Keywords
hematological - malignancies - acute leukemia - myeloid - lymphoid
Introduction
Leukemia is a group of hematological malignancy in which there is an uncontrolled,
unregulated, and rapid proliferation of leukemic cells resulting in replacement of
normal hematopoietic cells by abnormal proliferating cells in bone marrow and spilling
over peripheral blood as well.[1]
It accounts a major proportion of hematopoietic neoplasms that are diagnosed worldwide.
According to GLOBOCAN 2020, the worldwide estimates of cancer incidence and mortality
for developing countries in 2020 revealed 269,503 and 205,016 new cases of leukemia
in males and females, respectively.[2] Of these, the estimated deaths were 176,000 and 132,000 in males and females, respectively.
Approximately 23,660 deaths (13,900 males and 9,760 females) in the United States
are expected to be attributed to leukemia in 2021.[3] Hence, timely diagnosis and initiation of treatment in initial stages is required
for complete remission.
Leukemias are classified into two broad categories: acute and chronic. Acute leukemia
is further subdivided into myeloid and lymphoid. In adults, acute myeloid leukemia
(AML) accounts for 80 to 90% of cases of acute leukemias.[4] Acute lymphoblastic leukemia (ALL), common in childhood, comprises 12% of all leukemias.[5] Incidence rises again in the sixth decade, but this peak age is not commonly seen
in developing countries.
Immunophenotyping by flow cytometry (FCM) is an essential aid for accurately diagnosing
and prognosticating acute leukemias. It serves various purposes, mainly identifying
and differentiating neoplastic population from normal population, defining number
of neoplastic cells with their phenotypes, typing and subtyping of leukemias, and
minimal residual disease analysis.[6]
To the best of our knowledge, there is a lack of literature regarding distribution
and pattern of acute leukemia cases in Bihar. Population-based cancer registry (PBCR)
is still lacking in major eastern states of India including most populous Uttar Pradesh
and Bihar which can keep a track and notify regarding the prevalence and incidence
of various types of cancer including leukemias.[7] Because of population growth, changing dietary habits, increasing tobacco consumption,
and exposure to arsenic in the Gangetic belt, Bihar is anticipated to bear greater
cancer burden, including hematolymphoid malignancies.
Objective
-
To estimate the prevalence and clinico-hematological profile of acute leukemia cases.
-
To study immunophenotypic profile in patients with acute leukemia.
Materials and Methods
A retrospective study was conducted in the Department of Hematology, Indira Gandhi
Institute of Medical Sciences, Patna, over a period of 2 years from July 2019 to June
2021 and it was approved by the Institutional Ethical and Scientific committee. A
total of 176 cases with relevant clinical features and hematological findings were
involved in the study. A predesigned proforma was made in which patient characteristics
were entered. Medical records were studied and data were collected.
Inclusion and Exclusion Criteria
All patients presenting with relevant clinical features and hematological findings
suggestive of acute leukemia and those with myelodysplastic syndrome transformed to
acute leukemia were included in the study, whereas all those patients who were on
prior chemotherapy or radiotherapy were excluded from the study.
The required quantity of venous blood will be collected in EDTA vials. The collected
blood will be analyzed by using fully automated analyzer (SIEMENS ADVIA 2120i) having
six parts from which large unstained cell count will be estimated and subsequently
peripheral blood smears will be prepared in such cases on glass slides and stained
with Leishman's stain. Special stains like MPO and PAS will be done in all cases.
Bone marrow aspiration will be done and slides stained with Leishman's stain will
be evaluated and its findings will be correlated with immunophenotyping (done by BD
FACS Canto, based on the principle of hydrodynamic focusing) and cytogenetics for
further confirmation. Data were recorded and analyzed using Statistical Package for
Social Sciences version 25 (SPSS version 25). Chi-square test and Fisher's exact test
were applied to test the association of qualitative data and Student's t-test was applied to test the association of quantitative data. Results were recorded
as frequencies, means ± standard deviations, and p-values. For all purposes, a p-value of less than 0.05 (95% confidence level) was considered as the criteria of
significance.
Results
Out of 176 cases, 93 cases (52.8%) were of AML and 60 cases (34.1%) were of ALL. The
remaining 23 cases (13.1%) could not be classified into any type of acute leukemia
morphologically on bone marrow aspirate. FCM correlation was available in 150 cases.
Due to financial constraint, immunophenotyping was not available for the remaining
26 cases. On FCM out of 150 cases, 92 cases turned out to be AML and 56 cases were
ALL. Two cases were diagnosed as mixed phenotypic acute leukemia. There was concordance
between morphological and immunophenotypic diagnosis in 95% of cases.
Overall 62.5% (n = 110) of patients suffering from acute leukemias were males, while 37.5% (n = 66) were females. ALL in males accounted for 61.7% of cases, while in females,
it accounted for 38.3% of cases. 37.6% (n = 35) of AML cases were females, while 62.4% (n = 58) were male patients ([Table 1]).
Table 1
Sociodemographic characteristics of acute leukemia patients
|
Male
|
Female
|
Total
|
|
Age
|
0–10 y
|
16 (57.1%)
|
12 (42.8%)
|
28 (15.9%)
|
|
11–20 y
|
22 (61.1%)
|
14 (38.8%)
|
36 (20.4%)
|
|
21–30 y
|
14 (63.6%)
|
8 (36.3%)
|
22 (12.5%)
|
|
31–40 y
|
28 (65.1%)
|
15 (34.8%)
|
43 (24.4%)
|
|
41–50 y
|
15 (57.6%)
|
11 (42.3%)
|
26 (14.7%)
|
|
51–60 y
|
9 (75.0%)
|
3 (25.0%)
|
12 (6.8%)
|
|
61–70 y
|
4 (57.1%)
|
3 (42.8%)
|
7 (3.9%)
|
|
71–80 y
|
2 (100.0%)
|
0 (0.0%)
|
2 (1.1%)
|
|
Residency
|
Urban
|
41 (65.0%)
|
22 (34.9%)
|
63 (35.7%)
|
|
Rural
|
69 (61.0%)
|
44 (38.9%)
|
113 (64.2%)
|
|
Occupation
|
Student
|
21 (60.0%)
|
14 (40.0%)
|
35 (19.8%)
|
|
Farmer
|
28 (70.0%)
|
12 (30.0%)
|
40 (22.7%)
|
|
Government employee
|
11 (61.1%)
|
7 (38.8%)
|
18 (10.2%)
|
|
Private
|
7 (63.6%)
|
4 (36.3%)
|
11 (6.25%)
|
|
Unexplained
|
43 (59.7%)
|
29 (40.2%)
|
72 (40.9%)
|
Most of the study participants, 43 (24.4%), were within the age range of 31 to 40
years with a mean age of 28.8 ± 17.3 years. Among patients suffering from ALL, the
majority of patients were of pediatric age group (85.9%), while for AML, the majority
were adults (94.6%). For ALL, the mean age was 12.4 ± 10.1 years (ranging from 3 to
52 years). For AML, the mean age was 37.8 ± 13.3 years (ranging from 11 to 80 years).
Of the study participants, 113 (64.2%) were rural residents. From a total of 176 acute
leukemia patients, 104 (59.0%) have provided their occupational status, whereas 72
(40.9%) did not comply with any occupation. Of these patients, farmer and students
have the highest figures, 22.7 and 19.8%, respectively, while the least frequent were
private workers (11, 6.25%; [Table 1]).
Fever was the most common presenting complaint followed by pallor in both AML and
ALL. There was statistically significant difference in physical examination findings
between AML and ALL patients. Splenomegaly, lymphadenopathy, and sternal tenderness
were more often seen in ALL than in AML patients (p < 0.05). Pallor was more significantly associated with AML than with ALL patients
(p < 0.05; [Table 2], [Fig. 1]).
Fig. 1 Major clinical manifestations of the study population.
Table 2
Major clinical manifestations of the study population
|
Clinical features
|
AML
|
ALL
|
UC
|
Chi-square test
|
p-Value
|
|
Fever
|
86 (53.1%)
|
54 (33.3%)
|
22 (13.6%)
|
0.775
|
0.679
|
|
Easy fatigability
|
37 (49.3%)
|
29 (38.7%)
|
9 (12.0%)
|
1.221
|
0.543
|
|
Weakness
|
19 (46.3%)
|
18 (43.9%)
|
4 (9.8%)
|
2.385
|
0.303
|
|
Loss of appetite
|
29 (60.4%)
|
14 (29.2%)
|
5 (10.4%)
|
1.541
|
0.463
|
|
Weight loss
|
22 (50.0%)
|
16 (36.4%)
|
6 (13.6%)
|
0.193
|
0.908
|
|
Pallor
|
85 (55.9%)
|
52 (34.2%)
|
15 (9.9%)
|
10.739
|
0.005
|
|
Bleeding manifestations
|
19 (59.4%)
|
9 (28.1%)
|
4 (12.5%)
|
0.734
|
0.693
|
|
Lymphadenopathy
|
22 (34.4%)
|
37 (57.8%)
|
5 (7.8%)
|
25.216
|
0.000
|
|
Splenomegaly
|
34 (42.0%)
|
36 (44.4%)
|
11 (13.6%)
|
8.102
|
0.017
|
|
Hepatosplenomegaly
|
47 (49.5%)
|
36 (37.9%)
|
12 (12.6%)
|
1.349
|
0.509
|
|
Sternal tenderness
|
3 (23.1%)
|
9 (69.2%)
|
1 (7.7%)
|
7.748
|
0.021
|
Abbreviations: ALL, acute lymphoblastic leukemia; AML, acute myeloid leukemia; UC,
unclassified.
Out of 176 cases, 158 (89.7%) patients presented with leucocytosis. Only 7 cases (3.9%)
had normal leukocyte count and 11 cases (6.2%) had leucopenia ([Table 3]).
Table 3
Frequency of laboratory indices in acute leukemia patients
|
Parameter
|
Range
|
Number of cases
|
%
|
|
Total WBC count/mm3
|
< 4,000
|
11
|
6.2
|
|
4,000–11,000
|
07
|
3.9
|
|
11,000–50,000
|
28
|
15.9
|
|
50,000–100,000
|
72
|
40.9
|
|
100,000–200,000
|
45
|
25.5
|
|
> 200,000
|
13
|
7.3
|
|
Hemoglobin (g/dL)
|
< 6
|
54
|
35.5
|
|
6.1–10
|
90
|
59.2
|
|
10.1–12
|
08
|
5.2
|
|
Platelet count/mm3
|
< 20,000
|
13
|
7.3
|
|
20,000–50,000
|
53
|
30.1
|
|
50,000–100,000
|
101
|
57.3
|
|
> 100,000
|
09
|
5.1
|
|
Blast % on peripheral blood smear
|
< 20
|
15
|
8.5
|
|
21–89
|
104
|
59.0
|
|
90 or above
|
57
|
32.3
|
Abbreviation: WBC, white blood cell.
Anemia was detected in 100% of acute leukemia patients. There was severe degree of
anemia in 35.5% of cases (hemoglobin < 6 g/dL), and 59.2% had moderate degree of anemia
(hemoglobin: 6.1–10 g/dL), while 5.2% cases had mild anemia. No case showed percent
of hemoglobin more than 12 g. Mean hemoglobin in AML is 7.2 ± 2.0 gm/dL, while mean
hemoglobin in ALL is 7.4 ± 1.6 gm/dL ([Table 3]).
A total of 101 patients (57.3%) showed platelets count between 50,000 and 100,000/mm3, 53 cases (30.1%) showed 20,000 and 50,000/mm3, 13 cases (7.3%) showed platelets count less than 20,000/mm3, and only 9 cases (5.1%) showed platelets count more than 100,000/mm3 ([Table 3]).
Fifteen cases (8.5%) showed blasts less than 20%, 104 cases (59.0%) showed blasts
between 21 and 89%, and 57 cases (32.3%) showed blasts more than 90%. Mean blast percent
was 66.5 ± 27.4 in AML and 60.5 ± 29.6 in ALL ([Table 3]).
FCM was done in 150 (85.2%) patients of which 56 had ALL, 92 had AML, and 2 was classified
as mixed phenotypic acute leukemia ([Table 4]). Among the remaining 14.8% of study population, FCM was not done because of financial
constraints.
Table 4
Immunophenotypic findings in acute leukemia
|
Subtyping of AML on flow cytometry
|
No. of cases
|
|
AML M0
|
03
|
|
AML M1
|
18
|
|
AML M2
|
40
|
|
AML M3
|
05
|
|
AML M4
|
15
|
|
AML M5
|
09
|
|
AML M6
|
0
|
|
AML M7
|
02
|
|
Subtyping of ALL on flow cytometry
|
|
B-ALL
|
45
|
|
T-ALL
|
11
|
|
Mixed phenotypic acute leukemia
|
02
|
Abbreviations: ALL, acute lymphoblastic leukemia; AML, acute myeloid leukemia.
Cytogenetics was done in all five cases of AML M3 patients and PML RARA was found
positive.
Discussion
Leukemias have varied presentation. As there are no proper cancer registry programs
in Bihar, there is limited comprehensive assessment of clinical and laboratory profile
of acute leukemia in this region. As compared with the West, lower age standardized
incidence rates have been observed for most of the hematological malignancies.[8]
A total of 176 cases of acute leukemia were diagnosed on the basis of clinical course,
laboratory data, and morphological features of leukemic cells. In this study, the
AML-to-ALL ratio was approximately 1.5:1. The finding was consistent with the findings
of Kassahun et al, Idris et al, and Jatav et al.[1]
[9]
[10] This study is a hospital-based study and only those patients who visited us have
been included; hence, the findings are not externally valid.
In the present study, it was found that acute leukemias were more common in males,
with a male (62.5%) to female (37.5%) ratio of 1.6:1 (AML vs. ALL: 62.4 vs. 37.6%
and 61.7 vs. 38.3%, respectively). This is consistent with previous other studies.[9]
[11]
[12] Significant male preponderance reported in India can be partially explained by skewed
gender selection of cases at the time of presentation rather than true gender disparity.[13]
In the present study, maximum incidence of acute leukemia was found in 31 to 40 years
of age (24.4%), followed by 11 to 20 years of age (20.4%). Findings of our study were
not exactly comparable to that of Shahab and Raziq, where maximum incidence was seen
in 1 to 5 years of age group followed by over 35 years of age.[14] Kassahun et al reported maximum incidence of acute leukemia was seen among patients
older than 50 years (35.5%).[1]
A majority of patients suffering from acute leukemia belong to rural background. The
high preponderance may be because of varieties of chemical exposures such as pesticides,
herbicides, and fertilizers used in agricultural activities which may result in genetic
mutations leading to leukemogenesis.[15] Regarding the occupation distribution of patients who have AML, the highest proportion
of 22.7% was recorded in patients who were farmers (p < 0.001).[15] On the other hand, ALL was more common in students (p < 0.001). This may be due to the age distribution that most of the patients in the
age group of 0 to 20 years are students.
Clinical presentation of acute leukemia is very vague. In this study, most common
presenting clinical feature was fever followed by pallor, hepatosplenomegaly, easy
fatigability, and lymphadenopathy. These findings were consistent with several other
studies.[9]
[11]
[14] This result is supported by the fact that the infiltration of leukemic cells in
the bone marrow and their increased production will result in neutropenia (resulting
in fever), anemia, and thrombocytopenia (bleeding manifestations).[16]
Statistically significant difference was found in physical examination findings of
AML and ALL. Splenomegaly, lymphadenopathy, and sternal tenderness were more associated
with ALL compared with AML (p < 0.05). Pallor was more associated with AML than with ALL patients (p < 0.05). These findings were consistent with previous literature.[9]
[14]
[17] These data support the idea that patients in our region go to hospitals when their
condition has progressed to an advance stage.
In our study, anemia is the most common hematological abnormality followed by thrombocytopenia
and leucocytosis which has also been exemplified by Rathee et al.[18] Maximum patients (59.0%) had hemoglobin of 6.1 to 10 g/dL followed by less than
6 g/dL in 35.5% of cases. Thrombocytopenia and leucocytosis were most common hematological
abnormality in a study conducted by Preethi and Manisha et al.[19]
[20] Anemia and leucocytosis were found to be significantly associated with acute leukemia
patients (p < 0.05).
In this study, median blast percentage in AML and ALL was 79 and 63%, respectively.
Rathee et al found that the median blast proportion in AML was 45 and 38% in ALL.[18] In AML, Ghosh et al found mean values of 41.4% for peripheral blood blasts and 57.6%
for bone marrow blasts.[21]
AML and ALL patients were further categorized into subtypes by using the FAB classification.[6]
On FCM out of 150 cases, 92 cases were AML and 56 cases were ALL. Two cases were diagnosed
as mixed phenotypic acute leukemia. Most common subtype of AML in our study was AML
M2, followed by AML M1 and AML M4. In ALL, most common subtype was B-ALL. Minimal
screening panels recommended for immunophenotyping of acute leukemia include CD10,
CD19, CD7, CD5, CD13, CD33, CD117, CD34, HLA-DR, and CD45.[6] This finding was consistent with the study conducted by Preethi and Ghosh et al.[19]
[21] According to Shahab and Raziq, AML-M4 was the most frequent subtype.[14]
There were several flaws in this study as it was a hospital-based retrospective research.
The study is prone to selection bias and the findings of this study cannot be generalized.
Conclusion
In our study, AML was more common than ALL and males were frequently more involved
than females. AML M2 was the most common subtype of AML found in our study. To develop
our PBCR to know the future predictions of leukemia and for health surveillance, a
detailed clinico-hematological study is utmost needed which is still lacking from
our region that makes our study worth reporting.
