Diagnostics

Clonal hematopoiesis of indeterminate potential - CHIP in Hematology

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Classification
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Based on the current guidelines and the current state of research, different diagnostic recommendations arise for patients with clonal hematopoiesis of indeterminate potential. We have summarized the most important information on classification and diagnostic methods at MLL. In addition, we provide further links and literature on clonal hematopoiesis of undetermined potential.

CHIP in hematology: Classification

Clonal hematopoiesis of indeterminate potential (CHIP) is considered a myeloid precursor lesion according to the WHO 2022 classification.

CHIP definition (WHO 2022)

Detection of one or more somatic mutations with variant allele frequency (VAF) ≥2% (≥4% for X-linked gene mutations in males) in the DNA of blood or bone marrow cells involving selected genes (see Molecular Genetics, Table 2)
Absence of unexplained cytopenias
Absence of diagnostic criteria for defined myeloid neoplasms

CHIP is rare in individuals under 40 years of age and increases steadily after 65 years. In older individuals, it affects between 10-40%, with prevalence also depending on the sensitivity of the diagnostic sequencing method (Haferlach & Heuser 2022, WHO 2022).

Individuals with CHIP show an increased risk of developing hematologic neoplasia. The risk for other diseases, such as cardiovascular disease, is also increased in the presence of CHIP. You can learn more about CHIP in cardiology here.

VEXAS syndrome was only described for the first time a few years ago (Beck et al. 2020), which was classified by the WHO as a CH-associated disease in the clonal hematopoiesis section (WHO 2022). Further information on the diagnosis of VEXAS syndrome can be found here.

Differentiation of CHIP from CCUS and MDS

When clonality of hematopoiesis is detected, it is necessary to differentiate CHIP from CCUS (clonal cytopenia of undetermined significance) and full-blown myeloid neoplasms, especially myelodysplastic neoplasms (MDS).

If clonal hematopoiesis is accompanied by cytopenia of unclear cause, this is referred to as CCUS. If cytomorphological signs of dysplasia are also present, the diagnostic criteria of MDS are fulfilled.

Table 1: Differentiation of CHIP and CCUS from MDS (Hoermann 2022 [2])

	CHIP	CCUS	Low risk MDS	High risk MDS
Clonality	+	+	+	+
Dysplasia	-	-	+	+
Cytopenia	-	+	+	+
BM Blasts	<5%	<5%	<5%	5-20%
Cytogenetic aberrations	+/-	+/-	+	++
Molecular aberrations	+	+	++	+++
Risk of progression	+	++	++	+++

CHIP in hematology: Diagnostic methods and their relevance

If clonal hematopoiesis is present on the basis of the (molecular) genetic findings, a blood count analysis including cytomorphological examination should be performed to differentiate CHIP (absence of dysplasia and cytopenia) from CCUS (cytopenia but absence of dysplasia) or a hematologic neoplasia. If hematologic neoplasia is suspected, further diagnostic testing - usually bone marrow aspiration - is indicated.

Research data from microarray and WGS studies have demonstrated that clonal hematopoiesis can also manifest as copy number alterations or copy number-neutral loss of heterozygosity (CN-LOH) in individuals without hematologic neoplasia. Among the most common alterations in this regard are copy number-neutral loss of heterozygosity in regions 9p and 4q, trisomies of chromosomes 8, 12, and 15, and 20q deletions (Jacobs et al. 2012, Saiki et al. 2021, Haferlach & Heuser 2022). In practice, CHIP usually represents an incidental finding in individuals without blood count abnormalities based on molecular genetic studies of peripheral blood. Chromosome analysis is not indicated in CHIP diagnosis. However, if unexplained cytopenia is present, chromosome analysis from bone marrow should be performed. See also CCUS (clonal cytopenia of undetermined significance).

CHIP diagnosis is typically made by DNA sequencing (WHO 2022). Since an allele frequency of mutations of ≥2% is defining, the selected diagnostic method must still be able to reliably detect a VAF of 2% (Haferlach & Heuser 2022). The WHO 2022 definition of CHIP includes the following mutations:

Table 2: Driver mutations of clonal hematopoiesis (after WHO 2022)

Common / clinically significant genes	DNMT3A, TET2, ASXL1, JAK2, TP53, SF3B1, PPM1D, SRSF2, IDH1, IDH2, U2AF1, KRAS, NRAS, CTCF, CBL, GNB1, BRCC3, PTPN11, GNAS, BCOR, BCORL1
Additional genes	BRAF, CALR, CEBPA, CRBBP, CSF1R, CSF3R, CUX1, ETV6, EZH2, GATA2, JAK3, KDM6A, KIT, KMT2A, MPL, MYD88, NOTCH1, PHF6, PIGA, PRPF40B, PTEN, RAD21, RUNX1, SETBP1, SF1, SF3A1, SMC1A, SMC3, STAG2, STAT3, U2AF2, WT1, ZRSR2

CHIP can in principle progress to MDS or other hematologic neoplasms, but the risk of progression is low. If there is additional cytopenia (CCUS) in the presence of clonal hematopoiesis, this is associated with a significantly higher risk of hematologic progression than CHIP (Malcovati et al. 2017). The distinction between CHIP, CCUS, and MDS cannot be made on the basis of molecular genetic testing alone, but is currently based on differences in the presence of cytopenias and diagnostic morphologic criteria for MDS. A smooth transition between CHIP, CCUS and MDS is thought likely. An important indication of this is the increase in genetic complexity (see Table 3) in terms of the number of mutations as well as clone size (allele frequency) (Cargo et al. 2015, Bejar 2017, Malcovati et al. 2017, Bewersdorf et al. 2019).

Table 3: Comparison of molecular genetic characteristics between CHIP, CCUS and MDS (selection according to Bejar 2017)

	CHIP	CCUS (at diagnosis)	MDS (all risk groups)
Frequently mutated genes	DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53	TET2, DNMT3A, ASXL1, SRSF2, TP53	SF3B1, TET2, ASXL1, SRSF2, DNMT3A
Average number of mutations	1	1,6	2,6
Typical allele frequency	9-12%	30-40%	30-50%

Overall, MDS are molecularly more complex than CHIP: two or more mutations are usually present and the mutational burden is usually above 10% (Haferlach et al. 2014, Malcovati et al. 2017, Sperling et al. 2017). Since progression results in an accumulation of multiple genetic alterations, it is recommended that progression testing be performed in questionable cases.

Table 4 shows an overview of known mutations in MDS and CHIP. The presence of multiple mutations (e.g. SF3B1, SRSF2) increases the likelihood of an MDS diagnosis or the likelihood of developing MDS.

Table 4: Somatic gene mutations in MDS and CHIP (Valent et al. 2017)

Gene	Chromosome localization	Frequency (% of all patients)
Gene	Chromosome localization	MDS	CHIP
NRAS	1p13.2	1-10	<1
DNMT3A	2p23	>10	>10
SF3B1	2q33.1	>10	1-10
IDH1	2q33.3	1-10	<1
GATA2	3q21.3	<1	<1
KIT	4q11-12	1-10	<1
TET2	4q24	>10	>10
NPM1	5q35.1	<1	<1
EZH2	7q35-36	1-10	<1
JAK2	9p24	1-10	>10
CBL	11q23.3	1-10	1-10
KRAS	12p12-11	<1	<1
ETV6	12p13	<1	<1
FLT3	13q12	<1	<1
IDH2	15q26.1	<1	<1
TP53	17p13.1	1-10	1-10
PRPF8	17p13.3	<1	<1
SRSF2	17q25.1	>10	1-10
CEBPA	19q13.1	<1	<1
ASXL1	20q11	>10	>10
U2AF1	21q22.31	1-10	<1
RUNX1	21q22.12	1-10	<1
BCOR	Xp11.4	<1	<1
ZRSR2	Xp22.1	1-10	<1
STAG2	Xq25	1-10	<1

CHIP in hematology: Prognosis

The transformation rate in the presence of a CHIP is 0.17-0.22% per year (Haferlach & Heuser 2022). Large clones or the presence of multiple mutations increase the risk of progression to myeloid neoplasia. In particular, mutations in TP53, U2AF1, SRSF2, IDH1, IDH2, SF3B1, and ASXL1 increase the risk of progression. In addition, the simultaneous presence of mosaic clonal alterations (detected by microarray or WGS studies) is considered an independent risk factor for the development of leukemia (Haferlach & Heuser 2022, Hoermann 2022 [1], Hoermann 2022 [2], WHO 2022). In the age group of ≥80 years an association of CHIP with a lower survival probability has been described especially in the presence of ≥2 mutations. In this age group, in addition to specific mutation patterns, a mutation allele frequency of ≥9.6% has been identified as a risk factor for the development of myeloid neoplasia (Rossi et al. 2021).

The Clonal Hematopoiesis Risk Score (CHRS) was established in 2023 as a prognostic risk score for the development of a haematological neoplasia. This incorporates various variables that lead to classification into low, intermediate and high risk groups. The sole presence of a DNMT3A mutation is considered a prognostically favorable factor in CHRS. In contrast, the following parameters are considered to be prognostically unfavorable factors (Weeks et al. 2023):

defined high-risk mutation (SRSF2, SF3B1, ZRSR2, IDH1, IDH2, FLT3, RUNX1, JAK2 and TP53)
Detection of multiple mutations
Clone size of at least 20% variant allele frequency (VAF)
Red cell distribution width (RDW) of ≥15%
Mean corpuscular volume (MCV) of ≥100 fl
Presence of cytopenia (CCUS vs. CHIP)
Age ≥65 years

With "MN-predict", another model for risk stratification was developed in 2023. Using a web application, annual probabilities (up to 15 years) of transformation into a myeloid neoplasia can be provided on the basis of simple blood tests and genetic data (Gu et al. 2023).

Based on real-world data, the "Clonal Cytopenia Risk Score" (CCRS) was also recently developed, a risk score specifically tailored to patients with CCUS (Xie et al. 2024). This model integrates only three parameters:

Platelet count of <100×10⁹/L,

Presence of ≥2 mutations

Presence of a splicing mutation

Patients can thus be divided into groups with low (score <2.5), medium (score 2.5-<5) and high risk (score ≥5). The model is therefore easy to apply in everyday clinical practice.

CHIP is associated with an increased risk of all-cause mortality, atherosclerotic cardiovascular disease, and autoimmune disease (WHO 2022). CHIP is also considered a risk factor for advanced chronic obstructive pulmonary disease (COPD) and the development of various other non-hematologic diseases (Haferlach & Heuser 2022). A high number of genetic alterations significantly increases the likelihood of dying from hematologic neoplasia or cardiovascular disease (Saiki et al. 2021). See also CHIP in Cardiology and German CHIP Registry.

CHIP as a risk factor for the development of therapy-associated myeloid neoplasms

CHIP clones can gain a selective survival advantage and expand under hematologic stress, which can result from cytotoxic therapy, radiation, or stem cell transplantation, among others. In particular, TP53 and PPM1D mutant clones have been associated with the development of therapy-associated neoplasms following cytotoxic therapy (Wong et al. 2015, Coombs et al. 2017, Hsu et al. 2018, Kahn et al. 2018, Wong et al. 2018, Ortmann et al. 2019, Hoermann 2022 [1]).

CHIP in the context of autologous and allogeneic stem cell transplantation

A few studies have described an increased risk of developing therapy-associated myeloid neoplasia after autologous stem cell transplantation (Gibson et al. 2017). However, according to current knowledge, there is insufficient evidence to use CHIP in the context of autologous stem cell transplantation to make treatment decisions.

In allogeneic stem cell transplantation, donor CHIP status did not affect overall survival of recipients in a first study. In the context of CHIP-positive stem cell donors, an increased incidence of chronic graft-versus-host disease was observed, which was associated with a lower rate of relapse or progression (Frick et al. 2019). However, there are case reports of the occurrence of donor cell leukemia after allogeneic stem cell transplantation in the context of CHIP-positive older donors (Gondek et al. 2016, Frick et al. 2019).

CHIP in hematology: Recommendation

Also, in view of the lack of therapeutic intervention options, screening for the presence of CHIP in individuals with normal blood counts is not currently recommended from a hematologic perspective. Often, the detection of clonal hematopoiesis is an incidental finding. If the blood count is normal, patients with CHIP should have a blood count including differential blood count at regular intervals (initially after 3 months, later every 12 months) to detect possible progression. If the patient has unexplained peripheral cytopenia (CCUS), a bone marrow aspiration and differential blood count is recommended initially at 1, 2, and 3 months and subsequently every 3 months (Heuser et al. 2016). A recommendation on the diagnostic algorithm for CHIP is also given by Haferlach & Heuser 2022:

Fig. 1: Diagnostic algorithm for detection of CHIP, CCUS or unexplained blood count changes (modeled after Haferlach & Heuser 2022).

Beck DB et al. Somatic Mutations in UBA1 and Severe Adult-Onset Autoinflammatory Disease. N Engl J Med 2020;383(27):2628-2638.

Bejar R. Implications of molecular genetic diversity in myelodysplastic syndromes. Curr Opin Hematol 2017 [2];24(2):73-78.

Bewersdorf JP et al. From clonal hematopoiesis to myeloid leukemia and what happens in between: Will improved understanding lead to new therapeutic and preventive opportunities? Blood Rev 2019;37(100587.

Cargo CA et al. Targeted sequencing identifies patients with preclinical MDS at high risk of disease progression. Blood 2015;126(21):2362-2365.

Coombs CC et al. Therapy-Related Clonal Hematopoiesis in Patients with Non-hematologic Cancers Is Common and Associated with Adverse Clinical Outcomes. Cell Stem Cell 2017;21(3):374-382.e374.

Desai P et al. Somatic mutations precede acute myeloid leukemia years before diagnosis. Nat Med 2018;24(7):1015-1023.

Frick M et al. Role of Donor Clonal Hematopoiesis in Allogeneic Hematopoietic Stem-Cell Transplantation. J Clin Oncol 2019;37(5):375-385.

Gibson CJ et al. Clonal Hematopoiesis Associated With Adverse Outcomes After Autologous Stem-Cell Transplantation for Lymphoma. J Clin Oncol 2017;35(14):1598-1605.

Gondek LP et al. Donor cell leukemia arising from clonal hematopoiesis after bone marrow transplantation. Leukemia 2016;30(9):1916-1920.

Gu M et al. Multiparameter prediction of myeloid neoplasia risk. Nat Genet 2023;55(9):1523-1530.

Haferlach C, Heuser M. [Diagnostics in unclear cytopenia-How and when do we screen for clonal hematopoiesis?]. Inn Med (Heidelb) 2022.

Haferlach T et al. Landscape of Genetic Lesions in 944 Patients with Myelodysplastic Syndromes. Leukemia 2014;28(2):241-247.

Heuser M et al. Clonal Hematopoiesis of Indeterminate Potential. Dtsch Arztebl Int 2016;113(18):317-322.

Hoermann G. Clinical Significance of Clonal Hematopoiesis of Indeterminate Potential in Hematology and Cardiovascular Disease. Diagnostics (Basel) 2022 [1];12(7):

Hoermann G. Clonal hematopoiesis of indeterminate potential: clinical relevance of an incidental finding in liquid profiling. Journal of Laboratory Medicine 2022 [2];46(4):301-310.

Hsu JI et al. PPM1D Mutations Drive Clonal Hematopoiesis in Response to Cytotoxic Chemotherapy. Cell Stem Cell 2018;23(5):700-713.

Jacobs KB et al. Detectable clonal mosaicism and its relationship to aging and cancer. Nat. Genet 2012;44(6):651-658.

Kahn JD et al. PPM1D-truncating mutations confer resistance to chemotherapy and sensitivity to PPM1D inhibition in hematopoietic cells. Blood 2018;132(11):1095-1105.

Malcovati L et al. Clinical significance of somatic mutation in unexplained blood cytopenia. Blood 2017;129(25):3371-3378.

Ortmann CA et al. Functional Dominance of CHIP-Mutated Hematopoietic Stem Cells in Patients Undergoing Autologous Transplantation. Cell Rep 2019;27(7):2022-2028.e2023.

Rossi M et al. Clinical relevance of clonal hematopoiesis in persons aged ≥80 years. Blood 2021;138(21):2093-2105.

Saiki R et al. Combined landscape of single-nucleotide variants and copy number alterations in clonal hematopoiesis. Nat Med 2021;27(7):1239-1249.

Sperling AS et al. The genetics of myelodysplastic syndrome: from clonal haematopoiesis to secondary leukaemia. Nat Rev Cancer 2017;17(1):5-19.

Valent P et al. Proposed minimal diagnostic criteria for myelodysplastic syndromes (MDS) and potential pre-MDS conditions. Oncotarget 2017;8(43):73483-73500.

Weeks LD et al. Prediction of risk for myeloid malignancy in clonal hematopoiesis. NEJM Evid 2023;2(5).

WHO Classification of Tumours Editorial Board. Haematolymphoid tumours [Internet]. Lyon (France): International Agency for Research on Cancer; 2022. (WHO classification of tumours series, 5th ed.; vol. 11). Available from: https://tumourclassification.iarc.who.int/chapters/63.

Wong TN et al. Cellular stressors contribute to the expansion of hematopoietic clones of varying leukemic potential. Nat Commun 2018;9(1):455.

Wong TN et al. Role of TP53 mutations in the origin and evolution of therapy-related acute myeloid leukaemia. Nature 2015;518(7540):552-555.

Xie Z et al. Risk Prediction for Clonal Cytopenia: Multicenter Real-World Evidence. Blood 2024;2024024756.

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Clonal hematopoiesis of indeterminate potential - CHIP in Hematology

CHIP in hematology: Classification

CHIP definition (WHO 2022)

Differentiation of CHIP from CCUS and MDS

CHIP in hematology: Diagnostic methods and their relevance

Cytomorphology

Chromosome analysis

Molecular genetics

Table 2: Driver mutations of clonal hematopoiesis (after WHO 2022)

Table 3: Comparison of molecular genetic characteristics between CHIP, CCUS and MDS (selection according to Bejar 2017)

Table 4: Somatic gene mutations in MDS and CHIP (Valent et al. 2017)

CHIP in hematology: Prognosis

CHIP as a risk factor for the development of therapy-associated myeloid neoplasms

CHIP in the context of autologous and allogeneic stem cell transplantation

CHIP in hematology: Recommendation

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