PAK4 inhibition augments anti-tumour effect by immunomodulation in oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is one of the most common malignant tumours, warranting novel treatments. Here, we examined the therapeutic efficacy of inhibiting p21 activated kinase 4 (PAK4) in OSCC and determined its immunomodulatory effect by focusing on the enhancement of anti-tumour effects. We examined PAK4 expression in OSCC cells and human clinical samples and analysed the proliferation and apoptosis of OSCC cells following PAK4 inhibition in vitro. We also investigated the effects of in vivo administration of a PAK4 inhibitor on immune cell distribution and T-cell immune responses in OSCC tumour-bearing mice. PAK4 was detected in all OSCC cells and OSCC tissue samples. PAK4 inhibitor reduced the proliferation of OSCC cells and induced apoptosis. PAK4 inhibitor significantly attenuated tumour growth in mouse and was associated with increased proportions of IFN-γ-producing CD8+ T-cells. Furthermore, PAK4 inhibitor increased the number of dendritic cells (DCs) and up-regulated the surface expression of various lymphocyte co-stimulatory molecules, including MHC-class I molecules, CD80, CD83, CD86, and CD40. These DCs augmented CD8+ T-cell activation upon co-culture. Our results suggest that PAK4 inhibition in OSCC can have direct anti-tumour and immunomodulatory effects, which might benefit the treatment of this malignancy.

Rapamycin Induces Phenotypic Alterations in Oral Cancer Cells That May Facilitate Antitumor T Cell Responses.

OBJECTIVES: In this study, we investigated the antitumor immunomodulatory effects of rapamycin in oral cancer. STUDY DESIGN: We examined the proliferation, apoptosis, and migration of cancer cells and investigated the cell surface expression levels of immune accessory molecules and T cell immune responses in vitro. We investigated the effect of in vivo administration of rapamycin on immune cell distribution and T cell immune responses in oral tumor-bearing mice. RESULTS: Rapamycin treatment significantly inhibited OSCC cell proliferation and migration, increased apoptotic cell death, and upregulated cell surface expression of several immune accessory and adhesion molecules, including CD40, CD83, PD-L1, PD-L2, MHC class I, P-selectin, and VCAM-1. These cancer cells augmented T cell proliferation. In vivo rapamycin administration significantly attenuated mouse tumor growth with an increased proportion of immune cells, including CD4+ T cells, CD8+ T cells, and dendritic cells (DCs); decreased the proportion of immune suppressive cells, such as myeloid-derived suppressor cells and regulatory T cells; enhanced DC maturation and upregulated the surface expression of CD40, CD86, and ICAM-1. CONCLUSIONS: Our results suggest that the therapeutic effect of mTOR inhibition in oral cancer can cause direct antitumor and immunomodulatory effects.

Presurgical imaging of the subscapular artery with three-dimensional-computed tomography angiography: Application to harvesting subscapular system free-flaps.

A subscapular system free-flap is extremely useful for maxillofacial reconstruction since it facilitates the simultaneous harvesting of multiple flaps using one subscapular artery (SSA) alone. However, cases of aberrations in the SSAs have been reported. Therefore, the morphology of SSA needs to be confirmed preoperatively before harvesting the flaps. Recent developments in imaging, such as three-dimensional (3D) computed tomography angiography (3D CTA), facilitate obtain high-quality images of blood vessel images. Therefore, we examined the utility of 3D CTA in navigating the course of the SSA before harvesting subscapular system free-flaps. We examined the morphology and aberrations of the SSA using 39 sides of the 3D CTA data and 22 sides of Japanese cadavers. SSAs can be classified into types S, I, P, and A. Type S SSAs are significantly long (mean length = 44.8 mm). Types I and P SSAs have short mean lengths, measuring ≤2 cm in approximately 50% of cases. In type A, the SSA is absent. The frequency of types S, I, P, and A SSAs were 28.2%, 7.7%, 51.3%, and 12.8%, respectively. Type S can be advantageous for harvesting the SSA in subscapular system free-flaps, because it is significantly longer. In contrast, types I and P might be dangerous because their mean lengths are shorter. In type A, caution is needed not to injure the axillary artery because the SSA is absent. When surgeons need to harvest the SSA, presurgical 3D CTA is recommended.

Cleft Lip With Ankyloblepharon Filiforme Adnatum: A Case Report.

Ankyloblepharon filiforme adnatum (AFA) is a rare, benign congenital anomaly. Notably, it is characterized by the adhesion of the ciliary edges of the upper and lower eyelids at the trabecular line. AFA is usually a solitary malformation of sporadic occurrence; however, it can occur in conjunction with other congenital diseases. Herein, we report a case of cleft lip with AFA. A patient was referred to the ophthalmology department of our hospital. The ophthalmic diagnosis was AFA in both the eyes. The left eye was observed to have a fibrous adhesion in the center, and she underwent surgery to excise the fibrous adhesion of tissue with scissors. The right eye was observed to have a fibrous adhesion in the external canthus and was excised during lip plasty. After surgery, her eyes were able to fully open, and no other apparent disease was diagnosed. AFA is thought to be caused by an ectodermal-derived developmental abnormality. Notably, cases of AFA with a cleft lip are rare. Diagnosis and surgery should be performed promptly to minimize any risk of amblyopia and for the early detection of congenital diseases, including glaucoma.

Successful treatment with nivolumab in a patient with unresectable oral squamous cell carcinoma following ineffective chemoradiotherapy.

Key clinical message: Nivolumab has been clinically successful in prolonging the overall survival of patients with recurrent and metastatic head and neck cancer, complete remission is rare. Synergistic combinations of immunotherapy and conventional cancer treatments, such as radiotherapy or chemotherapy, are likely to be the most viable strategies for improving patient responses. Abstract: Immune checkpoint inhibitors have revolutionized recurrent, metastatic oral cancer treatment; however complete remission in advanced stages is unusual. We present a case of complete remission of advanced oral squamous cell carcinoma for >4 years in a 64-year-old Japanese woman, that responded poorly to chemoradiotherapy but well to subsequent nivolumab treatment.

Treatment of severe micrognathia in an adult with distraction osteogenesis: A case report.

Key Clinical Message: Distraction osteogenesis (DO) of the mandible is often performed at a young age, and there are few reports after age 30, as in this case. The Hybrid MMF used in this case was useful in that it allowed correction of fine directionality. Abstract: DO is often performed in young patients with a high capability of osteogenesis. We performed distraction surgery for a 35-year-old man who had severe micrognathia with serious sleep apnea syndrome. Four years postoperatively, suitable occlusion and improvement of apnea were observed.

Neutrophil-to-lymphocyte ratio as an early marker of outcomes in patients with recurrent oral squamous cell carcinoma treated with nivolumab.

The immune checkpoint inhibitor (ICI), nivolumab, has revolutionised the treatment of recurrent and metastatic oral cancer. However, the response rate to ICIs remains low, and identifying predictors of nivolumab response is critical. Although the neutrophil-to-lymphocyte ratio (NLR) has been suggested as a predictive marker of nivolumab response in patients with various types of cancer, its utility in oral squamous cell carcinoma (OSCC) has not been elucidated. In this retrospective multicentre cohort study, we evaluated the association between NLR and outcome of nivolumab treatment in 64 patients with OSCC treated between 2017 and 2020. The objective response and disease control rates were 25.1% and 32.9%, respectively. The rates for complete and partial responses were 15.7% (10/64) and 9.4% (6/64), respectively; stable and progressive disease rates were 7.8% (5/64) and 67.1% (43/64), respectively. Complete and partial responses were classified as responders, and stable and progressive diseases were classified as non-responders. The median (range) pre-treatment NLR among responders was 4.3 (2.8-8.0), which decreased to 4.0 (2.6-6.3) after nivolumab treatment, and the median (range) pre-treatment NLR among non-responders was 5.1 (2.7-7.9), which increased to 6.4 (4.0-14.0) with tumour growth. Moreover, overall survival was significantly worse in the group with a higher post-treatment NLR (≥5) than in the group with a lower NLR (<5). Patients with a post-treatment NLR of ≥6 had worse outcomes for salvage chemotherapy following nivolumab treatment. Thus, post-treatment NLR could be a useful marker for predicting the response to nivolumab treatment or salvage chemotherapy in patients with OSCC.

Clinical Application of Hyperdry Amniotic Membrane in Cleft Palate Repair.

OBJECTIVE: To examine the safety and efficacy of hyperdry amniotic membrane (HDAM) for wound closure after palatoplasty in cleft palate patients. METHODS: HDAMs were prepared by washing and drying under infrared rays and microwaves at temperatures less than 60°C using a hyperdrying device. A total of 16 cleft palate patients (8 males, 8 females), aged 1 to 3 years (mean age 1 year 9 months), received one-stage pushback palatoplasty. The remaining raw wound after surgery was covered by an HDAM and a plastic cover plate. The cover plate was removed 1 week after surgery and parameters including temperature, feeding, allergic reactions, postoperative bleeding, re-epithelialization, wound dehiscence, and infection were monitored during the follow-up period of 31.2 months. RESULTS: All patients could adequately ingest at 5 days postoperation and after removal of the cover plate. None of the patients had a persistent fever or allergic reactions. Ingestion was feasible immediately in all patients, and no postoperative bleeding was observed during ingestion. No secondary hemorrhages were observed during follow-up. No postoperative wound dehiscence on the midline of the palate was observed. No infections were observed after the removal of the cover plate. No patients suffered from severe scar formation or contracture of the wound in the follow-up period. Hemorrhage, undue epithelialization, and scar contracture did not occur in any patient. The mean evaluation score was 7.75 points. CONCLUSION: HDAM can be used safely and effectively for wound closure following palatoplasty in cleft palate infants. Future studies testing the safety of patient's own amnion for palatoplasty, are required.

Novel strategy to predict high risk of inferior alveolar nerve injury during extraction of lower third molars based on assessment of computed tomographic images of multiple anatomical features.

Preoperative assessment is essential to prevent inferior alveolar nerve (IAN) injury during surgical extraction of the lower third molar (LM3). Here, we aimed to establish an assessment system to predict IAN injury during surgical extraction of the LM3. We conducted a retrospective cohort study on 115 patients diagnosed as 'high-risk' based on our previous risk assessment method involving three anatomical features of the inferior alveolar canal using computed tomographic (CT) images. We evaluated the occurrence of neurosensory impairment in these high-risk patients, and its association with novel anatomic features based on CT images. Neurosensory impairments were observed in 19 patients (16.5%). The inferior alveolar canal major diameter (p < 0.0001) and lingual bone thickness (p = 0.0039) were significantly associated with the occurrence of neurosensory impairment during LM3 extraction. Receiver operating characteristic curves were used to determine cut-off values of these quantitative factors to specifically predict IAN injury. Preoperative risk assessment with quantitative factors based on anatomical features observed on CT images may facilitate more appropriate surgical planning for patients at a high risk of IAN injury.

[Neutrophil-to-Lymphocyte Ratio(NLR)as a Predictive Indicator of the Response to Nivolumab in Patients with Oral Squamous Cell Carcinoma].

BACKGROUND: The immune checkpoint inhibitor (ICI) nivolumab has revolutionized the treatment for recurrent or metastatic advanced oral cancer. Because the response rate remains low, the identification of predictive indicators of the response to nivolumab is among the most critical issues. The neutrophil-to-lymphocyte ratio(NLR)is a potential predictive marker of the response to nivolumab in patients with various cancer types. However, the utility of the NLR as a biomarker for predicting the response of oral cancer patients to ICIs is poorly understood. PATIENTS AND METHODS: In this retrospective cohort study, we evaluated the association between NLR and nivolumab treatment outcome in 13 patients diagnosed with recurrent or metastatic oral squamous cell carcinoma(OSCC)treated with nivolumab at the Toyama University Hospital between December 2017 and December 2019. RESULTS: Complete response(CR)and partial response(PR)rates of 38.5%(5/13)and 0% (0/13), respectively, were observed in responders; stable disease(SD)and progressive disease(PD)rates of 7.7%(1/13) and 53.8%(7/13), respectively, were observed in non-responders. After nivolumab treatment, the median NLR among responders decreased to 3.3(3.0-3.9)from 4.1(3.7-4.3)during pre-treatment assessment and increased from 5.6(3.2- 9.2)at pre-treatment to 9.4(5.3-17.9)among non-responders. Moreover, patients with higher NLRs(≥5)in the post- treatment group had a significantly worse overall survival than those with lower NLRs(<5). Specifically, patients with a higher post-treatment NLR(≥10)had significantly worse outcomes for post-nivolumab salvage chemotherapy. CONCLUSION: The NLR could be a useful marker for predicting the treatment response to nivolumab or post-nivolumab salvage chemotherapy in OSCC patients.

Central mucoepidermoid carcinoma of the maxilla developing from a calcifying odontogenic cyst: A rare case report.

Intraosseous mucoepidermoid carcinoma of the jaw is a rare lesion that has been suggested to originate from the odontogenic epithelium. We report an unusual case of central mucoepidermoid carcinoma in an 18-year-old Japanese man with an odontogenic cyst.

Durable complete response to immunotherapy with anti-PD-1 antibody nivolumab in a patient with oral squamous cell carcinoma presenting with lung metastasis: A case report.

Although the optimal treatment method for metastatic oral cancer remains largely unknown, the present case suggests that immunotherapy is a potentially promising alternative for metastatic oral cancer in which other therapies are no longer effective.

Protumor role of estrogen receptor expression in oral squamous cell carcinoma cells.

OBJECTIVE: Accumulating evidence has demonstrated the protumor role of estrogen receptor (ER)-mediated signaling in multiple cancer types, which is distinct from this signaling in sex steroid-dependent organs. However, its role in oral squamous cell carcinoma (OSCC) remains unclear. STUDY DESIGN: We assessed the expression of ERα and ERß in human OSCC tissues by immunohistochemistry and evaluated the expression of both receptors in OSCC cell lines by immunoblotting and flow cytometry. To further assess the contribution of ER-mediated signals to oral cancer progression, proliferation, invasion, and chemosensitivity, cell lines were stimulated with the ER agonist ß-estradiol. RESULTS: Immunohistochemical analysis of OSCC tissues showed that ERß was present in the cytoplasm and nuclei of OSCC cells. In contrast, ERα was not detected in any of the cases analyzed. Additionally, the proliferation and invasiveness of OSCC cells were significantly elevated following stimulation with ß-estradiol. Chemotherapeutic agent-induced apoptosis of cancer cells was attenuated by pretreatment with ß-estradiol. CONCLUSIONS: ER-mediated signaling plays a crucial role in oral cancer progression by facilitating the proliferation, invasion, and chemoresistance of OSCC cells, indicating its potential for developing novel targeted therapies for this type of cancer.

CD36 expression on oral squamous cell carcinoma cells correlates with enhanced proliferation and migratory activity.

OBJECTIVE: Recent studies have demonstrated the pro-tumour role of CD36 in multiple cancer types. However, its role has not been well elucidated in oral squamous cell carcinoma (OSCC). Here, we aimed to evaluate the role of CD36 in proliferation and migration of OSCC cells. METHODS: Human OSCC cell lines HSC-2, HSC-3, HSC-4 and Ca9-22 were assessed for proliferation by staining with the cell proliferation marker Ki-67. We also assessed migration activity, and the expression of cell adhesion molecules such as E-cadherin and ß-catenin and platelet-derived growth factor receptors (PDGFRs) of CD36-positive cells. RESULTS: CD36-positive cells showed increased expression of Ki-67 and migration activity compared with CD36-negative cells. Moreover, CD36-positive cells showed reduced expression of E-cadherin and ß-catenin, whereas the expression of PDGFRs increased compared with that in CD36-negative cells. CONCLUSIONS: Our results strongly suggest that CD36 has an important role in facilitating the proliferation and migration activity of OSCC cells, indicating its usefulness in the diagnosis of high-grade tumour and targeted therapy of oral cancer.

Alterations in composition of immune cells and impairment of anti-tumor immune response in aged oral cancer-bearing mice.

OBJECTIVES: Aging has been suggested to be associated with immune dysregulation. An understanding of alterations in the host immunity with advancing age is, therefore, important for designing immune therapy for elderly cancer patients. In this context, not much is known about age-associated alterations in the immune system in oral cancer. METHODS: To evaluate age-associated alterations in the immune system, which might affect anti-tumor immune responses in oral cancer, we performed a comparative analysis of the proportion of different immune cells, the proliferative capacity of T cell compartment, and the response against immune therapies targeting immune check point molecules between young and aged oral cancer-bearing mice. RESULTS: The proportion of immune regulatory cells, such as regulatory T cells and myeloid derived suppressor cells, was significantly increased in aged mice compared to that in young mice. Moreover, the expression of PD-1 and CTLA-4 on both CD4+ and CD8+ T cells was elevated in aged mice compared to that in young mice, and the proliferative abilities of CD4+ and CD8+ T cells derived from aged mice were significantly reduced following stimulation of T-cell receptors. Moreover, tumor growth was significantly enhanced in aged mice compared to that in young mice. However, immunotherapies targeting PD-1, CTLA-4, and PD-L1 resulted in faster tumor regression in aged mice than in young mice. CONCLUSIONS: Together, our results indicate that age-associated alterations in the immune system are directly associated with the impairment of anti-tumor immunity in aged mice bearing oral cancer, and might facilitate the progression of the tumor.

Roles of intracellular and extracellular ROS formation in apoptosis induced by cold atmospheric helium plasma and X-irradiation in the presence of sulfasalazine.

Sulfasalazine (SSZ) is a well-known anti-inflammatory drug and also an inhibitor of the cystine-glutamate antiporter that is known to reduce intracellular glutathione (GSH) level and increase cellular oxidative stress, indicating its anti-tumor potential. However, the combination of SSZ with other physical modalities remains unexplored. Here, the effects of SSZ on cold atmospheric helium plasma (He-CAP), which produces approximately 24 x higher concentration of hydroxyl radicals (. OH) compared to X-irradiation (IR) in aqueous solution, and on IR-induced apoptosis in human leukemia Molt-4 cells were studied to elucidate the mechanism of apoptosis enhancement. Both the Annexin V-FITC/PI and DNA fragmentation assay revealed that pre-treatment of cells with SSZ significantly enhanced He-CAP and IR-induced apoptosis. Similar enhancement was observed during the loss of mitochondrial membrane potential, intracellular Ca2+ ions, and mitochondria- and endoplasmic reticulum-related proteins. The concentration of intracellular reactive oxygen species (ROS) was much higher in He-CAP treated cells than in X-irradiated cells. On the other hand, strong enhancement of Fas expression and caspase-8 and -3 activities were only observed in X-irradiated cells. It might be possible that the higher concentration of intracellular and extracellular ROS suppressed caspase activities and Fas expression in He-CAP-treated cells. Notably, pretreating the cells with an antioxidant N-acetyl-L-cysteine (NAC) dramatically decreased apoptosis in cells treated by He-CAP, but not by IR. These results suggest that IR-induced apoptosis is due to specific and effective ROS distribution since intracellular ROS formation is marginal and the high production of ROS inside and outside of cells plays unique roles in He-CAP induced apoptosis. We conclude that our data provides efficacy and mechanistic insights for SSZ, which might be helpful for establishing SSZ as a future sensitizer in He-CAP or IR therapy for cancer.

Primary peri-implant oral intra-epithelial neoplasia/carcinoma in situ: a case report considering risk factors for carcinogenesis.

BACKGROUND: Major risk factors for oral squamous cell carcinoma (SCC) are tobacco smoking, a betel quid chewing habit, and heavy alcohol consumption. However, around 15% of oral SCCs cannot be explained by these risk factors. Although oral SCC associated with dental implants is quite rare, there has been a recent gradual accumulation of reports about it. Here, we report a case of primary peri-implant oral intra-epithelial neoplasia/carcinoma in situ (OIN/CIS) in a woman without the major risk factors for oral SCC. CASE PRESENTATION: A 65-year-old woman was referred to our clinic with a tumor in the right lower gingiva. She had no history of tobacco smoking and only drank socially. Ten years previously, mandibular right posterior teeth had been replaced with an implant-supported porcelain-fused-to-metal restoration in a dental clinic. About 7 years later, she noticed swelling on the lingual side of the gingiva around the implant-supported restoration, and was eventually referred to our clinic with the suspicion of a neoplasia around the dental implant. The upper part of the implant body was exposed on the implant corresponding to the first molar of the right side of the mandible; this was associated with painless, elastic soft, and relatively well circumscribed gingival swelling on the lingual site. A panoramic radiograph showed slight vertical bone resorption around the implants. An incisional biopsy was conducted under the suspicion of neoplasia. Pathological microscopic examination of the biopsy specimen revealed thickened squamous epithelia with slight nuclear atypism and disorders of the epithelial rete pegs. Immunohistochemical findings showed positive staining for keratin 17 and a negative staining mosaic pattern for keratin 13. High p53, p63, and Ki-67 reactivity was also observed. From these findings, OIN/CIS of the gingiva was pathologically diagnosed, and a wide local excision with rim resection of the mandible, including the implants, was performed. The pathological findings for the resected specimen were same as those for the biopsy specimen. After 1 year of follow-up, there was no evidence of recurrence. CONCLUSION: In this case, prolonged peri-implant mucositis or peri-implantitis may have been a plausible risk factor for carcinogenesis.

Conventional osteosarcoma of the mandible successfully treated with radical surgery and adjuvant chemotherapy after responding poorly to neoadjuvant chemotherapy: a case report.

BACKGROUND: Osteosarcoma, the most common primary bone malignancy, has an extremely poor prognosis and a high rate of local recurrence and distal metastases. Because osteosarcomas of the head and neck region are rare, accounting for less than 10% of all osteosarcoma cases, limited information is available about their treatment and prognosis. Because of the high rate of distal metastases associated with extragnathic osteosarcoma, surgery combined with chemotherapy is currently considered essential in its treatment. However, the role of chemotherapy has not been well elucidated in the treatment of head and neck osteosarcoma because of the rarity of this condition. CASE PRESENTATION: In this report, we present the case of a 58-year-old Japanese woman with osteosarcoma of the mandible that was treated with radical surgery combined with neoadjuvant and adjuvant chemotherapy. Because the tumor showed rapid growth during neoadjuvant chemotherapy, neoadjuvant chemotherapy was suspended and surgical resection was performed, followed by adjuvant chemotherapy. No evidence of local recurrence and distal metastasis was found 14 months after initial treatment. Local control is considered a principal prognostic factor for head and neck osteosarcoma. CONCLUSIONS: Wide surgical excision should be considered a primary goal even during neoadjuvant chemotherapy, especially in cases that respond poorly to neoadjuvant chemotherapy.

Successful multimodal treatment of intraoral salivary duct carcinoma in a patient with multiple lymph node metastases: a case report.

BACKGROUND: Salivary duct carcinoma (SDC) is a high-grade salivary gland malignancy that is associated with an aggressive clinical behavior and poor prognosis. Herein, we report on a long surviving case of SDC of the minor salivary gland with multiple lymph node metastases (LNMs). CASE PRESENTATION: An 83-year-old woman presented with a history of lymphadenopathy in the right side of the neck and recent onset and rapid growth of a mass in the right buccal region. Clinical examinations and biopsy findings were suggestive of a salivary gland malignant tumor with regional LNMs. The patient was treated with neoadjuvant chemotherapy. Tumor excision and ipsilateral radical neck dissection were performed, followed by adjuvant chemoradiotherapy. Postoperative histological examination revealed a tumor with irregular nests of atypical ductal epithelial cells, a cribriform growth pattern, and comedo-like central necrosis that lead to a final diagnosis of SDC. LNMs were observed in six lymph nodes of the right side of the neck. The patient underwent postoperative chemotherapy using single-agent cisplatin that was administered concurrently with radiotherapy (total, 65 Gy). There was no evidence of local recurrence or distant metastasis for >6 years. CONCLUSIONS: Although available data on treatment modalities for SDC remain limited, multimodal therapy may contribute to improved clinical outcomes in patients with advanced intraoral SDC.