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Yuan-jie Liu,1,2,* Mei Han,1,* Jie-pin Li,1– 3 Shu-hong Zeng,1,2 Qian-wen Ye,1,2 Zhong-hua Yin,1,2 Shen-lin Liu,1,2 Xi Zou1,2,4 1Department of Oncology, Affiliated Hospital of Nanjing University of Chinese Medicine, Jiangsu Province Hospital of Chinese Medicine, Nanjing, People’s Republic of China; 2No. 1 Clinical Medical College, Nanjing University of Chinese Medicine, Nanjing, People’s Republic of China; 3Department of Oncology, Zhangjiagang TCM Hospital Affiliated to Nanjing University of Chinese Medicine, Zhangjiagang, People’s Republic of China; 4Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine in Prevention and Treatment of Tumor, Nanjing, People’s Republic of China*These authors contributed equally to this workCorrespondence: Xi Zou; Shen-lin Liu, Email zxvery@126.com; lsljsszyy@126.comBackground: Gap junctions, as one of the major ways to maintain social connections between cells, are now considered as one of the potential regulators of tumor metastasis. However, to date, studies on the relationship between gap junctions and colorectal cancer (CRC) are limited.Methods: We synthesized connexins-coding gene expression data from public Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases. Bioinformatics analysis was performed using R software and several database resources such as MEXPRESS database, Gene Set Cancer Analysis (GSCA) database, Human Protein Atlas (HPA) database, Tumor Immune Single Cell Hub (TISCH) database, Search Tool for Retrieval of Gene Interaction Relationships (STRING), and Cytoscape software, etc., to investigate the biological mechanisms that may be involved in connexins. Immunofluorescence and immunohistochemical staining were used to validate the expression and localization of GJA4.Results: We found that CRC patients can be divided into two connexin clusters and that patients in cluster C1 had shorter survival than in cluster C2. The infiltration of M1 macrophages and NK cells was lower in cluster C1, while the levels of M2 macrophages and immune checkpoints were higher, indicating an immunosuppressed state in cluster C1. In addition, the epithelial–mesenchymal transition (EMT) phenotype was significantly activated in cluster C1. We observed that GJA4 was up-regulated in colorectal cancer tissues, which was related to poor prognosis. It was mainly expressed in fibroblasts, but the expression levels in normal intestinal epithelial cells were low. Finally, we found that GJA4 was associated with M2 macrophages and may be a potential immunosuppressive factor.Conclusion: We found that there is a significant correlation between abnormal connexins expression and patients’ prognosis, and connexins play an important role in stromal-tumor interactions. Connexins, especially GJA4, can help enhance our understanding of tumor microenvironment (TME) and may guide more effective immunotherapeutic strategies.Keywords: colorectal cancer, gap junction protein alpha 4, cancer-associated fibroblasts, M2 macrophage, bioinformatics

Breslow thickness (BT), a parameter measuring the depth of invasion of abnormally proliferating melanocytes, is a key indicator of melanoma severity and prognosis. However, the mechanisms underlying the increase in BT remain elusive. Utilizing data from The Cancer Genome Atlas (TCGA) human skin cutaneous melanoma (SKCM), we identified a set of BT-related molecules and analyzed their expression and genomic heterogeneity across pan-cancerous and normal tissues. Through consensus clustering, we identified two distinct BT phenotypes in melanoma, which exhibited significant differences in clinical, genomic, and immune infiltration characteristics. High BT molecular expression was associated with reduced CD8+ T cell infiltration and poor immunotherapy response, potentially mediated by the Macrophage Migration Inhibitory Factor (MIF) signaling pathway. In vitro experiments confirmed that BT molecules, including TRIM29, SERPINB5, and RAB25, promoted melanoma development through distinct mechanisms. Notably, fibroblast-derived TRIM29 and B-cell-derived RAB25 interacted with SPP1+ monocytes/macrophages via different pathways. Our findings suggest that genomic variations leading to imbalanced expression of BT molecules across cancers contribute to increased BT, which is closely linked to an immunosuppressive microenvironment. The involvement of multiple cell types and complex intercellular interactions underscores the importance of evaluating dynamic cellular crosstalk in the tumor microenvironment to better understand BT increases and develop more effective immunotherapeutic strategies., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The authors declare no conflicts of interest. All authors contributed to data analysis, drafting, or revising of the article; agree on the journal to which the article is being submitted; provided final approval of the version to be published; and agree to be accountable for all aspects of the work. Prior to the commencement of data collection, the first author obtained written consent from all participants concerning participation and subsequent publication of the study results., (Copyright © 2025 Elsevier B.V. All rights reserved.)

Photo-thermo catalysis that combines photo- and thermo-catalysis has emerged as an attractive approach for effective and economical elimination of pollutants. Herein, we fabricated a robust, highly water stable 2D Co(II)-MOF photo-thermo catalyst towards the decomposition of permanganate anions. It has been demonstrated that Co(II)-MOF exhibits excellent catalytic capacity, good selectivity and incredible reusability for degrading MnO 4 - . The reaction rate of the process was investigated for thermo-photo vs. thermo/photo processes to provide conclusive evidence that Co(II)-MOF shows a strong synergistic use as both an energy source and Lewis acid.

As one of the two main histologic subtypes of gastric cancer (GC), diffuse-type gastric cancer (DGC) containing poorly cohesive gastric carcinoma (PCC) components has a worse prognosis and does not respond well to typical therapies. Despite the large number of studies revealing the complex pathogenic network of DGC, the molecular heterogeneity of DGC is still not fully understood. We obtained single-cell RNA-seq data and bulk data from the tumor immune single cell hub, the public gene expression omnibus, and the cancer genome atlas databases. A series of bioinformatics analyses were performed using R software. Immunofluorescence staining, hematoxylin and eosin staining, western blot, and functional experiments were used for experimental validation. Caudin-3, -4 and -7 were lowly expressed in DGC and their expression levels were further reduced in PCC. The PCC components were mainly located in the deeper layers of the DGC and had a high level of hypoxic Wnt/β-catenin signaling and stemness. We further identified Insulin Like Growth Factor Binding Protein 7 (IGFBP7) as a marker for PCC components in the deep layer. IGFBP7 is stimulated by hypoxia and promotes cancer cell invasiveness and reduced claudin expression. In addition, programmed death-1 ligand (PD-L1) was specifically expressed in the deep layer, reflecting deep layer-specific immunosuppression. The PCC components are predominantly situated in the deeper layers of DGC. Initial molecular characterization of these PCC components revealed distinct features, including low expression of claudin-3, -4, and -7, high expression of IGFBP7, and the presence of PD-L1. These molecular traits may partially account for the pronounced tumor heterogeneity observed in GC., (© 2024. The Author(s).)

Immunotherapy has limited response rates in colorectal cancer (CRC) due to an immunosuppressive tumor microenvironment (TME). Combining transcriptome sequencing, clinical specimens, and functional experiments, we identified a unique group of CAF subpopulations (COX4I2 + ) with inhibited mitochondrial respiration and enhanced glycolysis. Through bioinformatics predictions and luciferase reporter assays, we determined that EBF1 can upstreamly regulate COX4I2 transcription. COX4I2 + CAFs functionally and phenotypically resemble myofibroblasts, are important for the formation of the fibrotic TME, and are capable of activating the M2 phenotype of macrophages. In vitro experiments demonstrated that COX4I2 + CAFs promote immunosuppressive TME by blocking CD8 + T cell infiltration and inducing CD8 + T cell dysfunction. Using multiple independent cohorts, we also found a strong correlation between the immunotherapy response rate of CRC patients and COX4I2 expression in their tumors. Our results identify a CAF subpopulation characterized by activation of the EBF1-COX4I2 axis, and this group of CAFs can be targeted to improve cancer immunotherapy outcomes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Colorectal cancer (CRC) is the third most common cancer worldwide. Connexin is a transmembrane protein involved in gap junctions (GJs) formation. Our previous study found that connexin 37 (Cx37), encoded by gap junction protein alpha 4 (GJA4), expressed on fibroblasts acts as a promoter of CRC and is closely related to epithelial-mesenchymal transition (EMT) and tumor immune microenvironment. However, to date, the mechanism concerning the malignancy of GJA4 in tumor stroma has not been studied., Methods: Hematoxylin-eosin (HE) and immunohistochemical (IHC) staining were used to validate the expression and localization of GJA4. Using single-cell analysis, enrichment analysis, spatial transcriptomics, immunofluorescence staining (IF), Sirius red staining, wound healing and transwell assays, western blotting (WB), Cell Counting Kit-8 (CCK8) assay and in vivo experiments, we investigated the possible mechanisms of GJA4 in promoting CRC., Results: We discovered that in CRC, GJA4 on fibroblasts is involved in promoting fibroblast activation and promoting EMT through a fibroblast-dependent pathway. Furthermore, GJA4 may act synergistically with M2 macrophages to limit T cell infiltration by stimulating the formation of an immune-excluded desmoplasic barrier. Finally, we found a significantly correlation between GJA4 and pathological staging (P < 0.0001) or D2 dimer (R = 0.03, P < 0.05)., Conclusion: We have identified GJA4 expressed on fibroblasts is actually a promoter of the tumor mesenchymal phenotype. Our findings suggest that the interaction between GJA4+ fibroblasts and M2 macrophages may be an effective target for enhancing tumor immunotherapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

The importance of the immune microenvironment in poorly cohesive carcinoma (PCC) has been highlighted due to its limited response rate to conventional therapy and emerging treatment resistance. A combination of clinical cohorts, bioinformatics analyses, and functional/molecular experiments revealed that high infiltration of Interferon Induced Protein with Tetratricopeptide Repeats 1 (IFIT1) + tumor-associated neutrophils (TANs) is a distinguishing feature of PCC patients. Upregulation of IFIT1 + TANs promote migration and invasion of gastric cancer (GC) cell lines (MKN45 and MKN74) and stimulates the growth of cell-derived xenograft models. Besides, by promoting macrophage secreted phosphoprotein 1 (SPP1) expression and facilitating cancer-associated fibroblast and endothelial cell recruitment and activation through TANs, IFIT1 promotes a mesenchymal phenotype, which is associated with a poor prognosis. Importantly, compared to non-PCC (NPCC), PCC tumors is more immunosuppressive. Mechanistically, IFIT1 can be stimulated by IFN-γ and contributes to the expression of Programmed Cell Death 1 Ligand (PDL1) in TANs. We demonstrated in mouse models that IFIT1 + PDL1 + TANs can induce acquired resistance to anti-PD-1 immunotherapy, which may be responsible for the difficulty of PCC patients to benefit from immunotherapy. This work highlights the role of IFIT1 + TANs in mediating the remodeling of the tumor immune microenvironment and immunotherapeutic resistance and introduces IFIT1 + TANs as a promising target for precision therapy of PCC., (© 2024. The Author(s).)

Cellular communication (CC) influences tumor development by mediating intercellular junctions between cells. However, the role and underlying mechanisms of CC in malignant transformation remain unknown. Here, we investigated the spatiotemporal heterogeneity of CC molecular expression during malignant transformation. It was found that although both tight junctions (TJs) and gap junctions (GJs) were involved in maintaining the tumor microenvironment (TME), they exhibited opposite characteristics. Mechanistically, for epithelial cells (parenchymal component), the expression of TJ molecules consistently decreased during normal-cancer transformation and is a potential oncogenic factor. For fibroblasts (mesenchymal component), the expression of GJs consistently increased during normal-cancer transformation and is a potential oncogenic factor. In addition, the molecular profiles of TJs and GJs were used to stratify colorectal cancer (CRC) patients, where subtypes characterized by high GJ levels and low TJ levels exhibited enhanced mesenchymal signals. Importantly, we propose that leiomodin 1 (LMOD1) is biphasic, with features of both TJs and GJs. LMOD1 not only promotes the activation of cancer-associated fibroblasts (CAFs) but also inhibits the Epithelial-mesenchymal transition (EMT) program in cancer cells. In conclusion, these findings demonstrate the molecular heterogeneity of CC and provide new insights into further understanding of TME heterogeneity., (© 2024. The Author(s).)

The TBC (Tre-2/Bub2/Cdc16, TBC) structural domain is now considered as one of the factors potentially regulating tumor progression. However, to date, studies on the relationship between TBC structural domains and tumors are limited. In this study, we identified the role of TBC1 domain family member 8 (TBC1D8) as an oncogene in colorectal cancer (CRC) by least absolute shrinkage and selection operator (LASSO) and Cox regression analysis, showing that TBC1D8 may independently predict CRC outcome. Functional enrichment and single-cell analysis showed that TBC1D8 levels were associated with hypoxia. TBC1D8 levels were also positively correlated with M2 macrophage infiltration, which may have a complex association with hypoxia. Taken together, these results show that the TBC1D8 gene is involved in colorectal carcinogenesis, and the underlying molecular mechanisms may include hypoxia and immune cell infiltration., (© 2022. The Author(s).)

Background: The downstaging of hepatocellular carcinoma (HCC) has been confirmed to benefit liver transplantation (LT) patients whose tumors are beyond the transplantation criteria. Milan criteria (MC), a tumor size and number-based assessment, is currently used as the endpoint in these patients. However, many studies believe that tumor biological behavior should be added to the evaluation criteria for downstaging efficacy. Hence, this study aimed to explore the feasibility of Hangzhou criteria (HC), which introduced tumor grading and alpha-fetoprotein in addition to tumor size and number, as an endpoint of downstaging., Methods: We performed a multicenter and retrospective study of 206 patients accepted locoregional therapy (LRT) as downstaging/bridge treatment prior to LT in three centers of China., Results: Recipients were divided into four groups: failed downstaging to the HC (group A, n = 46), successful downstaging to the HC (group B, n = 30), remained within the HC all the time (group C, n = 113), and tumor progressed (group D, n = 17). The 3-year HCC recurrence probabilities of groups B and C were not significantly different (10.3% vs. 11.6%, P = 0.87). The HCC recurrent rate was significantly higher in group A (52.3%) compared with that in group B/C (P < 0.05). Seven patients (7/76, 9.2%) whose tumor exceeded the the HC were successfully downstaged to the MC, and 39.5% (30/76) to the the HC. In group B, 23 patients remained beyond the MC and their survivals were as well as those of patients within the MC., Conclusions: Compared to the MC, HC downstaging criteria can give more HCC patients access to LT and furthermore, the outcome of these patients is the same as those matching MC downstaging criteria. Hangzhou downstaging criteria therefore is applicable in clinical practice., (Copyright © 2020. Published by Elsevier B.V.)

Background: We aim to investigate the safety and efficacy of radiofrequency ablation in the treatment of solitary hepatocellular carcinoma (3-5 cm) in comparison with surgical resection., Methods: Included in this study were 388 patients with primary and solitary hepatocellular carcinoma, of whom 196 patients underwent surgical resection and the other 192 patients received radiofrequency ablation. Clinicopathological characteristics, prognosis, post-treatment complications, hospital stay, and financial expenditures between the two groups were compared retrospectively., Results: The result of propensity score matching and subgroup analysis showed that the 1-, 3-, and 5-year overall survival and disease-free survival were comparable in patients with tumors of 3-4 cm in diameter between surgical resection and radiofrequency ablation groups. However, when the tumor size exceeded 4 cm in diameter, surgical resection exhibited a superior long-term prognosis compared with radiofrequency ablation. Nevertheless, hepatectomy was associated with high occurrences of postoperative complications, long hospital stay, and high hospitalization cost as compared with radiofrequency ablation. Further analysis of the relationship between tumor size and pathological features of hepatocellular carcinoma showed that tumors larger than 4 cm were positively correlated with a high rate of microvascular invasion and satellite nodule formation., Conclusion: For solitary hepatocellular carcinoma of 3-4 cm in diameter, radiofrequency ablation could achieve a comparable prognosis with a low incidence of post-treatment complications and low hospitalization costs, while surgical resection is recommended for solitary hepatocellular carcinoma tumors of 4-5 cm in diameter when long-term prognosis is considered.

A fast-track clinical pathway is designed to streamline patient care delivery and maximize cost effectiveness. It has decreased postoperative length of stay (LOS) and hospital charges for many surgical procedures. However, data on clinical pathways after liver surgery are sparse. This study examined whether use of a fast-track clinical pathway for patients undergoing elective liver resection affected postoperative LOS and hospital charges. A fast-track clinical pathway was developed and implemented by a multidisciplinary team for patients undergoing liver resection. Between July, 2007 and May, 2008, a total of 117 patients underwent elective liver resection: the fast-track clinical pathway (education of patients and families, earlier oral feeding, earlier discontinuation of intravenous fluid, no drains or nasogastric tubes, early ambulation, use of a urinary catheter for less than 24 h and planned discharge 6 days after surgery) was studied prospectively in 56 patients (postpathway group). These patients were compared with the remainder who had usual care (prepathway group). Outcome measures were postoperative LOS, perioperative hospital charges, intraoperative and postoperative complications, mortality, and readmission rate. Among all patients, 69 (59%) had complicating diseases and/or a history of surgery and 24 patients belonged to American Society of Anesthesiologists grade III-IV. Compared with the prepathway group, the postpathway group had a significantly shorter postoperative LOS (7 vs. 11 days, P < 0.01). The average perioperative hospital charges were RMB 26,626 for patients in the prepathway group and only RMB 21,004 for those in the postpathway group (P < 0.05), with no differences in intraoperative and postoperative complications (P = 0.814), mortality (P = 0.606), and readmission rate (P = 0.424). Implementation of the fast-track clinical pathway is an effective and safe method for reducing postoperative LOS and hospital charges for high-risk patients undergoing elective liver resection. The result supports the further development of fast-track clinical pathways for liver surgical procedures.

Purpose: Many physicians express a relatively nihilistic approach to the treatment of hepatocellular carcinoma (HCC) with portal vein tumor thrombus (PVTT). Consensus among surgeons regarding the indications for an aggressive approach has not been reached. Current study was aimed to determine whether an aggressive approach, with an extended resection with thrombectomy and adjuvant therapy, would lead to an improved survival for HCC patients with PVTT., Methods: A retrospective review of 116 HCC patients with PVTT admitted from 1996 to 2006 was conducted. Patients were divided into 2 time-period (TP) cohorts, of them, 51 cases in the first 5 years (TP1) and 65 in the last 5 years (TP2)., Results: Surgical operations were performed on 68 patients. Twenty-one surgical resections were performed in TP1 and forty-seven in TP2. The extent of liver resections, as well as the frequency of thrombectomy, was greater in TP2 (P = 0.039). During both time-periods, an aggressive therapy was associated with improved survival (P < 0.02 TP1, P < 0.001 TP2). Overall survival of all patients in TP2 was significantly greater than in TP1 (P < 0.001), with a median survival of 15 months in TP2, whereas in TP1, the survival was only 9 months. The median 1-, 3-year survivals in TP2 (54 and 34%, respectively) were also greater than that in TP1 (31 and 7%, respectively). A multiple logistic regression analysis revealed that radical resection and adjuvant therapy were the independent predictors of overall survival., Conclusions: An aggressive approach, combining extended liver resection with thrombectomy and adjuvant therapy, leads to an improved survival in the HCC patients with PVTT.

A hybridoma TB 21 clone was derived from fusion between Sp 2/0-Ag 14 myeloma cells and spleen cells of BALB/c mice immunized with transforming growth factor-beta 1 (TGF-beta 1) purified from human platelets. The TB 21 clone was identified to produce monoclonal antibody with IgG1 subclass and had sufficient titer for immunoreactivity to both human platelets-derived TGF-beta 1 and recombinant human TGF-beta 1 by enzyme-linked immunosorbent assay (ELISA). Western blotting studies demonstrated that two immunoreactive bands corresponding to 25 Kda and 12.5 Kda molecules were observed in the sample of acid/ethanol extracts from human platelets. The affinity constant (Kaff) was determined as 1.47 x 10(8) M-1 with non-competitive ELISA. Moreover, using bioassay for the effects of TGF-beta 1 on the growth of mink lung epithelial cells (CCL/64 cell line) and fibroblast cells (NRK 49 F cell line), TB 21 IgG was shown to be able to neutralize the action of TGF-beta 1 on the growth of these target cells. Therefore, this monoclonal antibody may be a useful probe for studying the growth modulatory activity of TGF-beta 1 in a variety of cells and tissues.

Following low-dose cyclophosphamide (CY) therapy (15 mg/kg) of mice bearing a large MOPC-315 tumor, the suppressive activity of their Sephadex G-10-adherent spleen cells (primarily macrophages) is overcome. Accordingly, when Sephadex G-10-adherent spleen cells from CY-treated tumor-bearing mice are added to the in vitro immunization culture of normal spleen cells, they do not suppress but actually bring about the generation of an augmented level of antitumor cytotoxicity. The ability to enhance the generation of antitumor cytotoxicity appears in the Sephadex G-10-adherent spleen cell population by Day 5 post-CY therapy of tumor-bearing mice and persists for at least 55 days; no such immunopotentiation is observed following administration of a low dose of CY to normal mice. In order for the immunopotentiating cells from CY-treated tumor-bearing mice to be effective in enhancing the generation of antitumor cytotoxicity, they must be added to the immunization culture of normal spleen cells no later than Day 3 (out of the 5 days) post-culture initiation. The CY-induced immunopotentiating activity resides in the T-cells, as is evident from the following observations. The immunopotentiating activity was abolished when the Sephadex G-10-adherent spleen cell population from CY-treated tumor-bearing mice was depleted of T-cells by anti-Thy 1.2 plus complement but not when this adherent spleen cell population was depleted of macrophages by carbonyl iron and magnet. Moreover, the immunopotentiating activity was also present in a population of CY-treated tumor-bearer spleen cells highly enriched for T-cells by passage through nylon wool columns. Thus, low-dose CY therapy overcomes the immunosuppressive activity of macrophages and induces the appearance of T-cell-mediated immunopotentiating activity, thereby leading to the development of an augmented level of antitumor cytotoxicity that can cooperate effectively with the tumoricidal activity of CY in the eradication of a late-stage, large s.c. tumor and extensive metastases.

We had shown previously that progression of MOPC-315 plasmacytoma growth is associated with an increase in the percentage of macrophages in the spleen as well as a decrease in the ability of tumor-bearer spleen cells to mount an antitumor cytotoxic response upon in vitro immunization. Here we provide evidence that macrophages in the MOPC-315 tumor-bearer spleen are responsible at least in part for the suppression of the generation of antitumor cytotoxicity. Accordingly, removal of most macrophages by depletion of phagocytic cells or Sephadex G-10-adherent cells from spleens of mice bearing a large tumor resulted in augmented antitumor immune potential. Also, Sephadex G-10-adherent spleen cells from tumor-bearing (but not normal) mice drastically suppressed the in vitro generation of antitumor cytotoxicity by normal spleen cells. The suppressive activity of these adherent cells did not reside in contaminating suppressor T cells, since it was not reduced by treatment with monoclonal anti-Thy 1.2 antibody plus complement. The Sephadex G-10-adherent cell population from the tumor-bearer spleen suppressed the in vitro generation of antitumor cytotoxicity against autochthonous tumor cells but not against allogeneic EL4 tumor cells, and hence the suppression was apparently specific. The suppressive activity of the Sephadex G-10-adherent cell population from tumor-bearer spleens was overcome by treatment of the tumor-bearing mice with a low curative dose of cyclophosphamide. This immunomodulatory effect of a low dose of the drug in overcoming the suppression mediated by the Sephadex G-10-adherent cell population enables the effector arm of the immune system of tumor-bearing mice to cooperate effectively with the drug's tumoricidal activity in tumor eradication.

We have shown previously (Ye, Q-W., and Mokyr, M. B. Cancer Res., 44: 3873-3879, 1984) that, following low-dose cyclophosphamide (CY) therapy (15 mg/kg) of mice bearing a large s.c. MOPC-315 tumor and extensive metastases, T-cell-dependent immunopotentiating activity appears in their hitherto immunosuppressive Sephadex G-10-adherent spleen cell population. Here we show that the CY-induced immunopotentiating T-cells express the Lyt 1, Lyt 2, and L3T4 phenotypes. The phenotype of the immunopotentiating T-cells was deduced from our observations that depletion of Lyt 1+, Lyt 2+, or L3T4+ cells from the Sephadex G-10-adherent spleen cell population of CY-treated tumor bearers abolished the ability of the adherent cells to enhance the generation of antitumor cytotoxicity when added to the in vitro immunization culture of normal spleen cells. Moreover, admixture of a Sephadex G-10-adherent cell population depleted of Lyt 2+ cells with a Sephadex G-10-adherent cell population depleted of L3T4+ cells failed to restore the immunopotentiating activity, indicating that T-cells that are apparently expressing simultaneously the Lyt 2 and L3T4 antigens are required for the exertion of the CY-induced immunopotentiating activity. The CY-induced immunopotentiating T-cells from MOPC-315 tumor bearers brought about the appearance of enhanced antitumor cytotoxicity not only against the MOPC-315 tumor cells, but also against two other syngeneic plasmacytomas, with surface immunoglobulin of a different class and antigenic specificity than the MOPC-315 tumor cells, as well as against a variant MOPC-315 tumor line which lacks surface immunoglobulin. The CY-induced immunopotentiating T-cells did not enhance the appearance of antitumor cytotoxicity against a syngeneic (WEHI 22.1) or an allogeneic (EL4) tumor of T-cell origin nor against the natural killer-sensitive YAC-1 cells. Thus, L3T4+, Lyt2+ T-cells from CY-treated MOPC-315 tumor bearers enhance the generation of antitumor cytotoxicity that is directed against plasmacytoma shared antigens other than immunoglobulins.