Leukemia, a group of cancers that affect the blood and bone marrow, is characterized by the uncontrolled proliferation of abnormal white blood cells (1). These malignant cells can impede the production of normal blood cells, leading to symptoms such as anemia, infection, and bleeding. Leukemia is classified into several types, including acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML), each with distinct clinical features, genetic abnormalities, and treatment approaches (2). Despite significant progress in medical research, the exact causes of leukemia are still not fully understood. It is generally believed that a combination of genetic factors, environmental influences, and personal lifestyle choices may play a role in its development.

Among the environmental factors, chemical exposures have been extensively studied due to their potential to induce genetic mutations and disrupt cellular processes. Chloroform (CHCl₃), a colorless (3), sweet-smelling organic compound, is one such chemical that has raised concerns (4). Historically used as an anesthetic, chloroform is now primarily utilized in the production of refrigerants, as a solvent in the pharmaceutical industry, and as a byproduct in water chlorination processes (5, 6). Consequently, human exposure to chloroform can occur in various settings, including industrial workplaces, healthcare facilities, and even homes through the consumption of chlorinated drinking water (7).

The potential health risks associated with chloroform exposure have been a topic of ongoing research and debate. Chloroform is metabolized in the liver to produce phosgene (8), a highly reactive and toxic metabolite that can induce cellular damage and DNA mutations (9). Animal studies have demonstrated that chronic exposure to high levels of chloroform can lead to liver and kidney tumors, prompting the International Agency for Research on Cancer (IARC) to classify chloroform as a Group 2B carcinogen (10), indicating it is possibly carcinogenic to humans. However, the evidence from human studies remains inconclusive, particularly regarding the risk of leukemia.

Epidemiological studies investigating the association between chloroform exposure and leukemia have yielded mixed results. Some studies have reported an increased risk of leukemia among individuals with high levels of chloroform exposure, particularly in occupational settings. For instance, workers in industries involving the use of chloroform or individuals living near industrial sites may be at higher risk (11). However, other studies have found no significant association, highlighting the need for a comprehensive evaluation of the available evidence.

The inconsistencies in the findings may be attributed to several factors, including variations in study design, differences in exposure assessment methods, and potential confounding variables. Additionally, the relatively low incidence of leukemia compared to other cancers can make it challenging to detect statistically significant associations. To address these challenges and provide a clearer understanding of the potential link between chloroform exposure and leukemia risk, a systematic review and meta-analysis is warranted.

Chloroform exposure can occur through various environmental routes, with air and drinking water being the most common sources (12). While air and drinking water are distinct exposure pathways, both represent significant routes through which humans can be exposed to chloroform, especially in industrial or residential settings where chlorinated water is consumed, and air may be contaminated with chloroform from indoor sources, such as from water disinfection byproducts (11).

The rationale for combining air and drinking water exposure pathways in this analysis lies in the fact that both pathways may contribute similarly to an individual’s overall chloroform burden. Individuals living in areas with significant air pollution from industrial sources or chlorinated water supplies may have overlapping or cumulative exposure to chloroform through both routes. Combining these pathways allows for a more holistic evaluation of chloroform’s potential role in leukemia risk. Additionally, different studies have measured exposure via these two pathways independently, but the combined analysis helps capture the overall environmental impact on health.

Despite their distinct origins, air and drinking water exposures share similarities in their underlying mechanisms, such as the potential for chloroform to enter the body and undergo metabolism to form toxic metabolites like phosgene, which are associated with genotoxic effects (13). Therefore, it is important to consider both exposure pathways to better understand the full scope of risk.

This article aims to systematically review the existing literature on the relationship between chloroform exposure and leukemia. By conducting a meta-analysis of the available studies, we aim to quantify the overall risk and identify potential sources of heterogeneity. Our objectives are to: (1) assess the strength and consistency of the evidence linking chloroform exposure to leukemia, (2) explore variations in risk estimates across different populations and exposure levels, and (3) identify gaps in the current research to inform future studies.

Understanding the potential role of chloroform in leukemia development is crucial for public health. If a significant association is confirmed, it would underscore the need for stricter regulatory measures to limit chloroform exposure, particularly in vulnerable populations. Moreover, it could lead to targeted prevention strategies and enhanced screening programs for individuals at higher risk. Through this comprehensive analysis, we aim to contribute to the growing body of knowledge on environmental carcinogens and their impact on leukemia, ultimately guiding efforts to reduce the burden of this devastating disease.

This systematic review and meta-analysis provides significant insights into the potential relationship between chloroform exposure and leukemia risk. Our pooled analysis revealed no statistically significant overall association between chloroform exposure and leukemia (pooled OR: 0.75, 95% CI: 0.25–2.27). This suggests that, at the exposure levels studied, chloroform may not pose a substantial risk for leukemia. However, the observed heterogeneity and subgroup differences suggest more complex interactions that warrant further investigation.

A key finding in our analysis was the high heterogeneity observed (I² = 95%), indicating considerable variability across the included studies. This heterogeneity likely arises from differences in study design, population characteristics, exposure assessment methods, and the confounding variables controlled for in individual studies. For instance, exposure assessment methods ranged from residential history and questionnaires to air monitoring, potentially introducing biases. Future studies should employ standardized methods for exposure assessment to improve comparability and enhance evidence quality.

Given the substantial heterogeneity, a subgroup analysis based on leukemia subtypes was conducted, revealing notable differences in associations. For Acute Myeloid Leukemia (AML), a significant association was found between chloroform exposure and increased AML risk (pooled OR: 1.98, 95% CI: 1.31–3.00), aligning with previous evidence of chloroform’s potential carcinogenic effects. Conversely, a significant inverse association was observed for Chronic Lymphocytic Leukemia (CLL) (pooled OR: 0.15, 95% CI: 0.09–0.25), a finding requiring further exploration to determine if it is biologically meaningful or methodological in origin. No significant association was found between chloroform exposure and Acute Lymphoblastic Leukemia (ALL) risk (pooled OR: 0.95, 95% CI: 0.63–1.27). The significant variability in effect sizes across leukemia subtypes [Qb(2) = 31.91, p = 0.00] emphasizes the necessity of subtype-specific analyses in future research. The overall lack of a significant association may obscure subtype-specific risks or protective effects.

The exposure window for childhood leukemia is crucial in interpreting these findings. Childhood leukemia risks may differ depending on whether exposures occur prenatally or postnatally, as both periods could influence leukemia development through distinct biological mechanisms. In the studies included in this meta-analysis, exposures during prenatal (maternal exposure to chloroform) and postnatal periods (early childhood environmental exposure) were both assessed. The precise timing and duration of exposure varied across studies, which likely contributed to heterogeneity in outcomes. Future studies should explicitly differentiate between prenatal and postnatal exposure windows to clarify critical periods of vulnerability and enhance our understanding of chloroform’s potential role in leukemogenesis.

The biological plausibility of chloroform’s role in leukemia involves its metabolism to toxic metabolites, such as phosgene, inducing oxidative stress and DNA damage. The observed increased AML risk aligns well with these genotoxic mechanisms. Future research should investigate molecular pathways further to clarify causality and mechanisms specific to leukemia subtypes.

While our study did not find an overall significant risk, subgroup analyses suggest potential subtype-specific susceptibility, particularly for AML. The inverse relationship with CLL also warrants exploration. Future studies should focus on specific leukemia subtypes, diverse populations, varied exposure levels, and detailed temporal exposure windows. Such targeted research can provide a clearer understanding of chloroform as a potential environmental carcinogen, aiding in developing public health policies and preventive strategies.

Several limitations should be noted. Variability in methods and populations limits generalizability. Diverse exposure assessment methods introduce potential bias. Limited cases for certain subtypes reduce statistical power. Additionally, the small number of studies (four) limits robust detection of publication bias.

This systematic review and meta-analysis found no overall significant association between chloroform exposure and leukemia risk at studied levels. However, significant subtype-specific associations highlight the complexity of leukemia etiology. Future research should emphasize standardized exposure assessments, clear delineation of prenatal and postnatal exposure windows, and detailed analysis of leukemia subtypes. Such efforts will enhance understanding and inform preventive measures for populations potentially at risk.