizole should be considered as an adjunct to acetylcysteine in patients with renal injury following acetaminophen overdose.

7. Acetylcysteine Administration, Stopping Criteria and Further Risk Analysis

Numerous one and two bag protocols have been devised to reduce the adverse effects of intravenous acetylcysteine, to reduce the complexity of the original three bag protocol and to shorten the time of hospitalization, especially in low-risk patients [35,36,53,54,55,56,57,58,59,60]. The Scottish and Newcastle anti-emetic pretreatment (SNAP) trial was quite rigorous and provides a methodology to stop treatment at 12 h [36]. An evaluation of adducts supports early cessation of acetylcysteine in appropriate circumstances [58].

Examples of acetylcysteine administration, as referenced above:

Oral—still utilized in some countries where intravenous use is not possible:

140 mg/kg loading dose followed by 17 doses 70 mg/kg every 4 h [26].

IV—original dose regimen: 150 mg/kg over 15 min, 50 mg/kg over 4 h and then 100 mg/kg over 16 h. Modified the initial infusion from 15 min to 1 h. No effect on anaphylactoid reactions, nausea or vomiting [34].

IV—one bag method: 30 g of NAC in one liter of IV fluid with a loading dose of 150 mg/kg over 1 h followed by a continuous infusion of 12.5 mg/kg/h [55].

IV—two bag 200 mg/kg over 4 h and 100 mg/kg over 16 h [56]

IV—SNAP Trial: 300 mg/kg over 12 h using a rapid infusion for 2 h (100 mg/kg) and a slower infusion for 10 h (200 mg/kg). Continue as needed [36].

IV—Hi-SNAP Trial: 300/kg, 450/kg and 600/kg study in progress [61].

Summary: At least 300 mg/kg during the first 20 to 24 h of treatment

(except SNAP or Hi-SNAP).

The length of treatment with acetylcysteine is sufficient to decrease toxicity as part of the 20.25 h intravenous or 72 h oral protocol. However, providing criteria related to the status of the patient at the end of each protocol may be more appropriate [62]. Generally, there is no reason to administer acetylcysteine after there is no more acetaminophen to be converted to NAPQI and produce toxicity. However, there is experimental evidence that indicates continued administration of acetylcysteine may interfere with hepatic regeneration [63]. A recent consensus document for the United States and Canada makes specific recommendations in terms of stopping acetylcysteine [64]. This consensus publication also contains a revised nomogram showing only the 150 mg/L at 4 h and the 300 mg/L at the 4 h high-risk line. (Figure 6)

Acetylcysteine stopping criteria include the following:

• Acetaminophen concentration <10 μg/mL (mg/L);
• International normalized ratio (INR) <2.0;
• ALT/AST normal for patient, or if elevated have decreased from peak (25–50%);
• Patient is clinically well.

If acetylcysteine stopping criteria are NOT met then acetylcysteine should be administered at a rate of 6.25 mg/kg/hr or greater and transaminases, INR and APAP levels drawn at 12-to-24 h intervals until stopping criteria are met.

The use of acetylcysteine in each patient is determined from a risk analysis by the treating physician. The use of the nomogram to plot the measured acetaminophen concentration versus time remains the primary method of risk determination where such laboratory analyses are rapidly available [64]. Several other methodologies have been developed to refine risk including a widely utilized multiplication product [65]. This simple calculation of the APAP concentration × ALT level provides a product estimating risk and is interpreted as follows:

Less than 1500 = low likelihood of hepatotoxicity with acetylcysteine-treatment (100% sensitivity and 100% specificity);

More than 10,000 = high likelihood of hepatotoxicity with acetylcysteine-treatment (sensitivity 80%, specificity 99.6%).

A more elegant approach to risk evaluation for use of acetylcysteine was developed known as the Psi parameter [66,67]. While the patient history of APAP dose ingested may be inaccurate, the Psi parameter utilizes the measured concentration to estimate the toxic metabolite (NAPQI) and estimate the quantity ingested during exposure using known pharmacokinetic principles.

Recently, an augmented method combining both the multiplication product and the Psi parameter was proposed [68]. This publication demonstrates that risk calculation from the acetaminophen concentration can be augmented with the inclusion of the timing of acetylcysteine treatment. Utilizing the proposed calculation, the authors were able to have both high sensitivity (96.5%) and high specificity (97.3%) based on an analysis of 421 case records. These authors previously provided a methodology for the evaluation of patients to receive acetylcysteine where the rapid availability of an acetaminophen level or transaminases is not possible [69]. This dose estimate method, using 150 mg/L as a cutoff, has a specificity of 55.3% when it is used, which results in 44.7% of patients being treated unnecessarily. The authors consider this acceptable in those circumstances where detailed rapid laboratory analysis is unavailable.

8. Acetylcysteine Adjunctive Treatment with Fomepizole and Other Therapies

The use of fomepizole as an adjunct to acetylcysteine should be considered especially in patients with delay until treatment, massive ingestions or renal injury. There is very solid experimental evidence for this adjunctive treatment [70]. Clinical use of fomepizole as an adjunct relies primarily on case reports and case series demonstrating use in humans [71,72]. It is hoped that current clinical trials underway will be able to answer the question as to use in a controlled setting. Fomepizole has been used for over 20 years effectively and safely with cases of methanol and ethylene glycol toxicity [73]. There is no known toxicity when utilized with acetylcysteine. There may be a place for adding fomepizole to acetylcysteine therapy to take advantage of its ability to inhibit CYP2E1 production of NAPQI and interfere further downstream with c-Jun N-terminal kinase [74,75]. Use of fomepizole in place of acetylcysteine is not recommended currently and although it is widely used, the results of clinical trials are yet to demonstrate absolute efficacy.

In addition to fomepizole, other therapies have been considered and some are under investigation. The following have been extensively reviewed in a recent publication [74].

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Calmangafodipir is a superoxide dismutase “mimetic” (manganese-dependent superoxide dismutase, Mn-SOD) 1. SOD is required in the cellular defense of reactive oxygen species and interferes with the formation of peroxynitrite, increases nitrotyrosine formation and hepatic toxicity. Mn-SOD mimetics, e.g., Mito-Tempo (mitochondria-targeted antioxidant) can enter mitochondria. Increasing Mn-SOD capacity can be effective in treating APAP toxicity. A phase IIb/III Albatross Trial of calmangafodipir for acetaminophen toxicity was to commence Q1 2024 but has not begun as of the writing of this article.

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Nrf2 binds to the ARE (antioxidant response element) and induces transcription of a large number of genes which all have protective roles in the APAP hepatotoxicity model. However, Nrf2 is activated by APAP and there may be no further benefit from further activation.

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Thrombopoietin Mimetic Peptide1 (PEG-TPOm) JNJ-26366821. PEG-TPO treatment appears to be beneficial when administered at 24 h after APAP overdose when NAC is ineffective. PEG-TPO arrests the progression of acetaminophen overdose-induced liver injury (AILI) and accelerates the onset of the proliferative response essential for liver recovery. Distinct from 4-MP which prevents injury but does not promote hepatocyte proliferation and liver recovery, PEG-TPO is a potential novel therapeutic for the enhancement of liver recovery after AILI1.

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Adenosine A2B receptor activators such as BAY 60-6583 decreased necrosis and enhanced infiltration of reparative macrophages when NAC would be ineffective. Clinical trials in cancer patients have begun.

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Whartons Jelly Mesenchymal Stem Cell (WJMSC) protected against liver injury at 6 h by preserving mitochondrial function despite JNK activation and its mitochondrial translocation accompanied by enhanced infiltration of macrophages with the reparative anti-inflammatory phenotype by 24 h. Clinical trials have begun.

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Lipid-nanoparticle-encapsulated mRNA of HGF (human grown factor) and EGF epidermal growth factor have been shown to be effective in the mouse model and may be further investigated.

9. Conclusions

Acetylcysteine has been the primary treatment for acetaminophen-overdose toxicity for over 50 years. Foundational research in the early 1970s played a critical role in the treatment of this overdose without which treatment could not have been devised. Acetylcysteine treatment, despite never having been part of a randomized controlled trial nor having a dose–response curve remains the successful treatment of choice to treat toxicity in most patients from this overdose. Standard intravenous treatment is 21 h and multiple methods of administration have been devised to reduce adverse reactions and deliver acetylcysteine safely. Patients with massive ingestions or delay in arrival in a health care facility may require longer treatment with acetylcysteine and/or at a higher dose. Individualized patient-focused treatment is considered the standard. Stopping criteria have been developed and these methods have been formalized in a recent consensus publication. Experimental work has shown that renal injury is based on a different mechanism and is characterized by apoptosis rather than necrosis as seen in the liver. Augmentation with fomepizole has been experimentally demonstrated and may be especially useful in patients with massive ingestions, delay until treatment or renal injury.