Clinical And Experimental Studies Of "Nasal Cleaning Formula" In Treating Nasopharyngeal Carcinoma

Nasopharyngeal carcinoma (NPC for short), one of the common malignant tumors in China, is ranked as top above all head and neck cancer, with a significant regional distribution of incidence rate. It is more common in China and Southeast Asia, and among all, Guangdong is the highest. In recent years, NPC incidence rate has increased steadily, because its' specific pathogenesis is not certain. Although recent combination chemotherapy showed significant effect, but its'serious toxicity of side effect, easily lead to sequelae of different degree, severely affecting the quality of life and prognosis of the patients. Chinese medicine, as an effective adjuvant therapy, not only could alleviate radiation therapy side effects, enhance immunity, improve the tolerance of chemotherapy and radiation therapy, but also could improve patients' quality of life. Thus it has been increasingly used in clinical, and has become an important research topic in medical field. At the moment, Chinese medicine research has been more in-depth in China and overseas, and the researchers found that single herbs or Chinese herbal formula is indeed therapeutic, especially in immune regulation and tumor suppressor. Domestic Chinese Medicine treatment of NPC normally according to the treatment with syndrome differentiation and prescription studies, its exact mechanism of action remains to be elucidated."Nasal Cleaning Formula"(NCF for short), is formulated by Professor Chen Ruishen based on the "Qi" deficiency and contamination, as well as years of clinical practice experience, and has performed excellent clinical efficacy. Current research of NPC pathogenesis in immunology has progressed rapidly; the treatment of diseases in Chinese medicine is applying multi-target synergy. The author has studied the clinical efficacy and its mechanism of action of "Nasal Cleaning Formula" in the treatment of nasopharyngeal carcinoma.1,Clinical Research Objective:To observe the clinical efficacy of the adjunctive therapy of "Nasal Cleaning Formula" in radiation therapy of nasopharyngeal cancer. Method:76cases of NPC patients were randomly divided into treatment group ("Nasal Cleaning Formula"+radiation therapy) and the control group (radiation therapy),38cases respectively. Control group using conventional radiation therapy, while treatment group using additional "Nasal Cleaning Formula", with a dose every day, a total of seven weeks. Skin mucosal damage and adverse events before and after treatment is observed. T cell subsets, IL-2, TNF-alpha and etc. before and after treatment are determined. Result:(1)No explicit difference (P<0.05) in term of age, gender, radiation dose, pathological staging, radiation treatment and other general information between two groups.(2) Oropharyngeal injury appeared mainly in1,2degrees among patients of treatment group after three weeks of radiation therapy, incidence rate of grade Ⅲ reaction was44.7%,57.9%in the control group; incidence of grade Ⅳ reaction in treatment group was0,15.8%in the control group, the difference is significant(P<0.05);after7weeks of radiation therapy, incidence of grade Ⅲ reaction in treatment group was52.63%,71.05%in the control group; incidence of grade IV reaction in treatment group was2.6%,31.57%in the control group, the difference is significant(P<0.01).(3)Reaction time for radioactive oropharyngeal to be appeared in the treatment group (14.3±1.76) d lower than the control group (19.3±1.56) d (P <0.01).(4) IL-2与TNF-α of treatment group after treatment higher than control group(P<0.01); CD3,CD4&CD4/CD8of treatment group significantly higher than control group.(5) The toxicity in skin, mucous membranes, gastrointestinal and bone marrow of the treatment group is lighter than the control group.Conclusion:"Nasal Cleaning Formula" has the effect of supplementing vital energy and nourishing yin, clearing away heat, detoxication and resolving blood stasis. It could alleviate and delay the injury of skin and mucus due to radiation therapy, and could also reduce the gastrointestinal, bone marrow suppression and other adverse reactions. Besides, it could mitigate the injury of immunity made by radiation therapy and regulate the immune function, leading to a better protection effect against injury after radiation therapy. Therefore, it could increase patients'quality of life by increasing the effect and reducing the toxins.2,Laboratory Research Objectives:On the basis of the establishment of the rat model with low immunity, applying "Nasal Cleaning Formula" in interventions of rat model, to study the mechanism of "Nasal Cleaning Formula" in enhancing immunity of organism. Based on cell line (CNE cells) isolated from the nasopharyngeal carcinoma in China and serum pharmacology, explore the anticancer effect of "Nasal Cleaning Formula" against CNE cells through cell biology, molecular biology techniques. Method:(1)The effect of "Nasal Cleaning Formula"(NCF for short) on non-specific immune functions of immunosuppressive rats induced by cyclophosphamide70healthy BALB/c rats (both male&female50%) were randomly divided into5groups:model control group,"NCF" High Dosage Group,"NCF" Low Dosage Group, interferon positive control medicine group, normal control(blank) group. Every group except normal control group is given subcutaneous injection of cyclophosphamide, with multiple-doses of80mg/kg according to weight. Model control group,"NCF" High Dosage Group,"NCF" Low Dosage Group and normal control group were given medicine by intragastric administration, while interferon group were given medicine by intramuscular administration every other day, all with the testing cycle of30days. Evaluate by the body weight of rat, spleen weight, spleen suspension cell count, IgA, IgG and IgM level of serum and T-cell subsets CD3/4/8expression levels.(2)"NCF" treatment effects on the proliferation of nasopharyngeal epithelial cells in rats Preparation of Chinese herbal formula "NCF" concentrated decoction12healthy SD rats (both male&female50%) were randomly divided into6model groups and6experimental groups. Model groups were given normal saline through intragastric administration, while experimental groups were given "NCF" concentrated decoction through intragastric administration, with the testing cycle of5days. Blood was collected from rat's abdominal aorta, supernatant was made through centrifugation to make up medicine-contained serum to be act upon CNE cells, and by using MTT method, flow cytometry, electron microscope for growth curve observation, we can study the inhibition of the growth of tumor cells when "NCF" act on CNE cells, and also tumor cells apoptosis induced by "NCF". Results:(1)Compared to blank group, each group has the body weight of rats, spleen weight, spleen index, spleen cell count significantly decreased (P<0.01), while model control group hits the lowest for all values. When comparing the quantity of bone marrow cells (BMC) of each group with the blank group, there is no statistical significance (P>0.05). IgA level for model control group decreased significantly, but IgA level of interferon group,"NCF" Low Dosage Group and "NCF" High Dosage Group increased significantly (P<0.01). For IgG level, model control group and interferon group showed significant decrease (P<0.01), but "NCF" High Dosage Group and "NCF" Low Dosage Group showed insignificant change (P>0.05). Although IgM level of model control group, interferon group and "NCF" Low Dosage Group increased, level of "NCF" High Dosage Group decreased, but none of them showed statistical discrepancy(P<0.05).(2) Compared to model control group, the quantity of bone marrow cells (BMC) of each group with medication changed insignificantly (P>0.05). If we compared the body weight, spleen weight, spleen index, spleen cells count between "NCF" Low Dosage Group and model control group, both showed no statistical significance (P>0.05); Whereas the body weight, spleen weight, spleen index of "NCF" High Dosage Group significantly higher than model control group (P<0.01); The difference of the body weight, spleen weight between interferon group and model control group both showed obvious statistical significance(P<0.01). IgA level of interferon group,"NCF" Low Dosage Group and "NCF" High Dosage Group have increased significantly (P<0.01); IgG level of interferon group changed insignificantly, while IgG level of "NCF" Low Dosage Group and "NCF" High Dosage Group showed significant increase(P<0.05); There is no statistical discrepancy (P<0.05) between interferon group and "NCF" Low Dosage Group, while "NCF" High Dosage Group significantly lower than model control group, difference between them are statistical significant (P<0.05).(3)The overall expression levels of rat peripheral blood CD3, CD4, CD8are relatively lower than spleen, and showed no significant drop after modeling, but expression levels of spleen CD3, CD4, CD8decreased significantly. After giving interferon, high and low dosage of "NCF", CD3, CD4, CD8levels indicated certain degrees of increment. Among this, interferon group nearly reached the level of normal control group, while both "NCF" Low and High Dosage Group did not achieve.(4) Color comparison result of MTT method showed, after acting on for24hours, the inhibition of "NCF" medicine-contained serum of different concentration towards CNE cells was not significant. But after48hours, the inhibition effect of serum of5%,10%and15%concentration upon CNE cells was significant. After72hours, inhibition effect of serum of5%,10%concentration upon CNE cells was significant.(5) The growth curve of CNE cells showed:Activated serums of normal rat or medicine-contained (10%and15%) both possess cells growth inhibition effect. Activated medicine-contained serum of15%concentration are stronger than activated medicine-contained serum of10%in inhibiting CNE cells growth. Inactivated serums of normal rat or medicine-contained (10%and15%) also showed similar effect, but overall performance was lower than activated serum group.(6) For cells apoptosis rate, after using serum of normal rat and of medicine-contained (10%) to process CNE cells, there is apoptotic cells (23%) in serum of normal rat, whereas the apoptosis rate in medicine-contained serum is much higher (31.0%). By processing CNE cells using15%concentrated serum of normal rat and medicine-contained, apoptotic cells appeared in the normal rat serum (25.3%), while medicine-contained serum reached a total of33.9%apoptosis rate. Conclusion:"Nasal Cleaning Formula" could alleviate the immunosuppression caused by chemotherapy, regulate the cellular immunity and humoral immunity of organism, so as to reverse the immunity disorder, and promotes NPC patients recovery;"NCF" medicine-contained serum could inhibit the growth of CNE cells, inducing cells apoptosis, and this could possibly be its important mechanism.