JAK3 targeted smart drugs and uses thereof

Abstract

Provided herein are inhibitor compounds or targeted smart drugs that are selective for a domain in a Janus kinase 3 enzyme, for example, a Janus kinase 3-Src homolog 2 domain. The targeted smart drugs are N-methylsulfonylmethanesulfonamide, N-(1-cyano-1-methyl- ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N-methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide. Also provided are methods for treating a lung cancer and / or a metastatic lung cancer and inhibiting lung cell proliferation using the inhibitor compounds or targeted smart drugs.

Description

[0001] JAK3 TARGETED SMART DRUGS AND USES THEREOF

[0002] Cross-Reference to Related

[0003] This international patent application claims benefit of priority under 35 U.S.C. §119(e) of provisional patent application U.S. Serial No. 63 / 646,285, filed May 13, 2024, the entirety of which is hereby incorporated in its entirety.

[0004] BACKGROUND OF THE INVENTION

[0005] Field of the Invention

[0006] The present invention relates generally to the fields of drug discovery and lung cancer treatment. More specifically, the present invention relates to inhibitor compounds, selective only for a Jak3 enzyme domain, as therapeutics to treat lung cancer. of the Related Art

[0007] According to the American Lung Association, lung cancer has the highest mortality rate among all cancers. The National Cancer Institute reports that 55% of lung cancers are metastatic with a five year relative survival of just 7%. In 2022, there were 236,740 new cases of lung cancer in the US and 130,180 lung cancer-associated deaths. The major problem is the current limitations in targeted therapeutics for lung cancer.

[0008] Current therapeutic challenges in lung cancer include whole-body radiation, surgery, and chemotherapy. These lead to challenges such as non-specificity with severe side effects and a high level of toxicity with multi-organ complications. As a result, patients respond poorly, have a low survival rate, and poor quality of life due to the aggressive nature of this type of cancer and the lack of targeted therapy.

[0009] The enzyme Janus kinase 3 (Jak3) is frequently overexpressed and is known to promote both cell proliferation and angiogenesis. Its overactivation in lung-cancer cells makes the cancer more aggressive and metastatic. The Jak family of kinases (JAKs) function as responders to cytokine or growth factor receptor activation. There are four members of the JAK family: JAK1 , JAK2, and tyrosine kinase 2 (Tyk2) that are ubiquitously expressed in vertebrates, including human, while the fourth, JAK3, is mostly limited to hematopoietic cells and mucosal epithelial cells.

[0010] In 2012, Tofacitinib (manufactured by Pfizer®), a Jak3 inhibitor, was approved by the FDA for the treatment of moderately to severely active rheumatoid arthritis. However, Tofacitinib shows cross reactivity by inhibiting other Jak members JAK1 and JAK2. Because other Jaks are ubiquitously expressed, such non-specific inhibition by Tofacitinib causes several severe side effects including anemia and neutropenia and has a warning label stating ’’Warning: Serious infections, mortality, malignancy, major adverse cardiovascular events (mace), and thrombosis”. This cross reactivity is likely because of the faulty screening technology used to discover this and all currently available Jak3 drugs. Firstly, a truncated kinase domain of Jak3 was used to screen the drug compounds while the human body has the full length of all Jak kinases. Secondly, an external synthetic substrate was used to monitor Jak3’s phosphorylation activity in the screening of the drug compounds while in the human body, Jak3 auto-phosphorylates itself first as the rate limiting step.

[0011] Thus, there is a need in the art for targeted therapeutics to treat lung cancer early, to decrease toxicity and to prevent advancement of a metastatic stage. Specifically, there is a need for therapeutic compounds that target only a specific domain in the Janus kinase 3 enzyme that is overexpressed in lung cancer and associated with lung cancer metastasis. The present invention fulfills this unmet need and desire in the art.

[0012] SUMMARY OF THE INVENTION

[0013] The present invention is directed to an inhibitor compound that selectively targets a domain in a Janus Kinase 3 enzyme (Jak3). Particularly, the inhibitor compound may be N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N- methyl-propane-1 -sulfonamide, or 5-methyl-2-prosylsulfonyl-benzamide or a combination thereof.

[0014] The present invention is further directed to a pharmaceutical composition comprising at least one of the compounds described herein and a pharmaceutically acceptable carrier, diluent or excipient.

[0015] The present invention is directed further to a method for treating a lung cancer in a patient in need thereof. In this method, a therapeutically effective amount of the pharmaceutical composition comprising at least one of the inhibitor compounds described herein is administered to inhibit a Jak3 enzyme activity associated with the lung cancer.

[0016] The present invention is directed further still to a method for inhibiting proliferation of lung cancer cells. In this method, the lung cancer cells are contacted with at least one of the inhibitor compounds described herein to inhibit at least one signaling in the lung cancer cells.

[0017] The present invention is directed further still to a targeted smart drug selective for a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain. The targeted smart drug is N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N- methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide or a combination thereof. The present invention is directed further still to a pharmaceutical composition comprising at least one of the targeted smart drugs described herein and a pharmaceutically acceptable carrier, diluent or excipient.

[0018] The present invention is directed further still to a method for treating a lung cancer in a patient in need thereof. In this method, proliferation of cancer cells associated with the lung cancer is inhibited via administration at least once of the pharmaceutical composition comprising the targeted smart drugs described herein.

[0019] The present invention is directed further still to a method for inhibiting an autophosphorylation activity by a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain in lung cancer cells or metastases therefrom. In the method the lung cancer cells are targeted with at least one targeted smart drug selective for the Jak3-SH2 domain as described herein.

[0020] BRIEF DESCRIPTION OF THE FIGURES

[0021] So that the matter in which the above-recited features, advantages and objects of the invention, as well as others that will become clear, are attained and can be understood in detail, more particular descriptions of the invention briefly summarized above may be had by reference to certain embodiments thereof that are illustrated in the appended drawings. These drawings form a part of the specification. It is to be noted, however, that the appended drawings illustrate preferred embodiments of the invention and therefore are not to be considered limiting in their scope.

[0022] FIGS. 1A-1 B show the ATP binding site in the kinase domain of Jak 3 (FIG. 1A) and the sequence alignment of Jak1 , Jak 2, Jak 3, and Tyk2 (FIG. 1 B).

[0023] FIGS. 2A-2D show Al-based smart druggable site screening for Jak3-targeted drugs where virtual compounds binding to human Jak 3 co-crystalized with a smart drug. FIG. 2A is a schematic diagram of a full length human Jak3 with location of the kinase domain shown by the broad arrow and of the SH2 domain shown by the narrow arrow. FIG. 2B illustrates a co-crystalized image and FIG. 2C illustrates the binding site defining the interactions between the inhibitor drug compound and the key amino acid residues in the kinase domain of Jak3 protein. FIG. 2D illustrates an Al-based virtual screening for the druggable sites in Jak3 through binding of virtual compounds.

[0024] FIGS. 3A-3B show a side-by-side target binding efficacy comparison between the currently marketed drug Tofacitinib (FIG. 3A) and the compound 1 targeted smart drug (FIG. 3B). FIGS. 3C-3D show a lung biopsy sample (FIG. 3C) and the inhibition of Jak3 (FIG. 3D) by the smart Jak3 inhibitor. FIGS. 3E-3F are immunofluorescence visualizations showing significant expression of Jak3 in LUAD patient biopsies (n=264) at 20X magnification (FIG. 3E) and a XZ visualization to show lateral expression (FIG. 3F). FIG. 4 shows that expression of JAK3 is higher in lung adenocarcinoma (LUAD) samples and those LUAD samples that have TP53 mutations compared to normal non-lung cancer tissues.

[0025] FIGS. 5A-5F show the expression of JAK3 in various lung adenocarcinoma patient types. JAK3 expression is higher in LUAD patients who have smoking habits compared to non-smokers (FIG. 5A), is higher in older LUAD patients (FIG. 5B), is slightly higher in LUAD female patients (FIG. 5C), is higher in LUAD patients compared to normal samples across all racial groups (FIG. 5D), is higher in LUAD patients at all the stages of lung cancer development compared to the surrounding normal tissues (FIG. 5E), and is higher in the primary tumors of LUAD patients compared to surrounding normal tissues (FIG. 5F).

[0026] FIG. 6 shows a comparative analysis for the expression of JAK3 across TCGA cancers (tumors and surrounding normal tissues) shows a higher Jak3 expressions in both lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) compared to surrounding normal tissues.

[0027] FIG. 7 shows that JAK3 expression increases with the presence and extent of nodal metastasis in LUAD patients.

[0028] FIG. 8 shows that higher Jak3 expression significantly decreases the 5-year survival in lung cancer patients. JAK3 expression increases with the presence and extent of nodal metastasis in LUAD patients.

[0029] DETAILED DESCRIPTION OF THE INVENTION

[0030] As used herein, the articles "a" and "an" when used in conjunction with the term “comprising” in the claims and / or the specification, may refer to “one”, but it is also consistent with the meaning of “one or more”, “at least one”, and “one or more than one”. Some embodiments of the invention may consist of or consist essentially of one or more elements, components, method steps, and / or methods of the invention. It is contemplated that any composition, component or method described herein can be implemented with respect to any other composition, component or method described herein.

[0031] As used herein, the term “or” in the claims refers to “and / or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or”.

[0032] As used herein "another" or “other” may mean at least a second or more of the same or different claim element or components thereof.

[0033] As used herein, the terms "comprise" and "comprising" are used in the inclusive, open sense, meaning that additional elements may be included. As used herein, the terms "consist of" and "consisting of" are used in the exclusive, closed sense, meaning that additional elements may not be included.

[0034] As used herein, the term “includes” or “including” refers to “including, but not limited to”. The terms “includes, “including” and “including, but not limited to” are used interchangeably.

[0035] As used herein, the term “about” refers to a numeric value, including, for example, whole numbers, fractions, and percentages, whether or not explicitly indicated. The term “about” generally refers to a range of numerical values (e.g., ± 5-10% of the recited value) that one of ordinary skill in the art would consider equivalent to the recited value (e.g., having the same function or result). In some instances, the term “about” may include numerical values that are rounded to the nearest significant figure.

[0036] As used herein, the term “targeted smart drug” refers to a therapeutic compound that targets only a specific domain in a Janus kinase 3 enzyme to selectively inhibit or interfere with at least one activity thereof, such as, but not limited to, autophosphorylation and / or overexpression in lung cancer or a metastasis thereof.

[0037] As used herein, the term “therapeutically effective amount” refers to a dosage sufficient to achieve a measurable improvement in the lung cancer and / or its metastatic cancers or progression, though not necessarily a cure.

[0038] As used herein, the term “pharmaceutically acceptable” refers to a composition that is safe and suitable for use in contact with cells and tissues in accordance with established medical practices.

[0039] As used herein, the term “treatment” refers to interventions aimed at alleviating, preventing, stabilizing, or resulting in remission of a cancer, for example, but not limited to, lung cancer or its metastatic cancers.

[0040] As used herein, the terms “targeted smart drug” and “inhibitor compound” are used interchangeably.

[0041] As used herein, the terms “inhibit” or “inhibitor" or “inhibitor compound" denotes a reduction in activity or a compound effective to enable a reduction in activity, for example, in JAK3 activity, such as, but not limited to, autophosphorylation activityorpreventing or reducing overexpression of the enzyme, which may range from partial to complete suppression. In certain embodiments, inhibition refers to a decrease of at least 10%, 20%, 50%, or even 100% compared to control levels.

[0042] As used herein, the term “contacting” refers to any suitable method of bringing a compound or a pharmaceutical composition into contact with a cell in vivo, in vitro or ex vivo. For in vivo applications, any known method of administration is suitable as described herein. As used herein, the term “subject" refers to a human that is the recipient of the inhibitor compounds described herein or is a control.

[0043] As used herein, the term “patient" refers to a subject who is undergoing clinical treatment for lung cancer and / or it metastases.

[0044] In one embodiment of the present invention, there is provided an inhibitor compound that selectively targets a domain in a Janus Kinase 3 enzyme (Jak3). In this embodiment the inhibitor compound may comprise N-methylsulfonylmethanesulfonamide, N-(1-cyano-1- methyl-ethyl)-2-methoxy-ethanesulfonamide, N-(2-cyanophenyl)methyl-N-ethyl- ethanesulfonamide, N-(2-cyanoethyl)-N-methyl-propane-1 -sulfonamide, or 5-methyl-2- propylsulfonyl-benzamide or a combination thereof. Particularly, the inhibitor compound is N- methylsulfonylmethanesulfonamide. In this embodiment, a preferred Jak3 domain may be a Jak3-Src homology 2 (Jak3-SH2) domain.

[0045] In another embodiment of the present invention, there is provided a pharmaceutical composition comprising at least one of the compounds as described supra and a pharmaceutically acceptable carrier, diluent or excipient.

[0046] In yet another embodiment of the present invention, there is provided a method for treating a lung cancer in a patient in need thereof, comprising administering at least once to the subject a therapeutically effective amount of the pharmaceutical composition as described supra to inhibit a Jak3 enzyme activity associated with the lung cancer.

[0047] In this embodiment, the administering step may comprise selectively targeting a single domain within the Jak3 enzyme. In this embodiment, a preferred domain is Jak3-SH2. In addition, a representative lung cancer includes but is not limited to a metastatic lung cancer. Particularly, the metastatic lung cancer may be a metastatic brain cancer or a metastatic nodal cancer. Furthermore the Jak3 enzyme activity may be autophosphorylation, overexpression or copy number increase or a combination thereof.

[0048] In yet another embodiment of the present invention, there is provided a method for method for inhibiting proliferation of lung cancer cells, comprising contacting the lung cancer cells with at least one of the inhibitor compounds as described supra effective to inhibit at least one signaling in the lung cancer cells.

[0049] In this embodiment, the signaling pathway may be a cell signaling proliferative pathway or an apoptotic signaling pathway or a combination thereof. Also, in this embodiment, the contacting step is in vitro or in vivo. In addition the lung cancer cells may be metastatic lung cancer cells. Particularly, the metastatic lung cancer cells may be metastatic brain cancer cells or metastatic nodal cancer cells.

[0050] In yet another embodiment of the present invention, there is provided a targeted smart drug selective for a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain that is N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N- methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide or a combination thereof. Particularly, the targeted smart drug may be N-methylsulfonylmethanesulfonamide.

[0051] In yet another embodiment of the present invention, there is provided a pharmaceutical composition comprising at least one of the targeted smart drugs as described supra and a pharmaceutically acceptable diluent or excipient.

[0052] In yet another embodiment of the present invention, there is provided a method for treating a lung cancer in a patient in need thereof, comprising inhibiting proliferation of cancer cells associated with the lung cancer via administration at least once of the pharmaceutical composition as described supra. In this embodiment the cancer cells may comprise a metastatic lung cancer. Particularly, the cancer cells may comprise a metastatic brain cancer or a metastatic nodal cancer.

[0053] In yet another embodiment of the present invention, there is provided a method for inhibiting an autophosphorylation activity by a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain in lung cancer cells or metastases therefrom, comprising contacting the lung cancer cells with at least one targeted smart drug selective for the Jak3-SH2 domain.

[0054] In this embodiment, the targeted smart drug may be N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N- methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide. Also in this embodiment, the contacting step may occur in vivo or in vitro. In addition, the metastases may be brain metastases or nodal metastases.

[0055] Provided herein are targeted smart drugs or inhibitor compounds designed and developed as Janus kinase 3 (Jak3) inhibitors that selectively bind to a Jak3 domain, for example, the Jak3-Src homology 2 (Jak3-SH2) domain. The targeted smart drugs and inhibit its autophosphorylation activity thereby decreasing the side effects caused by nonspecific binding of the drug.

[0056] The targeted smart drugs or inhibitor compounds described herein may be pharmaceutical compositions comprising one or more of the aforementioned inhibitors formulated with pharmaceutically acceptable diluents, carriers, excipients, salts, and / or adjuvants as are known in the art. Formulation and route of administration are well-known in the art and may depend on the stage of the lung cancer and therapeutic requirements.

[0057] Also provided are methods for treating a lung cancer and / or a metastatic lung cancer in a subject or patient in need of such treatment and for inhibiting the proliferation of lung cancer cells, either in vivo or in vitro, via administration of the inhibitor compounds or pharmaceutical compositions described herein or via contacting the lung cancer cells with these compounds. The lung cancer may be a lung adenocarcinoma or a lung squamous cell carcinoma. One of ordinary skill in the art is well-able to determine doses and treatment regimens for a specific subject or lung cancer patient depending at least on the age, sex, overall health thereof, progression of the lung cancer, for example, the presence of metastases, or remission thereof.

[0058] The following examples are given for the purpose of illustrating various embodiments of the invention and are not meant to limit the present invention in any fashion.

[0059] EXAMPLE 1

[0060] Targeted smart drug design and development

[0061] Artificial intelligence (Al)-based screening of druggable sites in Jak3 and virtual inhibitor compounds

[0062] The X-ray crystal structures for the kinase domain of JAK3 were used with the ATP binding regions (FIG. 1A) as the only druggable site, and potential inhibitors were first screened by the Al-based virtual screening system. Sequence alignment of the ATP binding site for the four JAK family members (SEQ ID NOS: 1-4; FIG. 1 B) illustrates that JAK3 has only two residues that could be utilized to design selective inhibitors for Jak3. In other Jak family members, JAK1 , JAK2, and Tyk2, these residues are serine and glycine, while in Jak3 they are cysteine and alanine.

[0063] The full length human Jak3 diagrammed in FIG. 2A shows the locations of the kinase domain and the SH2 (Src homolog 2) domain. Molecular modeling was conducted on the X- ray crystal structure (FIG. 2B) of the human JAK3, particularly, the SH2 domain, co-crystalized model with an inhibitor (PDB: 5LWN ). In FIGS. 2C-2D Al-based smart drug molecule bound within the kinase domain is highlighted in the center indicating the interactions with the noted amino acids of Jak3-kinase domain protein for the inhibitory functions.

[0064] The modeling site is defined by at least the following residues in the structure after removal of the co-crystallized inhibitor are the hydrophobic residues LEU-828, VAL-884 and LEU-905, the charged residues ASP-912 and ARG-953, the polar residue ASN-954, and a disulfide-capable residue CYS-909 that is possibly involved in binding or catalysis. Other residues defining the site are ALA-853, GLU-903, TYR-904, ARG-911 , LEU-956 and, ASP- 967. A molecular library of several million compounds using AtomNet technology was used for structure-based drug design and discovery. Top scoring compounds were clustered and subsequently filtered for favorable properties to arrive at a final subset of 79 compounds. Selection of top eight drug compounds with inhibition constant in the nanomolar range through direct inhibition of full-length Jak3 autophosphorylation

[0065] These 79 compounds were screened for direct binding to the full-length Jak3 and their kinetics of inhibitions of Jak3 autophosphorylation. Upon screening, the top eight compounds were selected that had an inhibition constant in the nanomolar range. Subsequently, these eight compounds that had the higher binding inhibition affinity with Jak3 protein were used for testing to find their potential for inhibition of lung-cancer cell proliferation and inhibition of metastasis using a lung cancer cell line. These potential kinase inhibitor drug compounds were vetted through artificial intelligence (Al)-based screening of druggable sites that are unique in Jak3. Virtual inhibitor compounds were subsequently discovered, synthesized, and validated.

[0066] The specific structures of these top eight compounds are presented in (Table 1 ). The inhibition constant as determined show that the Al-based Jak3 specific top eight Jak3 targeted potential drug compounds have an IC50 that ranges from 5.1 nM to 336 nM (Table 2). TABLE 1

[0067] Specific structures of Jak3-specific eight compounds

[0068] TABLE 2

[0069] Efficacy (IC50) of Al-vetted Jak3-specific top eight drug compounds Exploiting the non-Kinase-domain architecture of JAK3 for lung cancer-targeted anticancer agent

[0070] Though Jak3 is such an important target for different types of immunological diseases indication and several types of cancer, to date there is no FDA approved drug that selectively targets Jak3 and there are no non-kinase-domain targeted allosteric inhibitors of Jak3. Using a side-by-side target binding efficacy comparison, while Tofacitinib drug has IC50 of 128nM, (over 125-fold higher than reported by Pfizer), one of the Jak3-SH2 targeted potential drug compounds had an IC50 1.6 nM (FIGS. 3A-3D).

[0071] The specific structures of the top five JAk3-SH2 domain targeted allosteric inhibitor compounds are presented in (Table 3). The inhibition constant as determined show that the discovered SH2-domain based Jak3 specific top five Jak3 targeted potential drug compounds have an IC50 that ranged from 1 .6 nM to 200 nM (Table 4).

[0072] TABLE 3

[0073] Specific structures of Jak3-SH2 domain specific five drug compounds

[0074] TABLE 4 Efficacy (IC50) of SH2-domain targeted allosteric Jak3-inhibitor drug compounds

[0075] These drug compounds targeted to Jak3 are examined in inhibiting the metastatic potential of lung cancer cells in patient derived xenograft mouse model in vivo.

[0076] Jak3 overexpresses metastatic lung cancer patients and the metastatic lung cancer cell line FIGS. 3A-3D show the overexpression of Jak3 in biopsies of human lung cancer patients. Immunofluorescence illustrates this significant Jak3 expression in representative images taken from 264 patient biopsies in FIGS. 3E-3F. The drug compounds of the present invention developed through Artificial Intelligence (Al) technology targeting the ATP binding domain of JAK3 are highly selective and limit lung cancer brain metastasis by promoting apoptosis while decreasing the migration and invasion of the lung cancer cell. EXAMPLE 2

[0077] In vitro testing

[0078] Cytotoxicity assay

[0079] The eight compounds shown in Table 1 are dissolved in dimethyl sulfoxide (DMSO) and lung cancer-metastatic and non-metastatic cells are treated with selected compounds at; 1 , 3, 6, 12, 25, 50, and 100 pg / ml and the cell growth are quantified by MTT assay. The IC50 is determined based on dose-response curves of individual compounds and compared.

[0080] Cellular morphology, markers for apoptosis and Jak3-, STAT3, STATS-phosphorylation

[0081] After treatment with inhibitor, the impact of the inhibitors on the signaling pathways for lung cancer cell proliferation and apoptosis, is through western blot analysis for STAT3 / 5- phospphorylation, Jak3-phosphorylaiton and migration through scratch wound healing migration assay. For determining the blood brain permeability of the compounds, the trans well assay is performed. Phase contrast microscopy, confocal microscopy, and histological and immune histochemical analyses are used for confirmation.

[0082] EXAMPLE 3

[0083] Jak3 expression in primary lung cancer and metastatic lung cancer

[0084] Mutated genes in LUAD patients

[0085] To determine what types of LUAD patients will benefit the most from a JAk3 targeted drug, the top 10 frequently mutated genes in a samples of 6,418 LUAD patients was first determined. Table 5A shows that TP53, EGFR, and KRAS were the top 3 genes frequently mutated in LUAD. Then it was determined if any of these top three co-occurred of Jak3. Table 5B shows the cooccurrence of JAk3 with TP53, EGFR, and KRAS.

[0086] TABLE 5A

[0087] Top 10 frequently mutated genes in LUAD patients TABLE 5B Co-occurrence of Jak3 with top 10 frequently mutated genes in LUAD patients

[0088] A positive Log 2 odds ratio value here suggests that alterations in these genes cooccur with JAk3 in the same LUAD biopsied samples, while a negative value suggests that alterations in these genes are mutually exclusive and occur in different samples. These data suggest while Jak3 and EGFR were mutually exclusive indicating that LUAD patients with EGFR mutation likely would not benefit from the Jak3 targeted smart drugs. LUAD patients with the TP53 mutation co-occur with Jak3 indicating that LUAD patients with TP53 mutations may benefit the most with the Jak3 targeted smart drug. Together, Tables 5A and 5B enable stratification of the LUAD patients who would benefit the most from the Jak3 targeted smart drugs. FIG. 4 illustrates that the expression of JAK3 is higher in LUAD samples, particularly in those with TP53 mutations, compared to normal lung tissue.

[0089] Jak3 expression among patient types

[0090] In FIGS. 5A-5F Jak3 expression is demonstrated to be higher in various types of lung adenocarcinoma patients selected from the TCGA (The Cancer Genome Atlas) project. While FIG. 5A illustrates that LUAD patients with a smoking habit have a higher Jak3 expression compared to normal tissue, Jak3 expression is higher regardless of smoking status. There is no significant difference in JAK3 expression between current smokers, non-smokers, and reformed smokers, indicating that smoking habits may not significantly impact JAK3 expression in LUAD. This indicates that the LUAD patients with a smoking history would benefit the most from treatment with the Jak3 targeted smart drugs.

[0091] FIG. 5B demonstrates that the expression of JAK3 in LUAD is slightly higher in individuals between the age of 61-80 years compared to individuals between the ages 41-60 and 81 -100 years. This indicates that LUAD patients with a higher age would benefit the most from treatment with Jak3 targeted smart drugs. FIG. 5C shows that both male and female LUAD patients have higher JAK3 expression levels compared to normal samples. Female LUAD patients show a slightly higher median JAK3 expression compared to male LUAD patients. These indicate that the female LUAD patients may respond better to treatment with Jak3 targeted smart drugs compared to male counterparts.

[0092] FIG. 5D illustrates that JAK3 expression is higher in LUAD patients compared to normal samples across all racial groups. Among LUAD patients, African-Americans show a slightly higher median JAK3 expression compared to Caucasians and Asians. As a caution, the small sample size of the Asian group should be taken into consideration when interpreting these results. This indicates that African American and Caucasian LUAD patients may respond better to treatment with Jak3 targeted smart drugs compared other ethnic groups.

[0093] FIG. 5E illustrates that JAK3 expression is higher in LUAD patients at all the stages of lung cancer development, however, stage-1 showed relatively higher Jak3 expression compared to other stages of LUAD. Thus, LUAD patients with any stage of cancer may respond to to treatment with Jak3 targeted smart drugs without impact to the surrounding healthy tissues. The stage 1 LUAD patients may respond better than during other stages. In FIG. 5F, JAK3 expression is shown to be higher in the primary tumors of LUAD patients, thereby indicating that the Jak3 targeted smart drug may help in remission of primary tumors in LUAD patients.

[0094] Comparative JAK3 expression in TCGA project cancers

[0095] In FIG. 6, a comparative analysis of JAK3 expression in TCGA cancers, both tumors and surrounding tissues, demonstrates that JAK3 expression is higher in primary tumors in LUAD patients. This indicates that the Jak3 targeted smart drug may have a therapeutic effect on lung small cell carcinoma (LUSC) as well as lung adenocarcinoma.

[0096] JAK3 expression in metastases

[0097] In FIG. 7, JAK3 expression is shown to increase with the presence and extent of nodal metastasis in LUAD patients, suggesting that higher JAK3 expression may be associated with more advanced disease stages involving lymph node metastasis. These indicate that the Jak3 targeted smart drug may help in remission of nodal metastatic tumors in LUAD patients.

[0098] Effect of JAK3 expression on 5-vear survival

[0099] FIG. 8 shows the first progression survival, i.e., the length of time during and after lung cancer treatment that a patient lives with the disease without it worsening. In a clinical trial, measuring the first progression-free survival is one way to see how well a new treatment works. The curves for high JAK3 expression groups decline over time, indicating that the survival probability decreases as time progresses. The p-value is 1.4xe'06In survival analysis, a p-value less than 0.05 is generally considered statistically significant. This p-value indicates that there is a statistically significant difference in survival between patients with high JAK3 expression and those with low JAK3 expression. It appears that the group with high JAK3 expression, as labeled, has a significantly worse survival rate, particularly in the long-term follow-up period. The curve labeled high expression is below the low expression curve in the later stages, indicating a potentially lower survival probability at those time points. This indicates that treatment with the Jak3 targeted smart drug may increase the 5-year survival in both LUAD and LUSC patients.

[0100] EXAMPLE 4

[0101] In vivo testing

[0102] Patient selection

[0103] Prior IRB approval is obtained before the study. Patients with newly diagnosed or recurrent lung cancer brain metastases, undergoing surgical resection, are consented to participate. Only those patients with Jak3 overexpression in the cancer biopsies are selected. Immunodeficient NOD SCID gamma (NSG) (Envigo, Indianapolis) and Hsd:athymic Nude- Foxnl nu (Envigo, Indianapolis) mice are used for patient derived xenograft (PDX) development.

[0104] Establishment of xenografts

[0105] Patient tissue collected during surgery are added to a tube with media (DMEM+10%FBS+1 %PS) and minced with sharp, sterile scissors into a slurry. Matrigel is added to the media in a 1 :1 ratio and injected subcutaneously into NSG mice with an 18- gauge needle. Subsequent passages are made in a similar fashion into either NSG or athymic nude mice. Cryopreservation of xenografts is accomplished by mixing minced tumor pieces with complete transport media supplemented with 10% DMSO. Tumors are frozen in controlled rate freezers to -80°C overnight and transferred to liquid nitrogen for long-term storage. To thaw tumors, aliquots are warmed to 37°C in a heated water bath and tumor tissue are washed twice in transport complete media (without DMSO) and immediately implanted into mice.

[0106] Establishment of cell lines

[0107] Tumors taken from early PDX passages are dissociated into single cell suspensions and seeded in T-25 flasks in media (DMEM+10%FBS+1 %PS+2.5 ug / mL amphotericin B) and incubated at 37°C in a humidified atmosphere of 5% CO2. Media is changed regularly, and cells are passaged once they reached 70% confluence. Mycoplasma testing and short tandem repeat profiling is performed during passaging.

[0108] Xenograft and drug Injection

[0109] Control LC or Brain mLC is injected (175,000 cells / injection, once) subcutaneously of 8-12-week-old control NSG mice or mice administered @1 mg / kg of 1 mg / mL of inhibitor compounds dissolved in 10% sucrose solution. The injection of the testing compound inhibitor is performed at (I) from two days before cell injection, or (II) once every 7 days after intraperitoneal injection up to 19 days when applicable.

[0110] In vivo toxicity study of Jak3-inhibitor directed drug compound

[0111] Three compounds are tested for the in vivo toxicity. The compounds are dissolved in pharmaceutical grade sucrose buffer and are injected intraperitoneally in the 8-10-week-old female mice at increasing concentration (0.025-1.0 mg / Kg) in a 100 ul volume (once every 7 days for 3 weeks) and loss of body weight (weighed everyday), signs of fatigue (grooming, reaching for food, and water. Compared to non-treated mice), and loss of appetite (food intake by weighing the food everyday) is observed. At any loss of more than 10% bodyweight, the experiment is stopped, and the mouse is euthanized. Metastatic burden is quantified in postsacrificed brain sections of the mice at day 0, 7, 14, and 19 following intracardiac injection.

[0112] To assess the proliferative state of BMs, mice are injected intraperitoneally @ 100 mg / kg with 10 mg / mL Brdllrd solution, 2 hours before euthanasia. Following euthanasia, primary and brain metastatic lesions are excised and used for analysis. Western analysis to determine the expression and phosphorylation status of Jak3 and its downstream signaling partner STAT3, and STAT5 is done in brain tissue lysates. H&E staining of brain section, microscopic observation for tumor in brain and the breast is done in control and inhibitor compound treated humanized mice.

Claims

1. WHAT IS CLAIMED IS:

1. An inhibitor compound that selectively targets a domain in a Janus Kinase 3 enzyme (Jak3).

2. The inhibitor compound of claim 1 , comprising N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy- ethanesulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)- N-methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide or a combination thereof.

3. The inhibitor compound of claim 2 that is N-methylsulfonylmethanesulfonamide.

4. The inhibitor compound of claim 1 , wherein the Jak3 domain is a Jak3-Src homology 2 (Jak3-SH2) domain.

5. A pharmaceutical composition comprising at least one of the compounds of claim 1 and a pharmaceutically acceptable carrier, diluent or excipient.

6. A method for treating a lung cancer in a patient in need thereof, comprising: administering at least once to the subject a therapeutically effective amount of the pharmaceutical composition of claim 5 to inhibit a Jak3 enzyme activity associated with the lung cancer.

7. The method of claim 6, said administering step comprising selectively targeting a single domain within the Jak3 enzyme.

8. The method of claim 7, wherein the domain is Jak3-SH2.

9. The method of claim 6, wherein the lung cancer is a metastatic lung cancer.

10. The method of claim 9, wherein the metastatic lung cancer is a metastatic brain cancer or a metastatic nodal cancer.11 . The method of claim 6, wherein the Jak3 enzyme activity is autophosphorylation, overexpression or copy number increase or a combination thereof.

12. A method for inhibiting proliferation of lung cancer cells, comprising: contacting the lung cancer cells with at least one of the inhibitor compounds of claim 1 effective to inhibit at least one signaling in the lung cancer cells.

13. The method of claim 12, wherein the signaling pathway is a cell signaling proliferative pathway or an apoptotic signaling pathway or a combination thereof.

14. The method of claim 12, wherein the contacting step is in vitro or in vivo.

15. The method of claim 12, wherein the lung cancer cells are metastatic lung cancer cells.

16. The method of claim 15, wherein the metastatic lung cancer cells are metastatic brain cancer cells or metastatic nodal cancer cells.

17. A targeted smart drug selective for a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain that is N-methylsulfonylmethanesulfonamide, N-(1-cyano-1-methyl-ethyl)-2-methoxy- ethanesulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)- N-methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide or a combination thereof.

18. The targeted smart drug of claim 17 that is N-methylsulfonyl methanesulfonamide.

19. A pharmaceutical composition comprising at least one of the targeted smart drugs of claim 17 and a pharmaceutically acceptable diluent or excipient.

20. A method for treating a lung cancer in a patient in need thereof, comprising: inhibiting proliferation of cancer cells associated with the lung cancer via administration at least once of the pharmaceutical composition of claim 19.

21. The method of claim 20, wherein the cancer cells comprise a metastatic lung cancer.

22. The method of claim 21 , wherein the metastases are brain metastases or nodal metastases.

23. A method for inhibiting an autophosphorylation activity by a Janus kinase 3-Src homolog 2 (Jak3-SH2) domain in lung cancer cells or metastases therefrom, comprising: contacting the lung cancer cells or metastatic lung cancer cells with at least one targeted smart drug selective for the Jak3-SH2 domain.

24. The method of claim 23, wherein the targeted smart drug is N- methylsulfonylmethanesulfonamide, N-(1 -cyano-1 -methyl-ethyl)-2-methoxy-ethane sulfonamide, N-(2-cyanophenyl)methyl-N-ethyl-ethanesulfonamide, N-(2-cyanoethyl)-N- methyl-propane-1 -sulfonamide, or 5-methyl-2-propylsulfonyl-benzamide.

25. The method of claim 23, wherein the contacting step occurs in vivo or in vitro.

26. The method of claim 23, wherein the metastases are brain metastases or lymph node metastases.

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