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\begin{document}

\title{Therapeutic potential of mega-dose vitamin C to reverse organ
dysfunction in sepsis and COVID-19}



\author[1]{Clive May}%
\author[2]{Rinaldo Bellomo}%
\author[1]{Yugeesh Lankadeva}%
\affil[1]{The Florey Institute of Neuroscience and Mental Health}%
\affil[2]{The University of Melbourne}%


\vspace{-1em}



  
  \date{May 20, 2024}


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\selectlanguage{english}
\begin{abstract}
Sepsis causes multi-organ dysfunction and is a major cause of death in
intensive care units, but there are no treatments that reverse the
pathophysiological effects of sepsis. Vitamin C has antioxidant,
anti-inflammatory, anticoagulant and immune modulatory actions, so is a
potential treatment for sepsis. Recent clinical trials of high-doses of
intravenous vitamin C (6-16 g/day) had variable effects. Since much
higher doses are without side-effects in cancer and burns patients, we
studied the effects of a mega-dose of intravenous sodium ascorbate (150
g/40 kg) in a clinically relevant ovine model of sepsis. This treatment
dramatically improved the clinical state and over 3-7-h improved
cardiovascular, pulmonary, hepatic and renal function and reduced body
temperature. In a critically ill COVID-19 patient, intravenous sodium
ascorbate (60 g) restored arterial pressure, improved renal function and
increased arterial blood oxygen levels. Clinical trials are testing the
effectiveness of mega-dose vitamin C in septic patients.%
\end{abstract}%



\sloppy


\textbf{Introduction}

Sepsis is a pathophysiological syndrome characterised by an overwhelming
inflammatory and immune response to a bacterial, viral or fungal
infection that can lead to multi-organ dysfunction and death (Singer et
al., 2016). Sepsis is the leading cause of mortality in intensive care
units with an annual global incidence of 49 million cases and 11 million
deaths (Rudd et al., 2020). Standard of care treatment for sepsis
consists of antibiotics, fluid resuscitation and vasopressors (Rhodes et
al., 2017), with continuous renal replacement therapy being increasingly
used in critically ill patients (Bellomo, Baldwin, Ronco \& Kellum,
2021). These interventions are mostly aimed towards keeping the patient
alive in the expectation that organ function should recover following
resolution of the infection. However, patients who recover from severe
sepsis frequently exhibit a degree of chronic organ dysfunction.
Currently there are no treatments that reverse sepsis-induce organ
dysfunction.

\textbf{Pathophysiology of sepsis-induced cardiovascular and renal
dysfunction}

The factors causing sepsis-induced organ dysfunction remain unclear due
to the complex pathophysiology of sepsis that changes as the response to
the infection progresses. There is evidence that redox homeostasis is
disrupted in sepsis resulting in oxidative stress, which together with
excessive inflammation is thought to cause mitochondrial, endothelial
and microvascular dysfunction, resulting in vasoplegia,
inflammation-mediated tissue injury, tissue hypoxia and multi-organ
dysfunction (Joffre \& Hellman, 2021) (Lankadeva, Okazaki, Evans,
Bellomo \& May, 2019).

Hypotension secondary to peripheral vasodilatation is a hallmark of
sepsis that is treated with aggressive fluid resuscitation and
vasopressor therapy to restore target mean arterial pressure (Rhodes et
al., 2017). Reduced vascular responsiveness to noradrenaline, the
primary vasopressor used clinically, is common in sepsis resulting in
persistent and sometimes refractory hypotension (Annane et al., 1998),
which, in itself, can result in tissue hypoperfusion and hypoxia,
mitochondrial dysfunction and multi-organ failure.

Indeed, tissue hypoperfusion and hypoxia in the renal medulla are
critical pathophysiological features of ovine hyperdynamic sepsis that
precede the development of acute kidney injury (AKI) by 8 to 12-h
(Calzavacca, Evans, Bailey, Bellomo \& May, 2015). Renal medullary
hypoxia can lead to mitochondrial dysfunction, initiating a progressive
loss of renal function culminating in AKI (Lankadeva, Okazaki, Evans,
Bellomo \& May, 2019; Nourbakhsh \& Singh, 2014). Therapies that target
these sepsis-induced pathophysiological processes may confer better
circulatory management and mitigate AKI.

\textbf{Antioxidants as a therapy for sepsis-induced organ dysfunction}

In view of the damaging effects of tissue hypoxia and oxidative stress,
there has been interest over many decades in the use of antioxidants as
a treatment for sepsis. The antioxidant N-acetylcysteine, which has both
antioxidant and anti-inflammatory properties, showed some promise in
experimental studies, but clinical studies have yielded largely
disappointing findings (Chertoff, 2018). The antioxidant tempol has been
shown to reduce the level of AKI in a porcine model of sepsis (Matejovic
et al., 2005), but its effects have not been examined clinically.
Recently, however, there has been increasing interest in the effects of
vitamin C as a treatment for sepsis.

\textbf{Rational for vitamin C therapy in sepsis}

Vitamin C, ascorbic acid, is an essential vitamin in humans that must be
obtained in the diet; it cannot be synthesised because of mutations in
the gene for gluconolactone oxidase, the final biosynthetic enzyme in
its synthesis. Vitamin C is essential for collagen synthesis, accounting
for the symptoms of scurvy caused by its deficiency. Vitamin C also has
numerous pleiotropic effects that would be predicted to be of benefit in
sepsis, including as an anti-oxidant, anti-inflammatory, anticoagulant,
immune-modulator and stimulant of noradrenaline and vasopressin
synthesis (Holford et al., 2020). Furthermore, septic patients have
abnormally low plasma vitamin C levels, likely due to increased
metabolic turnover (de Grooth et al., 2018) and downregulation of
cellular sodium-dependent vitamin C transporters (SVCTs) (Subramanian,
Sabui, Moradi, Marchant \& Said, 2018), which is compounded by the
inability of humans to synthesise vitamin C. Importantly, intravenous
administration is required to produce high plasma levels of vitamin C as
there is a limit on the intestinal absorption (Padayatty et al., 2004).
These observations provided the impetus for clinical trials examining
the effects of intravenous vitamin C in sepsis.

\textbf{Controversies with high-dose vitamin C therapy in human sepsis}

Single-centre controlled randomised clinical trials (RCT) showed that
intravenous vitamin C reduced inflammatory biomarkers and reduced
sequential organ failure assessment (SOFA) scores (50, 100, 200
mg/kg/day, n=24) (Fowler et al., 2014) and improved vasopressor
sensitivity (2 g 4 times/day, n=28) (Zabet, Mohammadi, Ramezani \&
Khalili, 2016). A widely publicised single-centre before and after study
(n=47), using a combination therapy of vitamin C (1.5 g 4 times/day)
with hydrocortisone and thiamine, reduced organ failure and mortality
from 40.4\% to 8.5\% (Marik, Khangoora, Rivera, Hooper \& Catravas,
2017). However, subsequent multi-centre RCTs, VITAMINS (Fujii et al.,
2020), ACTS (Moskowitz et al., 2020) and ATESS (Hwang et al., 2020) that
trialled a maximum dose of vitamin C of 6 g/day for up to 10 days with
thiamine \selectlanguage{ngerman}± corticosteroid, had no significant benefit above placebo
(Fujii et al., 2020) (Moskowitz et al., 2020) (Hwang et al., 2020). The
CITRUS-ALI trial, that used 200 mg/kg/day (16 g/day in an 80 kg patient)
of vitamin C for 4 days, however, reduced 28-day mortality from 46\% to
30\% (Fowler et al., 2019).

We hypothesized that the lack of consistent benefit in the clinical
trials of vitamin C in sepsis might be due to the use of inadequate
doses. In view of these findings, and the fact that very high doses of
intravenous vitamin C have been shown to be safe in burns and cancer
patients (Yanase et al., 2020), we recently investigated the safety and
efficacy of a much larger dose (mega-dose) of vitamin C in experimental
sepsis.

\textbf{Rationale for mega-dose vitamin C treatment in sepsis}

Intravenous administration of a mega-dose of sodium ascorbate (150
g/\textasciitilde{}40 kg 7-h) in established ovine hyperdynamic sepsis,
induced by infusion of live \emph{Escherichia. coli} , caused a
remarkable improvement in the clinical state from malaise, lethargy and
somnolence to an alert, responsive, mobile state (Lankadeva et al.,
2021). MAP was restored to pre-septic levels with reduced noradrenaline
requirements, which decreased to zero in 4 of 5 cases. Mega-dose vitamin
C increased arterial blood oxygen levels, indicative of improvements in
lung function, restored body temperature from febrile to normal levels
and reduced arterial blood lactate indicating improved metabolic
function. The treatment also reversed renal medullary hypoperfusion and
hypoxia, accompanied by a reversal in septic AKI, as shown by dramatic
increases in urine flow and creatinine clearance leading to a
normalisation of plasma creatinine levels.

The re-distribution of intra-renal perfusion in ovine septic AKI
(Calzavacca, Evans, Bailey, Bellomo \& May, 2015) is accompanied by
reduced gene expression of renal medullary endothelial nitric oxide
synthase (eNOS) (Langenberg, Bagshaw, May \& Bellomo, 2008). The
reversal of renal medullary microcirculatory dysfunction by vitamin C
may result from its ability to increase eNOS activity and thus nitric
oxide bioavailability (Ladurner et al., 2012). Furthermore, the
antioxidant effects of vitamin C are likely to reduce mitochondrial
dysfunction and cellular injury.

\textbf{Treatment of COVID-19 with vitamin C}

Critically ill COVID-19 patients develop an excessive inflammatory
response, disseminated intravascular coagulation and multi-organ
dysfunction. Immunosuppressive agents like tocilizumab (a humanized
monoclonal antibody against the interleukin-6 receptor) (Salama et al.,
2021) and dexamethasone (Group et al., 2021) have been shown to be
beneficial treatments for such patients. The known actions of vitamin C
indicate that it would also be a plausible adjunct treatment for
COVID-19. The findings that plasma vitamin C levels are low in COVID-19
patients (Chiscano-Camon, Ruiz-Rodriguez, Ruiz-Sanmartin, Roca \&
Ferrer, 2020), and that vitamin C lowers expression of angiotensin
converting enzyme 2, the entry point for SARS-CoV-2 into cells (Ivanov,
Goc, Ivanova, Niedzwiecki \& Rath, 2021), further indicate that it may
have beneficial actions in COVID-19. Intravenous vitamin C (1.5-14.0 g)
has been investigated in COVID-19 patients with mild beneficial effects,
but the effects of mega-doses have not been studied.

Following our finding of potent beneficial effects of mega-dose vitamin
C in ovine sepsis, a critically ill patient with COVID-19 induced acute
respiratory distress syndrome, hypotension and AKI was treated with
intravenous sodium ascorbate (60 g over 7 hours) (Lankadeva et al.,
2021). As in septic sheep, sodium ascorbate restored arterial pressure
in the face of withdrawal of noradrenaline, accompanied by reduced
plasma creatinine, increased urine flow and reduced heart rate. Arterial
blood oxygen levels improved while fractional inspired oxygen was
reduced. The patient was subsequently extubated and discharged from
hospital 22 days after mega-dose vitamin C treatment.

\textbf{Clinical trials of mega-dose vitamin C}

A pilot placebo-controlled RCT is currently underway examining the
effects of intravenous mega-dose vitamin C treatment (sodium ascorbate,
60 g) in 30 septic patients (ACTRN12620000651987p) in Australia with an
additional RCT in COVID-19 patients scheduled to commence in Columbia.

\textbf{Conclusions}

Our findings demonstrate a potent ability of mega-dose intravenous
vitamin C to reverse organ dysfunction and improve the clinical state in
a clinically relevant ovine model of sepsis. We also demonstrated the
safety of this treatment and its benefit in a COVID-19 patient. It is
now critical to complete dose-response studies in which the plasma
levels of vitamin C are measured to determine the optimum dosing
regimen. Further studies are necessary to determine the mechanisms by
which vitamin C reverses multi-organ dysfunction in sepsis, with a focus
on its anti-oxidant and anti-inflammatory actions. Such studies are
essential to provide the scientific rationale for the design of large
double-blinded multi-centre RCTs.

\textbf{Figure legend}

Overview of the systemic and organ specific effects of sepsis, the
effects of treatment we have observed with intravenous mega-dose vitamin
C in ovine sepsis and the possible mechanisms of action.

\textbf{Conflicts of Interest:} YRL, RB and CNM have a provisional
patent on Vitamin C use in sepsis (\textbf{2020901120).}

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