Hemolysis attributed to high dose vitamin C: Two case reports

Abstract

BACKGROUND

High-dose vitamin C treatment (HVCT) can reduce the adverse effect of chemotherapy and enhance the effect of antitumor therapy, which has been considered one of the safest alternative treatments. However, the severity of its adverse effects may have been underestimated. The most serious adverse effect is hemolysis, which may result in acute kidney injury or death. Although glucose-6-phosphate dehydrogenase (G6PD) deficiency is considered to be the main cause, the probability and pathological mechanism are not completely understood, leading to a lack of effective and standardized treatment methods.

CASE SUMMARY

Two patients with colorectal cancer developed hemolytic anemia after using 1 g/kg HVCT. In contrast to previous cases, the lowest hemoglobin level in the two cases was < 50 g/L, which was lower than previously reported. This may be because Case 1 had chronic hepatitis B for many years, which caused abnormal liver reserve function, and Case 2 had grade II bone marrow suppression. Both patients improved and were discharged after blood replacement therapy. Our cases had the most severe degree of hemolysis but the best prognosis, suggesting that our treatment may be helpful for rescue of drug-induced hemolysis. This is the first review of the literature on hemolysis caused by HVCT, and we found that all patients with G6PD deficiency developed hemolysis after HVCT.

CONCLUSION

G6PD deficiency should be considered as a contraindication to HVCT, and it is not recommended for patients with bone marrow suppression, moderate-to-severe anemia, hematopoietic abnormalities, or abnormal liver and kidney function. Early blood purification and steroid therapy may avoid acute kidney injury or death caused by HVCT-related hemolytic anemia.

Key Words: Hemolysis, Vitamin C, Adverse effects, Acute kidney injury, Cancer, Case report

Core Tip: Two patients suffered from extremely severe hemolysis after high dose vitamin C treatment (HVCT), and both patients had glucose-6-phosphatase dehydrogenase (G6PD) deficiency. By reviewing the literature and summarizing the findings of available case reports, we conclude for the first time that patients who develop hemolytic anemia after HVCT may have G6PD deficiency, thus G6PD deficiency is an absolute contraindication for HVCT. Our treatment protected the patients from acute kidney injury in the context of extremely severe hemolysis. Our experience may be helpful for the treatment of similar cases.

INTRODUCTION

We report two cases of hemolytic anemia induced by high dose vitamin C treatment (HVCT) in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. This is the first report of this condition in patients with combined myelosuppression and liver dysfunction.

We conducted a literature search in PubMed, CNKI and OVID, and found a total of 7 articles, and the data in them were compared with our patients’ data. Combined with our case, the 9 patients with hemolysis after HVCT reported to date were summarized, and G6PD deficiency was found in all cases. We analyzed the reasons for this occurrence. We recommend strict control of the contraindications of HVCT, and HVCT should be prohibited for G6PD deficiency patients[1,2].

Our patients had more severe hemolysis after HVCT, which may be related to their comorbidities, but they recovered best after treatment. Therefore, we also compared and summarized the treatment regimens of these 9 patients, and our treatment experience may be helpful for the treatment of drug-induced hemolysis.

CASE PRESENTATION

Chief complaints

Both Case 1 and Case 2 presented with jaundice after HVCT used for anti-tumor therapy.

History of present illness

Case 1: A 48-year-old man from Yongfeng County, Jiangxi Province, China, received FOLFOX (oxaliplatin + calcium folinate +fluorouracil) chemotherapy + bevacizumab with concurrent HVCT (1 g/kg, 90 g, qd, ivgtt for 3 consecutive days) for colon cancer. After finishing treatment, the patient was admitted to the emergency department because of jaundice of the skin and sclera and fatigue for 1 d, which worsened for 3 h. On the day of presentation, the patient had severe fatigue, inability to walk, and nonexertional shortness of breath, with hematuria that rapidly progressed to soy-colored urine within 1 d (Figure 1A). He had no fever, chills, hematemesis, hematochezia, cough, chest pain or back pain.

Figure 1 Photographs of Case 1 during hemolysis. A: Hematuria; B: Plasma exchange line; C: Discarded patient plasma; D and E: Skin jaundice.

Case 2: The patient was a 54-year-old woman from Yongfeng County, Jiangxi Province, China. She was given single HVCT (10 g, qd, ivgtt, days 1 and 2) to establish tolerance, and the dose increased to 1 g/kg, 60 g, qd, ivgtt, on days 3 and 4, after no discomfort was observed. On the morning of day 5 of infusion, the patient complained of fatigue, dark yellow urine, skin jaundice and icteric sclera. She had no other complaints and her general condition was fair.

History of past illness

Case 1: The patient reported no jaundice, but he had stopped anti-tumor treatment because of abnormal liver function, and was treated with intermittent oral antiviral drug entecavir previously. In addition, the patient developed COVID-19 with tachycardia as the main manifestation 3 months previously, without dyspnea and standard antiviral treatment.

Case 2: She received anti-tumor treatment with one cycle of Xelox (oxaliplatin and capecitabine); nine cycles of FOLFOX + cetuximab; eight cycles of FOLFOXIRI (Fluorouracil + leucovorin + oxaliplatin + irinotecan)+ cetuximab; and four cycles of TAS-102 (trifluridine tipiracil tablets) + bevacizumab for rectal cancer with liver and lung metastases for > 1 year. At the time of the last chemotherapy in April 2023, no signs of tumor remission were observed, and grade II bone marrow suppression occurred.

Personal and family history

Case 1: His son had jaundice but the cause was not established. He had not previously received any food or medication that might have induced hemolysis, except HVCT.

Case 2: The patient and her next of kin had no history of jaundice or hemolysis, and she had not previously received any food or medication that might have induced hemolysis, except HVCT.

Physical examination

Case 1: The patient had obvious skin and scleral jaundice and poor consciousness. After emergency admission, the patient underwent electrocardiographic monitoring, which showed a heart rate 110 beats/min, blood pressure 115/80 mmHg, oxygen inhalation 10 L/min, and peripheral oxygen saturation 62% (Figure 2).

Figure 2 Blood smear. Red blood cells of different sizes with signs of rupture (arrow).

Case 2: Her vital signs were stable, but the skin and the sclera were jaundiced, with soy sauce colored urine.

Laboratory examinations

Case 1: Blood gas analysis showed partial pressure of oxygen 83 mmHg, partial pressure of carbon dioxide 42 mmHg, pH 7.43, and total hemoglobin 5.8 g/dL. Routine blood tests showed white blood cell count 23.65 × 10⁹/L, red blood cell count 1.75 × 10¹²/L, hemoglobin 62 g/L, total bilirubin 179.9 μmol/L, direct bilirubin 31.2 μmol/L, indirect bilirubin 148.7 μmol/L, and serum iron 42.5 μmol/L (Table 1). The direct antiglobulin test was negative.

Table 1 Hemoglobin changes during high-dose vitamin C and rescue treatment.

Items	Case 1				Case 2				Reference ranges	Units
	Before VC	After VC	Minimum hemoglobin	Post-treatment	Before VC	After VC	Minimum hemoglobin	Post-treatment
RBC	4.81	1.75	1.06	3.54	2.69	1.26	1.16	2.69	4.3-5.8	× 1012/L
HGB	152	63	37	112	96	44	41	86	130-175	g/L
HCT	0.444	0.173	0.105	0.335	0.285	0.144	0.129	0.274	0.40-0.50	L/L
MCV	92.3	98.9	99.1	94.7	105.9	114.8	110.9	101.9	82-100	fL
MCH	31.6	36	34.9	31.6	35.7	35.4	35.2	32	27-34	pg
MCHC	342	364	352	334	2.69	1.26	1.16	2.69	316-354	g/L

VC: Vitamin C; RBC: Red blood cell; HGB: Hemoglobin; HCT: Hematocrit; MCV: Mean corpuscular volume; MCH: Mean corpuscular hemoglobin; MCHC: Mean corpuscular hemoglobin concentration.

Case 2: The blood examination showed red blood cell count 1.26 × 10¹²/L, hemoglobin concentration 44 g/L, total bilirubin 148.8 μmol/L, direct bilirubin 48.4 μmol/L, and indirect bilirubin 100.4 μmol/L (Table 1).

FINAL DIAGNOSIS

Case 1: The patient was diagnosed with intravascular hemolysis, hemolytic anemia, hemolytic jaundice, methemoglobinemia, and malignancy of the sigmoid colon.

Case 2: The patient was diagnosed with intravascular hemolysis, hemolytic anemia, hemolytic jaundice, rectal malignancy, secondary liver malignancy, and secondary lung malignancy.

TREATMENT

Case 1: The patient was in critical condition and was transferred to intensive care unit for plasma exchange (Figure 1B–E). On the next day, the disease continued to progress and the patient became unconscious. He was treated with endotracheal intubation and mechanical ventilation, and then plasmapheresis, blood transfusion, hemofiltration and high-dose steroid pulse therapy. The endotracheal tube was extubated 33 h after intubation, and the patient was transferred back to the general ward 1 d later. He was discharged after a step-down treatment with high-dose steroids.

Case 2: HVCT was stopped immediately and the patient was transferred to intensive care unit. Electrocardiographic monitoring, oxygen inhalation, gammaglobulin 15 g, dexamethasone 10 g, and methylprednisolone 40 mg pulse therapy were administered for two consecutive days. A total of 7 U red blood cells were transfused and hemofiltration was performed once.

OUTCOME AND FOLLOW-UP

In both cases, hemoglobin levels gradually increased after treatment, their condition gradually stabilized, and they were discharged safely. No acute kidney injury occurred during the follow-up period.

DISCUSSION

After clarifying the two polarities of vitamin C dose effect, which is, reductive at low dose and oxidative high dose[3], HVCT has been shown to prolong survival and improve quality of life of patients with colorectal cancer harboring KRAS mutation[4], and has a good synergistic effect in some antitumor therapies[5]. This is closely related to the production of a large number of reactive oxygen species (ROS), such as hydrogen peroxide (H₂O₂) and superoxide anion by HVCT, which directly or indirectly kill tumor cells.

G6PD is a key enzyme in the pentose phosphate pathway (PPP), which is widely expressed in the body to assist glucose metabolism. Under normal conditions of G6PD, glucose can produce nicotinamide adenine dinucleotide phosphate (NADPH) (reduced coenzyme II) through the PPP to protect red blood cells from the threat of oxidized substances[1,2]. When G6PD gene mutation leads to the reduction of expression, the PPP metabolism of red blood cells is abnormal. At this time, when the body is exposed to food or drugs with strong oxidation, H₂O₂ produced by oxidation cannot be reduced to water in time, and excessive H₂O₂ can cause oxidative damage to hemoglobin and membrane proteins. Finally, oxidative damage and hemolysis of erythrocyte membrane are caused[6,7]. G6PD deficiency is the most common inherited enzyme deficiency disease in humans.

There are 400 million people around the world who carry mutations in the G6PD gene associated with enzyme deficiency. G1388A, G1376T and A95G are the most common mutations in the Chinese population. The degree of enzyme deficiency triggered by genetic defects varies. It is more common in male than female patients, with obvious racial and geographic differences, and it is common in Africa, Asia and the Middle East. It is often found in areas with high incidence of malaria, thalassemia and abnormal blood protein diseases[8-12]. The distribution in China shows a trend of high in the south and low in the north, especially in Guangdong, Hainan, Guangxi, Yunnan, Jiangxi, Guizhou, and Sichuan, with incidence rates of 4%–15% and up to 40% in some areas[13]. The occurrence of hemolytic anemia in patients with G6PD deficiency after intravenous HVCT may be related to the strong oxidation of HVCT, which produces a large amount of ROS, especially H₂O₂, inside and outside the cells, causing oxidative damage to the cells[5]. When G6PD is deficient, red blood cells fail to produce sufficient NADPH, a key molecule for restoring glutathione depletion due to vitamin C-induced oxidative stress, which leads to fragmentation of red blood cells and thus hemolysis[14] (Figure 3).

Figure 3 Mechanism of high dose vitamin C-induced hemolysis. In glucose-6-phosphatase dehydrogenase deficient erythrocytes, Pentose Phosphate Pathway metabolism is a disorder due to oxidative stress, and reduced substances in erythrocytes are depleted under oxidative damage, leading to rupture of a large number of erythrocytes; however, in normal or aged erythrocytes, there is also a chance of lipid peroxidation on the partial phospholipid bilayer on the surface of the erythrocyte membrane. Reactive oxygen species produced by high-dose vitamin C treatment induces lipid peroxidation, and ferroptosis of erythrocytes occurs, followed by rupture of a small number of erythrocytes. RBC: Red blood cells; GSH: Glutathione; GSSG: Oxidized glutathione; HVCT: High-dose vitamin C treatment; G6PD: Glucose-6-phosphatase dehydrogenase; NADPH: Nicotinamide adenine dinucleotide phosphate.

The key words "ascorbic acid", "vitamin C" and "hemolysis" were used for our literature search, and 7 cases of hemolysis caused by HVCT were screened out. Together with the 2 patients reported here, a total of 9 cases were recorded, all of whom had G6PD deficiency. Among such patients, the probability of acute kidney injury or methemoglobinemia at the time of hemolysis are 44% respectively, and the probability of death is 11% (Table 2).

Table 2 Cases of hemolysis attributed to intravenous high-dose vitamin C treatment.

Year	Nationality	Sex	G6PD deficiency	Dose, g/d	The lowest level of hemoglobin, g/L	Treatment	Complications and prognosis	Ref.
1975	United States	M	Yes	80	58	Blood transfusion + glucocorticoids (day 3) + hemodialysis (after anuria)	AKI, DIC, death	[19]
1993	United Kingdom	M	Yes	80	67	Drink plenty of water + folic acid orally	AKI	[20]
2014	Taiwan, China	M	Yes	75	54	Supportive treatment	None	[21]
2017	United States	M	Yes	60	59	Intravenous fluids + glucocorticoids	AKI, methemoglobinemia, acute oxalate nephropathy	[22]
2019	United States	M	Yes	65-200	-	-	None	[23]
2020	Hong Kong, China	F	Yes	30	63	Blood transfusion	Methemoglobinemia	[6]
2022	Taiwan, China	M	Yes	50	71	-	AKI	[24]
2023	Mainland China	M	Yes	90	37	Plasmapheresis + glucocorticoids	Methemoglobinemia	This text
2023	Mainland China	F	Yes	60	41	Hemofiltration + glucocorticoids	Methemoglobinemia	This text

All patients who developed hemolysis after HVCT had G6PD deficiency, and two patients reported here had lower hemoglobin levels than previous patients, and no serious complications occurred after early blood transfusion and hormone therapy. G6PD: Glucose 6 phosphate dehydrogenase; M: Male; F: Female; AKI: Acute kidney injury; DIC: Disseminated intravascular coagulation.

In clinical application of HVCT, it is necessary to check the G6PD related indicators to detect G6PD enzyme activity for patients in advance, for example, methemoglobin reduction test, fluorescent spot test, nitrotetrazolium disk method. G6PD deficiency should also be considered as one of the exclusion criteria in the clinical experimental study of HVCT[5]. During the infusion, patients should be paid attention to whether they have adverse reactions and hemoglobin content should be monitored.

HVCT is now mostly used as adjuvant therapy for antitumor therapy, such as combined chemotherapy, radiotherapy, immune or targeted therapy. When combined, the killing effect on normal cells increases[15], which may be one of the risk factors for hemolysis induced by HVCT. Therefore, special attention should be paid to the hematopoietic and metabolic functions of patients when combined. If the patient has bone marrow suppression and reticulocytosis, indicating reduced hematopoietic function and tolerance to injury, HVCT may be more likely to induce hemolytic reaction (Table 3). Therefore, when HVCT is used in combination, especially with antitumor drugs that may lead to autoimmune hemolytic anemia, HVCT may be more likely to induce a hemolytic reaction. When used with anti-programmed cell death protein-1 (e.g., nivolumab, pembrolizumab), cytotoxic T lymphocyte-associated protein 4 (e.g., ipilimumab), the dosage should be reduced as appropriate[16]. Patients with impaired liver function have a reduced ability to deal with schistocytes, and the degree of hemolysis is more severe than that of patients with normal liver function, and the condition is more difficult to control. For patients with hepatitis, cirrhosis, liver cancer and other diseases that affect liver metabolism, the indocyanine green (ICG) test should be supplemented under the condition of normal liver function to determine whether the liver reserve capacity can cope with hemolysis induced by HVCT.

Table 3 Reticulocyte analysis during hemolysis.

Items	Results		Units	Reference ranges
	Case 1	Case 2
RET1	1.8	6.9	%	0.59-2.07
RET2	32.2	207.8	´109/L	22.4-93.5
LFR	92.4	77.3	%	87.8-99.5
MFR	7.4	15.7	%	1.8-14.4
HFR	0.2	7	%	0-2.4
IRF	7.6	22.7	%	2.1-17.5

Low LFR indicates a lack of hematopoietic raw materials, inducing more severe hemolysis. Results in Case 2 with myelosupression was significantly different from Case 1 without myelosuppression in reticulocyte analysis, which shows that hematopoietic capacity is significantly reduced in patients with myelosuppression, which may be the cause of more severe hemolysis. RET¹: Reticulocyte percentage; RET²: Total reticulocytes; LFR: Low fluorescence intensity reticulocytes; MFR: Medium fluorescence intensity reticulocytes; HFR: High fluorescence intensity reticulocytes; IRF: Percentage of immature reticulocytes.

Since HVCT also have a certain destructive effect on about 5% G6PD normal erythrocytes[17], may be related to the ferroptosis of lipid peroxidation on the surface of red blood cells caused by HVCT. The manifestation of this effect in patient is an extremely slight decrease in hemoglobin. Therefore, in addition to G6PD detection, routine blood and biochemical tests, reticulocyte count and liver function storage test (ICG test) are necessary before administration of HVCT. At the same time, the above blood indicators should be measured on the second day before administration of HVCT to determine whether the patient can tolerate the treatment, and the infusion cannot be continued until there is no significant change, otherwise the infusion should be stopped permanently.

In patients with acute hemolysis, all oxidative drugs including HVCT should be stopped immediately. During the hemolytic phase, adequate hydration, correction of electrolyte imbalance, oral or intravenous supplementation of sodium bicarbonate, and alkaline urine should be maintained to prevent the deposition of hemoglobin in the renal tubules. Renal function should be closely monitored to prevent acute kidney injury and acute oxalate nephropathy. Mild anemia caused by HVCT (hemoglobin 60-90 g/L) generally recovers within 1 wk after withdrawal in patients without serious cardiopulmonary comorbidities[18]. The patients with severe anemia (hemoglobin < 60 g/L) or decreased hemoglobin, rapid increase in bilirubin, fatigue, shortness of breath, tachycardia and other symptoms of anemia, especially those with coronary heart disease, pulmonary heart disease or cerebrovascular disease, can be transfused with red blood cells to correct anemia. Patients with severe hemolysis should be treated with blood purification as soon as possible to prevent the occurrence of bilirubin encephalopathy or acute kidney injury or renal failure. Early blood purification and hormone pulse therapy may be the methods to avoid acute kidney injury caused by HVCT-related hemolysis.

CONCLUSION

HVCT is a mild antitumor therapy for most patients, except for those with G6PD deficiency and those with a risk of adverse effects such as renal insufficiency, recurrent urolithiasis, or hematopoietic abnormalities. Our cases were unique due to their complications and severity of the hemolysis. With the early blood purification and hormone therapy that we administered, these patients were spared acute kidney injury, renal failure, or death that occurred in the other patients, suggesting that these treatments may be effective in patients with similar conditions. In addition, we suggest that patients should be screened for enzyme activity to exclude G6PD deficiency before HVCT, and changes in hemoglobin content and hemolysis-related indicators should be detected in all patients during administration, especially after the first day of treatment. A standardized administration regimen is the best solution to avoid HVCT-related hemolysis.