Diagnostic value of serum LDH in pre - Treatment Cancer Patients

Haya Salameh¹_,Firas Hussein²

¹Postgraduate student, Department of Biochemistry and Microbiology, Faculty of Pharmacy,

Tishreen University, Lattakia, Syria

²Assistant Professor, Department of Internal Medicine and Heamatology, Faculty of Medicine,

Tishreen University, Lattakia, Syria.

*Corresponding Author E-mail: ph.hayasalameh@gmail.com

ABSTRACT:

Purpose: Cancer is defined as a group of diseases, It involve abnormal cell growth with the potential of invade and spread, it also involve with metabolic changes such as a metabolic switch from oxidative phosphorylation to increased glycolysis, even under normal oxygene conditions, this phenomenon is termed the Warburg effect[6]. Lactate dehydrogenase (LDH) is a metabolic enzyme widely expressed in almost all tissues and is detectable in serum, which catalyzes the interconversion of pyruvate and lactate during glycolysis and gluconeogenesis, and is considered to be a key checkpoint of anaerobic glycolysis. LDH levels is elevated in many types of cancer in compare to normal tissues, and it has been linked to tumor growth, maintenance, and invasion; This enzyme is receiving a great deal of attention as a potential diagnostic marker or a predictive biomarker for many types of cancer and as a therapeutic target for new therapies.[5] Lactate dehydrogenase (LDH) is regulated by the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (mTOR)–containing complex 1 (PI3K/Akt/TORC1) pathway as well as tumor hypoxia induced factor (HIF1). This study aimed to investigate whether lactate dehydrogenase (LDH) level in diagnostic is a marker of hypoxia, had clinical value in determining the kind of cancer or the stage of the disease in pretreatment cancer patients.[7, 9] Patients and Methods: We evaluated pretreatment serum lactate dehydrogenase (LDH) in 285 newly diagnosed cancer patients. the study samples included both men and women, It also included all kind of cancer. statistical analysis: The Shapiro- Wilk test was used to determine the distribution of continuous variables. Student’s t-test, Mann-Whitney U test, chi square, and Ficher exact test were used too.[2] Results: Baseline LDH levels 210 IU/L was an independent prognosticator for recurrence-free survival The predictive value of baseline LDH value remained significant in the subgroup analysis. LDH level <227 IU/L was identified as an independent predictor of complete remission after treatment. It helps distinguish patients with different prognosis and select patients who are more likely to benefit from normal treatment regimens. A highly significant difference was found between the survival patterns of patients with LDH levels of 233 U or less and those with LDH levels greater than 233 U. (recurrence free survival rates were 63% and 74%, respectively. Discussion: The association of LDH level with the stage of the disease in Lung cancer, Brest cancer, Germcell cancer (prostate and testicular cancer, ovarian and uterus cancer), lymphoma and leukemia was evident even after adjustment for other factors that might affect the diagnosis. Pretreatment serum LDH determinations may provide a useful means of staging in cancer types mentioned previously.[30]

KEYWORDS: cancer, Lactate dehydrogenase, metabolism, anaerobic analysis

1. INTODUCTION:

Cancer is defined as a group of diseases involve several changes in proliferation, metabolic and other changes, cancer cells undergo unregulated growth associated with metabolic reprogramming.

in contrast to normal cells, most cancer cells display an increased rate of glycolysis instead of oxidative phosphorylation to generate sufficient energy for the demands of rapidly proliferating tumor cells, although the normal and sufficient oxygen concentrations. This phenomenon is known as the Warburg effect and is one of the most predominant metabolic alterations that occur during malignant transformation .[4]

increased attention in research in cell biology, biochemistry, and molecular biology has provided astonishingly detailed information about the molecules and processes that allow cells to divide, grow, differentiate, and perform their essential functions. This basic knowledge of cell biology has also led to practical discoveries about the mechanisms of cancer. Specific molecules that control the progression of a cell through the cell cycle regulate cell growth.[5]

An understanding of normal cell cycle processes and how those processes go awry provides key information about the mechanisms that trigger cancer. Loss of control of the cell cycle is one of the critical steps in the development of cancer.

Serum markers would be useful for diagnosis, monitoring the course of disease and evaluating the effectiveness of treatment Fisher and colleagues emphasized as poor prognostic factors The serum level of lactate dehydrogenase (LDH), commonly elevated in neoplastic disorders, has been suggested as a possible tumor marker for many years.[30]

Although cancer comprises at least 100 different diseases, all cancer cells share one important characteristic: they are abnormal cells in which the processes regulating normal cell division are disrupted.

That is, cancer develops from changes that cause normal cells to acquire abnormal functions. These changes are often the result of inherited mutations or are induced by environmental factors such as UV light, X-rays, chemicals, tobacco products, and viruses. All evidence suggests that most cancers are not the result of one single event or factor. Rather, around four to seven events are usually required for a normal cell to evolve through a series of premalignant stages into an invasive cancer. Often many years elapse between the initial event and the development of cancer. The development of molecular biological techniques may help in the diagnosis of potential cancers in the early stages, long before tumors are visible.[15]

Cancer results from a series of molecular events that fundamentally alter the normal properties of cells. In cancer cells the normal control systems that prevent cell overgrowth and the invasion of other tissues are disabled. These altered cells divide and grow in the presence of signals that normally inhibit cell growth; therefore, they no longer require special signals to induce cell growth and division. As these cells grow they develop new characteristics, including changes in cell structure, decreased cell adhesion, and production of new enzymes.[12]

These heritable changes allow the cell and its progeny to divide and grow, even in the presence of normal cells that typically inhibit the growth of nearby cells. Such changes allow the cancer cells to spread and invade other tissues[15].

For locally advanced solid tumors, hypoxia is a characteristic property due to rapid cancer cell proliferation, high metabolic demands, and functional angiogenesis.

There is clear evidence that hypoxia can promote cancer development and it is involved in the resistance to treatment via the formation of new blood vessels. Lactate dehydrogenase (LDH) is known to be a marker of hypoxia, which plays an important role in the proliferation and metastasis of tumor cells. Moreover, pretreatment serum LDH levels have been found to correlate with the prognosis of patients with malignant diseases.[5]

There is a good relationship between neoplasia and increased serum lactate dehydrogenase (LDH) level.[3]

Serum LDH level has also significant correlation with stage of the disease. There is a good relationship between serum LDH level and the stage of neoplasia. The patient with neoplasia, serum LDH level correlates with the stage as the pretreatment serum LDH level is elevated with the higher stages.

Pretreatment serum LDH level which may reflect the mass of tumor present and it is lowest in patient with localized disease. High proliferation with high cellular turnover rate could explain the markedly elevated levels of serum LDH in untreated patient with large tumor. Serum LDH level estimation not serves as an enzymatic indicator and a diagnostic factor of the tumor.

A definite and consistent shift in the pattern of molecular form of LDH has been found in a large series of malignant human neoplasm as compared with benign tumors and normal controls. Serum LDH activity approximate to normality when the tumor regression is near completion so that its reliability as an indicator is evident[8]

In this report, we examine the relationship between pretreatment LDH levels and the stage of several kinds of cancer in patients.

2. MATERIALS AND METHODS:

2.1 Patients:

Between January 2018 and December 2018, 285 consecutive patients newly diagnosed with cancer were entered onto the study, (134 men and 151 women), diagnosed with different kinds of cancer.

Blood samples were collected for laboratory tests within 1 week before initiationof therapy, directly after diagnosing. Laboratory personnel performing these assays were blinded to study outcomes.

The diagnosis was determined by clinical examination, routine laboratory tests and Biobsy.

Lactate dehydrogenase assay was performed by the Central Laboratory in Biochemistry Department In Tishreen Hospital, using Mindray Biochemical spectrophotometer (normal values, 165-240 U/liter). The LDH value used in analysis was that obtained just prior to therapy.

Patients were divided into subgroups, based on the kind of cancer they were diagnosed with (Lung cancer, Breast cancer, colorectal cancer, prostate cancer, uterus cancer and the other kinds in one group).

No patient included in the study had active hepatitis, Inflammation, Infarctions (myocardial, pulmonary, cerebral), pulmonary embolism, hemolysis. The study had not include patients who had already suffered of the disease, or patient who have previously been treated or been underwent surgery, It also had not include patients have any chronic or severe disease.

The primary endpoint of this study was to investigate whether the serum LDH was a diagnostic factor for the kind of cancer or the stage of the disease.

2.2: Statistical analyses

Data were analyzed using Microsoft Excel 2010, and SPSS version 14.0 statistical software, with lsd test we determine the LDH value that correlate with stage of the disease.

Continuous variables were presented as the median and range, while Categorical variables were presented as the number and percentages.[2]

The Shapiro- Wilk test was used to determine the distribution of continuous variables. Student’s t-test was used to compare normally distributed continuous variables, and MannßWhitnez U test, whereas Chi-square test (χ2) or Fisher exact test were used to analyze the frequency distribution between categorical variables where appropriate.

a P value of <0.05 was considered to be statistically significant.

Throughout the analysis, P values <.05 were considered statistically significant.

Serum LDH values collected within 7 days before randomisation were taken into consideration. To avoid the exclusion of cases with missing data, the multiple imputation method was used.

3. RESULTS:

3.1: Characteristics of the study population:

The study included 285 patients The median age at the diagnosis of cancer was 53 years (range: 22–80 years). The median serum LDH level for the entire cohort was 410.57±190 IU/L at diagnosis with a range 189.9 to 1326 IU/L Of the included patients, 52.98% of the patients were women and 47.02% of the patients were men, there was 71.57% with solid tumors which contains lung cancer (7%), breast cancer (25%), colorectal (12%), prostate and testicular (6%), uterus cancer(7%), other kinds (15%). and (28%) of patients with blood tumors, 61.73% of the patients with Blood cancers were diagnosed with leukemia and 38.27% were diagnosed with lymphoma.

There were 52.98% of patients diagnosed with stage 2, 30.88% of patients diagnosed with stage 3, 15.09% of patients diagnosed with stage 1 and 1.05% of patients diagnosed with stage 4.

42.45% of patients had serum LDH level < 230 IU/L, wherease the rest of patients had serum LDH level ≥ 230 IU/L..

3.2: Correlation between serum LDH levels and therapeutic response:

Pretreatment serum LDH level is a usefull diagnostic factor when it is combined with other diagnostic factors such as Tumor Markers and Blood tests. The study showed that LDH help distinguish between Blood Tumor and Solid Tumor (as one group) (Pvalue= 0.05), pretreatment serum LDH levels were elevated in Stage 1 in Blood cancer twice as much as the normal values, wherease in the Stage 1 in solid tumors the enzyme levels were within the normal values.

In all solid tumors groups, there were weak correlation between diagnostic serum LDH level and the kind of cancer, therfore LDH was not a diagnostic factor (Pvalue>0.005).

Table(1): cancer groups data

group	Stage 1	Stage 2	Stage 3	Stage 4	Pvalue	Correlation coefficient
Lymphoma	[209-253]	[364-414]	[692-1093]	-	0.05	0.915
Leukemia	[227-249]	[344-425]	[605-817]	-	0.041	0.895
Lung	[210-243]	[356-485]	[618-925]	-	0.045	0.918
Breast	[198-220]	[235-309]	[294-748]	-	0.044	0.645
Prostate	[244-274]	[348-443]	[713-919]	-	0.018	0.9
Uterus	[231-250]	[350-437]	[597-657]	-	0.029	0.904
Colorectal	-	[265-357]	[288-590]	1034	0.065	0.629
Other	[197-257]	[282-440]	[397-767]	[1322-1326]	0.31	0.776

3.3: Correlation between serum LDH levels and staging:

Pretretment serum Lactate dehydrogenase were a diagnostic factor usefull in staging of the disease in Lymphoma (P=0.05), Leukemia (P=0.041), Lung cancer (P=0.045) Fig(2), Breast cancer (P=0.044) Fig (3), Prostate cancer (P=0.018) Fig(6), Uterus cancer (P=0.029)Fig(4). Other cancers Fig(5), Colorectal cancer.

The t-test revealed a strong diagnostic impact of LDH. Patients with high LDH expression were associated with a higher stage of the disease compared with those having low LDH.

Lymphoma and Leukemia Stage 1 patients had LDH levels within normal range and slightly higher, in Stage 2, both Lymphoma and Leukemia patients had elevated LDH levels to one and half times the normal range, whereas in Stage 3 in Lymphoma, LDH levels is elevated to five times the normal value an in Leukemia it is elevated to four times the normal value.

Lung cancer in Stage 1 of the disease, patients had LDH levels within normal range and slightly higher, in Stage 2, LDH levels are twice as the normal value, whereas in Stage 3 LDH levels are tripled.

Breast cancer in Stage 1 and Stage 2 of the disease, patients had LDH levels within normal range and slightly higher, whereas in Stage 3 LDH levels are elevated up to four times the normal values.

Prostate and Testicular cancer in Stage 1 of the disease, patients had elevated LDH levels in range between (244-274 UI/L), in Stage 2, LDH levels are twice as the normal value, whereas in Stage 3 LDH levels elevated up to four times the normal values.

Uterus cancer in Stage 1 of the disease, patients had LDH levels within normal range and slightly higher, in Stage 2, LDH levels are twice as the normal values,

whereas in Stage 3 LDH levels are tripled.whereas in Stage 3 LDH levels are elevated up to four times the normal values.

In Colorectal cancer and Other kind of cancer (Pvalue>0.05), It was noticed that the pretreatment LDH levels were higher as the stage of the disease progressed.

Fig (1): Relation between Stage and LDH level blood cancer

Fig (2): Relation between Stage and LDH level in lung cancer

Fig (3): Relation between Stage and LDH level in Breast cancer

Fig (4): Relation between Stage and LDH level in Prostate

Fig (5): Relation between Stage and LDH level in Uterus

4. DISCUSSION:

A correlation between diagnostic serum LDH levels and stage of the disease in lung cancer, breast cancer, germcells cancers, lymphoma and leukemia.[1]

Pretreatment serum LDH level is elevated with the advanced stage. Furthermore, compared with patients with baseline LDH less than 210 IU/L, patients with LDH ≥210 IU/L had a statistically significant 3.6-fold risk of cancer recurrence and 3.1-fold risk of cancer-specific death, and this association was independent of other potential prognostic factor.[3]

We found that pretreatment serum LDH levels was an independent diagnostic factor for staging.

Previous research has explored the biological mechanisms that are responsible for the association between elevated LDH levels and staging in patients with several kinds of cancer.[8,9]

Possible explanations are as follows. First, tumor cells utilize glycolysis instead of mitochondrial oxidative phosphorylation to generate ATP that is required for the increased energy demand of the rapidly proliferating tumor cells. As a key enzyme in the process of glycolysis, LDH converts pyruvate and NADH to lactate and NAD+, determining the maintenance of glycolytic flow and consequently the production of ATP [24, 28]. Because LDH can be transcriptionally up regulated by hypoxia inducible factor1α (HIF-1α) and hypoxia in the tumor microenvironment is sufficient to stimulate the activation of HIF, there is a positive feedback loop between HIF and LDH under hypoxic conditions. Therefore, elevated levels of LDH indicate an aggressive phenotype. Second, an increased serum LDH has been reported to reflect a heavy tumor burden. Owing to the heterogeneity of tumor cells, tumors with heavier load contain tumor cells with greater diversities.[11,25]

Additionally, the vascular density is significantly higher in patients with elevated LDH levels which suggest an aggressive angiogenesis.[10,23]

As angiogenesis is essential for tumor proliferation and metastasis, patients with increased LDH levels are more likely to have a higher Stage of the disease. What is noteworthy is that cutoffs for LDH were heterogeneous in previous studies. In the study, we used a 230 IU/L as the normal value, and a range [150-240] as normal range. The impact of variations in LDH cutoffs has been evaluated in a published meta-analysis.

The importance of the pretreatment serum LDH levels as a diagnostic factor and direct indicator of tumor burden has emerged in several recent clinical studies It has been suggested that the tumor cells are the source of the elevated level of LDH, survival, response to therapy, and diagnostic which have been found to be associated with the pretreatment LDH level in patients with Lymphoma, Lung cancer and other cancer's types.

In our study the pretreatment serum LDH levels predicted Stage of the disease according to the elevation in its value, and the range it reached.

The data further suggest that tumors with a high rate of LDH production behave in a consistent manner; they follow a more aggressive course than do tumors with low LDH production. The observations presented here are consistent with the often made suggestion that rapidly proliferating populations of neoplastic cells depend largely upon glycolytic mechanisms for derivation of energy, with a high requirement for LDH to regenerate NAD from NADH in order to support continued glycolysis. Although often contemplated, a useful therapeutic exploitation of this metabolic peculiarity of neoplastic cells has yet to appear [29].

The present study indicates that pretreatment serum levels of LDH can provide a useful parameter of disease Stage in patients with several kinds of cancer. If these data are confirmed in analyses of patients entered into other treatment protocols at this and at other centers, the serum LDH may provide significant assistance in comparing the effectiveness of various treatment regimens in the “Diagnosis” of the disease in cancer patients. [20, 18]

This study have several strengths including: (1) it was not only the first one to specifically explore the diagnostic value of pretreatment serum LDH levels in all cancer's types but also the largest one to test the diagnostic values of LDH in patients with cancer in all stages; (2) all patients were from a single institution, so uniform treatment protocol can be ensured. This study had several limitations. First, its observational design prevents us from discounting completely any residual factors of confusion that may influence the levels of LDH such as bone disease. Second, data about serial dynamic serum LDH levels are lacking. Finally, the findings of this study may be specific to Asian populations.

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Received on 30.07.2019 Modified on 20.10.2019

Research J. Pharm. and Tech 2020; 13(3):1303-1308.

DOI: 10.5958/0974-360X.2020.00240.1