Comparisons of Colistin-Induced Nephrotoxicity Between Two Different Formulations of Colistin in Critically Ill Patients: A Retrospective Cohort Study

Colistin is widely used for the treatment of nosocomial infections caused by carbapenem-resistant gram-negative bacilli (CR-GNB). Colistin-induced nephrotoxicity is one of the major adverse reactions during colistin treatment. Comparisons of colistin-induced nephrotoxicity between different formulations of colistin are rarely reported. We conducted a retrospective cohort study that enrolled ICU-admitted patients with cultured isolates of CR-GNB and treatment with intravenous colistin. Occurrences of acute kidney injury (AKI) during treatment with intravenous colistin were recorded. Colistin-induced nephrotoxicity between two formulations of colistin, Locolin® and Colimycin®, were compared. The treatment outcomes associated with the occurrence of colistin-induced nephrotoxicity were also investigated.

Colistin is a polypeptide antibiotic with speci c action against gram-negative bacteria [5]. Despite the increasing use in the management of nosocomial infection, the use of intravenous colistin is frequently limited by its adverse reactions, including nephrotoxicity and neurotoxicity [6][7][8][9]. It is proposed that the interaction between colistin and phospholipid in the cell membrane can lead to increased membrane permeability of tubular epithelial cells and acute tubular necrosis [10]. The presentations of colistininduced nephrotoxicity include a decrease in creatinine clearance, proteinuria, cylindruria, or oliguria and usually occur in the rst 5-7 days of treatment [11][12][13][14]. If discontinued early, the acute kidney injury (AKI) is mostly alleviated within 10 days from discontinuation [13].
Several clinical factors, including old age, hypoalbuminemia, baseline renal function impairment, underlying comorbidities, concomitant nephrotoxins, higher colistin dose, and presence of septic shock, have been proposed to be related to colistin-induced nephrotoxicity [15][16][17][18]. However, the risk of nephrotoxicity by different brands of colistin has rarely been compared until now. Different products of colistin may have their speci c pharmacological characteristics. Their lipopeptide components may also be different, which may lead to a different risk of nephrotoxicity. In the present study, we retrospectively enrolled critically ill patients who were treated with two different formulations of intravenous colistin. The occurrence of AKI between patients who were administered with the two different formulations of colistin and the clinical factors associated with AKI were investigated. The effect of AKI on treatment outcomes of critically ill patients was explored as well.

Patients and settings
This was a retrospective study conducted in a referral medical center in Taiwan. From Jan 2016 to Oct. 2018, intensive care units (ICU)-admitted patients were enrolled if CR-GNB, including CRAB, CRE, and CRPA isolates was cultured from their clinical specimens and if they received intravenous colistin for ≥ 48 hours. In patients with more than one treatment course, only the rst treatment course was included for analysis. Patients were excluded if < 20 years old, with a history of ESRD, under regular dialysis when initiation of intravenous colistin, missing baseline creatinine results, without at least three follow-up creatinine data, and treated with different brands of colistin in one treatment course. The study was approved by the Institutional Review Board of Taipei Veterans General Hospital and informed consents were waived. (IRB Nos: 2019-11-009AC).

Data collection
The demographic characteristics (age, gender, BMI, smoking status) and underlying comorbidities (diabetes, malignancies, renal insu ciency, chronic liver disease, and heart failure) were obtained from hospital chart review. The infection sources were determined according to the site of specimens collected. In patients with multiple samples with CR-GNB, the sample which was obtained most close to the date of prescription of colistin was used to de ne infection source. The disease severity was evaluated by Acute Physiology and Chronic Health Evaluation (APACHE) II score on the day of ICU admission. The presence of respiratory failure and septic shock on the day of specimen collection were also recorded.

Intravenous colistin administration and concomitant nephrotoxins
All the enrolled patients received intravenous colistin for ≥ 48 hours. The intravenous colistin was initiated within seven days of CR-GNB cultured from clinical specimens. For patients with multiple intravenous colistin treatment courses, only the rst treatment course was recorded. Two formulations of colistin, Locolin® (Gentle, Taiwan) and Colimycin® (T.T.Y., Taiwan), were available during the study period. Both the formulations were supplied as 66.8mg of colistin base activity per vial, which was considered equivalent to 2 million IU of sodium colistin methanesulfonate. The recommended loading and maintenance dosing of intravenous colistin was based on previous suggestion [19], and adjusted based on body weight and renal function (Supplementary Table 1). Estimated glomerular ltration rate (eGFR) was estimated with the Chronic Kidney Disease Epidemiology Collaboration equation [20].
Maintenance dose of intravenous colistin above the recommended dose was determined as inappropriate colistin dosage.
Concomitant nephrotoxins, including aminoglycoside, vancomycin, and intravenous contrast, that were administered within 28 days of colistin treatment were also recorded.

Outcome de nition
The primary outcome evaluated in the present study was colistin-induced nephrotoxicity, which was de ned as the occurrence of AKI during colistin treatment. Serum creatinine levels were recorded at baseline (day 1 of intravenous colistin administration) and thereafter until the end of colistin treatment or death. AKI was determined based on the de nition of KDIGO recommendation by creatinine criteria [21]. The occurrence of stage 1 (with 1.5-1.9 fold increase or ≥ 0.3mg/dL increase in serum creatinine), stage 2 (with 2-2.9 fold increase in serum creatinine than baseline) and stage 3 (≥ 3 fold increase in creatinine than baseline, or ≥ 4.0mg/dl) AKI, and newly-onset dialysis were recorded.
Other clinical outcomes evaluated in the present study included mechanical ventilator days, ICU stays, hospital stays, and all-cause mortality at day-28 and upon discharge.

Statistical analysis
Statistical analyses were performed using SPSS version 20.0 software (SPSS, Inc., Chicago, IL, USA).
Participants were categorized into Locolin and Colimycin groups, and analyzed accordingly. Continuous variables such as APACHE II and hospital stays between sub-groups were compared using the Mann-Whitney U test, and categorical variables were compared using Pearson's chi-square or Fisher's exact tests, as appropriate. We used mean imputation for missing data.
The occurrences of KDIGO stage 2, stage 3 AKI, and newly onset dialysis, were compared between patients treated with Locolin® and Colimycin®. Kaplan-Meier curves were constructed to evaluate difference in occurrence of AKI between subgroups of patients. Cox regression analysis was performed to identify independent variables associated with KDIGO 3 AKI. Treatment outcomes, including mechanical ventilator days, ICU stays, hospital stays, and all-cause mortality were also compared between patients with and without the occurrence of KDIGO stage 3 AKI. All tests were two-tailed and p-value < 0.05 was considered statistically signi cant.

Patient characteristics
During the study period, the medical records of 195 ICU-admitted patients who were administered with intravenous colistin were analyzed. Among these patients, 95 received Locolin® and 100 received Colimycin®. A ow diagram of the case numbers and reasons for exclusion is shown in Fig. 1 proportions of CRAB, CRE, and CRPA were 72.3%, 23.1%, and 4.6% respectively. The median daily colistin maintenance dose was 8 MIU (IQR 4-10), and the median treatment duration of colistin was 7 days (IQR 4-12). Twenty-three patients (11.8%) were administered with colistin at an inappropriate maintenance dosage. Comparatively, patients who were administered with Locolin® had higher serum albumin level (2.8 g/dL vs. 2.6 g/dL, p = 0.017) and were more likely to receive concomitant aminoglycoside treatment (15.8% vs. 7%, p = 0.052) and less likely to have colistin at inappropriate dosage (7.4% vs. 16%, p = 0.062) than those administered with Colimycin. Otherwise, the two groups of patients, had similar demographic characteristics, underlying comorbidities, and disease severities. There were no signi cant differences in the dosage and duration of colistin treatment between the two groups of patients. The rates of occurrence of various severities of AKI and newly initiated dialysis within 28 days after colistin administration are shown in Fig. 2. Overall, the rate of occurrence of KDIGO stage 1, stage 2, stage 3 AKI, and newly initiated dialysis was 49.7%, 39%, 21%, and 9.2%, respectively. Comparatively, patients who were administered with Colimycin® had a signi cantly lower occurrence rate of KDIGO stage 2 AKI (32% vs. 46.3%, p = 0.040) and KDIGO stage 3 AKI (13% vs. 29.5%, p = 0.005) than those administered with Locolin®. The rate of occurrence of newly initiated dialysis was comparable between the two groups of patients.
Kaplan-Meier analysis of the occurrence of KDIGO stage 2 and stage 3 AKI in both groups of patients is shown in Fig. 3. Patients who were administered with Colimycin® had a signi cantly lower occurrence of KDIGO 3 AKI than those administered with Locolin® (log rand p = 0.008

Discussion
This retrospective study involved critically ill patients who underwent intravenous colistin treatment for CR-GNB and evaluated the occurrence of AKI. During the colistin treatment period, the occurrence of KDIGO stage 1, stage 2, stage 3 AKI was 49.7%, 39%, and 21%, respectively. Meanwhile, 9.2% of the patients had newly initiated dialysis. Comparatively, patients who were administered with Colimycin® had a lower rate of occurrence of KDIGO stage 2 and stage 3 AKI than those administered with Locolin®.
In the multivariate analysis, we found that independent factors associated with KDIGO stage 3 AKI included the presence of septic shock and inappropriate colistin dosage. In contrast, Colimycin® use was an independent factor associated with a lower rate of occurrence of KDIGO stage 3 AKI. We also found that the occurrence of KDIGO stage 3 AKI during colistin treatment was associated with longer mechanical ventilator using days, but not related to increased all-cause mortality.
Nephrotoxicity and neurotoxicity are well-documented adverse reactions associated with the treatment with intravenous colistin. Nephrotoxicity in colistin is dose-dependent and usually reversible [16,18,22,23]. The nephrotoxicity of colistin is mainly related to its d-aminobutyric acid and fatty acid component. Similar to its bactericidal effects, colistin increases the membrane permeability of tubular epithelial cells, which in turn leads to cell swelling and lysis [10]. Colistin is a multicomponent lipopeptide that contains colistin A and colistin B, which differ in the fatty acid chain attached to the cyclic decapeptide moiety of the drug [24]. The proportion of colistin A and colistin B can have a large difference in commercial preparations of colistin [25]. Although there are comparable bactericidal effects between colistin A and colistin B, colistin A has been shown to have a higher nephrotoxic effect than colistin B in an animal model study [24]. The different compositions of colistin A and colistin B in various formulations of colistin might lead to different risks of colistin-induced nephrotoxicity. In the present study, we demonstrated a signi cant difference in the rate of occurrence of AKI between two different formulations of colistin. To our knowledge, this is the rst study to evaluate the nephrotoxicity between different formulations of colistin. Although the exact mechanisms remain uncertain, we speculate that the composition of colistin A and B in various colistin products could play a pivotal role. Clinicians should therefore be aware of the possible difference in the risks of nephrotoxicity in various formulations of colistin. Further studies are also warranted to verify our ndings.
The rate of occurrence of AKI reported in previous studies ranged from 27-51% and varied with the different study methodology and de nition of nephrotoxicity used [7,15,16,22,26,27]. In the present study, the overall rate of occurrence of KDIGO stage 2 and stage 3 AKI were 39% and 21%, which were consistent with previous reports. It is worth noting that all the enrolled patients were ICU-admitted cases and 90% of them had respiratory failure, which may further increase the rate of occurrence of nephrotoxicity during colistin treatment. We found that patients with KDIGO stage 3 AKI were more likely to have CR-GNB isolated from respiratory specimens and have a septic shock at the onset of colistin treatment. It may imply that patients with a respiratory infection and unstable hemodynamic status are more vulnerable to the nephrotoxicity of colistin. In addition, we found that more than 10% of our patients had inappropriate colistin dosage, which was rarely analyzed in previous studies, and it was an independent factor associated with AKI in the multivariate analysis. In contrast, both the accumulated dose and treatment duration of colistin were not signi cantly associated with AKI. Our ndings suggest that the nephrotoxicity of colistin is more likely correlated with the disease severity and inappropriate colistin dose adjustment according to renal function. Clinicians should therefore monitor renal function rigorously at the onset and during colistin treatment and adjust colistin dose carefully to keep the risk of nephrotoxicity as low as possible. Some strategies, including N-acetylcysteine (NAC) treatment and plasma volume expansion, have been proposed to prevent or ameliorate colistin-related AKI and deserve further veri cation [28,29].
We further evaluated the impact of the occurrence of colistin-related AKI on treatment outcomes, which have rarely been evaluated until now. We found that patients with colistin-induced AKI may have prolonged mechanical ventilator dependence; there were no differences in mortality and hospital stays between patients with and without colistin-related AKI. Our ndings were consistent with a previous study, which prospectively enrolled patients infected by extensively drug-resistant Acinetobacter baumannii and treated by colistin [30]. However, ventilator dependence and hospital stays were noted in that study. A study involving patients infected by drug-resistant Pseudomonas aeruginosa reported the presence of AKI as an independent factor associated with increased mortality [31]. Another study reported that patients who experienced AKI had higher mortality if kidney function failed to return to the baseline level [6]. Although the ndings remain controversial, we believe that close monitoring of renal function during colistin treatment and discontinuation of colistin early in patients with AKI is the best way to reduce the effect of AKI on treatment outcomes in these critically ill patients.
There are some limitations to this study. First, as a retrospective study, the demographic characteristics and disease severities were not equal between patients who were treated with Colimycin® and Locolin®.
Although we had used multivariate analysis to adjust the effects from clinical factors, our ndings still should be interpreted with caution. Second, all the enrolled patients had CR-GNB isolated from clinical specimens and some of them may have colonization, rather than true infection. However, its effect on our analysis was limited because the present study aimed to investigate colistin-induced nephrotoxicity, rather than treatment effectiveness. Third, exposure to concomitant nephrotoxins, including vancomycin, aminoglycosides, and contrast, were not rare in our patients. Therefore, the risk of colistin-induced nephrotoxicity could be overestimated. Finally, we enrolled critically ill patients with ICU admission and high APACHE II scores. Most of them had respiratory failure and nearly one-third of them received inotropic agents. Therefore, the ndings in our study may not be applicable in patients with low disease severities.

Conclusion
This retrospective study involved critically ill patients who were treated with intravenous colistin. We