Xylene-Free Papanicolaou Staining.

INTRODUCTION: Xylene (XL) is the most commonly used clearing agent in Papanicolaou staining. XL is hazardous and toxic chemical and prolonged exposure to XL can cause many ill-health effects. The health risk due to XL can be minimized by substituting XL with less hazardous clearing reagents such as Pine Oil (PO), Eucalyptus Oil (EO) or Limonene (LM). The objective of this study was to compare the clearing ability, staining quality, preservation of morphology, physical properties and cost of XL, PO, EO and LM. METHODS: Four smears were prepared from each of 50 serous effusions and were subjected to Papanicolaou stain. Out of four smears, one each was exposed to clearing specifically with XL (Control), PO (Test), EO (Test) and LM (Test). Test smears were compared with control for clearing, staining and morphology; graded as excellent, good or fair and further scored as 3, 2, 1 and the Quality Index (QI) was calculated. Statistical analysis was performed and the p-value was calculated. In addition, the physical properties and cost of all the reagents were compared. RESULTS: QI was 0.96 for both XL and PO, whereas 0.92 and 0.54 for EO & LM respectively. Compared to XL, the quality of staining, clearing and morphology of PO and EO were statistically not significant, whereas the difference was statistically significant with LM (p=0.005). Physical properties such as volatility, flammability, miscibility with alcohol and DPX and the refractive-indices of all the reagents were almost similar and all were recyclable. Odour was pungent for XL and EO but was pleasant for PO and LM. The cost was less for PO as compared to others. CONCLUSION: PO was a natural, less hazardous, less toxic and economical clearing agent and can be considered as a substitute for XL.

Biomarker expression and impact on clinical outcomes in an international study of chemoradiation and MRI-based image-guided brachytherapy for locally advanced cervical cancer: BIOEMBRACE.

PURPOSE: BIOEMBRACE-I was designed to study the impact of biomarkers in addition to clinic-pathological factors on disease outcomes in patients treated with chemoradiation and MRI-guided brachytherapy (BT) for locally advanced cervical cancer in EMBRACE study. PATIENT AND METHODS: Between 2018-2021, eight EMBRACE-I sites contributed tumour tissue for immunohistochemistry of p16, PD-L1 and L1CAM. These biomarkers and clinicopathological factors (FIGO 2009 stage, nodal status, histology, necrosis on MRI) were analysed to predict poor response at brachytherapy (BT) (high-risk clinical target volume [HR-CTV] ≥40cc) at BT), and 5-year local control, pelvic control and disease-free survival (DFS). Interaction between p16, PD-L1, radiotherapy dose (HR-CTV D90) and disease outcomes was investigated. Univariable and multivariable analysis were performed. RESULTS: Two-hundred sixty-four patients were included. The median HR-CTV D90 was 89 (86-95) Gy. p16 positive (pos), PD-L1>1% and L1CAM ≥ 10% was noted in 86.6%, 20.1% and 17.8% respectively. P16 negative (neg) status (OR 2.0 (1.0-5.7), p=0.04), necrosis on MRI (OR 2.1 (1.1-4.3), p<0.02) independently predicted for HR-CTV≥40cc, as did FIGO stage and tumour width >5cm. PDL1>1% was associated with reduced local (82% vs. 94%, p=0.02) and pelvic control (79% vs. 89%, p=0.02). HR-CTV D90 <85Gy was associated with inferior 5-year local control in p16+ patients especially if PD-L1 was co-expressed. On multivariable analysis, PD-L1>1% was the only independent factor for 5-year local control (HR 3.3, p=0.04) and L1CAM ≥50% for pelvic control (HR 5.5 (1.3-23.3), p =0.02). CONCLUSIONS: P16 neg status and tumor necrosis on MRI are independently associated with poor response to chemoradiation, whereas PD-L1>1% and L1CAM≥50% have an independent impact on local and pelvic control suggesting impact of biomarker expression on outcomes. Further validation is needed.

An octad of fumarate hydratase-deficient uterine leiomyomas: Case series with review of literature from a single institution.

BACKGROUND: Fumarate hydratase (FH)-deficient (FH-d) leiomyomas are included in the recent World Health Organization fascicle of the female genital tumors. These are known to be associated with hereditary leiomyomatosis and renal cell cancer (HLRCC) syndrome. The tumors can be diagnosed based on certain histopathological features, along with loss of immunohistochemical expression of FH immunostain in most tumors. Currently, there is no documentation on these tumors from our subcontinent. AIMS: We analyzed eight FH-d leiomyomas diagnosed at our institute. RESULTS: The most common presentation was vaginal bleeding (menorrhagia). Pelvic ultrasonogram revealed multiple fibroids in most patients except in two, who harbored a single fibroid. The size of these fibroids ranged from 3 to 19 cm. Five patients underwent myomectomies, while three underwent a total abdominal hysterectomy and bilateral salphino-ophorectomy. The most consistently observed histopathological features were hemangiopericytomatous vascular patterns, cytoplasmic globules, increased cellularity, distinct eosinophilic nucleoli, and cytological atypia (8/8, 100% tumors), followed by multinucleate giant cells and perivascular edema, seen in 62% and 50% tumors, respectively. Immunohistochemically, all tumors were positive for desmin, smooth muscle actin, and h-caldesmon and showed loss of FH immunostain, along with low Ki-67/MIB1. None of those patients had any renal or cutaneous manifestations. CONCLUSIONS: This constitutes the first such study from the Indian subcontinent and reinforces that although uterine leiomyomas constitute an integral component of the diagnosis of HLRCC syndrome, these occur in the absence of renal or cutaneous manifestations. FH-d uterine leiomyomas are more likely sporadic and could be a false alarm to raise the possibility of HLRCC with their exclusive presence.

Clear cell adenocarcinoma of the cervix and its mimickers - A series of 19 cases from a tertiary care referral centre in India.

ABSTRACT: Clear cell adenocarcinoma (CCAC) of cervix is a rare subtype of endocervical adenocarcinoma that accounts for 4% of all cervical adenocarcinoma with many morphological mimickers. Retrospectively study cases of cervical clear cell adenocarcinoma of the cervix. Clinical profile and pathological features of CCAC of the cervix diagnosed between 2018-2022 were retrospectively analyzed.The database of the Department of Pathology of our institute was systematically searched for patients diagnosed with clear cell adenocarcinoma of the cervix during 2018-2022.A total of 19 patients were studied with the mean age of patients being 53.72 years (range 25 -84 yrs,standard deviation-25.9) and median tumor size being 5.6cm. Lymph node metastasis was identified in 33.3% and distant metastasis were seen in 20% of the cases. Staging could not be done in 4 cases.FIGO staging of the cases included IB1(2 cases), IB2(2 cases), IIB (3 cases),IIIA (1 case)IIIB(4 cases),and IV(3 cases). On histopathological evaluation, heterogeneous architectural pattern comprising of tubulocystic, solid, and papillary patterns were seen in 13 cases (13/19,68.4%). Pure tubulocystic (3/19,15.7%), pure papillary (2/19,10.5%), and pure solid patterns (1/19,5.3%) were also identified. Tumor cells with clear cytoplasm ranged from 5% to 95%. Nuclear atypia was moderate to marked in all the cases (19/19,100%). Mitotic activity varied from 1/10hpf to 20-22/10hpf. By immunohistochemistry, tumor was positive for Napsin A in all the cases,p16INK4a was negative in majority of cases (15/19,78.9%) and ER was negative in 14 cases (14/19,73.7%) .p53 showed wild type staining except for one case . Clear cell adenocarcinoma being a rare subtype of cervical adenocarcinoma, needs to be differentiated from other Human Papilloma Virus(HPV) independent adenocarcinomas (gastric and mesonephric types) and benign entities such as endocervical glandular Arias-Stella reaction. Judicious use of a panel of immunostains is often helpful.

Role of Intraoperative Frozen Section Assessment of Proximal Bile Duct Margins and the Impact of Additional Re-Resection in Perihilar Cholangiocarcinomas.

Intraoperative frozen section (FS) analysis to assess the bile duct margin status is commonly used to assess the completeness of resection during surgery for perihilar cholangiocarcinoma (pCCA) resection. However, the impact of additional re-section on the long-term outcome after obtaining an initial positive margin remains unclear. Patients diagnosed as pCCA on preoperative imaging and subjected to curative intent surgery from May 2013 to June 2021 with a minimum follow-up of 2 years were included. Intraoperative FS analysis of the proximal bile duct margin was performed in all patients. A positive margin was defined by the presence of invasive cancer. Out of the 62 patients with a preoperative diagnosis of pCCA on imaging, 35 patients were included for final analyses after excluding patients with inoperable disease (on staging laparoscopy or local exploration) and other/benign pathology on the final histopathology report. Out of the 35 patients, patients with postoperative 90-day mortality were excluded from the final survival analysis. FS analysis revealed an initial positive margin in 10 (28.5%) patients. Among 10 patients who underwent re-resection to achieve negative proximal margins, only 5 patients achieved a negative margin (secondary R0). An initial positive margin was associated with poor long-term outcomes. Median disease-free survival (DFS) and overall survival (OS) were 16 and 19.6 months for patients with an initial positive margin, but 36 and 58.2 months for patients with an initial negative margin, respectively (p = 0.012). The median DFS and OS were significantly lower for those with secondary R0 as compared to primary R0 (16 vs. 36 months for DFS, p = 0.117 and 19.6 vs. 58.2 months for OS, p = 0.027, respectively). An intraoperative FS positive proximal hepatic duct margin dictates poor long-term outcomes for patients with resectable pCCA. Additional resection has a questionable benefit on survival, when a secondary negative margin is achieved.

Impact of pCR after neoadjuvant chemotherapy and radical D2 dissection in locally advanced gastric cancers: Analysis of 1001 cases.

BACKGROUND: Advances in perioperative chemotherapy have improved outcomes in patients with gastric cancers (GC). This strategy leads to tumour downstaging and may result in a pathologic complete response (pCR). The study aimed to evaluate the predictors of pCR and determine the impact of pCR on long-term survival. METHODS: At the Department of Gastrointestinal and HPB Oncology at the Tata Memorial Centre, Mumbai, 1001 consecutive patients with locally advanced GCs undergoing radical resection following neoadjuvant chemotherapy from January 2005 to June 2022 were included. RESULTS: At a median follow-up of 61 months, the median OS was 53 months with a 5-year OS of 46.8 %. Ninety-five patients (9.49 %) realized pCR. Non-signet and well-differentiated histology were associated with pCR. pCR was significantly associated with improved OS, 5-year OS 79.2 % vs 43.2 % (HR 0.30, p < 0.001). On multivariable analysis, the realization of pCR and completion of adjuvant chemotherapy had superior OS. Whereas, signet-ring histology, linitis-like tumours, and high lymph node ratio had adverse outcomes. CONCLUSION: Tumour grade and signet-ring histology predict achievement of pCR in locally advanced GCs after neoadjuvant chemotherapy. Patients with pCR have significantly improved survival. Future neoadjuvant strategies should focus on enhancing pCR rates to improve overall outcomes.