Clinicopathologic consequences following discontinuation of rifaximin in patients with sickle cell disease
Authors: Seah H. Lim, Dibyendu Dutta
Journal: American Journal of Hematology (2020)

Patients with sickle cell disease (SCD) exhibit altered microbial compositions,1 evidence of enterocyte injury,2 and a breach in the intestinal barrier,2 leading to an increase in the translocation of bacteria/bacterial products across the intestine into the systemic circulation. The increased translocation of intestinal bacteria/bacterial products into the systemic circulation could explain the increased baseline neutrophil activation in SCD.2, 3 It may also provide the mechanistic basis for how intestinal microbiota regulates aged neutrophils in these patients.4 Both activated and aged neutrophils adhere to endothelium and form the nidus on which sickled erythrocytes may aggregate to induce vaso-occlusive crises (VOC). Based on the above considerations and the favorable clinical outcome to tumor necrosis factor α-induced fatal VOC in sickle cell mice administered with a combination of oral ampicillin, streptomycin, metronidazole, and vancomycin,4 we treated 12 SCD subjects with daily oral rifaximin, a minimally absorbed broad-spectrum antibiotic. This was done for 6 months in a Phase II study (Clinicaltrials.gov: NCT03719729) in our attempt to reduce the number of activated and aged neutrophils by reducing the intestinal microbial load. Rifaximin was chosen for its established safety profile for long-term use seen in patients with liver cirrhosis and resultant encephalopathy.5 We found that rifaximin not only attenuated enterocyte injury and decreased translocation of bacteria/bacterial products into the systemic circulating,6 it also reduced circulating aged neutrophils (CANs).6, 7 Correspondingly, these subjects experienced fewer episodes of painful VOC and days needing intravenous opioid, and a better quality of life during the six-month study period.8 To further examine the impact of rifaximin on SCD, we have investigated the clinicopathologic consequences in this cohort of patients 6 to 8 months after completing the Phase II study and stopping the rifaximin. Venous blood specimens were obtained from six of the 12 subjects (Two subjects switched their care to institutions closer to their homes, two relocated to other states, one was incarcerated, and one was lost for follow-up). Clinical data were obtained in person from 11 subjects, either at our sickle cell clinic or by telephone calls/emails. We first examined changes in enterocyte injury by comparing the serum intestinal fatty acid binding protein (iFABP) at the end of the six-month period of rifaximin with those 6 to 8 months after discontinuation of the antibiotic. In all six subjects, iFABP increased uniformly, by a median of nearly 50%, from 0.70 ng/mL to 1.09 ng/mL (P = .039) (Figure 1A). Increase in enterocyte injury was associated with breaches in the intestinal barrier in all six subjects. Serum lipopolysaccharides (LPS) rose correspondingly by a median of nearly 100%, from 0.51 μg/mL to 0.98 μg/mL (P = .024) (Figure 1B). Since pathogen-associated molecular patterns (PAMPs) interact with Myd88 and toll-like receptor (TLR) 2 and 4 that regulate CANs,4 changes in the serum LPS upon discontinuation of the rifaximin were associated with dramatic increases in CANs. This was from a median of 1.35% at the end of the Phase II study to 9.43% 6 to 8 months after stopping the antibiotic (P < .001) (Figure 1C). We next determined the clinical relevance of these intestinally-related pathologic sequelae by examining changes in the frequency of VOC and days needing intravenous opioid analgesia (IOA) during the six-month periods after stopping rifaximin. Among the 11 subjects treated and followed, the median number of VOC per 6 months before entering the study was 2 (range: 1-6.5). During the study period the median VOC per 6 months dropped to 1 (range: 0-2) (P = .007) but this reverted back to 2 VOC per 6 months (range: 0-6) (P = .03) during the 6 months after completing the study (Figure 1D). Five of the nine subjects who developed VOC did so within 2 months of stopping the rifaximin (three within 1 month), and two of the five subjects who did not develop any VOC during the study period remained VOC-free 6 months after stopping the rifaximin. Changes in the frequency of VOC were clinically translated into corresponding changes in the days needing IOA. The total number of days needing IOA among these 11 subjects 6 months prior to being started on rifaximin was 160 (median = 9; range: 4-33). The IOA days in 6 months dropped to 47 (median = 4; range: 0-16) (P < .001) during the study period but the reverted to 124 (median = 11; range: 0-30) (P = .04) during the 6 months after discontinuation of the rifaximin (Figure 1E). Combining the current data with the results that we previously reported on the laboratory changes in these subjects when they were treated with rifaximin,7 the current study supports further the notion that rifaximin mediates intestinal pathologic changes that favor those with SCD. However, there still remains many unanswered questions. It is unclear whether the reduction in enterocyte injury observed during rifaximin therapy is the result of decreased frequency of VOC that affects the splanchnic vasculature or the local anti-inflammatory effect of the antibiotic.9 Closer serial measurements of serum iFABP may provide the answer. Furthermore, although the clinical benefits of rifaximin in SCD is most likely related to changes in the levels of CANs during rifaximin, the mechanisms responsible for the drops in CANs during rifaximin treatment still need to be established. Although we previously found changes in the intestinal microbiome, primarily in a shift towards increased abundance in Bacteroides, in these subjects during rifaximin therapy,6 the changes were modest. Coupled with the rapidity of response to the rifaximin and the recurrence of painful VOC soon after discontinuation of the antibiotic in a number of the subjects, we speculate that rifaximin most probably exerts its beneficial effects in SCD by reducing the intestinal microbial load Hence, decreasing bacteria/bacterial products being translocated across the intestinal barrier to mediate aging and activation of neutrophils, rather than via altering the intestinal metabolomics. If the primary mechanism of action of rifaximin in reducing VOC in SCD is reducing the microbial load, it is possible that other antibiotics could confer similar benefits. However, since our study suggests that the antibiotic will need to be on a long-term basis, rifaximin appears to be the ideal antibiotic. It has an excellent safety profile for long-term use, at least in patients with liver cirrhosis,5 and has activity against Clostridium difficile, thereby avoiding the risks for Clostridium difficile infection associated with long-term antibiotic use. Finally, it is possible that effective antibiotic therapy for SCD is better achieved through a slow-release formulation of rifaximin that provides a sustained antimicrobial effect on the intestinal microbiota. This may also reduce the frequency of administration and improve patient compliance. These many questions suggest that in further development of rifaximin for SCD, a clinical study supported by some very carefully designed laboratory correlative investigations directed at answering some of these questions are warranted. None. Supported by a grant from Bausch Health Companies, Inc.