Oral mucositis (OM) one of the most significant acute toxicity of Hematopoietic Stem Cell Transplantation (HSCT). There is already scientific evidence suporting OM the use of: low-level laser therapy (LLLT), cryotherapy (in autologous HSCT when conditioning regimens include high-dose melphalan), and keratinocyte growth factor (in autologous HSCT with high-dose chemotherapy associated with RT). In this sense, an alternative treatment becomes necessary. Based on the preliminary results of the phase I study, a double-blind, randomized phase II study is suggested to evaluate the effectiveness of copaiba-based mouthwash in the prevention of oral mucositis in HSCT was proposed. Patients will be randomized into 2 groups: A (copaíba) and B (placebo) and will use the mouthwash 4x/day. Each group will have 38 patients and be blind to the group in which they are included. They will be evaluated daily by a dental surgeon about oral mucositis, pain in the oral cavity and oropharynx, and dysphagia and will undergo daily laser therapy sessions, until neutropenic recovery.
INTRODUCTION Hematopoietic Stem Cell Transplantation (HSCT) involves the infusion of hematopoietic progenitor cells from a donor into a patient with compromised bone marrow, aiming to replace the marrow and restore immune function. This bone marrow impairment may result from neoplastic infiltration or malfunction due to benign, malignant, hereditary, or acquired conditions. The progenitor cells to be infused may originate from a compatible donor or from the patient themself. These cells can be harvested directly from the iliac crest through bone marrow aspirations and punctures; from peripheral blood using apheresis machines; or, less commonly, from umbilical cord blood. In allogeneic HSCT (allo-HSCT), the infused cells come from a compatible donor; in autologous HSCT (auto-HSCT), the cells are from the patient and are re-infused after collection and processing. Allo-HSCT can be used to treat a variety of conditions, including leukemia, lymphoma, myeloproliferative disorders, myelodysplastic syndromes, bone marrow failure syndromes, congenital immunodeficiencies, enzymatic deficiencies, and hemoglobinopathies. Auto-HSCT, on the other hand, is commonly used in neoplastic diseases and autoimmune conditions. Its purpose is to enable the administration of high-dose chemotherapy (HDC) in diseases that are particularly sensitive to increased chemotherapy intensity. Since the dose-limiting toxicity of HDC is myelosuppression, the reinfusion of hematopoietic progenitor cells allows the use of higher doses that would otherwise be unfeasible. Current evidence supports the use of auto-HSCT in diseases such as Hodgkin's lymphoma (HL), non-Hodgkin's lymphoma (NHL), advanced neuroblastoma, Ewing sarcoma, acute myeloid leukemia (AML), Wilms tumor, germ cell tumors, high-risk medulloblastoma, and multiple myeloma (MM). Hematological malignancies collectively accounted for 6.6% of the total estimated new cancer cases globally, according to The Global Cancer Observatory (GLOBOCAN) in 2021: NHL: 544,352 cases (2.8%); leukemia: 474,519 (2.5%); MM: 176,404 (0.9%); HL: 83,087 (0.4%). In Brazil, according to the National Cancer Institute (INCA), NHL is the ninth most common cancer in men and the tenth in women. Considering the most prevalent hematological malignancies, 26,600 new cases are expected annually between 2023-2025, with 14,170 in men and 12,490 in women (NHL: 6,420 in men and 5,620 in women; HL: 1,500 in men and 1,580 in women; leukemias: 6,250 in men and 5,290 in women). It's worth noting that MM is not included in INCA's estimates due to its classification as a rare disease. According to the Brazilian Transplant Registry, over the last 10 years (2013-2023), a total of 36,432 HSCT procedures were performed. In 2023 alone, 4,262 procedures were registered: 2,568 autologous and 1,694 allogeneic transplants. LITERATURE REVIEW Hematopoietic Stem Cell Transplantation (HSCT) The first step in the HSCT process is mobilization, which involves stimulating the bone marrow to release hematopoietic progenitor cells into the peripheral blood. Various regimens of chemotherapeutic agents and granulocyte colony-stimulating factors can be used during this phase. The second phase is cell collection. Once progenitor cells are present in the peripheral blood, they are harvested: in allogeneic HSCT, this typically occurs via bone marrow aspiration from the donor; in autologous HSCT, collection is done via apheresis from peripheral blood. The third phase is conditioning, a critical step for successful HSCT. This phase consists of a chemotherapy regimen, with or without total body irradiation (TBI), designed to eliminate the recipient's hematopoietic system, thereby preparing the bone marrow niche to receive the donor cells. In addition, the conditioning regimen may exert anti-tumor effects against the underlying disease. Conditioning regimens are classified as follows: myeloablative (high-intensity): leads to prolonged pancytopenia, typically indicated for younger patients (under 55 years) with few or no comorbidities; reduced-intensity (intermediate); non-myeloablative (low-intensity): induces minimal cytopenia and is suitable for older or medically fragile patients. The fourth phase is the infusion of hematopoietic cells, which marks Day 0 (D0) of the transplant process. The days preceding this event-during which conditioning occurs-are referred to as negative days (D-3, D-2, D-1), while subsequent days are denoted as positive (D+1, D+2, D+3, etc.). After infusion, the transplanted bone marrow typically enters a phase of aplasia, lasting approximately 2-3 weeks. Depending on the conditioning regimen and underlying disease, white blood cell counts often drop below 100 cells/mm³ around D+4. The fifth phase is neutrophil recovery, defined as the first of three consecutive days with an absolute neutrophil count of ≥500 cells/μL (0.5 × 10⁹/L). The sixth phase is platelet recovery, defined as the first of three days with unsupported platelet counts ≥20 × 10⁹/L. The seventh phase involves graft-versus-host disease (GVHD) prophylaxis, applicable specifically to allogeneic HSCT. This includes administration of immunosuppressive therapy-most commonly calcineurin inhibitors such as cyclosporine or tacrolimus-to prevent GVHD and graft rejection. An HSCT is considered successful when engraftment occurs-i.e., when blood cell counts remain above 500 cells/mm³ for three consecutive days. Despite advances in HSCT techniques, the procedure is still associated with a range of complications. Patients undergoing HSCT comprise a highly specific population, whose most prominent characteristic is profound immunosuppression, resulting from the underlying disease, the oncologic treatment, or both. All blood cell lines are often affected, leading to neutropenia, thrombocytopenia, and anemia. These hematological changes, combined with immunosuppression, result in a common set of complications, including infections, bleeding, and oral manifestations. One of the most critical phases for complications in autologous HSCT is conditioning, due not only to immunosuppression, but also to the direct and indirect effects of cytotoxic therapies on oral tissues, leading to a range of oral toxicities such as oral mucositis (OM), xerostomia, dysgeusia, and pain. These toxicities affect up to 80% of patients, significantly impairing oral function, including the ability to eat, drink, and speak. Oral Mucositis Oral mucositis (OM) is a process triggered by the generation of reactive oxygen species (ROS) and oxidative stress caused by chemotherapy (CT) and total body irradiation (TBI) on epithelial cells and connective tissues. These agents cause direct cellular and tissue damage and stimulate the activation of transcription factors, leading to an acute inflammatory response in the mucosal tissues. ROS cause DNA damage and cell death in the epithelial layer and stimulate the release of pro-inflammatory cytokines, which further exacerbate tissue injury and apoptosis. Additionally, ROS indirectly stimulates the cyclooxygenase-2 (COX-2) pathway, promoting angiogenesis. Pro-inflammatory cytokines also amplify tissue injury caused by CT or CT+TBI by inducing the production of further inflammatory mediators. The final stage of this pathophysiological process is ulceration, resulting from tissue damage, apoptosis, and necrosis of the basal epithelial layer, ultimately leading to mucosal breakdown. Previous studies have established that ROS play a central role in the pathogenesis of CT- and RT-induced OM, by causing direct DNA damage, increasing transcription factor expression, and triggering a cascade of pro-inflammatory cytokines, such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α). This results in a self-perpetuating cycle of tissue damage and inflammation, which may persist even after the cessation of CT or CT+TBI. The studies by Martins (2021), Kiyomi (2022), and Bossi (2016) indicated that increased levels of certain cytokines-namely IL-1β, IL-6, and TNF-α-were associated with greater inflammatory responses and higher OM severity in patients compared to control groups. The classical clinical signs of OM include mucosal erythema, which may or may not be accompanied by ulcerations. These lesions are often associated with patient-reported symptoms of pain, burning sensation, dysgeusia, and dysphagia, which significantly impair swallowing, oral hygiene, speech, and mastication. The disruption of the mucosal barrier also increases the risk of secondary and opportunistic infections, which in extreme cases may lead to septic shock. Collectively, these factors result in a significant decline in quality of life during and immediately following cancer treatment. The most widely used OM grading system is that of the World Health Organization (WHO), which classifies OM into the following grades: grade 0 - no change; grade 1 - erythema only; grade 2 - erythema and ulcers, but the patient can maintain a solid diet; grade 3 - erythema and ulcers, with the patient limited to a liquid or semi-liquid diet; grade 4 - severe ulceration requiring enteral or parenteral nutritional support. Several studies have identified laboratory parameters as potential predictors for the development of OM. Wardill et al. (2020), in a systematic review, found that lymphopenia in laboratory tests may serve as a predictor of OM. Similarly, Nishii (2019), in a multicenter retrospective study involving 326 patients, showed that low leukocyte and lymphocyte counts were associated with a higher risk of OM development. However, it is important to note that these studies did not specify at which point during cancer therapy these lab values were assessed, and results should therefore be interpreted with caution. The management of oral mucositis can be both preventive and therapeutic, and various strategies have been proposed and studied. These include the use of topical anesthetics, anti-inflammatory agents, systemic analgesics (opioids and non-opioids), growth factors, reinforcement of oral hygiene, low-level laser therapy (LLLT), cryotherapy, and natural products - among which zinc, propolis, aloe vera, and chamomile are notable. There is already scientific evidence, particularly in the context of hematopoietic stem cell transplantation (HSCT), supporting the use of: LLLT, cryotherapy (in autologous HSCT when conditioning regimens include high-dose melphalan), and keratinocyte growth factor (in autologous HSCT with high-dose chemotherapy associated with RT). However, even within the specific context of autologous HSCT, preventive measures for OM remain limited to specific conditioning regimens. LLLT has become widely used in the context of OM due to its ease of application and demonstrated efficacy. Its mechanism is based on the interaction of low-energy-density light with cells and tissues, producing photochemical, photophysical, and photobiological effects. The effect is primarily due to the activation of mitochondrial respiratory chain receptors by light, leading to cellular stimulation or inhibition, depending on the dosage used - importantly, without producing heat. To date, nine randomized clinical trials have demonstrated the efficacy of LLLT in preventing OM in the HSCT setting. Of these, six studies used red-spectrum diode lasers with a 660 nm wavelength. In a randomized clinical trial, Antunes et al. (2007) evaluated 38 patients undergoing either autologous or allogeneic HSCT using both the Oral Mucositis Assessment Scale (OMAS) and WHO OM scale. Patients were randomized into two groups: one group (n=19) received prophylactic LLLT, the other (n=19) did not receive LLLT, and the laser parameters were: InGaAlP diode laser, 660 nm, 50 mW, 4 J/cm², applied daily from D-7 to the day of marrow recovery (D+). Once OM lesions were identified, therapeutic LLLT (8 J/cm²) was applied directly to affected areas. 63.2% of patients receiving prophylactic LLLT had no ulceration (Grades 0 and 1), compared to only 10.5% in the control group (p \< 0.001). Since this was the first study to use prophylactic LLLT from the beginning of conditioning until marrow recovery, it significantly influenced clinical practice, and LLLT is now used as a preventive measure in various HSCT centers, including the National Cancer Institute (INCA) in Brazil. However, despite the efficacy of LLLT, some patients still develop OM, highlighting the need for additional preventive strategies that are scalable and easily incorporated into standard care. Copaiba In 2002, the World Health Organization (WHO) acknowledged the importance of traditional medicine as part of comprehensive patient care. Since then, the use of phytotherapeutic agents has increased significantly due to their efficacy, low toxicity, biocompatibility, and low cost. The copaiba tree (Copaifera spp.) is native to Latin America, with several species found in the Brazilian flora. Some of these species are used to produce copaiba oil, which is traditionally employed in folk medicine for its anti-inflammatory and wound-healing properties, either through oral ingestion or topical application. Given that oral mucositis (OM) is a toxicity induced by antineoplastic therapy, the use of natural compounds for its prevention and management is attractive. These agents are expected to have fewer side effects compared to synthetic drugs. A systematic review by Menezes et al. (2022) assessed the anti-inflammatory and wound-healing effects of copaiba oil (CPB) in oral lesions, including five preclinical studies conducted in animal models. Two studies demonstrated wound-healing activity, evidenced by a reduction in wound area and increased formation of immature bone tissue. Two other studies revealed anti-inflammatory effects, such as reduced acute inflammatory response, advancement in tissue repair stages, early collagen fibril formation, stronger anti-inflammatory response, reduced edema, and decreased concentration of CD68+ macrophages. The anti-inflammatory effect is primarily attributed to β-caryophyllene, a sesquiterpene that reduces matrix metalloproteinase production, decreases leukocyte count in circulating blood, and blocks cytokine receptors on leukocytes, thus reducing their recruitment to inflamed areas. This effect is further enhanced by inhibition of nuclear factor kappa-B (NF-κB), which consequently reduces the secretion of pro-inflammatory cytokines. The wound-healing activity is linked to increased vascular network formation, granulation tissue development, and fibroblast proliferation, all of which support the second phase of the healing process. Other studies have suggested that copaiba may also possess: antioxidant, osteogenic (bone-stimulating), cytotoxic, gastroprotective, analgesic, antimicrobial, antileishmanial, anti-edematous, antifungal, antigonorrheal, anthelmintic, and antiseptic properties. Its analgesic potential has been associated with kaurenoic acid, which interacts with opioid receptors. The antimicrobial effect - effective against both Gram-positive and Gram-negative bacteria, including Streptococcus mutans and other plaque-forming bacteria - appears to result from the compound's ability to penetrate or disrupt bacterial cell membranes. Although inhibition of fungal growth - particularly of Candida species - has also been observed, the antifungal effect of copaiba oil has not yet been conclusively proven. Despite the lack of clinical studies specifically investigating the use of copaiba in HSCT patients, its demonstrated anti-inflammatory, analgesic, antimicrobial, antifungal, and wound-healing effects make it a promising candidate for the prevention and supportive management of OM. Its use may help relieve symptoms and improve quality of life during oncologic treatment. This research line aims to propose an alternative and adjunctive methodology for the prevention of OM in patients undergoing HSCT. While LLLT is effective, some patients still develop OM, underscoring the importance of developing complementary preventive strategies. A Phase I dose-escalation clinical trial was previously conducted by the present research group to evaluate the safety and tolerability of copaiba-based mouthwash in patients with oral cancer undergoing radiotherapy (RT). In that study: * No dose-limiting toxicity (DLT) was observed at any tested concentration. * The maximum tested dose (used in the 6th cohort), which was 15% copaiba oil, administered four times daily, was deemed safe for use in a Phase II trial. * None of the patients reported pain or burning associated with the mouthwash in the absence of OM lesions, suggesting it is safe and non-toxic for this application. * No participant required interruption of cancer therapy (RT or chemotherapy) due to copaiba-related toxicity. JUSTIFICATION Based on studies available up to the start date of this study, it is observed that Hematopoietic Stem Cell Transplantation (HSCT) presents significant toxicity to oral cavity tissues, with oral mucositis (OM) being the most limiting acute effect. There is scientific evidence supporting the use of Low-Level Laser Therapy (LLLT), cryotherapy in autologous HSCT with high-dose melphalan conditioning, and keratinocyte growth factor, also in autologous HSCT with high-dose chemotherapy or chemotherapy plus total body irradiation conditioning. However, it is known that despite LLLT, patients still develop OM. Regarding cryotherapy and keratinocyte growth factor, their applications are limited to specific regimens and do not apply to all patients in need. Our group's experience with the use of Copaiba Oil-based Mouthwash (CPB) for preventing OM in radiotherapy for head and neck cancer patients in a phase I study defined the appropriate dose to propose a phase II study aimed at providing a treatment that reduces morbidity in these patients and can be widely applied within the Brazilian Public Health System (SUS). Thus, the proposed project constitutes a decisive tool in reducing the incidence of OM. In this regard, it is expected that Copaiba mouthwash, combined with LLLT, will be effective in preventing OM in patients undergoing autologous HSCT.
Study Type
INTERVENTIONAL
Allocation
RANDOMIZED
Purpose
PREVENTION
Masking
QUADRUPLE
Enrollment
76
An aqueous solution of copaíba (Copaifera Officinalis Resin) at 15% concentration will be used, manufactured by Interativo Pharmacy, according to the formula: copaíba oil (Copaifera Officinalis Resin) 15%, liquid mint aroma 5%, Tween 80 1%, aqueous solution (distilled water + Nipagin 0.1%) q.s.p. 100%. The product will be stored at room temperature and kept in the pharmacy of INCA. Patients will receive the solution once a week. They will be instructed to shake the bottle before use to homogenize the solution. They should use 10 mL of the solution per mouthwash (measured with the 10 mL dosing cup provided with each bottle), 4 times daily, performing vigorous mouthwashes for 1 minute and then spitting out all the solution after use. Swallowing the mouthwash is not recommended.
Placebo will be used, formulated as: liquid mint aroma 5%, Nipagin 0.1%, and distilled water q.s.p. 60 mL, packaged in a container identical to the investigational product to maintain physical characteristics and blinding. The product will be stored at room temperature and kept in the INCA pharmacy. Patients will receive the solution once a week. They will be instructed to shake the bottle before use to homogenize the solution. They should use 10 mL per mouthwash (measured with the 10 mL dosing cup provided), 4 times daily, performing vigorous mouthwashes for 1 minute and then spitting out all the solution after use. Patients will be instructed not to swallow the mouthwash.
Instituto Nacional de Cancer, Brazil
Rio de Janeiro, Rio de Janeiro, Brazil
RECRUITINGSafety and efficacy evaluation through adverse event related to the use of the product (Common Terminology Criteria for Adverse Events v5.0 and the Visual Analogic Scale)
Evaluate the safety and efficacy of Copaiba mouthwash combined with LLLT in patients undergoing autologous Hematopoietic Stem Cell Transplantation by evaluating the occurrence of an adverse event related to the use of the product under investigation (pain 4 degrees above the Visual Analogic Scale score previously reported by the patient, nausea or vomiting for 7 consecutive days exclusively associated with the times of use of the mouthwash) The Common Terminology Criteria for Adverse Events v5.0 ranks the events from 1 to 5, with higher scores meaning worse outcome. The Visual Analogic Scale score ranks the eventos from 0 to 10, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
OM (World Health Organization scale)
Compare the incidence of the maximum grade of oral mucositis between the two groups. The World Health organization scale is used for mucositis assessment and ranks the lesions from 1 to 4, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
OM ulcerated area (Sonis scale)
Compare the average ulcerated area between the two groups according to the Sonis scale. The Sonis scale is used for mucositis assessment and ranks the ulcers from 0 to 2 and the erithema from 0 to 1, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
OM erythema area (Sonis scale)
Compare the average erythema area between the two groups according to the Sonis scale. The Sonis scale is used for mucositis assessment and ranks the ulcers from 0 to 2 and the erithema from 0 to 1, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
OM (Sonis scale)
Compare the average sum of ulcerated and erythema areas between the two groups according to the Sonis scale. The Sonis scale is used for mucositis assessment and ranks the ulcers from 0 to 2 and the erithema from 0 to 1, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
Oral pain
Compare the oral cavity pain index between the two groups according to the Visual Analog Scale and the Common Terminology Criteria for Adverse Events v5.0. The Common Terminology Criteria for Adverse Events v5.0 ranks the events from 1 to 3, with higher scores meaning worse outcome. The Visual Analogic Scale score ranks the eventos from 0 to 10, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
Oropharyngeal pain
Compare the oropharyngeal pain index between the two groups according to Visual Analogic Scale and Common Terminology Criteria for Adverse Events v5.0. The Common Terminology Criteria for Adverse Events v5.0 ranks the events from 1 to 3, with higher scores meaning worse outcome. The Visual Analogic Scale score ranks the eventos from 0 to 10, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
Dysphagia
Compare the dysphagia index between the two groups according to Common Terminology Criteria for Adverse Events v5.0. The Common Terminology Criteria for Adverse Events v5.0 ranks the events from 1 to 3, with higher scores meaning worse outcome. The Visual Analogic Scale score ranks the eventos from 0 to 10, with higher scores meaning worse outcome.
Time frame: From the first day (day 1) of conditioning and continuing until engraftment (defined as the third consecutive day with neutrophil count ≥ 500 cells/mm³), approximately 28 days.
Pro-inflammatory cytokines
Compare the levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α between the two groups at the three specified time points (in pg/mL)
Time frame: On the first day (+/- 2 days) of conditioning, on day +6 (+/- 2 days), and on the day of "engraftment" (patient presenting 500 neutrophils for three consecutive days) (+/- 2 days).
Laboratorial analysis
Compare, in patients who develop oral mucositis lesions, the leukocyte and neutrophil counts (by cell/mm³)
Time frame: On day one, on the day oral mucositis lesions are detected (up to 2 weeks after the HSCT), and on the day oral mucositis lesions heal (up to 4 weeks after the HSCT).
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