Diagnostic value of procalcitonin, C-reactive protein and lactate dehydrogenase in paediatric malignant solid tumour concurrent with infection and tumour progression

Infection is a fatal complication in cancer patients that sometimes is not distinguished from tumour progression. We compared the diagnostic value of procalcitonin (PCT), C-reactive protein (CRP) and lactate dehydrogenase (LDH) in paediatric malignant solid tumour concurrent with infection and tumour progression. The 152 children enrolled were divided into infection and control groups. Each group was divided further into stable and progression groups. An intergroup comparison was made in terms of serum PCT, CRP and LDH in all children. PCT, CRP and LDH levels were significantly higher in the infection than in the control groups (P < 0.05). Among the controls, PCT, CRP and LDH levels were significantly higher in the progression than in the stable groups (P < 0.05). In diagnosing infection, the sensitivity and specificity of PCT and CRP at the cutoff values of 0.296 ng/mL and 28.13 mg/L were relatively better than those at 0.5 ng/mL and 10 mg/L, respectively. LDH had the highest correlation with tumour progression, whereas PCT had the lowest (LDH, r = 0.684; CRP, r = 0.570; PCT, r = 0.322). Thus, PCT has the highest value in diagnosing infection and is less susceptible to tumour progression than CRP. LDH has obvious advantages in judging tumour progression.

serum pCt, CRp and LDH levels between the stable and progression groups among the infection and control groups. In the infection group, there was no significant difference in terms of PCT and CRP levels between the stable and progression groups (P > 0.05), but serum LDH levels were statistically significantly increased in the progression group (P < 0.05). In the control group, serum PCT, CRP and LDH levels were statistically significantly increased in the progression compared with those in the stable groups (P < 0.05; Table 4). www.nature.com/scientificreports www.nature.com/scientificreports/

Value of pCt, CRp and LDH in diagnosing tumour progression in children with malignant solid tumour.
The ROC curve of serum PCT, CRP and LDH was drawn for children in the control group (Fig. 1)   www.nature.com/scientificreports www.nature.com/scientificreports/ Value of pCt, CRp and LDH in combined diagnosis. The ROC curve of PCT combined with CRP in diagnosing infection, LDH combined with CRP in diagnosing tumour progression, and the combination of three indices in diagnosing infection with tumour progression were drawn, respectively (Fig. 2). The area under the ROC curve was 0.996 (95% CI, 0.990~1.000), 0.944 (95% CI, 0.901~0.987) and 0.885 (95% CI, 0.817~0.952) for PCT combined with CRP, LDH combined with CRP and the combination of three indices, respectively. Sensitivity/specificity was 72.0%/100% and 98.6%/73.7% for PCT combined with CRP in series and parallel methods (PCT ≥ 0.5 ng/mL and CRP ≥ 10 mg/L), respectively; 42.8%/97.8% and 91.3%/71.1% for LDH combined with CRP in series and parallel methods (LDH ≥ 295 U/L and CRP ≥ 10 mg/L), respectively and 57.1%/91.3% for combination of three indices in series method (PCT ≥ 0.5 ng/mL, CRP ≥ 10 mg/L, and LDH ≥ 295 U/L) ( Table 5).

Discussion
Infection is the most common and one of the most fatal complications during long-term chemotherapy in children with malignant solid tumour. Early diagnosis and treatment of infection facilitate control of the patient's  www.nature.com/scientificreports www.nature.com/scientificreports/ conditions and improve prognosis. In cancer patients treated with myelosuppressive chemotherapy, infection often shows lack of corresponding inflammatory symptoms and signs due to the chemotherapy-induced neutropaenia 1 , with no increase in body temperature and no easy tendency to form infectious foci, such as suppuration at infection sites, so that patients often are misdiagnosed. CRP is a common clinically used infection diagnosis biomarker that is rapid and inexpensive and may be a good partner to refine the diagnosis of infection 20 . Nonetheless, neoplastic fever often is accompanied by increased CRP, so that it may not be distinguished from infection fever 21 . Early empirical antibacterial therapy often leads to fungal infection and produces antimicrobial resistance 22 . In children with malignant solid tumour accompanied by early infection, a sensitive and specific infection diagnosis method is vital to improve survival rate and prognosis. Recent studies have indicated that, as an inactive propeptide substance of calcitonin and an indicator for systemic inflammatory reaction, PCT has a high specificity and sensitivity and is superior to other inflammatory factors in the diagnosis of bacterial infection, and its concentration is not susceptible to immunodeficiency conditions and use of corticosteroids; its diagnostic value has been significantly superior to CRP and other cytokines [11][12][13][14][15] . In addition, recent studies also have indicated that LDH is closely associated with infectious and tumour diseases, and increased LDH level may be related to tissue damages caused by infection or tumour cells [16][17][18] , so that LDH detection and analysis in children with tumour progression plus infection also can provide some evidence for clinical differentiation. The combined detection of serum CRP, PCT and LDH provides a new idea for diagnosis of children with malignant solid tumour and infection and further guides the clinical treatment in a better manner.
We detected PCT, CRP and LDH in serum specimens of children and investigated their correlation with infection and tumour progression. The serum PCT, CRP and LDH levels and positive detection rates of PCT and CRP were significantly higher (P < 0.05 and P < 0.05, respectively) in the infection than in the control groups. Therefore, PCT and CRP can serve as infective indicators for diagnosis of paediatric tumour with infection. The area under the ROC curves for PCT, CRP and LDH showed that PCT was superior to CRP and that LDH did not show much value in terms of infection diagnosis. Additional studies indicated relatively low (76.5%) sensitivity and relatively good (100%) specificity at a PCT cutoff value of 0.5 ng/mL and relatively good sensitivity/specificity (94.1%/97.5%) at a cutoff value of 0.296 ng/mL. Therefore, we deduced that using 0.3 ng/mL as the threshold value for diagnosing infection in children with malignant tumour seemed reliable. The reason might be related with age, immunosuppression, tumour factors or other nonspecific factors and also with the small sample size. The sample size will be expanded, or multicentre studies will be performed for further validation.
Our study also found relatively good (94.1%) sensitivity and relatively low (73.7%) specificity at a CRP cutoff value of 10 mg/L and relatively good sensitivity/specificity (88.2%/87.3%) at a cutoff of 28.13 mg/L. Some studies have indicated that CRP level is positively correlated with tumour progression 23 . Therefore, considering the impact of tumour factors, we also deduced that using 25 to 30 mg/L as the threshold value in children with solid tumour seemed more reliable when diagnosing infection, and a prospective study should be conducted for further validation.
Regarding the relationship between these three markers and tumour progression, in the control group, PCT, CRP and LDH levels were significantly higher in the progression than in the stable groups, which indicated that the levels of the three markers were affected by tumour progression. However, in the infection group, LDH levels were significantly increased in the progression compared with those in the stable groups, and there was no significant difference in PCT and CRP levels between these groups. It may be that PCT and CRP levels were significantly affected by the infection factors and, thus, led to no difference in results. LDH levels were less affected by infection, which suggested that LDH levels in the two groups were significantly different. The results were in line with expectations. To avoid the influence of infection factors, we performed ROC curves on PCT, CRP and LDH in the control group. The area under the ROC curve was 0.693 (95% CI, 0.591~0.795), 0.837 (95% CI, 0.764~0.910) and www.nature.com/scientificreports www.nature.com/scientificreports/ 0.908 (95% CI, 0.853~0.963), respectively. Sometimes, when unexplained increases in CRP and LDH were found in non-infected patients, clinicians often needed to consider whether it was caused by a definite tumour progression, so this study chose the cutoff values at 100% specificity and relatively highest sensitivity as a reference. When the PCT, CRP and LDH cutoff value was greater than 0.252 ng/mL, 38.3 mg/L and 391.5 U/L, respectively, 100% of non-infected patients had tumour progression.
This study established a combined diagnosis model. PCT combined with CRP was used for the diagnosis of infection, LDH combined with CRP was used for the diagnosis of tumour progression in non-infected children, and the combination of three indices were used for the diagnosis of infection combined with tumour progression. In the combined diagnosis, the series method improved the specificity, while the parallel method improved the sensitivity. We found that the parallel method was more reliable in the combined diagnosis of infection, and the optimal cutoff was 0.296 ng/mL and 28.13 mg/L for PCT and CRP, respectively. When LDH and CRP were combined to diagnose tumour progression, the parallel method was also better, and the optimal cutoff was 295 U/L and 10 mg/L for LDH and CRP, respectively. Considering that the parallel method has no practical application value when using the combination of three indices in diagnosing infection combined with tumour progression, this study only used series method for diagnosis. Sensitivity and specificity was relatively better at the cutoff of 0.296 ng/mL, 28.13 mg/L, and 300.5 U/L for PCT, CRP and LDH, respectively. The results showed that sensitivity and specificity could be improved by selecting the appropriate method for combined diagnosis, which was helpful for the differentiation of infection from tumour progression in children with malignant tumour.
Analysis indicated that their PCT, CRP and LDH levels were positively correlated with infection (PCT, r = 0.717, P = 0.000; CRP, r = 0.628, P = 0.000; LDH, r = 0.239, P = 0.003), with PCT and CRP having a relatively high and LDH a relatively low correlation. Serum PCT, CRP and LDH levels were positively correlated with tumour progression; LDH had the highest and PCT the lowest correlation. Analysis of these results indicated that PCT and CRP had significantly higher values than LDH in diagnosing infection, and LDH and CRP also had significant advantages in judging tumour progression. Despite the positive correlation with tumour progression, PCT was less impacted by the tumour itself, so there was almost no significance when using it to judge tumour progression.
In summary, our results indicated that serum PCT and CRP are important indicators in diagnosing infection in children with malignant solid tumour. PCT is superior to CRP in diagnosing infection, and it is significantly less susceptible to tumour factors than CRP. LDH and CRP have obvious advantages in judging whether the tumour has progressed compared with PCT. The combined detection of PCT, CRP and LDH is of high diagnostic value in identifying paediatric malignant solid tumour concurrently with infection and tumour progression and is helpful in the early detection of tumour conditions. Therefore, combined detection provides the basis for the early and reasonable implementation of clinical treatment.  (Table 1). All children were divided into infection (34 cases) and control (without infection, 118 cases) groups. Of 34 children in the infection group who had nosocomial bacterial or bacterial and viral infections, 23 had lower respiratory, 9 catheter-associated and 2 urinary tract infections. The 34 children in the infection group were subdivided into stable tumour (20) and tumour progression (14) groups. The 118 children in the control group were subdivided into stable tumour (74) and tumour progression (44) groups. All serum specimens were collected before the start of the chemotherapy cycle, and children who had taken antibiotics and corticosteroids were ruled out.

Diagnosis criteria of tumour and infection.
All malignant solid tumours were pathologically diagnosed by pathological biopsy or surgical resection. The judgement criteria for tumour progression were a 25% increase in one or more measurable lesions and appearance of new lesions 24 . The infection was diagnosed according to the definitions of the Centers for Disease Control and Prevention (CDC) 25 .

Methods
This study was approved by the hospital's Ethic Committee, and the informed consent was approved from the patients and their legal guardians. All experiments were performed in accordance with relevant guidelines and regulations. All children had 4 mL fasting venous blood collected before chemotherapy after admission. Serum PCT and routine biochemical tests (including CRP and LDH) were performed via the conventional detection methods at this hospital: electrochemiluminescence method to detect PCT, immunoturbidimetry to detect CRP and enzyme kinetics method to detect LDH. Infection results were judged according to PCT ≥ 0.5 (positive) and <0.5 (negative) ng/mL [26][27][28] and CRP ≥ 10 (positive) and <10 (negative) mg/L 20,29 . The normal reference range of serum LDH was 110-295 U/L. The sensitivity and specificity of the combined diagnosis were evaluated by both series and parallel methods. Diagnostic rule of series method: the combined diagnosis result is considered positive when the results of all individual assays are positive. Diagnostic rule of parallel method: the combined diagnosis result is considered positive when the result of any individual assay is positive.