Histopathological Evaluation and Molecular Diagnostic Tests for Peritoneal Metastases with Unknown Primary Site-a Review.

Cancer of unknown primary (CUP) is a well-studied entity with guidelines available for the management of patients with CUP. The peritoneum represents one of the metastatic sites in CUP, and peritoneal metastases (PM) could present as CUP. PM of unknown origin remains a poorly studied clinical entity. There is only one series of 15 cases, one population-based study, and few other case reports on this subject. Studies on CUP, in general, cover some common tumour histological types like adenocarcinomas and squamous carcinomas. Some of these tumours may have a good prognosis though majority have high-grade disease with a poor long-term outcome. Some of the histological tumour types commonly seen in the clinical scenario of PM like mucinous carcinoma have not been studied. In this review, we divide PM into five histological types-adenocarcinomas, serous carcinomas, mucinous carcinomas, sarcomas and other rare varieties. We provide algorithms to identify the primary tumour site using immunohistochemistry when imaging, and endoscopy fails to establish the primary tumour site. The role of molecular diagnostic tests for PM or unknown origin is also discussed. Current literature on site-specific systemic therapy based on gene expression profiling does not show a clear benefit of this approach over empirical systemic therapies.

Distribution of residual disease in the peritoneum following neoadjuvant chemotherapy in advanced epithelial ovarian cancer and its potential therapeutic implications.

INTRODUCTION: Residual disease in 'normal appearing' peritoneum is seen in nearly 30% of the patients following neoadjuvant chemotherapy (NACT) for advanced ovarian cancer. The goal was to study the sequence of response in different regions, the commonest sites of occult residual disease, its incidence in different peritoneal regions and the potential therapeutic implications of these. METHODS: This was a prospective multi-centre study (July 2018-June 2019). Pathological evaluation of cytoreductive surgery specimens was performed according to a fixed protocol. Prevalence of residual disease in different regions was used to study patterns of response and distribution of residual disease. RESULT: In 85 patients treated between July 2018 to June 2019, microscopic disease in 'normal appearing' peritoneal regions was seen in 22 (25.2%) and in normal peritoneum around tumor nodules in 30 (35.2%) patients. Regions 4 and 8 of Sugarbaker's PCI had the highest incidence of occult disease and regions 9 and 10 the lowest. The response to chemotherapy occurred in a similar manner in over 95%- the least common site of residual disease was the small bowel mesentery, followed by upper regions (regions 1-3), omentum and middle regions (regions 0, 4, 8), lower regions (regions 5-7) and lastly the ovaries. CONCLUSIONS: During interval CRS, based on the disease mapping provided in this manuscript, regions that have a high probability of residual disease should be explored and dissected. Complete resection of involved the peritoneal region can completely address the occult disease. The role of resection of the entire region as well as 'normal appearing' parietal peritoneal regions should be prospectively evaluated.

Essentials for Pathological Evaluation of Peritoneal Surface Malignancies and Synoptic Reporting of Cytoreductive Surgery Specimens-A review and evidence-based guide.

Peritoneal surface oncology has emerged as a subspecialty of surgical oncology, with the growing popularity of surgical treatment of peritoneal metastases comprising of cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC). Pathological evaluation plays a key role in multidisciplinary management but there are still many areas where there are no guidelines or consensus on reporting. Some tumors presenting to a peritoneal surface oncology unit are rare and pathologists my not be familiar with diagnosing and classifying those. In this manuscript, we have reviewed the evidence regarding various aspects of histopathological evaluation of peritoneal tumors. It includes establishing a diagnosis, appropriate classification and staging of common and rare tumors and evaluation of pathological response to chemotherapy. In many instances, the information captured is of prognostic value alone with no direct therapeutic implications. But proper capturing of such information is vital for generating evidence that will guide future treatment trends and research. There are no guidelines/data set for reporting cytoreductive surgery specimens. Based on the authors' experience, a format for handling/grossing and synoptic reporting of these specimens is provided.

The Pathologic Peritoneal Cancer Index (PCI) Strongly Differs From the Surgical PCI in Peritoneal Metastases Arising From Various Primary Tumors.

BACKGROUND: The surgical peritoneal cancer index (sPCI) is calculated based on a subjective evaluation of the extent of peritoneal disease during surgery. The pathologic PCI (pPCI) may be a more accurate and objective method for determining the PCI. This study aimed to compare the sPCI and pPCI and to study the potential pitfalls and clinical implications of using the pPCI. METHODS: This prospective study (July to December 2018) included all patients undergoing cytoreductive surgery (CRS). The pPCI was calculated for each patient and compared with the sPCI. The impact of potential confounding factors on the difference between pPCI and sPCI was evaluated. RESULTS: Among 191 patients undergoing CRS at four centers, the pPCI and sPCI were concordant for 37 patients (19.3%). The pPCI was lower than the sPCI for 125 patients (65.4%) and higher for 29 patients (15.1%). The concordance between the two groups was maximum for gastric cancer (38.8%) and colorectal cancer (27.6%) and least for mesothelioma (6.7%) and rare primary tumors (5.6%) (p = 0.04). The difference was 0 to 3 points for 119 patients (62.3%), 4 to 5 points for 27 patients (14.1%), and more than 5 points for 45 patients (23.5%). The rate of concordance was not influenced by the use of neoadjuvant chemotherapy (NACT) (p = 0.4), but the difference was greater when NACT was used (p = 0.03). CONCLUSIONS: The pPCI strongly differs from the sPCI for patients undergoing CRS for peritoneal disease and may provide a more accurate evaluation of the peritoneal disease extent. Further studies are needed to determine its prognostic value compared with sPCI, and consensus guidelines are needed for calculating it.

Target region resection in patients undergoing cytoreductive surgery for peritoneal metastases-is it necessary in absence of visible disease?

BACKGROUND: The aim was to study the patterns of target region (greater omentum, lesser omentum, falciform and umbilical round ligament) involvement in patients undergoing cytoreductive surgery (CRS) from various primary tumors, factors affecting involvement and implications on surgical practice. METHODS: All patients undergoing CRS from July 2018 to December 2018 were included in this prospective study. The incidence of target region involvement in presence and absence of visible disease and the impact of primary tumor site, PCI and other variables on target region involvement was evaluated. RESULTS: In 191 patients, greater omentum was involved in over 15% of patients irrespective of the primary tumor type and in 15.7% in absence of visible disease. 75% of these had PCI <20. The involvement of the other three target regions was higher than 20% in ovarian cancer, appendiceal tumors and peritoneal mesothelioma. Involvement of these 3 regions was associated with a higher PCI (p < 0.001 for all) and omental involvement (p < 0.001for all). 2.1% of colorectal cancer patients had umbilical round ligament involvement, 4.2% had falciform ligament involvement and none had lesser omentum involvement. CONCLUSIONS: Target region involvement varies according to primary tumour site and disease extent. Resection of the greater omentum should be performed during CRS for PM arising from all primary sites. Resection of other target organs may be performed for selected patients with ovarian cancer, peritoneal mesothelioma and mucinous appendiceal tumors in absence of visible disease. For other patients, it should be done only in presence of visible disease.

Pathological assessment of cytoreductive surgery specimens and its unexplored prognostic potential-a prospective multi-centric study.

BACKGROUND AND AIM: The grade/histological subtype is one of the most important prognostic markers in patients undergoing cytoreductive surgery (CRS). Our aim was to study other potential prognostic information that can be derived from the pathological evaluation of CRS specimens and provide a broad outline for evaluation of these. METHODS: This prospective study (July to December 2018) included all patients undergoing cytoreductive surgery (CRS). A protocol for pathological evaluation was laid down which was based on existing practices at the participating centers and included evaluation of the pathological PCI, regional node involvement, response to chemotherapy, morphology of peritoneal metastases (PM) and distribution in the peritoneal cavity. RESULTS: In 191 patients undergoing CRS at 4 centers, the pathological and surgical PCI differed in over 75%. Nodes in relation to peritoneal disease were positive in 13.6%. Disease in normal peritoneum adjacent to tumor nodules was seen in >50% patients with ovarian cancer and mucinous apppendiceal tumors. 23.8% of evaluated colorectal PM patients had a complete response and 25.0% ovarian cancer patients had a near complete pathological response to chemotherapy. CONCLUSIONS: Pathological evaluation of extent and distribution of peritoneal disease differs from the surgical evaluation in majority of the patients. Lymph node involvement in relation of peritoneal disease is common. The morphological presentation of PM in ovarian cancer and mucinous appendiceal tumors merits evaluation of more extensive resections in these patients. Standardized methods of synoptic reporting of CRS specimens could help capture vital prognostic information that may in future influence how these patients are treated.

Pressurized Intraperitoneal Aerosol Chemotherapy (PIPAC): Initial Experience from Indian Centers and a Review of Literature.

Cytoreductive surgery and HIPEC is a therapeutic option that benefits only selected patients with peritoneal metastases (PM). New treatments like pressurized intraperitoneal aerosol chemotherapy (PIPAC) have been developed to overcome some limitations of intraperitoneal chemotherapy and treat patients who are not eligible for a curative approach. The safety and feasibility of the procedure in the first few Indian patients treated with PIPAC, and the technique and the set-up required for PIPAC are described here. From May 2017 to August 2017, data was collected prospectively for all patients undergoing PIPAC at three Indian centers. The patients' characteristic, operative findings, and perioperative outcomes were recorded. Seventeen procedures were performed in 16 patients with peritoneal metastases from various primary sites using standard drug regimens developed for the procedure. The median hospital stay was 1 day, minor and major complications were seen in two patients each (11.7%), and there was one post-operative death. Of the six patients who completed at least 6 weeks of follow-up, there was disease progression in two, unrelated problems in two patients, and a second procedure was performed in one patient. One patient underwent subsequent CRS and HIPEC. Our results show the feasibility and safety of PIPAC in Indian patients with a low morbidity and mortality and short hospital stay. While clinical trials will determine its role in addition to systemic chemotherapy, it can be used in patients who have progressed on one or more lines of systemic chemotherapy and those who have chemotherapy-resistant ascites.

Patterns of pathological response to neoadjuvant chemotherapy and its clinical implications in patients undergoing interval cytoreductive surgery for advanced serous epithelial ovarian cancer- A study by the Indian Network for Development of Peritoneal Surface Oncology (INDEPSO).

OBJECTIVES: The goals were to study • The pattern of pathological response to neoadjuvant chemotherapy (NACT) and its clinical implications • The impact of chemotherapy response grade (CRG) on survival METHODS: A retrospective analysis of patients undergoing interval cytoreductive surgery (CRS) between January 2013 to December 2017 was performed. The surgical and pathological reports were analyzed and surgical and pathological PCI compared. The pathological response to chemotherapy was assessed using the score developed by Bohm. et al. RESULTS: In 79 patients, it was observed that sites involved by disease first like ovaries and pelvic peritoneum (lower region) were the last to respond preceded by the omentum, right upper quadrant (RUQ) peritoneum (upper region) and parietal peritoneum (middle region). Microscopic residual disease was seen in 20.2% in normal looking areas of peritoneum and in 20% with no gross residual disease in the RUQ. Visual inspection during surgery overestimated the disease extent in 40.5% and underestimated it in 15.1%. There was no difference in the progression free (p = 0.587) and overall survival (p = 0.157) between patients with CRG 1, 2 and 3 (poor, moderate, and complete/near complete response, respectively). Retroperitoneal nodes were positive in 0% with CRG 3, 27.5% with CRG 2 and 72.7% with CRG 1 (p < 0.0001). CONCLUSIONS: The pathological response to NACT follows a specific pattern. Visual inspection is of limited value in assessing disease extent following NACT. Surgery following NACT should target sites involved before NACT and not just residual disease. The response in regional nodes should be included in chemotherapy response scores.

Treading the beaten path with old and new obstacles: a report from the Indian HIPEC registry.

BACKGROUND: The Indian HIPEC registry is a self-funded registry instituted by a group of Indian surgeons for patients with peritoneal metastases (PM) undergoing surgical treatment. This work was performed to • Evaluate outcomes of cytoreductive surgery ± HIPEC in patients enrolled in the registry. • Identify operational problems. METHODS: A retrospective analysis of patients enrolled in the registry from March 2016 to September 2017 was performed. An online survey was performed to study the surgeons' attitudes and existing practices pertaining to the registry and identify operational problems. RESULTS: During the study period, 332 patients were enrolled in 8 participating centres. The common indication was ovarian cancer for three centres and pseudomyxoma peritonei for three others. The median PCI ranged from 3 to 23. A CC-0/1 resection was obtained in 94.7%. There was no significant difference in the morbidity (p = .25) and mortality (p = .19) rates between different centres. There was a high rate of failure-to-rescue (19.3%) patients with complications and the survival in patients with colorectal PM was inferior. A lack of dedicated personnel for data collection and entry was the main reason for only 10/43 surgeons contributing data. The other problem was the lack of complete electronic medical record systems at all centres. CONCLUSIONS: These results validate existing practices and identify country-specific problems that need to be addressed. Despite operational problems, the registry is an invaluable tool for audit and research. It shows the feasibility of fruitful collaboration between surgeons in the absence of any regulatory body or funding for the project.

Can low grade PMP be divided into prognostically distinct subgroups based on histological features? A retrospective study and the importance of using the appropriate classification.

INTRODUCTION: The pathological classification of PMP of appendiceal origin has prognostic and treatment implications. Our goals were to • Classify low grade mucinous carcinoma peritonei (LGMCP) into prognostically distinct subgroups based on histological features. • Compare the reproducibility of the WHO and the PSOGI classifications for both PMP and the appendiceal primary tumor. PATIENTS AND METHODS: A retrospective analysis of patients undergoing CRS and HIPEC or debulking surgery was done. All the tumors were re-classified according to the PSOGI classification. LGMCP was further classified into three histological subgroups and the impact on survival was evaluated. RESULTS: From Jun 2011 to June 2016, 101 patients underwent CRS with HIPEC (n = 89) or debulking surgery (n=12). The median PCI was 28 (3-39) and 74.1% patients had CC-0/1 resections. Of the 76.2% patients who had LGMCP, 4 patients (5.1%) were classified as group 1, 54 (70.1%) as group 2 and 19 patients (24.6%) as group 3. At a median follow up of 21 months, the disease free survival was not reached, 30 months and 14 months for groups 1, 2 and 3 respectively (p = 0.09). There was no difference in overall survival. Using the WHO classification, there was a discordance in the grade of the primary tumor and the peritoneal lesions in 19.8% and conflicting terminology was used in 62% of patients. CONCLUSIONS: The subgroups of LGMCP described here are prognostically different though this needs further prospective evaluation in larger series. The PSOGI classification is more uniformly reproducible and should be preferred to the WHO classification.

Sequential cultivation of human epidermal keratinocytes and dermal mesenchymal like stromal cells in vitro.

Human skin has continuous self-renewal potential throughout adult life and serves as first line of defence. Its cellular components such as human epidermal keratinocytes (HEKs) and dermal mesenchymal stromal cells (DMSCs) are valuable resources for wound healing applications and cell based therapies. Here we show a simple, scalable and cost-effective method for sequential isolation and propagation of HEKs and DMSCs under defined culture conditions. Human skin biopsy samples obtained surgically were cut into fine pieces and cultured employing explant technique. Plated skin samples attached and showed outgrowth of HEKs. Gross microscopic examination displayed polygonal cells with a granular cytoplasm and H&E staining revealed archetypal HEK morphology. RT-PCR and immunocytochemistry authenticated the presence of key HEK markers including trans-membrane protein epithelial cadherin (E-cadherin), keratins and cytokeratin. After collection of HEKs by trypsin-EDTA treatment, mother explants were left intact and cultured further. Interestingly, we observed the appearance of another cell type with fibroblastic or stromal morphology which were able to grow up to 15 passages in vitro. Growth pattern, expression of cytoskeletal protein vimentin, surface proteins such as CD44, CD73, CD90, CD166 and mesodermal differentiation potential into osteocytes, adipocytes and chondrocytes confirmed their bonafide mesenchymal stem cell like status. These findings albeit preliminary may open up significant opportunities for novel applications in wound healing.

Intraoperative Frozen Section Evaluation of Sentinel Lymph Nodes in Breast Carcinoma: Single-Institution Indian Experience.

Sentinel lymph node biopsy is an established way of predicting axillary nodal metastasis in early breast cancer. Intraoperative frozen sections (FS) of sentinel lymph nodes (SLNs) can be used to detect metastatic disease, allowing immediate axillary lymph node dissection. The purpose of this study was to evaluate the accuracy of intraoperative frozen sections in evaluation of sentinel lymph nodes in cases of breast cancer. Between March 2006 and August 2010, a total of 164 patients with clinically node-negative operable breast cancer were subjected to sentinel lymph node biopsy of axillary lymph nodes using preoperative peritumoral injection of radioactive colloid and methylene blue. Intraoperative identification of sentinel nodes was done using a handheld gamma probe and identification of blue-stained nodes. The nodes were sent for frozen section examination. The results of frozen section were compared with the final histopathology. Out of the 164 cases, metastases were detected in SLN by frozen section in 38 cases. There were three false-negative cases (all showing micrometastasis on final histopathology). FS had sensitivity of 92.6 %, specificity of 100 %, and overall accuracy of 98.1 %. The positive predictive value was 100 %, and the negative predictive value was 97.6 %. FS for diagnosis of metastasis of SLNs is reliable. Patients with negative SLNs by the FS diagnosis can avoid reoperation for axillary lymph node dissection. However, FS may fail to detect micrometastases, especially in cases with small tumors.

Ovarian serous carcinoma: relationship of p53 and bcl-2 with tumor angiogenesis and VEGF expression.

The aim of the study was to assess the microvessel density (MVD) and vascular endothelial growth factor (VEGF) expression in ovarian serous carcinoma and to examine their relation with apoptosis.Paraffin-embedded specimens of 41 cases of ovarian serous carcinomas were evaluated by immunohistochemistry for VEGF, p53, and bcl-2 expression. MVD was assessed with CD31 staining. We investigated the association of tumor angiogenesis (MVD and VEGF) with clinicopathologic factors, p53 overexpression, and bcl-2 expression.There was a significant correlation between high MVD and suboptimal debulking and advanced stage disease. A significant negative correlation was expressed between bcl-2 and VEGF expression. In univariate analysis, only stage had a significant impact on disease-free survival.The results of this study suggest that higher degree of angiogenesis is associated with suboptimal debulking and advanced-stage disease. Expression of VEGF had negative association with VEGF expression.

An immunohistochemical comparison of P53 and Bcl-2 as apoptotic and MIB1 as proliferative markers in low-grade and high-grade ovarian serous carcinomas.

BACKGROUND: Traditionally, ovarian serous carcinomas (OSCs) have been graded using a 3-tiered system, as well differentiated, moderately differentiated, and poorly differentiated. Recently, a 2-tiered system has been proposed depending on the nuclear atypia and mitotic count. The dualistic pathway of ovarian serous carcinogenesis with accumulating molecular genetic evidence forms the basis for this grading system. OBJECTIVE: To investigate the expression of apoptotic proteins, p53 and bcl-2, and MIB1 index in low-grade and high-grade OSCs. METHODS: Eighteen cases of low-grade OSCs and 28 cases of high-grade OSCs were stained immunohistochemically with antibodies against p53, bcl-2, and MIB1. For p53 and bcl-2, staining was evaluated on a semiquantitative scale depending on the number of cells showing positivity. For MIB1, the percentage of positive nuclei was calculated. RESULTS: Of 28 cases of high-grade OSCs, 15 (53.6%) showed 5+ staining with p53 compared with 3 (16.7%) of 18 cases of low-grade OSCs. Of 28 cases of high-grade OSCs, 10 (35.7%) showed 5+ staining with bcl-2 compared with 2 (11.1%) of 18 cases of low-grade OSCs. The mean MIB1 index was 42.1% in high-grade OSCs (range, 10%-90%) in contrast to the 19.4% (range, 10%-40%) in low-grade OSCs. The differences in apoptotic protein expression and proliferative index between the low-grade and high-grade OSCs were statistically significant (P < 0.05). CONCLUSIONS: The expression of apoptotic markers is higher in high-grade OSCs, which also have a higher proliferative activity compared with those in low-grade OSCs, which supports the dualistic pathway of ovarian serous carcinogenesis.

Coracoclavicular and costoclavicular joints at a common juncture: a rare phenomenon / Articulaciones coracoclavicular y costoclavicular en una unión común: un raro fenómeno

Movements at the human shoulder girdle are the result of complex interplay of glenohumeral, acromioclavicular, sternoclavicular and scapulothoracic articulations. Clavicle apart from articulating with the scapula and sternum is also connected with first rib by costoclavicular ligament and with coracoid process by coracoclavicular ligament. At times the area of attachment of these ligaments on clavicle, first rib and scapula show faceted apophysis suggesting the presence of additional diarthrodial articulations. Costoclavicular joint exists between clavicle and first rib and coracoclavicular joint between clavicle and coracoids process. Both these joints are described in the literature, but the concurrent occurrence of them in the same bone has not been reported yet. We found two clavicles, one of right and other of left side, both of them showed faceted apophysis for costoclavicular and coracoclavicular joint simultaneously, which is rare phenomenon.

Los movimientos de la cintura escapular humana son el resultado de la interacción compleja de las articulaciones glenohumeral, acromioclavicular, esternoclavicular y escapulotorácica. La clavícula, aparte de la articulación con la escápula y el esternón también está conectada con la primera costilla por el ligamento costoclavicular y con el proceso coracoides por el ligamento coracoclavicular. A veces la zona de unión de estos ligamentos de la clavícula, la primera costilla y la escápula muestran procesos facetarios que sugieren la presencia de nuevas articulaciones diartrodiales. La articulación costoclavicular existe entre la primera costilla y la clavícula y la articulación coracoclavicular entre la clavícula y proceso coracoides. Ambas articulaciones están descritas en la literatura, pero la aparición simultánea de ellas en el mismo hueso no se ha informado aún. Se encontraron dos clavículas, uno del lado derecho y otra del lado izquierdo, las dos presentaron procesos facetarios para las articulaciones costoclavicular y coracoclavicular simultáneamente, lo cual es un fenómeno raro.